Molecular Mechanisms of Increased Heart Rate in Shenxianshengmai-treated Bradycardia Rabbits

PubMed Central

Liu, Zhou-Ying; Huang, Jian; Liu, Na-Na; Zheng, Min; Zhao, Tao; Zhao, Bu-Chang; Wang, Yi-Min; Pu, Jie-Lin

2017-01-01

Background: The molecular mechanisms of Shenxianshengmai (SXSM), a traditional Chinese medicine, on bradycardia have been incompletely understood. The study tried to investigate the gene expression profile and proteomics of bradycardia rabbits’ hearts after SXSM treatment. Methods: Twenty-four adult rabbits were randomly assigned in four groups: sham, model, model plus SXSM treatment, and sham plus SXSM treatment groups. Heart rate was recorded in all rabbits. Then, total RNA of atria and proteins of ventricle were isolated and quantified, respectively. Gene expression profiling was conducted by gene expression chip, and quantitative real-time reverse transcription-polymerase chain reaction (RT-PCR) was performed to confirm the results of gene expression chip. We used isobaric tags for elative and absolute quantitation and Western blotting to identify altered proteins after SXSM treatment. Results: There was a constant decrease in the mean heart rate (32%, from 238 ± 6 beats/min to 149 ± 12 beats/min) after six weeks in model compared with that in sham group. This effect was partially reversed by 4-week SXSM treatment. Complementary DNA microarray demonstrated that the increased acetylcholinesterase and reduced nicotinic receptor were take responsibility for the increased heart rate. In addition, proteins involved in calcium handling and signaling were affected by SXSM treatment. Real-time RT-PCR verified the results from gene chip. Results from proteomics demonstrated that SXSM enhanced oxidative phosphorylation and tricarboxylic acid (TCA) cycle in ventricular myocardium to improve ATP generation. Conclusions: Long-term SXSM stimulates sympathetic transmission by increasing the expression of acetylcholinesterase and reduces the expression of nicotinic receptor to increase heart rate. SXSM also restored the calcium handling genes and altered genes involved in signaling. In addition, SXSM improves the ATP supply of ventricular myocardium by increasing proteins involved in TCA cycle and oxidation-respiratory chain. PMID:28091410

[Effect of Electroacupuncture on Expression of Apelin-APJ System of Cerebral Vascular Endothelial Cell in Rats with Cerebral Infarction].

PubMed

Yang, Li-Hong; Du, Yuan-Hao; Li, Jing

2017-02-25

To observe the regulation of APJ and its ligand Apelin on the angiogenesis pathway after cerebral infarction and the intervention effect of acupuncture. Wistar rats were randomly divided into model group( n =90), electroacupuncture(EA) group( n =90), sham operation group( n =90) and control group( n =10). The first three groups were further divided into 1,3,6,9,12,24 h and 3,7, 12 d subgroups( n =10 in each subgroup). The cerebral infarction model was established by middle cerebral artery occlusion (MCAO). EA(15 Hz, 2 mA) was applied to "Shuigou" (GV 26) for 20 min in the EA group. The 1, 3, 6, 9, 12, 24 h subgroups were treated immediately after modeling, the 3, 7, 9 d subgroups were treated once daily for 3, 7 or 9 days. Real-time fluorescent quantitative (RT-PCR) and Western blot were applied to detect the changes of Apelin and APJ in cerebrovascular endothelial cells, respectively. Compared with the control group, the expression of Apelin-APJ mRNA was decreased in the model group(12 h, 12 d, P <0.05, P <0.01); After EA, the Apelin mRNA expression was increased in the 12 h and 7 d subgroups ( P <0.01), while the APJ mRNA expression was increased in the 6, 9, 12 h subgroups( P <0.05, P <0.01). Compared with the control group, the Apelin(1, 3, 6, 24 h and 3, 7, 12 d) and APJ(1, 3, 6, 9 h and 3 d) protein expressions were decreased in the model group( P <0.01, P <0.05); After EA, the Apelin protein expression was increased in the 6, 24 h and 3, 7, 12 d subgroups ( P <0.05, P <0.01), while the APJ protein expression was increased in the 1, 9, 12, 24 h and 3, 7, 12 d subgroups ( P <0.05, P <0.01). EA can up-regulate the expression of Apelin-APJ mRNA and protein of cerebral vascular endothelial cell in MCAO rats which has an important role in the establishment of blood vessel regeneration and collateral circulation.

Gene expression during blow fly development: improving the precision of age estimates in forensic entomology.

PubMed

Tarone, Aaron M; Foran, David R

2011-01-01

Forensic entomologists use size and developmental stage to estimate blow fly age, and from those, a postmortem interval. Since such estimates are generally accurate but often lack precision, particularly in the older developmental stages, alternative aging methods would be advantageous. Presented here is a means of incorporating developmentally regulated gene expression levels into traditional stage and size data, with a goal of more precisely estimating developmental age of immature Lucilia sericata. Generalized additive models of development showed improved statistical support compared to models that did not include gene expression data, resulting in an increase in estimate precision, especially for postfeeding third instars and pupae. The models were then used to make blind estimates of development for 86 immature L. sericata raised on rat carcasses. Overall, inclusion of gene expression data resulted in increased precision in aging blow flies. © 2010 American Academy of Forensic Sciences.

Factors Expressed in an Animal Model of Anteroinferior Glenohumeral Instability

PubMed Central

Mulcahey, Mary K.; Marshall, Mindy; Gallacher, Stacey E.; Kaback, Lee A.; Blaine, Theodore A.

2015-01-01

Background: There is little information on the molecular factors important in healing and changes that occur in the glenoid labrum in response to injury. Using a novel animal model of acute anterior shoulder dislocation, this study characterizes the factors expressed in the glenoid labrum in response to injury and correlates their expression to glenohumeral stability. Purpose: To study the response of the glenoid labrum to injury both biomechanically and with immunohistochemical testing. Methods: An injury to the anteroinferior labrum was surgically induced in 50 male Lewis rats. Rats were sacrificed at 3, 7, 14, 28, or 42 days. Immunolocalization experiments were performed to localize the expression of growth factors and cytokines. For biomechanical testing, dynamic stiffness for anterior and posterior laxity, load to failure, stiffness, and maximum load were recorded. Statistical differences were determined at P < .05. Study Design: Descriptive laboratory study. Results: Expression of interleukin–1 beta (IL-1β), transforming growth factor–beta 1 (TGF-β1), matrix metalloproteinase 3 (MMP3), and matrix metalloproteinase 13 (MMP13) were increased in injured compared with uninjured specimens. Collagen III expression was increased early and decreased with time. Biomechanical testing verified instability by demonstrating increased anterior displacement and decreased stiffness in injured shoulders at all time points. Conclusion: This novel animal model of acute anterior shoulder dislocation showed increased expression of IL-1β, TGF-β1, MMP3, MMP13, and collagen III in the injured labral tissue at early time points. Increased anterior laxity and decreased stiffness and maximum load to failure were seen after anterior labral injury, supporting the model’s ability to re-create anterior glenohumeral instability. These data provide important information on the temporal changes occurring in a rat model of anterior glenohumeral dislocation. Clinical Relevance: Identification of factors expressed in the anterior capsule and glenoid labrum in response to injury may lead to the development of novel agents that can be used to augment glenoid labrum healing and ultimately improve both surgical and nonsurgical treatment of this common shoulder injury. PMID:26535392

Nuclear receptor TLX stimulates hippocampal neurogenesis and enhances learning and memory in a transgenic mouse model

PubMed Central

Murai, Kiyohito; Qu, Qiuhao; Sun, GuoQiang; Ye, Peng; Li, Wendong; Asuelime, Grace; Sun, Emily; Tsai, Guochuan E.; Shi, Yanhong

2014-01-01

The role of the nuclear receptor TLX in hippocampal neurogenesis and cognition has just begun to be explored. In this study, we generated a transgenic mouse model that expresses TLX under the control of the promoter of nestin, a neural precursor marker. Transgenic TLX expression led to mice with enlarged brains with an elongated hippocampal dentate gyrus and increased numbers of newborn neurons. Specific expression of TLX in adult hippocampal dentate gyrus via lentiviral transduction increased the numbers of BrdU+ cells and BrdU+NeuN+ neurons. Furthermore, the neural precursor-specific expression of the TLX transgene substantially rescued the neurogenic defects of TLX-null mice. Consistent with increased neurogenesis in the hippocampus, the TLX transgenic mice exhibited enhanced cognition with increased learning and memory. These results suggest a strong association between hippocampal neurogenesis and cognition, as well as significant contributions of TLX to hippocampal neurogenesis, learning, and memory. PMID:24927526

Increased activation of NADPH oxidase 4 in the pulmonary vasculature in experimental diaphragmatic hernia.

PubMed

Gosemann, Jan-H; Friedmacher, Florian; Hunziker, Manuela; Alvarez, Luis; Corcionivoschi, Nicolae; Puri, Prem

2013-01-01

Persistent pulmonary hypertension remains a major cause of mortality and morbidity in congenital diaphragmatic hernia (CDH). NADPH oxidases (Nox) are the main source of superoxide production in vasculature. Nox4 is highly expressed in the smooth muscle and endothelial cells of the vascular wall and increased activity has been reported in the pulmonary vasculature of both experimental and human pulmonary hypertension. Peroxisome proliferator-activated receptor (PPARγ) is a key regulator of Nox4 expression. Targeted depletion of PPARγ results in pulmonary hypertension phenotype whereas activation of PPARγ attenuates pulmonary hypertension and reduces Nox4 production. The nitrofen-induced CDH model is an established model to study the pathogenesis of pulmonary hypertension in CDH. It has been previously reported that PPARγ-signaling is disrupted during late gestation and H(2)O(2) production is increased in nitrofen-induced CDH. We designed this study to investigate the hypothesis that Nox4 expression and activation is increased and vascular PPARγ is decreased in nitrofen-induced CDH. Pregnant rats were treated with either nitrofen or vehicle on gestational day 9 (D9). Fetuses were sacrificed on D21 and divided into control and CDH. RT-PCR, western blotting and confocal-immunofluorescence-double-staining were performed to determine pulmonary expression levels of PPARγ, Nox4 and Nox4-activation (p22(phox)). There was a marked increase in medial and adventitial thickness in pulmonary arteries of all sizes in CDH compared to controls. Pulmonary Nox4 levels were significantly increased whereas PPARγ levels were decreased in nitrofen-induced CDH compared to controls. Western blotting revealed increased pulmonary protein expression of the Nox4-activating subunit p22(phox) and decreased protein expression of PPARγ in CDH compared to controls. Confocal-microscopy confirmed markedly increased pulmonary expression of the Nox4 activating subunit p22(phox) accompanied by decreased perivascular PPARγ expression in lungs of nitrofen-exposed fetuses compared to controls. To our knowledge, the present study is the first to report increased Nox4 production in the pulmonary vasculature of nitrofen-induced CDH. Down-regulation of the PPARγ-signaling pathway may lead to increased superoxide production, resulting in pulmonary vascular dysfunction and contributing to pulmonary hypertension in the nitrofen-induced CDH model. PPARγ-activation inhibiting Nox4 production may therefore represent a potential therapeutic approach for the treatment of pulmonary hypertension in CDH.

Proteomic identification of altered cerebral proteins in the complex regional pain syndrome animal model.

PubMed

Nahm, Francis Sahngun; Park, Zee-Yong; Nahm, Sang-Soep; Kim, Yong Chul; Lee, Pyung Bok

2014-01-01

Complex regional pain syndrome (CRPS) is a rare but debilitating pain disorder. Although the exact pathophysiology of CRPS is not fully understood, central and peripheral mechanisms might be involved in the development of this disorder. To reveal the central mechanism of CRPS, we conducted a proteomic analysis of rat cerebrum using the chronic postischemia pain (CPIP) model, a novel experimental model of CRPS. After generating the CPIP animal model, we performed a proteomic analysis of the rat cerebrum using a multidimensional protein identification technology, and screened the proteins differentially expressed between the CPIP and control groups. Results. A total of 155 proteins were differentially expressed between the CPIP and control groups: 125 increased and 30 decreased; expressions of proteins related to cell signaling, synaptic plasticity, regulation of cell proliferation, and cytoskeletal formation were increased in the CPIP group. However, proenkephalin A, cereblon, and neuroserpin were decreased in CPIP group. Altered expression of cerebral proteins in the CPIP model indicates cerebral involvement in the pathogenesis of CRPS. Further study is required to elucidate the roles of these proteins in the development and maintenance of CRPS.

Alpha-fetoprotein-targeted reporter gene expression imaging in hepatocellular carcinoma.

PubMed

Kim, Kwang Il; Chung, Hye Kyung; Park, Ju Hui; Lee, Yong Jin; Kang, Joo Hyun

2016-07-21

Hepatocellular carcinoma (HCC) is one of the most common cancers in Eastern Asia, and its incidence is increasing globally. Numerous experimental models have been developed to better our understanding of the pathogenic mechanism of HCC and to evaluate novel therapeutic approaches. Molecular imaging is a convenient and up-to-date biomedical tool that enables the visualization, characterization and quantification of biologic processes in a living subject. Molecular imaging based on reporter gene expression, in particular, can elucidate tumor-specific events or processes by acquiring images of a reporter gene's expression driven by tumor-specific enhancers/promoters. In this review, we discuss the advantages and disadvantages of various experimental HCC mouse models and we present in vivo images of tumor-specific reporter gene expression driven by an alpha-fetoprotein (AFP) enhancer/promoter system in a mouse model of HCC. The current mouse models of HCC development are established by xenograft, carcinogen induction and genetic engineering, representing the spectrum of tumor-inducing factors and tumor locations. The imaging analysis approach of reporter genes driven by AFP enhancer/promoter is presented for these different HCC mouse models. Such molecular imaging can provide longitudinal information about carcinogenesis and tumor progression. We expect that clinical application of AFP-targeted reporter gene expression imaging systems will be useful for the detection of AFP-expressing HCC tumors and screening of increased/decreased AFP levels due to disease or drug treatment.

Alpha-fetoprotein-targeted reporter gene expression imaging in hepatocellular carcinoma

PubMed Central

Kim, Kwang Il; Chung, Hye Kyung; Park, Ju Hui; Lee, Yong Jin; Kang, Joo Hyun

2016-01-01

Hepatocellular carcinoma (HCC) is one of the most common cancers in Eastern Asia, and its incidence is increasing globally. Numerous experimental models have been developed to better our understanding of the pathogenic mechanism of HCC and to evaluate novel therapeutic approaches. Molecular imaging is a convenient and up-to-date biomedical tool that enables the visualization, characterization and quantification of biologic processes in a living subject. Molecular imaging based on reporter gene expression, in particular, can elucidate tumor-specific events or processes by acquiring images of a reporter gene’s expression driven by tumor-specific enhancers/promoters. In this review, we discuss the advantages and disadvantages of various experimental HCC mouse models and we present in vivo images of tumor-specific reporter gene expression driven by an alpha-fetoprotein (AFP) enhancer/promoter system in a mouse model of HCC. The current mouse models of HCC development are established by xenograft, carcinogen induction and genetic engineering, representing the spectrum of tumor-inducing factors and tumor locations. The imaging analysis approach of reporter genes driven by AFP enhancer/promoter is presented for these different HCC mouse models. Such molecular imaging can provide longitudinal information about carcinogenesis and tumor progression. We expect that clinical application of AFP-targeted reporter gene expression imaging systems will be useful for the detection of AFP-expressing HCC tumors and screening of increased/decreased AFP levels due to disease or drug treatment. PMID:27468205

Design of optimal hyperthermia protocols for prostate cancer by controlling HSP expression through computer modeling (Invited Paper)

NASA Astrophysics Data System (ADS)

Rylander, Marissa N.; Feng, Yusheng; Diller, Kenneth; Bass, J.

2005-04-01

Heat shock proteins (HSP) are critical components of a complex defense mechanism essential for preserving cell survival under adverse environmental conditions. It is inevitable that hyperthermia will enhance tumor tissue viability, due to HSP expression in regions where temperatures are insufficient to coagulate proteins, and would likely increase the probability of cancer recurrence. Although hyperthermia therapy is commonly used in conjunction with radiotherapy, chemotherapy, and gene therapy to increase therapeutic effectiveness, the efficacy of these therapies can be substantially hindered due to HSP expression when hyperthermia is applied prior to these procedures. Therefore, in planning hyperthermia protocols, prediction of the HSP response of the tumor must be incorporated into the treatment plan to optimize the thermal dose delivery and permit prediction of overall tissue response. In this paper, we present a highly accurate, adaptive, finite element tumor model capable of predicting the HSP expression distribution and tissue damage region based on measured cellular data when hyperthermia protocols are specified. Cubic spline representations of HSP27 and HSP70, and Arrhenius damage models were integrated into the finite element model to enable prediction of the HSP expression and damage distribution in the tissue following laser heating. Application of the model can enable optimized treatment planning by controlling of the tissue response to therapy based on accurate prediction of the HSP expression and cell damage distribution.

Uric acid upregulates the adiponectin-adiponectin receptor 1 pathway in renal proximal tubule epithelial cells

PubMed Central

Yang, Qingmei; Fu, Chensheng; Xiao, Jing; Ye, Zhibin

2018-01-01

Adiponectin (APN) is a protein hormone that is primarily derived from adipocytes. It can also be secreted by renal cells. Hypoadiponectinemia has been documented in patients with hyperuricemia, however, whether soluble uric acid (SUA) regulates the expression of APN and APN receptor 1 (AdipoR1) in renal proximal tubule epithelial cells (PTECs) remains to be elucidated. The present study investigated the expression of APN and AdipoR1 in cultured PTECs that were exposed to SUA through immunofluorescence and western blot analysis. In addition, Sprague-Dawley rats with oxonic acid-induced hyperuricemia (HUA) with or without febuxostat treatment were employed as an animal model to measure 24 h urine protein, serum creatinine, urea nitrogen, uric acid and homeostasis model assessment of insulin resistance. Renal pathology was evaluated using hematoxylin and eosin and immunohistochemical staining. APN and AdipoR1 expression in the renal cortex were evaluated by western blotting. The results demonstrated that, in PTECs, the expression of APN and AdipoR1 was constant and increased upon SUA exposure. Similar observations were made within the proximal renal tubules of rats, and the oxonic acid-induced increases in APN and AdipoR1 were offset by febuxostat treatment. Furthermore, SUA-treated PTECs exhibited an increase in the expression of NLR family pyrin domain-containing (NLRP) 3, which was dose-dependent. NLRP3 expression was also significantly increased in the renal cortex of HUA rats compared with control and febuxostat-treated rats. In conclusion, SUA enhanced the expression of APN and AdipoR1 in PTECs, which was associated with an increase in NLRP3 expression. The APN-AdipoR1 pathway was demonstrated to have an important role in in vitro and in vivo models of renal proximal tubule inflammatory injury. Therefore, this pathway may be a potential therapy target in urate nephropathy. PMID:29359786

C/EBPβ regulates delta-secretase expression and mediates pathogenesis in mouse models of Alzheimer's disease.

PubMed

Wang, Zhi-Hao; Gong, Ke; Liu, Xia; Zhang, Zhentao; Sun, Xiaoou; Wei, Zheng Zachory; Yu, Shan Ping; Manfredsson, Fredric P; Sandoval, Ivette M; Johnson, Peter F; Jia, Jianping; Wang, Jian-Zhi; Ye, Keqiang

2018-05-03

Delta-secretase cleaves both APP and Tau to mediate the formation of amyloid plaques and neurofibrillary tangle in Alzheimer's disease (AD). However, how aging contributes to an increase in delta-secretase expression and AD pathologies remains unclear. Here we show that a CCAAT-enhancer-binding protein (C/EBPβ), an inflammation-regulated transcription factor, acts as a key age-dependent effector elevating both delta-secretase (AEP) and inflammatory cytokines expression in mediating pathogenesis in AD mouse models. We find that C/EBPβ regulates delta-secretase transcription and protein levels in an age-dependent manner. Overexpression of C/EBPβ in young 3xTg mice increases delta-secretase and accelerates the pathological features including cognitive dysfunctions, which is abolished by inactive AEP C189S. Conversely, depletion of C/EBPβ from old 3xTg or 5XFAD mice diminishes delta-secretase and reduces AD pathologies, leading to amelioration of cognitive impairment in these AD mouse models. Thus, our findings support that C/EBPβ plays a pivotal role in AD pathogenesis via increasing delta-secretase expression.

Increased trophoblastic apoptosis mediated by neutrophil gelatinase-associated lipocalin (NGAL) activation in the nitrofen model of congenital diaphragmatic hernia.

PubMed

Kutasy, Balazs; Gosemann, Jan H; Duess, Johannes W; Puri, Prem

2013-01-01

Retinoids play a key role in fetal lung development. It has been suggested that the maternal-fetal retinol transport is disrupted by trophoblastic apoptosis. The mechanism underlying nitrofen-induced apoptosis in placenta is not fully understood. Neutrophil gelatinase-associated lipocalin (NGAL) is expressed in the fetal part of the maternal-fetal interface. NGAL is part of the immune barrier and serves primarily as a transport protein transferring biologically hazardous molecules in a safe and controlled way. It has been shown that over-activation of NGAL induces apoptosis. We hypothesized that increased placental NGAL expression induces trophoblastic apoptosis in the nitrofen model of CDH. Pregnant rats were exposed to either olive oil or nitrofen on day 9 of gestation (D9). Placenta harvested on D21 and divided into two groups: control and nitrofen with CDH. Immunohistochemistry was performed to evaluate trophoblasts (by cytokeratin expression), NGAL expression, and apoptotic trophoblastic cells (using TUNEL assay). Total RNA was extracted from each placenta and the relative mRNA expression levels of NGAL were analyzed using RT-PCR. Immunohistochemistry showed NGAL immunoreactivity both in control and CDH in the fetal part of the fetal-maternal interface of placenta. Markedly increased NGAL expression was detected in CDH group compared to controls. Relative mRNA expression levels of NGAL gene were significantly increased in the CDH group compared to control in the placenta (5.924 ± 0.93 vs. 1.895 ± 0.54, p < 0.001). Markedly increased numbers of apoptotic trophoblastic cells were seen in the maternal-fetal interface in the CDH group compared to controls. NGAL activation may lead to increased trophoblastic apoptosis in the maternal-fetal interface in the nitrofen model of CDH. These changes may therefore cause disturbance in maternal-fetal retinol transport affecting fetal lung morphogenesis.

Central control of interlimb coordination and speed‐dependent gait expression in quadrupeds

PubMed Central

Danner, Simon M.; Wilshin, Simon D.; Shevtsova, Natalia A.

2016-01-01

Key points Quadrupeds express different gaits depending on speed of locomotion.Central pattern generators (one per limb) within the spinal cord generate locomotor oscillations and control limb movements. Neural interactions between these generators define interlimb coordination and gait.We present a computational model of spinal circuits representing four rhythm generators with left–right excitatory and inhibitory commissural and fore–hind inhibitory interactions within the cord.Increasing brainstem drive to all rhythm generators and excitatory commissural interneurons induces an increasing frequency of locomotor oscillations accompanied by speed‐dependent gait changes from walk to trot and to gallop and bound.The model closely reproduces and suggests explanations for multiple experimental data, including speed‐dependent gait transitions in intact mice and changes in gait expression in mutants lacking certain types of commissural interneurons. The model suggests the possible circuit organization in the spinal cord and proposes predictions that can be tested experimentally. Abstract As speed of locomotion is increasing, most quadrupeds, including mice, demonstrate sequential gait transitions from walk to trot and to gallop and bound. The neural mechanisms underlying these transitions are poorly understood. We propose that the speed‐dependent expression of different gaits results from speed‐dependent changes in the interactions between spinal circuits controlling different limbs and interlimb coordination. As a result, the expression of each gait depends on (1) left–right interactions within the spinal cord mediated by different commissural interneurons (CINs), (2) fore–hind interactions on each side of the spinal cord and (3) brainstem drives to rhythm‐generating circuits and CIN pathways. We developed a computational model of spinal circuits consisting of four rhythm generators (RGs) with bilateral left–right interactions mediated by V0 CINs (V0D and V0V sub‐types) providing left–right alternation, and conditional V3 CINs promoting left–right synchronization. Fore and hind RGs mutually inhibited each other. We demonstrate that linearly increasing excitatory drives to the RGs and V3 CINs can produce a progressive increase in the locomotor speed accompanied by sequential changes of gaits from walk to trot and to gallop and bound. The model closely reproduces and suggests explanations for the speed‐dependent gait expression observed in vivo in intact mice and in mutants lacking V0V or all V0 CINs. Specifically, trot is not expressed after removal of V0V CINs, and only bound is expressed after removal of all V0 CINs. The model provides important insights into the organization of spinal circuits and neural control of locomotion. PMID:27633893

Antenatal retinoic acid administration increases trophoblastic retinol-binding protein dependent retinol transport in the nitrofen model of congenital diaphragmatic hernia.

PubMed

Kutasy, Balazs; Friedmacher, Florian; Pes, Lara; Coyle, David; Doi, Takashi; Paradisi, Francesca; Puri, Prem

2016-04-01

Low pulmonary retinol levels and disrupted retinoid signaling pathway (RSP) have been implicated in the pathogenesis of congenital diaphragmatic hernia (CDH) and associated pulmonary hypoplasia (PH). It has been demonstrated that nitrofen disturbs the main retinol-binding protein (RBP)-dependent trophoblastic retinol transport. Several studies have demonstrated that prenatal treatment with retinoic acid (RA) can reverse PH in the nitrofen-induced CDH model. We hypothesized that maternal administration of RA can increase trophoblastic RBP-dependent retinol transport in a nitrofen model of CDH. Pregnant rats were treated with nitrofen or vehicle on gestational day 9 (D9) and sacrificed on D21. RA was given i.p. on D18, D19, and D20. Retinol and RA levels were measured using high-performance liquid chromatography. Immunohistochemistry was performed to evaluate trophoblastic expression of RBP. Expression levels of the primary RSP genes were determined using quantitative real-time PCR and immunohistochemistry. Markedly increased trophoblastic RBP immunoreactivity was observed in CDH+RA compared to CDH. Significantly increased serum and pulmonary retinol and RA levels were detected in CDH+RA compared to CDH. Pulmonary expression of RSP genes and proteins were increased in CDH+RA compared to CDH. Increased trophoblastic RBP expression and retinol transport after antenatal administration of RA suggest that retinol-triggered RSP activation may attenuate CDH-associated PH by elevating serum and pulmonary retinol levels.

Not changes in membrane fluidity but proteotoxic stress triggers heat shock protein expression in Chlamydomonas reinhardtii.

PubMed

Rütgers, Mark; Muranaka, Ligia Segatto; Schulz-Raffelt, Miriam; Thoms, Sylvia; Schurig, Juliane; Willmund, Felix; Schroda, Michael

2017-12-01

A conserved reaction of all organisms exposed to heat stress is an increased expression of heat shock proteins (HSPs). Several studies have proposed that HSP expression in heat-stressed plant cells is triggered by an increased fluidity of the plasma membrane. Among the main lines of evidence in support of this model are as follows: (a) the degree of membrane lipid saturation was higher in cells grown at elevated temperatures and correlated with a lower amplitude of HSP expression upon a temperature upshift, (b) membrane fluidizers induce HSP expression at physiological temperatures, and (c) membrane rigidifier dimethylsulfoxide dampens heat-induced HSP expression. Here, we tested whether this holds also for Chlamydomonas reinhardtii. We show that heat-induced HSP expression in cells grown at elevated temperatures was reduced because they already contained elevated levels of cytosolic HSP70A/90A that apparently act as negative regulators of heat shock factor 1. We find that membrane rigidifier dimethylsulfoxide impaired translation under heat stress conditions and that membrane fluidizer benzyl alcohol not only induced HSP expression but also caused protein aggregation. These findings support the classical model for the cytosolic unfolded protein response, according to which HSP expression is induced by the accumulation of unfolded proteins. Hence, the membrane fluidity model should be reconsidered. © 2017 John Wiley & Sons Ltd.

Receptor for Advanced Glycation End Products (RAGE) is Expressed Predominantly in Medium Spiny Neurons of tgHD Rat Striatum.

PubMed

Shi, Dian; Chang, Joshua W; Choi, Jaimin; Connor, Bronwen; O'Carroll, Simon J; Nicholson, Louise F B; Kim, Joo Hyun

2018-06-01

Receptor for advanced glycation end products (RAGE) is a multi-ligand receptor involved in the pathology of several progressive neurodegenerative disorders including Huntington's disease (HD). We previously showed that the expression of RAGE and its colocalization with ligands were increased in the striatum of HD patients, increasing with grade severity, and that the pattern of RAGE expression coincided with the medio-lateral pattern of neurodegeneration. However, the exact role of RAGE in HD remains elusive. In order to address the necessity for a direct functional study, we aimed to characterize the pattern of RAGE expression in the transgenic rat model of HD (tgHD rats). Our results showed that RAGE expression was expanded laterally in tgHD rat caudate-putamen (CPu) compared to wildtype littermates, but the expression was unchanged by disease severity. The rostro-caudal location did not affect RAGE expression. RAGE was predominantly expressed in the medium spiny neurons (MSN) where it colocalized most extensively with N-carboxymethyllysine (CML), which largely contradicts with observations from human HD brains. Overall, the tgHD rat model only partially recapitulated the pattern in striatal RAGE expression in human brains, raising a question about its reliability as an animal model for future functional studies. Copyright © 2018 IBRO. Published by Elsevier Ltd. All rights reserved.

Iron regulation of hepcidin despite attenuated Smad1,5,8 signaling in mice without transferrin receptor 2 or Hfe

PubMed Central

Corradini, Elena; Rozier, Molly; Meynard, Delphine; Odhiambo, Adam; Lin, Herbert Y.; Feng, Qi; Migas, Mary C.; Britton, Robert S.; Babitt, Jodie L.; Fleming, Robert E.

2011-01-01

Background & Aims HFE and transferrin receptor 2 (TFR2) are each necessary for the normal relationship between body iron status and liver hepcidin expression. In murine Hfe and Tfr2 knockout models of hereditary hemochromatosis (HH), signal transduction to hepcidin via the bone morphogenetic protein 6 (Bmp6)/Smad1,5,8 pathway is attenuated. We examined the effect of dietary iron on regulation of hepcidin expression via the Bmp6/Smad1,5,8 pathway using mice with targeted disruption of Tfr2, Hfe, or both genes. Methods Hepatic iron concentrations and mRNA expression of Bmp6 and hepcidin were compared with wild-type mice in each of the HH models on standard or iron-loading diets. Liver phospho-Smad (P-Smad)1,5,8 and Id1 mRNA levels were measured as markers of Bmp/Smad signaling. Results While Bmp6 expression was increased, liver hepcidin and Id1 expression were decreased in each of the HH models compared with wild-type mice. Each of the HH models also demonstrated attenuated P-Smad1,5,8 levels relative to liver iron status. Mice with combined Hfe/Tfr2 disruption were most affected. Dietary iron loading increased hepcidin and Id1 expression in each of the HH models. Compared with wild-type mice, HH mice demonstrated attenuated (Hfe knockout) or no increases in P-Smad1,5,8 levels in response to dietary iron loading. Conclusions These observations demonstrate that Tfr2 and Hfe are each required for normal signaling of iron status to hepcidin via Bmp6/Smad1,5,8 pathway. Mice with combined loss of Hfe and Tfr2 up-regulate hepcidin in response to dietary iron loading without increases in liver BMP6 mRNA or steady-state P-Smad1,5,8 levels. PMID:21745449

Regulation of hepatic cardiolipin metabolism by TNFα: Implication in cancer cachexia.

PubMed

Peyta, Laure; Jarnouen, Kathleen; Pinault, Michelle; Coulouarn, Cedric; Guimaraes, Cyrille; Goupille, Caroline; de Barros, Jean-Paul Pais; Chevalier, Stephan; Dumas, Jean-François; Maillot, François; Hatch, Grant M; Loyer, Pascal; Servais, Stephane

2015-11-01

Cardiolipin (CL) content accumulation leads to an increase in energy wasting in liver mitochondria in a rat model of cancer cachexia in which tumor necrosis factor alpha (TNFα) is highly expressed. In this study we investigated the mechanisms involved in liver mitochondria CL accumulation in cancer cachexia and examined if TNFα was involved in this process leading to mitochondrial bioenergetics alterations. We studied gene, protein expression and activity of the main enzymes involved in CL metabolism in liver mitochondria from a rat model of cancer cachexia and in HepaRG hepatocyte-like cells exposed to 20 ng/ml of TNFα for 12 h. Phosphatidylglycerolphosphate synthase (PGPS) gene expression was increased 2.3-fold (p<0.02) and cardiolipin synthase (CLS) activity decreased 44% (p<0.03) in cachectic rat livers compared to controls. CL remodeling enzymes monolysocardiolipin acyltransferase (MLCL AT-1) activity and tafazzin (TAZ) gene expression were increased 30% (p<0.01) and 50% (p<0.02), respectively, in cachectic rat livers compared to controls. Incubation of hepatocytes with TNFα increased CL content 15% (p<0.05), mitochondrial oxygen consumption 33% (p<0.05), PGPS gene expression 44% (p<0.05) and MLCL AT-1 activity 20% (p<0.05) compared to controls. These above findings strongly suggest that in cancer cachexia, TNFα induces a higher energy wasting in liver mitochondria by increasing CL content via upregulation of PGPS expression.

Blood-Brain Barrier Permeability Is Exacerbated in Experimental Model of Hepatic Encephalopathy via MMP-9 Activation and Downregulation of Tight Junction Proteins.

PubMed

Dhanda, Saurabh; Sandhir, Rajat

2018-05-01

The present study was designed to investigate the mechanisms involved in blood-brain barrier (BBB) permeability in bile duct ligation (BDL) model of chronic hepatic encephalopathy (HE). Four weeks after BDL surgery, a significant increase was observed in serum bilirubin levels. Masson trichrome staining revealed severe hepatic fibrosis in the BDL rats. 99m Tc-mebrofenin retention was increased in the liver of BDL rats suggesting impaired hepatobiliary transport. An increase in permeability to sodium fluorescein, Evans blue, and fluorescein isothiocyanate (FITC)-dextran along with increase in water and electrolyte content was observed in brain regions of BDL rats suggesting disrupted BBB. Increased brain water content can be attributed to increase in aquaporin-4 mRNA and protein expression in BDL rats. Matrix metalloproteinase-9 (MMP-9) mRNA and protein expression was increased in brain regions of BDL rats. Additionally, mRNA and protein expression of tissue inhibitor of matrix metalloproteinases (TIMPs) was also increased in different regions of brain. A significant decrease in mRNA expression and protein levels of tight junction proteins, viz., occludin, claudin-5, and zona occluden-1 (ZO-1) was observed in different brain regions of BDL rats. VCAM-1 mRNA and protein expression was also found to be significantly upregulated in different brain regions of BDL animals. The findings from the study suggest that increased BBB permeability in HE involves activation of MMP-9 and loss of tight junction proteins.

Retinal hypoxia induces vascular endothelial growth factor through induction of estrogen-related receptor γ

DOE Office of Scientific and Technical Information (OSTI.GOV)

Do, Ji Yeon; Choi, Young Keun; Kook, Hyun

2015-05-01

Ischemic retinopathies causing overexpression of pro-angiogenic factors, including vascular endothelial growth factor (VEGF), are the most common cause of blindness. Thus, understanding the pathophysiology of targetable pathways that regulate retinal VEGF is of great interest. A conserved binding site for estrogen-related receptor γ (ERRγ) has been identified in the promoter of the Vegfa gene. ERRγ is a constitutively active orphan nuclear receptor and its expression is increased by hypoxic stimuli in metabolically active tissues. This study evaluated the role of ERRγ in the ischemic retina and the anti-VEGF potential of GSK5182, a selective inverse agonist of ERRγ. In an oxygen-inducedmore » retinopathy (OIR) mouse model, immunohistochemistry showed significantly increased ERRγ expression in the ganglion cell layer at postnatal day (P) 17. In a ganglion cell line (RGC-5), mRNA and protein levels of ERRγ were increased by desferrioxamine treatment and hypoxic conditions (1% O{sub 2}). Transient transfection of RGC-5 cells revealed that ERRγ regulated Vegfa expression and this was inhibited by GSK5182. Intravitreal injection of GSK5182 into the OIR model at P14 inhibited retinal Vegfa mRNA expression at P17. GSK5182 suppresses hypoxia-induced VEGF expression via ERRγ; therefore, ERRγ could be a treatment target for ischemic retinopathies. - Highlights: • OIR mice exhibited increased ERRγ expression in the ganglion cell layer. • Hypoxia-induced ERRγ expression was observed in retinal ganglion cells. • ERRγ overexpression increased VEGFA expression in retinal ganglion cells. • An ERRγ inverse agonist suppressed VEGFA expression in retinal ganglion cells. • Intravitreal injection of an ERRγ inverse agonist suppressed VEGFA in OIR mice.« less

Human serum activates CIDEB-mediated lipid droplet enlargement in hepatoma cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Singaravelu, Ragunath; National Research Council of Canada, Ottawa, Ontario K1A 0R6; Lyn, Rodney K.

Highlights: •Human serum induced differentiation of hepatoma cells increases cellular lipid droplet (LD) size. •The observed increase in LD size correlates with increased PGC-1α and CIDEB expression. •Induction of CIDEB expression correlates with rescue of VLDL secretion and loss of ADRP. •siRNA knockdown of CIDEB impairs the human serum mediated increase in LD size. •This system represents a cost-efficient model to study CIDEB’s role in lipid biology. -- Abstract: Human hepatocytes constitutively express the lipid droplet (LD) associated protein cell death-inducing DFFA-like effector B (CIDEB). CIDEB mediates LD fusion, as well as very-low-density lipoprotein (VLDL) maturation. However, there are limitedmore » cell culture models readily available to study CIDEB’s role in these biological processes, as hepatoma cell lines express negligible levels of CIDEB. Recent work has highlighted the ability of human serum to differentiate hepatoma cells. Herein, we demonstrate that culturing Huh7.5 cells in media supplemented with human serum activates CIDEB expression. This activation occurs through the induced expression of PGC-1α, a positive transcriptional regulator of CIDEB. Coherent anti-Stokes Raman scattering (CARS) microscopy revealed a correlation between CIDEB levels and LD size in human serum treated Huh7.5 cells. Human serum treatment also resulted in a rapid decrease in the levels of adipose differentiation-related protein (ADRP). Furthermore, individual overexpression of CIDEB was sufficient to down-regulate ADRP protein levels. siRNA knockdown of CIDEB revealed that the human serum mediated increase in LD size was CIDEB-dependent. Overall, our work highlights CIDEB’s role in LD fusion, and presents a new model system to study the PGC-1α/CIDEB pathway’s role in LD dynamics and the VLDL pathway.« less

Expression of the filaggrin gene in umbilical cord blood predicts eczema risk in infancy: A birth cohort study.

PubMed

Ziyab, A H; Ewart, S; Lockett, G A; Zhang, H; Arshad, H; Holloway, J W; Karmaus, W

2017-09-01

Filaggrin gene (FLG) expression, particularly in the skin, has been linked to the development of the skin barrier and is associated with eczema risk. However, knowledge as to whether FLG expression in umbilical cord blood (UCB) is associated with eczema development and prediction is lacking. This study sought to assess whether FLG expression in UCB associates with and predicts the development of eczema in infancy. Infants enrolled in a birth cohort study (n=94) were assessed for eczema at ages 3, 6, and 12 months. Five probes measuring FLG transcripts expression in UCB were available from genomewide gene expression profiling. FLG genetic variants R501X, 2282del4, and S3247X were genotyped. Associations were assessed using Poisson regression with robust variance estimation. Area under the curve (AUC), describing the discriminatory/predictive performance of fitted models, was estimated from logistic regression. Increased level of FLG expression measured by probe A_24_P51322 was associated with reduced risk of eczema during the first year of life (RR=0.60, 95% CI: 0.38-0.95). In contrast, increased level of FLG antisense transcripts measured by probe A_21_P0014075 was associated with increased risk of eczema (RR=2.02, 95% CI: 1.10-3.72). In prediction models including FLG expression, FLG genetic variants, and sex, discrimination between children who will and will not develop eczema at 3 months of age was high (AUC: 0.91, 95% CI: 0.84-0.98). This study demonstrated, for the first time, that FLG expression in UCB is associated with eczema development in infancy. Moreover, our analysis provided prediction models that were capable of discriminating, to a great extent, between those who will and will not develop eczema in infancy. Therefore, early identification of infants at increased risk of developing eczema is possible and such high-risk newborns may benefit from early stratification and intervention. © 2017 John Wiley & Sons Ltd.

A HER2-targeting antibody-drug conjugate, trastuzumab deruxtecan (DS-8201a), enhances antitumor immunity in a mouse model.

PubMed

Iwata, Tomomi Nakayama; Ishii, Chiaki; Ishida, Saori; Ogitani, Yusuke; Wada, Teiji; Agatsuma, Toshinori

2018-04-27

Trastuzumab deruxtecan (DS-8201a), a HER2-targeting antibody-drug conjugate with a topoisomerase I inhibitor exatecan derivative (DX-8951 derivative, DXd), has been reported to exert potent antitumor effects in xenograft mouse models and clinical trials. In this study, the immune system-activating ability of DS-8201a was assessed. DS-8201a significantly suppressed tumor growth in an immunocompetent mouse model with human HER2-expressing CT26.WT (CT26.WT-hHER2) cells. Cured immunocompetent mice rejected not only re-challenged CT26.WT-hHER2 cells, but also CT26.WT-mock cells. Splenocytes from the cured mice responded to both CT26.WT-hHER2 and CT26.WT-mock cells. Further analyses revealed that DXd up-regulated CD86 expression on bone marrow-derived DCs in vitro, and that DS-8201a increased tumor-infiltrating DCs and up-regulated their CD86 expression in vivo. DS-8201a also increased tumor-infiltrating CD8+ T cells and enhanced PD-L1 and MHC class I expression on tumor cells. Furthermore, combination therapy with DS-8201a and anti-PD-1 antibody was more effective than either monotherapy. In conclusion, DS-8201a enhanced antitumor immunity, as evidenced by the increased expression of DC markers, augmented expression of MHC class I in tumor cells, and rejection of re-challenged tumor cells by adaptive immune cells, suggesting that DS-8201a enhanced tumor recognition by T cells. Furthermore, DS-8201a treatment benefited from combination with anti-PD-1 antibody, possibly due to increased T cell activity and up-regulated PD-L1 expression induced by DS-8201a. Copyright ©2018, American Association for Cancer Research.

Remarkable alterations of Nav1.6 in reactive astrogliosis during epileptogenesis.

PubMed

Zhu, Hongyan; Zhao, Yuxiao; Wu, Hao; Jiang, Nan; Wang, Ziyi; Lin, Weide; Jin, Jiahui; Ji, Yonghua

2016-12-01

Voltage-gated sodium channels (VGSCs) play a vital role in controlling neuronal excitability. Nav1.6 is the most abundantly expressed VGSCs subtype in the adult central nervous system and has been found to contribute to facilitate the hyperexcitability of neurons after electrical induction of status epilepticus (SE). To clarify the exact expression patterns of Nav1.6 during epileptogenesis, we examined the expression of Nav1.6 at protein and mRNA levels in two distinct animal models of temporal lobe epilepsy (TLE) including a post-SE model induced by kainic acid (KA) intrahippocampal injection and a kindling model evoked by pentylenetetrazole (PTZ). A prominent, seizure intensity-dependent increase of Nav1.6 expression in reactive astrocytes was observed in ipsilateral hippocampus of post-SE rats, reaching the peak at 21 days after SE, a time point during the latent stage of epileptogenesis. However, Nav1.6 with low expression level was selectively expressed in the hippocampal neurons rather than astrocytes in PTZ-kindled animals. This seizure-related increase of a VGSCs subtype in reactive astrocytes after SE may represent a new mechanism for signal communication between neuron and glia in the course of epileptogenesis, facilitating the neuronal hyperexcitability.

microRNA as a Potential Vector for the Propagation of Robustness in Protein Expression and Oscillatory Dynamics within a ceRNA Network

PubMed Central

Gérard, Claude; Novák, Béla

2013-01-01

microRNAs (miRNAs) are small noncoding RNAs that are important post-transcriptional regulators of gene expression. miRNAs can induce thresholds in protein synthesis. Such thresholds in protein output can be also achieved by oligomerization of transcription factors (TF) for the control of gene expression. First, we propose a minimal model for protein expression regulated by miRNA and by oligomerization of TF. We show that miRNA and oligomerization of TF generate a buffer, which increases the robustness of protein output towards molecular noise as well as towards random variation of kinetics parameters. Next, we extend the model by considering that the same miRNA can bind to multiple messenger RNAs, which accounts for the dynamics of a minimal competing endogenous RNAs (ceRNAs) network. The model shows that, through common miRNA regulation, TF can control the expression of all proteins formed by the ceRNA network, even if it drives the expression of only one gene in the network. The model further suggests that the threshold in protein synthesis mediated by the oligomerization of TF can be propagated to the other genes, which can increase the robustness of the expression of all genes in such ceRNA network. Furthermore, we show that a miRNA could increase the time delay of a “Goodwin-like” oscillator model, which may favor the occurrence of oscillations of large amplitude. This result predicts important roles of miRNAs in the control of the molecular mechanisms leading to the emergence of biological rhythms. Moreover, a model for the latter oscillator embedded in a ceRNA network indicates that the oscillatory behavior can be propagated, via the shared miRNA, to all proteins formed by such ceRNA network. Thus, by means of computational models, we show that miRNAs could act as vectors allowing the propagation of robustness in protein synthesis as well as oscillatory behaviors within ceRNA networks. PMID:24376695

[Differential expression genes of bone tissues surrounding implants in diabetic rats by gene chip].

PubMed

Wang, Xin-xin; Ma, Yue; Li, Qing; Jiang, Bao-qi; Lan, Jing

2012-10-01

To compare mRNA expression profiles of bone tissues surrounding implants between normal rats and rats with diabetes using microarray technology. Six Wistar rats were randomly selected and divided into normal model group and diabetic group. Diabetic model condition was established by injecting Streptozotocin into peritoneal space. Titanium implants were implanted into the epiphyseal end of the rats' tibia. Bone tissues surrounding implant were harvested and sampled after 3 months to perform comprehensive RNA gene expression profiling, including 17983 for genome-wide association study.GO analysis was used to compare different gene expression and real-time PCR was used to confirm the results on core samples. The results indicated that there were 1084 differential gene expression. In the diabetic model, there were 352 enhanced expression genes, 732 suppressed expression genes. GO analysis involved 1154 different functional type. Osteoblast related gene expressions in bone tissue samples of diabetic rats were decreased, and lipid metabolism pathway related gene expression was increased.

Inhibiting effects of rhynchophylline on methamphetamine-dependent zebrafish are related with the expression of tyrosine hydroxylase (TH).

PubMed

Zhu, Chen; Liu, Wei; Luo, Chaohua; Liu, Yi; Li, Chan; Fang, Miao; Lin, Yingbo; Ou, Jinying; Chen, Minting; Zhu, Daoqi; Yung, Ken Kin-Lam; Mo, Zhixian

2017-03-01

In this study, to study the effect of rhynchophylline on TH in midbrain of methamphetamine-induced conditioned place preference (CPP) adult zebrafish, place preference adult zebrafish models were established by methamphetamine (40μg/g) and the expression of TH was observed by immunohistochemistry technique and Western blot. Ketamine (150μg/g), high dose of rhynchophylline (100μg/g) group can significantly reduce the place preference; immunohistochemistry results showed that the number of TH-positive neurons in midbrain was increased in the methamphetamine model group, whereas less TH-positive neurons were found in the ketamine group and high dosage rhynchophylline group. Western blot results showed that the expression of TH protein was significantly increased in the model group, whereas less expression was found in the ketamine group, high dosage rhynchophylline group. Our data pointed out that TH plays an important role in the formation of methamphetamine-induced place preference in adult zebrafish. Rhynchophylline reversed the expression of TH in the midbrain demonstrates the potential effect of mediates methamphetamine induced rewarding effect. Copyright © 2017 Elsevier B.V. All rights reserved.

BAG3 regulates ECM accumulation in renal proximal tubular cells induced by TGF-β1.

PubMed

Du, Feng; Li, Si; Wang, Tian; Zhang, Hai-Yan; Li, De-Tian; Du, Zhen-Xian; Wang, Hua-Qin; Wang, Yan-Qiu

2015-01-01

Previously we have demonstrated that Bcl-2-associated athanogene 3 (BAG3) is increased in renal fibrosis using a rat unilateral ureteral obstruction model. The current study investigated the role of BAG3 in renal fibrosis using transforming growth factor (TGF)-β1-treated human proximal tubular epithelial (HK-2) cells. An upregulation of BAG3 in vitro models was observed, which correlated with the increased synthesis of extracellular matrix (ECM) proteins and expression of tissue-type plasminogen activator inhibitor (PAI)-1. Blockade of BAG3 induction by shorting hairpin RNA suppressed the expression of ECM proteins but had no effect on PAI-1 expression induced by TGF-β1. Forced overexpression of BAG3 selectively increased collagens. TGF-β1-induced BAG3 expression in HK-2 cells was attenuated by ERK1/2 and JNK MAPK inhibitors. In addition, forced BAG3 overexpression blocked attenuation of collagens expression by ERK1/2 and JNK inhibitors. These data suggest that ERK1/2 and JNK signaling events are involved in modulating the expression of BAG3, which would ultimately contribute to renal fibrosis by enhancing the synthesis and deposition of ECM proteins.

BAG3 regulates ECM accumulation in renal proximal tubular cells induced by TGF-β1

PubMed Central

Du, Feng; Li, Si; Wang, Tian; Zhang, Hai-Yan; Li, De-Tian; Du, Zhen-Xian; Wang, Hua-Qin; Wang, Yan-Qiu

2015-01-01

Previously we have demonstrated that Bcl-2-associated athanogene 3 (BAG3) is increased in renal fibrosis using a rat unilateral ureteral obstruction model. The current study investigated the role of BAG3 in renal fibrosis using transforming growth factor (TGF)-β1-treated human proximal tubular epithelial (HK-2) cells. An upregulation of BAG3 in vitro models was observed, which correlated with the increased synthesis of extracellular matrix (ECM) proteins and expression of tissue-type plasminogen activator inhibitor (PAI)-1. Blockade of BAG3 induction by shorting hairpin RNA suppressed the expression of ECM proteins but had no effect on PAI-1 expression induced by TGF-β1. Forced overexpression of BAG3 selectively increased collagens. TGF-β1-induced BAG3 expression in HK-2 cells was attenuated by ERK1/2 and JNK MAPK inhibitors. In addition, forced BAG3 overexpression blocked attenuation of collagens expression by ERK1/2 and JNK inhibitors. These data suggest that ERK1/2 and JNK signaling events are involved in modulating the expression of BAG3, which would ultimately contribute to renal fibrosis by enhancing the synthesis and deposition of ECM proteins. PMID:26885277

GFRA1 promotes cisplatin-induced chemoresistance in osteosarcoma by inducing autophagy

PubMed Central

Kim, Mihwa; Jung, Ji-Yeon; Choi, Seungho; Lee, Hyunseung; Morales, Liza D.; Koh, Jeong-Tae; Kim, Sun Hun; Choi, Yoo-Duk; Choi, Chan; Slaga, Thomas J.; Kim, Won Jae; Kim, Dae Joon

2017-01-01

ABSTRACT Recent progress in chemotherapy has significantly increased its efficacy, yet the development of chemoresistance remains a major drawback. In this study, we show that GFRA1/GFRα1 (GDNF family receptor α 1), contributes to cisplatin-induced chemoresistance by regulating autophagy in osteosarcoma. We demonstrate that cisplatin treatment induced GFRA1 expression in human osteosarcoma cells. Induction of GFRA1 expression reduced cisplatin-induced apoptotic cell death and it significantly increased osteosarcoma cell survival via autophagy. GFRA1 regulates AMPK-dependent autophagy by promoting SRC phosphorylation independent of proto-oncogene RET kinase. Cisplatin-resistant osteosarcoma cells showed NFKB1/NFκB-mediated GFRA1 expression. GFRA1 expression promoted tumor formation and growth in mouse xenograft models and inhibition of autophagy in a GFRA1-expressing xenograft mouse model during cisplatin treatment effectively reduced tumor growth and increased survival. In cisplatin-treated patients, treatment period and metastatic status were associated with GFRA1-mediated autophagy. These findings suggest that GFRA1-mediated autophagy is a promising novel target for overcoming cisplatin resistance in osteosarcoma. PMID:27754745

GFRA1 promotes cisplatin-induced chemoresistance in osteosarcoma by inducing autophagy.

PubMed

Kim, Mihwa; Jung, Ji-Yeon; Choi, Seungho; Lee, Hyunseung; Morales, Liza D; Koh, Jeong-Tae; Kim, Sun Hun; Choi, Yoo-Duk; Choi, Chan; Slaga, Thomas J; Kim, Won Jae; Kim, Dae Joon

2017-01-02

Recent progress in chemotherapy has significantly increased its efficacy, yet the development of chemoresistance remains a major drawback. In this study, we show that GFRA1/GFRα1 (GDNF family receptor α 1), contributes to cisplatin-induced chemoresistance by regulating autophagy in osteosarcoma. We demonstrate that cisplatin treatment induced GFRA1 expression in human osteosarcoma cells. Induction of GFRA1 expression reduced cisplatin-induced apoptotic cell death and it significantly increased osteosarcoma cell survival via autophagy. GFRA1 regulates AMPK-dependent autophagy by promoting SRC phosphorylation independent of proto-oncogene RET kinase. Cisplatin-resistant osteosarcoma cells showed NFKB1/NFκB-mediated GFRA1 expression. GFRA1 expression promoted tumor formation and growth in mouse xenograft models and inhibition of autophagy in a GFRA1-expressing xenograft mouse model during cisplatin treatment effectively reduced tumor growth and increased survival. In cisplatin-treated patients, treatment period and metastatic status were associated with GFRA1-mediated autophagy. These findings suggest that GFRA1-mediated autophagy is a promising novel target for overcoming cisplatin resistance in osteosarcoma.

Translating natural genetic variation to gene expression in a computational model of the Drosophila gap gene regulatory network

PubMed Central

Kozlov, Konstantin N.; Kulakovskiy, Ivan V.; Zubair, Asif; Marjoram, Paul; Lawrie, David S.; Nuzhdin, Sergey V.; Samsonova, Maria G.

2017-01-01

Annotating the genotype-phenotype relationship, and developing a proper quantitative description of the relationship, requires understanding the impact of natural genomic variation on gene expression. We apply a sequence-level model of gap gene expression in the early development of Drosophila to analyze single nucleotide polymorphisms (SNPs) in a panel of natural sequenced D. melanogaster lines. Using a thermodynamic modeling framework, we provide both analytical and computational descriptions of how single-nucleotide variants affect gene expression. The analysis reveals that the sequence variants increase (decrease) gene expression if located within binding sites of repressors (activators). We show that the sign of SNP influence (activation or repression) may change in time and space and elucidate the origin of this change in specific examples. The thermodynamic modeling approach predicts non-local and non-linear effects arising from SNPs, and combinations of SNPs, in individual fly genotypes. Simulation of individual fly genotypes using our model reveals that this non-linearity reduces to almost additive inputs from multiple SNPs. Further, we see signatures of the action of purifying selection in the gap gene regulatory regions. To infer the specific targets of purifying selection, we analyze the patterns of polymorphism in the data at two phenotypic levels: the strengths of binding and expression. We find that combinations of SNPs show evidence of being under selective pressure, while individual SNPs do not. The model predicts that SNPs appear to accumulate in the genotypes of the natural population in a way biased towards small increases in activating action on the expression pattern. Taken together, these results provide a systems-level view of how genetic variation translates to the level of gene regulatory networks via combinatorial SNP effects. PMID:28898266

Effect of Nicotine on CYP2B1 Expression in a Glioma Animal Model and Analysis of CYP2B6 Expression in Pediatric Gliomas.

PubMed

Nava-Salazar, Sonia; Gómez-Manzo, Saúl; Marcial-Quino, Jaime; Marhx-Bracho, Alfonso; Phillips-Farfán, Bryan V; Diaz-Avalos, Carlos; Vanoye-Carlo, America

2018-06-16

Cyclophosphamide (CPA) is a pro-drug commonly used in the chemotherapeutic schemes for glioma treatment but has high toxicity and the side effects include brain damage and even death. Since CPA is activated mainly by CY2B6, over-expression of the enzyme in the tumor cells has been proposed to enhance CPA activation. In this study, we explored the induction of the Cyp2b1 (homologous to CYP2B6 ) by nicotine in an animal rat model with glioma. Gene expression and protein levels were analyzed by RT-PCR and Western blot. Nicotine treatment increased CYP2B1 protein levels in the healthy animals’ brain tissue. In the brain tissue of animals with glioma, the CYP2B1 showed a high expression, even before nicotine treatment. Nicotine did not increase significantly the CYP2B1 protein expression in the tumor, but increased its expression in the tumor vicinity, especially around blood vessels in the cortex. We also explored CY2B6 expression in glioma samples derived from pediatric patients. Tumor tissue showed a variable expression of the enzyme, which could depend on the tumor malignancy grade. Induction of the CYP2B6 in pediatric gliomas with lower expression of the enzyme, could be an alternative to improve the antitumoral effect of CPA treatment.

The Expressive Gaze Model: Using Gaze to Express Emotion

DTIC Science & Technology

2010-07-01

World of Warcraft or Oblivion , have thou- sands of computer-controlled nonplayer characters with which users can interact. Producing hand- generated...increasing to the right and the vertical increasing upward. In both cases, 0 degrees is straight ahead. Although the mechani- cal limits of human eye...to gaze from a target directly in front of her to one 60 degrees to her right , while performing these behaviors in a manner that expressed the de

Facial Expression Recognition Deficits and Faulty Learning: Implications for Theoretical Models and Clinical Applications

ERIC Educational Resources Information Center

Sheaffer, Beverly L.; Golden, Jeannie A.; Averett, Paige

2009-01-01

The ability to recognize facial expressions of emotion is integral in social interaction. Although the importance of facial expression recognition is reflected in increased research interest as well as in popular culture, clinicians may know little about this topic. The purpose of this article is to discuss facial expression recognition literature…

Heterozygous disruption of activin receptor-like kinase 1 is associated with increased renal fibrosis in a mouse model of obstructive nephropathy.

PubMed

Muñoz-Félix, José M; López-Novoa, José M; Martínez-Salgado, Carlos

2014-02-01

Tubulointerstitial fibrosis is characterized by an accumulation of extracellular matrix in the renal interstitium, myofibroblast activation, cell infiltration, and tubular cell apoptosis, leading to chronic renal failure. Activin receptor-like kinase 1 (ALK1) is a transforming growth factor-β1 type I receptor with a pivotal role in endothelial proliferation and migration, but its role in the development of renal fibrosis is unknown. To assess this we used the unilateral ureteral obstruction model of tubulointerstitial fibrosis in ALK1 haploinsufficient (ALK1(+/-)) and wild-type mice. After 15 days, there was an increase in extracellular matrix protein expression in the obstructed kidneys from both ALK1(+/+) and ALK1(+/-) mice, but obstructed kidneys from ALK1(+/-) mice showed significantly higher expression of type I collagen than those from wild-type mice. Ureteral obstruction increased kidney myofibroblasts markers (α-smooth muscle actin and S100A4), without differences between mouse genotypes. ALK1 expression was increased after ureteral obstruction, and this increased expression was located in myofibroblasts. Moreover, cultured renal fibroblasts from ALK1(+/-) mice expressed more collagen type I and fibronectin than fibroblasts derived from wild-type mice. Thus, ALK1 modulates obstruction-induced renal fibrosis by increased extracellular matrix synthesis in myofibroblasts, but without differences in myofibroblast number.

Knockdown of EphB1 receptor decreases medulloblastoma cell growth and migration and increases cellular radiosensitization

PubMed Central

Timofeeva, Olga; Pasquale, Elena B.; Hirsch, Kellen; MacDonald, Tobey J.; Dritschilo, Anatoly; Lee, Yi Chien; Henkemeyer, Mark; Rood, Brian; Jung, Mira; Wang, Xiao-Jing; Kool, Marcel

2015-01-01

The expression of members of the Eph family of receptor tyrosine kinases and their ephrin ligands is frequently dysregulated in medulloblastomas. We assessed the expression and functional role of EphB1 in medulloblastoma cell lines and engineered mouse models. mRNA and protein expression profiling showed expression of EphB1 receptor in the human medulloblastoma cell lines DAOY and UW228. EphB1 downregulation reduced cell growth and viability, decreased the expression of important cell cycle regulators, and increased the percentage of cells in G1 phase of the cell cycle. It also modulated the expression of proliferation, and cell survival markers. In addition, EphB1 knockdown in DAOY cells resulted in significant decrease in migration, which correlated with decreased β1-integrin expression and levels of phosphorylated Src. Furthermore, EphB1 knockdown enhanced cellular radiosensitization of medulloblastoma cells in culture and in a genetically engineered mouse medulloblastoma model. Using genetically engineered mouse models, we established that genetic loss of EphB1 resulted in a significant delay in tumor recurrence following irradiation compared to EphB1-expressing control tumors. Taken together, our findings establish that EphB1 plays a key role in medulloblastoma cell growth, viability, migration, and radiation sensitivity, making EphB1 a promising therapeutic target. PMID:25879388

Knockdown of EphB1 receptor decreases medulloblastoma cell growth and migration and increases cellular radiosensitization.

PubMed

Bhatia, Shilpa; Baig, Nimrah A; Timofeeva, Olga; Pasquale, Elena B; Hirsch, Kellen; MacDonald, Tobey J; Dritschilo, Anatoly; Lee, Yi Chien; Henkemeyer, Mark; Rood, Brian; Jung, Mira; Wang, Xiao-Jing; Kool, Marcel; Rodriguez, Olga; Albanese, Chris; Karam, Sana D

2015-04-20

The expression of members of the Eph family of receptor tyrosine kinases and their ephrin ligands is frequently dysregulated in medulloblastomas. We assessed the expression and functional role of EphB1 in medulloblastoma cell lines and engineered mouse models. mRNA and protein expression profiling showed expression of EphB1 receptor in the human medulloblastoma cell lines DAOY and UW228. EphB1 downregulation reduced cell growth and viability, decreased the expression of important cell cycle regulators, and increased the percentage of cells in G1 phase of the cell cycle. It also modulated the expression of proliferation, and cell survival markers. In addition, EphB1 knockdown in DAOY cells resulted in significant decrease in migration, which correlated with decreased β1-integrin expression and levels of phosphorylated Src. Furthermore, EphB1 knockdown enhanced cellular radiosensitization of medulloblastoma cells in culture and in a genetically engineered mouse medulloblastoma model. Using genetically engineered mouse models, we established that genetic loss of EphB1 resulted in a significant delay in tumor recurrence following irradiation compared to EphB1-expressing control tumors. Taken together, our findings establish that EphB1 plays a key role in medulloblastoma cell growth, viability, migration, and radiation sensitivity, making EphB1 a promising therapeutic target.

Interferon-γ-Induced Unfolded Protein Response in Conjunctival Goblet Cells as a Cause of Mucin Deficiency in Sjögren Syndrome.

PubMed

Coursey, Terry G; Tukler Henriksson, Johanna; Barbosa, Flavia L; de Paiva, Cintia S; Pflugfelder, Stephen C

2016-06-01

Goblet cells (GCs) are specialized secretory cells that produce mucins and a variety of other proteins. Significant conjunctival GC loss occurs in both experimental dry eye models and patients with keratoconjunctivitis sicca due to the induction of interferon (IFN)-γ. With the use of a primary murine culture model, we found that GCs are highly sensitive to IFN-γ with significantly reduced proliferation and altered structure with low concentrations. GC cultures treated with IFN-γ have increased gene expression of Muc2 and Muc5AC but do not express these mucin glycoproteins. We hypothesized that IFN-γ induces endoplasmic reticulum stress and the unfolded protein response (UPR) in GCs. Cultures treated with IFN-γ increased expression of UPR-associated genes and proteins. Increased GRP78 and sXBP1 expression was found in experimental dry eye and Sjögren syndrome models and was GC specific. Increased GRP78 was also found in the conjunctiva of patients with Sjögren syndrome at the gene and protein levels. Treatment with dexamethasone inhibited expression of UPR-associated genes and increased mucin production. These results indicate that induction of UPR by IFN-γ is an important cause of GC-associated mucin deficiency observed in aqueous-deficient dry eye. Therapies to block the effects of IFN-γ on the metabolically active endoplasmic reticulum in these cells might enhance synthesis and secretion of the protective GC mucins on the ocular surface. Copyright © 2016 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

[Piperine regulates glucose metabolism disorder in HepG2 cells of insulin resistance models via targeting upstream target of AMPK signaling pathway].

PubMed

Wan, Chun-Ping; Wei, Ya-Gai; Li, Xiao-Xue; Zhang, Li-Jun; Yang, Rui; Bao, Zhao-Ri-Ge-Tu

2017-02-01

To investigate the effect of piperine on the disorder of glucose metabolism in the cell model with insulin resistance (IR) and explore the molecules mechanism on intervening the upstream target of AMPK signaling pathway. The insulin resistance models in HepG2 cells were established by fat emulsion stimulation. Then glucose consumption in culture supernatant was detected by GOD-POD method. Enzyme-linked immunosorbent assay(ELISA) was used to measure the levels of leptin(LEP) and adiponectin(APN) in culture supernatant; Real-time quantitative PCR was used to assess the mRNA expression of APN and LEP; and the protein expression levels of LepR, AdipoR1, AdipoR2 and the activation of AMPK signaling pathway were detected by Western blot analysis. The results showed that piperine, rosiglitazone and AMPK agonist AICAR could significantly elevate the glucose consumption in insulin resistance cell models, enhance the level of APN, promote APN mRNA transcripts and increase the protein expression of Adipo receptor. Meanwhile,AMPKα mRNA and р-AMPKα protein expressions were also increased in piperine treated cells, but both LEP mRNA expression and LepR protein expressions were decreased in piperine treated group. The results indicated that piperine could significantly ameliorate the glucose metabolism disorder in insulin resistance cell models through regulating upstream molecules (APN and LEP) of AMPK signaling pathway, and thus activate the AMPK signaling pathway. Copyright© by the Chinese Pharmaceutical Association.

P-gp Protein Expression and Transport Activity in Rodent Seizure Models and Human Epilepsy.

PubMed

Hartz, Anika M S; Pekcec, Anton; Soldner, Emma L B; Zhong, Yu; Schlichtiger, Juli; Bauer, Bjoern

2017-04-03

A cure for epilepsy is currently not available, and seizure genesis, seizure recurrence, and resistance to antiseizure drugs remain serious clinical problems. Studies show that the blood-brain barrier is altered in animal models of epilepsy and in epileptic patients. In this regard, seizures increase expression of blood-brain barrier efflux transporters such as P-glycoprotein (P-gp), which is thought to reduce brain uptake of antiseizure drugs, and thus, contribute to antiseizure drug resistance. The goal of the current study was to assess the viability of combining in vivo and ex vivo preparations of isolated brain capillaries from animal models of seizures and epilepsy as well as from patients with epilepsy to study P-gp at the blood-brain barrier. Exposing isolated rat brain capillaries to glutamate ex vivo upregulated P-gp expression to levels that were similar to those in capillaries isolated from rats that had status epilepticus or chronic epilepsy. Moreover, the fold-increase in P-gp protein expression seen in animal models is consistent with the fold-increase in P-gp observed in human brain capillaries isolated from patients with epilepsy compared to age-matched control individuals. Overall, the in vivo/ex vivo approach presented here allows detailed analysis of the mechanisms underlying seizure-induced changes of P-gp expression and transport activity at the blood-brain barrier. This approach can be extended to other blood-brain barrier proteins that might contribute to drug-resistant epilepsy or other CNS disorders as well.

Role of ROCK expression in gallbladder smooth muscle contraction.

PubMed

Wang, Bin; Ding, You-Ming; Wang, Chun-Tao; Wang, Wei-Xing

2015-08-01

Cholelithiasis is a common medical condition whose incidence rate is increasing yearly, while its pathogenesis has yet to be elucidated. The present study assessed the expression of Rho-kinase (ROCK) in gallbladder smooth muscles and its effect on the contractile function of gallbladder smooth muscles during gallstone formation. Thirty male guinea pigs were randomly divided into three groups: The control group, the gallstone model group and the fasudil interference group. The fasting volume (FV) and bile capacity of the gallbladder (FB) as well as the total cholesterol (TC) and triglyceride (TG) contents of the gallbladder bile were determined. In addition, the gallbladder was dissected to identify whether any gallstones had formed. Part of the gallbladder tissue specimens were used for immunohistochemical analysis of ROCK expression in gallbladder smooth muscles. The results showed that four guinea pigs in the model group and eight in the fasudil group displayed gallstone formation, while there was no gallstone formation in the control group. The FV and FB were significantly increased in the model and fasudil groups. Similarly, the TC and TG contents of gallbladder bile were increased in these groups. The positive expression rate of ROCK in gallbladder smooth muscles in the model and fasudil groups was significantly reduced compared with that in the control group (P<0.05). The results of the present study indicated that the reduction of ROCK expression in guinea pig gallbladder smooth muscles weakened gallbladder contraction and thereby promoted gallstone formation.

Neuroprotective effects of Bcl-2 overexpression on nerve cells of rats with acute cerebral infarction.

PubMed

Zhang, H R; Peng, J H; Zhu, G Y; Xu, R X

2015-07-13

We aimed to investigate the influence of lentiviral-mediated Bcl-2 overexpression in cerebral tissues of rats with acute cerebral infarction. Forty-five rats were randomly divided into sham, model, and treatment groups. The sham and model groups were administered a control lentiviral vector via the intracranial arteries 10 days before surgery, while the treatment group received lentivirus encoding a Bcl-2 overexpression vector. We induced cerebral artery infarction using a suture-occlusion method and analyzed the cerebral expression levels of apoptosis-related genes (caspase-3, Bax), total cerebral apoptosis, range of cerebral tissue infarction, and changes in nerve cell function after 72 h. The Bcl-2-encoding lentivirus was well expressed in rat cerebral tissues. The treatment group had significantly higher expression levels of Bcl-2 than the other two groups. After cerebral infarction, the model group had significantly increased expression levels of caspase-3 and Bax protein in cerebral tissues than the sham (P < 0.05). Expression of these apoptosis-related proteins in the treatment group was obviously lower than that in the model group (P < 0.05), but significantly higher than in the sham group (P < 0.05). Compared to sham, neuronal apoptosis levels and infarction range of cerebral tissues was increased in the model and treatment groups; however, these values in the treatment group were significantly lower than that in the model group (P < 0.05). Importantly, the treatment group had significantly decreased neurological impairment scores (P < 0.05). In conclusion, Bcl-2 over-expression can decrease neuronal apoptosis in rat cerebral tissue, and thus is neuroprotective after cerebral ischemia.

Fibroblast growth factor 9 is a novel modulator of negative affect

PubMed Central

Aurbach, Elyse L.; Inui, Edny Gula; Turner, Cortney A.; Hagenauer, Megan H.; Prater, Katherine E.; Li, Jun Z.; Absher, Devin; Shah, Najmul; Blandino, Peter; Bunney, William E.; Myers, Richard M.; Barchas, Jack D.; Schatzberg, Alan F.; Watson, Stanley J.; Akil, Huda

2015-01-01

Both gene expression profiling in postmortem human brain and studies using animal models have implicated the fibroblast growth factor (FGF) family in affect regulation and suggest a potential role in the pathophysiology of major depressive disorder (MDD). FGF2, the most widely characterized family member, is down-regulated in the depressed brain and plays a protective role in rodent models of affective disorders. By contrast, using three microarray analyses followed by quantitative RT-PCR confirmation, we show that FGF9 expression is up-regulated in the hippocampus of individuals with MDD, and that FGF9 expression is inversely related to the expression of FGF2. Because little is known about FGF9’s function in emotion regulation, we used animal models to shed light on its potential role in affective function. We found that chronic social defeat stress, an animal model recapitulating some aspects of MDD, leads to a significant increase in hippocampal FGF9 expression, paralleling the elevations seen in postmortem human brain tissue. Chronic intracerebroventricular administration of FGF9 increased both anxiety- and depression-like behaviors. In contrast, knocking down FGF9 expression in the dentate gyrus of the hippocampus using a lentiviral vector produced a decrease in FGF9 expression and ameliorated anxiety-like behavior. Collectively, these results suggest that high levels of hippocampal FGF9 play an important role in the development or expression of mood and anxiety disorders. We propose that the relative levels of FGF9 in relation to other members of the FGF family may prove key to understanding vulnerability or resilience in affective disorders. PMID:26351673

A multi-Poisson dynamic mixture model to cluster developmental patterns of gene expression by RNA-seq.

PubMed

Ye, Meixia; Wang, Zhong; Wang, Yaqun; Wu, Rongling

2015-03-01

Dynamic changes of gene expression reflect an intrinsic mechanism of how an organism responds to developmental and environmental signals. With the increasing availability of expression data across a time-space scale by RNA-seq, the classification of genes as per their biological function using RNA-seq data has become one of the most significant challenges in contemporary biology. Here we develop a clustering mixture model to discover distinct groups of genes expressed during a period of organ development. By integrating the density function of multivariate Poisson distribution, the model accommodates the discrete property of read counts characteristic of RNA-seq data. The temporal dependence of gene expression is modeled by the first-order autoregressive process. The model is implemented with the Expectation-Maximization algorithm and model selection to determine the optimal number of gene clusters and obtain the estimates of Poisson parameters that describe the pattern of time-dependent expression of genes from each cluster. The model has been demonstrated by analyzing a real data from an experiment aimed to link the pattern of gene expression to catkin development in white poplar. The usefulness of the model has been validated through computer simulation. The model provides a valuable tool for clustering RNA-seq data, facilitating our global view of expression dynamics and understanding of gene regulation mechanisms. © The Author 2014. Published by Oxford University Press. For Permissions, please email: journals.permissions@oup.com.

Altered fatty acid-binding protein 4 (FABP4) expression and function in human and animal models of hepatocellular carcinoma.

PubMed

Thompson, Kyle J; Austin, Rebecca Garland; Nazari, Shayan S; Gersin, Keith S; Iannitti, David A; McKillop, Iain H

2017-11-24

Hepatocellular carcinoma (HCC) is the third leading cause of cancer-related mortality. Risk factors for developing HCC include viral hepatitis, alcohol and obesity. Fatty acid-binding proteins (FABPs) bind long-chain free fatty acids (FFAs) and are expressed in a tissue-specific pattern; FABP1 being the predominant hepatic form, and FABP4 the predominant adipocyte form. The aims of this study were to investigate the expression and function of FABPs1-9 in human and animal models of obesity-related HCC. FABP1-9 expression was determined in a mouse model of obesity-promoted HCC. Based on these data, expression and function of FABP4 was determined in human HCC cells (HepG2 and HuH7) in vitro. Serum from patients with different underlying hepatic pathologies was analysed for circulating FABP4 levels. Livers from obese mice, independent of tumour status, exhibited increased FABP4 mRNA and protein expression concomitant with elevated serum FABP4. In vitro, FABP4 expression was induced in human HCC cells by FFA treatment, and led to FABP4 release into culture medium. Treatment of HCC cells with exogenous FABP4 significantly increased proliferation and migration of human HCC cells. Patient serum analysis demonstrated significantly increased FABP4 in those with underlying liver disease, particularly non-alcoholic fatty liver disease (NAFLD) and HCC. These data suggest FABP4, an FABP not normally expressed in the liver, can be synthesized and secreted by hepatocytes and HCC cells, and that FABP4 may play a role in regulating tumour progression in the underlying setting of obesity. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Modeling and validation of autoinducer-mediated bacterial gene expression in microfluidic environments

PubMed Central

Austin, Caitlin M.; Stoy, William; Su, Peter; Harber, Marie C.; Bardill, J. Patrick; Hammer, Brian K.; Forest, Craig R.

2014-01-01

Biosensors exploiting communication within genetically engineered bacteria are becoming increasingly important for monitoring environmental changes. Currently, there are a variety of mathematical models for understanding and predicting how genetically engineered bacteria respond to molecular stimuli in these environments, but as sensors have miniaturized towards microfluidics and are subjected to complex time-varying inputs, the shortcomings of these models have become apparent. The effects of microfluidic environments such as low oxygen concentration, increased biofilm encapsulation, diffusion limited molecular distribution, and higher population densities strongly affect rate constants for gene expression not accounted for in previous models. We report a mathematical model that accurately predicts the biological response of the autoinducer N-acyl homoserine lactone-mediated green fluorescent protein expression in reporter bacteria in microfluidic environments by accommodating these rate constants. This generalized mass action model considers a chain of biomolecular events from input autoinducer chemical to fluorescent protein expression through a series of six chemical species. We have validated this model against experimental data from our own apparatus as well as prior published experimental results. Results indicate accurate prediction of dynamics (e.g., 14% peak time error from a pulse input) and with reduced mean-squared error with pulse or step inputs for a range of concentrations (10 μM–30 μM). This model can help advance the design of genetically engineered bacteria sensors and molecular communication devices. PMID:25379076

Coupled Dictionary Learning for the Detail-Enhanced Synthesis of 3-D Facial Expressions.

PubMed

Liang, Haoran; Liang, Ronghua; Song, Mingli; He, Xiaofei

2016-04-01

The desire to reconstruct 3-D face models with expressions from 2-D face images fosters increasing interest in addressing the problem of face modeling. This task is important and challenging in the field of computer animation. Facial contours and wrinkles are essential to generate a face with a certain expression; however, these details are generally ignored or are not seriously considered in previous studies on face model reconstruction. Thus, we employ coupled radius basis function networks to derive an intermediate 3-D face model from a single 2-D face image. To optimize the 3-D face model further through landmarks, a coupled dictionary that is related to 3-D face models and their corresponding 3-D landmarks is learned from the given training set through local coordinate coding. Another coupled dictionary is then constructed to bridge the 2-D and 3-D landmarks for the transfer of vertices on the face model. As a result, the final 3-D face can be generated with the appropriate expression. In the testing phase, the 2-D input faces are converted into 3-D models that display different expressions. Experimental results indicate that the proposed approach to facial expression synthesis can obtain model details more effectively than previous methods can.

Effects of obesity on liver cytochromes P450 in various animal models.

PubMed

Tomankova, Veronika; Anzenbacher, Pavel; Anzenbacherova, Eva

2017-06-01

The prevalence of obesity and other obesity-related diseases is increasing worldwide. Obesity is a disease characterized by increased body weight, or a condition resulting from excessive accumulation of body fat. Due to increased body fat deposits, obesity has also been associated with increased mortality resulting from higher incidence rates of hypertension, diabetes, or various types of cancer, such as breast, colorectal, cervical and prostate cancer. Physiological changes associated with obesity are likely to result in altered drug biotransformation. The main enzymes enabling the oxidative biotransformation of most drugs are cytochromes P450 (CYPs). The review summarizes how pathophysiological factors, especially obesity, affect properties (e.g. enzyme activity, protein expression, gene expression) of CYP enzymes in various experimental models of human obesity. Results reported by various authors suggest that obesity is associated with a decrease of CYP activities (except for the CYP2C and CYP2E1 enzymes). The only exception is mouse obesity induced by monosodium glutamate (administered to newborn mice) as it usually leads to increased CYP expression. Selecting an animal model that is as close as possible to the properties of human obesity is of paramount importance.

Inhibited Carnitine Synthesis Causes Systemic Alteration of Nutrient Metabolism in Zebrafish

PubMed Central

Li, Jia-Min; Li, Ling-Yu; Qin, Xuan; Degrace, Pascal; Demizieux, Laurent; Limbu, Samwel M.; Wang, Xin; Zhang, Mei-Ling; Li, Dong-Liang; Du, Zhen-Yu

2018-01-01

Impaired mitochondrial fatty acid β-oxidation has been correlated with many metabolic syndromes, and the metabolic characteristics of the mammalian models of mitochondrial dysfunction have also been intensively studied. However, the effects of the impaired mitochondrial fatty acid β-oxidation on systemic metabolism in teleost have never been investigated. In the present study, we established a low-carnitine zebrafish model by feeding fish with mildronate as a specific carnitine synthesis inhibitor [0.05% body weight (BW)/d] for 7 weeks, and the systemically changed nutrient metabolism, including carnitine and triglyceride (TG) concentrations, fatty acid (FA) β-oxidation capability, and other molecular and biochemical assays of lipid, glucose, and protein metabolism, were measured. The results indicated that mildronate markedly decreased hepatic carnitine concentrations while it had no effect in muscle. Liver TG concentrations increased by more than 50% in mildronate-treated fish. Mildronate decreased the efficiency of liver mitochondrial β-oxidation, increased the hepatic mRNA expression of genes related to FA β-oxidation and lipolysis, and decreased the expression of lipogenesis genes. Mildronate decreased whole body glycogen content, increased glucose metabolism rate, and upregulated the expression of glucose uptake and glycolysis genes. Mildronate also increased whole body protein content and hepatic mRNA expression of mechanistic target of rapamycin (mtor), and decreased the expression of a protein catabolism-related gene. Liver, rather than muscle, was the primary organ targeted by mildronate. In short, mildronate-induced hepatic inhibited carnitine synthesis in zebrafish caused decreased mitochondrial FA β-oxidation efficiency, greater lipid accumulation, and altered glucose and protein metabolism. This reveals the key roles of mitochondrial fatty acid β-oxidation in nutrient metabolism in fish, and this low-carnitine zebrafish model could also be used as a novel fish model for future metabolism studies. PMID:29867554

Inhibited Carnitine Synthesis Causes Systemic Alteration of Nutrient Metabolism in Zebrafish.

PubMed

Li, Jia-Min; Li, Ling-Yu; Qin, Xuan; Degrace, Pascal; Demizieux, Laurent; Limbu, Samwel M; Wang, Xin; Zhang, Mei-Ling; Li, Dong-Liang; Du, Zhen-Yu

2018-01-01

Impaired mitochondrial fatty acid β-oxidation has been correlated with many metabolic syndromes, and the metabolic characteristics of the mammalian models of mitochondrial dysfunction have also been intensively studied. However, the effects of the impaired mitochondrial fatty acid β-oxidation on systemic metabolism in teleost have never been investigated. In the present study, we established a low-carnitine zebrafish model by feeding fish with mildronate as a specific carnitine synthesis inhibitor [0.05% body weight (BW)/d] for 7 weeks, and the systemically changed nutrient metabolism, including carnitine and triglyceride (TG) concentrations, fatty acid (FA) β-oxidation capability, and other molecular and biochemical assays of lipid, glucose, and protein metabolism, were measured. The results indicated that mildronate markedly decreased hepatic carnitine concentrations while it had no effect in muscle. Liver TG concentrations increased by more than 50% in mildronate-treated fish. Mildronate decreased the efficiency of liver mitochondrial β-oxidation, increased the hepatic mRNA expression of genes related to FA β-oxidation and lipolysis, and decreased the expression of lipogenesis genes. Mildronate decreased whole body glycogen content, increased glucose metabolism rate, and upregulated the expression of glucose uptake and glycolysis genes. Mildronate also increased whole body protein content and hepatic mRNA expression of mechanistic target of rapamycin ( mtor ), and decreased the expression of a protein catabolism-related gene. Liver, rather than muscle, was the primary organ targeted by mildronate. In short, mildronate-induced hepatic inhibited carnitine synthesis in zebrafish caused decreased mitochondrial FA β-oxidation efficiency, greater lipid accumulation, and altered glucose and protein metabolism. This reveals the key roles of mitochondrial fatty acid β-oxidation in nutrient metabolism in fish, and this low-carnitine zebrafish model could also be used as a novel fish model for future metabolism studies.

Exercise reverses OVA-induced inhibition of glucocorticoid receptor and increases anti-inflammatory cytokines in asthma.

PubMed

Silva, R A; Almeida, F M; Olivo, C R; Saraiva-Romanholo, B M; Martins, M A; Carvalho, C R F

2016-01-01

The purpose of this study was to determine the effect of aerobic exercise training (AT) on the expression of glucocorticoid receptors (GR) and anti-inflammatory cytokines in an asthma model. BALB/c mice were divided into groups control (CT; nonsensitized/nontrained), aerobic training (AT; nonsensitized/trained), ovalbumin (OVA; sensitized/not trained), and OVA+AT (sensitized/trained). OVA groups received OVA by inhalation, and the AT groups completed 1, 3, or 7 days of exercise (60 min/session). Expression of GR, IL-4, IL-5, IL-10, IL-1ra, NF-κB, TGF-β, VEGF, ICAM-1, VCAM-1; eosinophils counting; and airway remodeling (AR) features [airway smooth muscle (ASM) and epithelial thickness and collagen fiber deposition] were quantified. OVA sensitization induced a decrease in the expression of GR and increases in the eosinophil, IL-4, IL-5, NF-κB, TGF-β, VEGF, ICAM-1, VCAM-1, and AR features (P < 0.05). After 3 days, AT reversed the OVA-induced reduction in the expression of GR, and subsequently induced increases in the expression of IL-10 and IL-1ra (seventh day). In contrast, the eosinophil migration, the expression of NF-κB, IL-4, IL-5, TGF-β, RANTES, VEGF, ICAM-1, VCAM-1, and the AR features (P < 0.05) were reduced. AT increases the expression of GR and anti-inflammatory cytokines (IL-10 and IL-1ra) and reduces the expression of inflammatory mediators and airway inflammation in an animal model of asthma. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Enhancing biomass and ethanol production by increasing NADPH production in Synechocystis sp. PCC 6803.

PubMed

Choi, Yun-Nam; Park, Jong Moon

2016-08-01

This study demonstrates that increased NADPH production can improve biomass and ethanol production in cyanobacteria. We over-expressed the endogenous zwf gene, which encodes glucose-6-phosphate dehydrogenase of pentose phosphate pathway, in the model cyanobacterium Synechocystis sp. PCC 6803. zwf over-expression resulted in increased NADPH production, and promoted biomass production compared to the wild type in both autotrophic and mixotrophic conditions. Ethanol production pathway including NADPH-dependent alcohol dehydrogenase was also integrated with and without zwf over-expression. Excessive NADPH production by zwf over-expression could improve both biomass and ethanol production in the autotrophic conditions. Copyright © 2016 Elsevier Ltd. All rights reserved.

Transgenic expression of cyclooxygenase-2 (COX2) causes premature aging phenotypes in mice.

PubMed

Kim, Joohwee; Vaish, Vivek; Feng, Mingxiao; Field, Kevin; Chatzistamou, Ioulia; Shim, Minsub

2016-10-07

Cyclooxygenase (COX) is a key enzyme in the biosynthesis of prostanoids, lipid signaling molecules that regulate various physiological processes. COX2, one of the isoforms of COX, is highly inducible in response to a wide variety of cellular and environmental stresses. Increased COX2 expression is thought to play a role in the pathogenesis of many age-related diseases. COX2 expression is also reported to be increased in the tissues of aged humans and mice, which suggests the involvement of COX2 in the aging process. However, it is not clear whether the increased COX2 expression is causal to or a result of aging. We have now addressed this question by creating an inducible COX2 transgenic mouse model. Here we show that post-natal expression of COX2 led to a panel of aging-related phenotypes. The expression of p16, p53, and phospho-H2AX was increased in the tissues of COX2 transgenic mice. Additionally, adult mouse lung fibroblasts from COX2 transgenic mice exhibited increased expression of the senescence-associated β-galactosidase. Our study reveals that the increased COX2 expression has an impact on the aging process and suggests that modulation of COX2 and its downstream signaling may be an approach for intervention of age-related disorders.

JiangTang XiaoKe granule attenuates cathepsin K expression and improves IGF-1 expression in the bone of high fat diet induced KK-Ay diabetic mice.

PubMed

Guo, Yubo; Wang, Lili; Ma, Rufeng; Mu, Qianqian; Yu, Na; Zhang, Yi; Tang, Yuqing; Li, Yu; Jiang, Guangjian; Zhao, Dandan; Mo, Fangfang; Gao, Sihua; Yang, Meijuan; Kan, Feifei; Ma, Qun; Fu, Min; Zhang, Dongwei

2016-03-01

To assess the beneficial effects of JiangTang XiaoKe (JTXK) granule on the bone metabolism in high fat diet (HFD) fed KK-Ay diabetic mice. The KK-Ay mice were used as a diabetic model, while C57BL/6 mice were utilized as the non-diabetic control. The left tibia was used for determining bone mineral density (BMD) and bone ash coefficient. The HE and alizarin red S staining of femur were employed to evaluate bone pathology and calcium deposition. The expressions of alkaline phosphatase (ALP), insulin growth factor 1 (IGF-1) and cathepsin K were assessed by western blotting and immunohistochemical staining. JTXK granule significantly improved the bone ash coefficient, the distribution of trabecular bone and the calcification nodules deposition in KK-Ay mice with diabetes. IGF-1 and ALP expressions were significantly decreased, and cathepsin K expression was dramatically increased in the HFD fed KK-Ay diabetic model mice, which can be reversed by JTXK granule treatment. JTXK granule at medium or high dosage was more efficient in improving diabetic bone quality when compared with that in mice with a low dosage. However, the BMD values in each group of KK-Ay diabetic mice were not significantly different. We demonstrate that cathepsin K expression is increased in KK-Ay diabetic mouse model. JTXK granule treatment inhibits osteoclastic bone resorption and promotes the new bone formation by decreasing cathepsin K activity and increasing IGF-1 and ALP levels. These changes may contribute to the increase of bone strength and thus reducing the risk of bone fractures. Copyright © 2016 Elsevier Inc. All rights reserved.

Change of Rin1 and Stathmin in the Animal Model of Traumatic Stresses

PubMed Central

Han, Fang; Jiang, Jingzhi; Ding, Jinlan; Liu, Hong; Xiao, Bing; Shi, Yuxiu

2017-01-01

The molecular mechanism of fear memory is poorly understood. Therefore, the pathogenesis of post-traumatic stress disorder (PTSD), whose symptom presentation can enhance fear memory, remains largely unclear. Recent studies with knockout animals have reported that Rin1 and stathmin regulate fear memory. Rin1 inhibits acquisition and promotes memory extinction, whereas stathmin regulates innate and basal fear. The aim of our study was to examine changes in the expression of Rin1 and stathmin in different animal models of stress, particluarly traumatic stress. We used three animal traumatic stresses: single prolonged stress (SPS, which is a rodent model of PTSD), an immobilization-stress (IM) and a Loud sound stress (LSS), to examine the change and uniqueness in Rin1/stathmin expression. Behavioral tests of SPS rats demonstrated increased anxiety and contextual fear-conditioning. They showed decreased long-term potentiation (LTP), as well as decreased stathmin and increased Rin1 expression in the hippocampus and the amygdala. Expression of the stathmin effector, tubulin, and downstream molecules Rin1, Rab5, and Abl, appeared to increase. Rin1 and EphA4 were endogenously coexpressed in primary neurons after SPS stimulation. IM rats exhibited increased anxiety behavior and enhanced fear-conditioning to contextual and auditory stimuli. Similar changes in expression of Rin1/stathmin were observed in IM rats whereas no changes were observed in rats exposed to a loud sound. These data suggest that changes in expression of the Rin1 and stathmin genes may be involved in rodents with SPS and IM stresses, which provide valuable insight into fear memories under abnormal conditions, particularly in PTSD. PMID:28491025

Trichostatin A increases SMN expression and survival in a mouse model of spinal muscular atrophy

PubMed Central

Avila, Amy M.; Burnett, Barrington G.; Taye, Addis A.; Gabanella, Francesca; Knight, Melanie A.; Hartenstein, Parvana; Cizman, Ziga; Di Prospero, Nicholas A.; Pellizzoni, Livio; Fischbeck, Kenneth H.; Sumner, Charlotte J.

2007-01-01

The inherited motor neuron disease spinal muscular atrophy (SMA) is caused by mutation of the telomeric survival motor neuron 1 (SMN1) gene with retention of the centromeric SMN2 gene. We sought to establish whether the potent and specific hydroxamic acid class of histone deacetylase (HDAC) inhibitors activates SMN2 gene expression in vivo and modulates the SMA disease phenotype when delivered after disease onset. Single intraperitoneal doses of 10 mg/kg trichostatin A (TSA) in nontransgenic and SMA model mice resulted in increased levels of acetylated H3 and H4 histones and modest increases in SMN gene expression. Repeated daily doses of TSA caused increases in both SMN2-derived transcript and SMN protein levels in neural tissues and muscle, which were associated with an improvement in small nuclear ribonucleoprotein (snRNP) assembly. When TSA was delivered daily beginning on P5, after the onset of weight loss and motor deficit, there was improved survival, attenuated weight loss, and enhanced motor behavior. Pathological analysis showed increased myofiber size and number and increased anterior horn cell size. These results indicate that the hydroxamic acid class of HDAC inhibitors activates SMN2 gene expression in vivo and has an ameliorating effect on the SMA disease phenotype when administered after disease onset. PMID:17318264

Electroconvulsive stimulation (ECS) increases the expression of neuropeptide Y (NPY) in rat brains in a model of neuropathic pain: a quantitative real-time polymerase chain reaction (RT-PCR) study.

PubMed

Okabe, Tadashi; Sato, Chiyo; Matsumoto, Keisuke; Ozawa, Hitoshi; Sakamoto, Atsuhiro

2009-11-01

Electroconvulsive shock therapy (ECT) has been widely used as an effective and established treatment for refractory depression and schizophrenia. Some reports have shown that ECT is also effective for treating refractory neuropathic pain. In a rat model of neuropathic pain produced by chronic constrictive injury (CCI) of the sciatic nerve, thermal hyperalgesia, and mechanical allodynia were observed from day 2 after surgery. An electroconvulsive shock (ECS) was administered to rodents once daily for 6 days on days 7-12 after CCI operation using a pulse generator. Thermal and mechanical stimulation tests were performed to assess pain thresholds. Real-time polymerase chain reaction was used to measure the gene expression levels for 5HT(1A)R, 5HT(2A)R, neuropeptide Y (NPY), and GABAA(alpha1)R in the brain. After ECS, the latency to withdrawal from thermal stimulation was significantly increased; however, pain withdrawal thresholds in response to mechanical stimulation were not significantly changed. Expression ratios of NPY were significantly greater after ECS. Symptoms of neuropathic pain improved and expression of NPY in the brain was increased in CCI model rats after ECS, suggesting that changes in the expression of NPY in the brain may be related to the mechanism of action of ECT in treating neuropathic pain.

Changes in gravitational force induce alterations in gene expression that can be monitored in the live, developing zebrafish heart

NASA Astrophysics Data System (ADS)

Gillette-Ferguson, I.; Ferguson, D. G.; Poss, K. D.; Moorman, S. J.

2003-10-01

Little is known about the effect of microgravity on gene expression, particularly in vivo during embryonic development. Using transgenic zebrafish that express the gfp gene under the influence of a β-actin promoter, we examined the affect of simulated-microgravity on GFP expression in the heart. Zebrafish embryos, at the 18-20 somite-stage, were exposed to simulated-microgravity for 24 hours. The intensity of GFP fluorescence associated with the heart was then determined using fluorescence microscopy. Our measurements indicated that simulated-microgravity induced a 23.9% increase in GFP-associated fluorescence in the heart. In contrast, the caudal notochord showed a 17.5% increase and the embryo as a whole showed only an 8.5% increase in GFP-associated fluorescence. This suggests that there are specific effects on the heart causing the more dramatic increase. These studies indicate that microgravity can influence gene expression and demonstrate the usefulness of this in vivo model of "reporter-gene" expression for studying the effects of microgravity.

Overcoming food allergy through acquired tolerance conferred by transfer of Tregs in a murine model.

PubMed

Yamashita, H; Takahashi, K; Tanaka, H; Nagai, H; Inagaki, N

2012-02-01

The number of food allergy patients is increasing. Some children outgrow their food allergies through tolerance, whereas others remain susceptible throughout their lives. We aimed to contribute to food allergy therapeutics by understanding induction of oral tolerance in a murine food allergy model. We modified an existing murine food allergy model by using ovalbumin (OVA) to induce oral tolerance, either by pretreating mice with OVA or by transferring mesenteric lymph node (MLN) cells or T cells derived from mice treated with OVA. Pretreatment with OVA prevented food allergy, with complete suppression of OVA-specific immunoglobulin (Ig)E and IgA antibody production and interleukin (IL)-4, IL-10, and IL-9 mRNA expression. The proportion of regulatory T cells (Tregs) in MLN cells and expression of transforming growth factor-β mRNA increased. In the transfer model, anaphylaxis secondary to OVA intake was suppressed by transfer of whole MLN cells and Tregs from OVA-treated mice. However, OVA-specific IgE and IgA expressions were partially attenuated by transfer of antigen-specific and nonspecific Tregs, but not by whole MLN cells from OVA-treated mice. In the Treg transfer model, IL-4 and IL-10 mRNA expression decreased, but IL-9 mRNA expression increased. We concluded that oral tolerance for food antigens is induced in two ways: (i) by initial exposure to antigen, or inherent tolerance, and (ii) by transfer of Tregs, or acquired tolerance. Because food allergies occur when inherent tolerance is absent, understanding of acquired tolerance is important for the development of therapies for food allergy. © 2011 John Wiley & Sons A/S.

Intermittent Fasting Alleviates the Increase of Lipoprotein Lipase Expression in Brain of a Mouse Model of Alzheimer's Disease: Possibly Mediated by β-hydroxybutyrate.

PubMed

Zhang, Jingzhu; Li, Xinhui; Ren, Yahao; Zhao, Yue; Xing, Aiping; Jiang, Congmin; Chen, Yanqiu; An, Li

2018-01-01

Intermittent fasting has been demonstrated to protect against Alzheimer's disease (AD), however, the mechanism is unclear. Histone acetylation and lipoprotein lipase (LPL) are involved in AD progression. Importantly, LPL has been documented to be regulated by histone deacetylases (HDACs) inhibitors (increase histone acetylation level) in adipocyte and mesenchymal stem cells, or by fasting in adipose and muscle tissues. In brain, however, whether histone acetylation or fasting regulates LPL expression is unknown. This study was designed to demonstrate intermittent fasting may protect against AD through increasing β-hydroxybutyrate, a HDACs inhibitor, to regulate LPL. We also investigated microRNA-29a expression associating with regulation of LPL and histone acetylation. The results showed LPL mRNA expression was increased and microRNA-29a expression was decreased in the cerebral cortex of AD model mice (APP/PS1), which were alleviated by intermittent fasting. No significant differences were found in the total expression of LPL protein (brain-derived and located in capillary endothelial cells from peripheral tissues) in the cerebral cortex of APP/PS1 mice. Further study indicated that LPL located in capillary endothelial cells was decreased in the cerebral cortex of APP/PS1 mice, which was alleviated by intermittent fasting. LPL and microRNA-29a expression were separately increased and down-regulated in 2 μM Aβ 25-35 -exposed SH-SY5Y cells, but respectively decreased and up-regulated in 10 μM Aβ 25-35 -exposed cells, which were all reversed by β-hydroxybutyrate. The increase of HDAC2/3 expression and the decrease of acetylated H3K9 and H4K12 levels were alleviated in APP/PS1 mice by intermittent fasting treatment, as well in 2 or 10 μM Aβ 25-35 -exposed cells by β-hydroxybutyrate treatment. These findings above suggested the results from APP/PS1 mice were consistent with those from cells treated with 2 μM Aβ 25-35 . Interestingly, LPL expression was reduced (0.2-folds) and microRNA-29a expression was up-regulated (1.7-folds) in HDAC2-silenced cells, but respectively increased (1.3-folds) and down-regulated (0.8-folds) in HDAC3-silenced cells. Furthermore, LPL expression was decreased in cells treated with microRNA-29a mimic and increased with inhibitor treatment. In conclusion, intermittent fasting inhibits the increase of brain-derived LPL expression in APP/PS1 mice partly through β-hydroxybutyrate-mediated down-regulation of microRNA-29a expression. HDAC2/3 may be implicated in the effect of β-hydroxybutyrate on microRNA-29a expression.

Multi-cellular human bronchial models exposed to diesel exhaust particles: assessment of inflammation, oxidative stress and macrophage polarization.

PubMed

Ji, Jie; Upadhyay, Swapna; Xiong, Xiaomiao; Malmlöf, Maria; Sandström, Thomas; Gerde, Per; Palmberg, Lena

2018-05-02

Diesel exhaust particles (DEP) are a major component of outdoor air pollution. DEP mediated pulmonary effects are plausibly linked to inflammatory and oxidative stress response in which macrophages (MQ), epithelial cells and their cell-cell interaction plays a crucial role. Therefore, in this study we aimed at studying the cellular crosstalk between airway epithelial cells with MQ and MQ polarization following exposure to aerosolized DEP by assessing inflammation, oxidative stress, and MQ polarization response markers. Lung mucosa models including primary bronchial epithelial cells (PBEC) cultured at air-liquid interface (ALI) were co-cultured without (PBEC-ALI) and with MQ (PBEC-ALI/MQ). Cells were exposed to 12.7 μg/cm 2 aerosolized DEP using XposeALI ® . Control (sham) models were exposed to clean air. Cell viability was assessed. CXCL8 and IL-6 were measured in the basal medium by ELISA. The mRNA expression of inflammatory markers (CXCL8, IL6, TNFα), oxidative stress (NFKB, HMOX1, GPx) and MQ polarization markers (IL10, IL4, IL13, MRC1, MRC2 RETNLA, IL12 andIL23) were measured by qRT-PCR. The surface/mRNA expression of TLR2/TLR4 was detected by FACS and qRT-PCR. In PBEC-ALI exposure to DEP significantly increased the secretion of CXCL8, mRNA expression of inflammatory markers (CXCL8, TNFα) and oxidative stress markers (NFKB, HMOX1, GPx). However, mRNA expressions of these markers (CXCL8, IL6, NFKB, and HMOX1) were reduced in PBEC-ALI/MQ models after DEP exposure. TLR2 and TLR4 mRNA expression increased after DEP exposure in PBEC-ALI. The surface expression of TLR2 and TLR4 on PBEC was significantly reduced in sham-exposed PBEC-ALI/MQ compared to PBEC-ALI. After DEP exposure surface expression of TLR2 was increased on PBEC of PBEC-ALI/MQ, while TLR4 was decreased in both models. DEP exposure resulted in similar expression pattern of TLR2/TLR4 on MQ as in PBEC. In PBEC-ALI/MQ, DEP exposure increased the mRNA expression of anti-inflammatory M2 macrophage markers (IL10, IL4, IL13, MRC1, MRC2). The cellular interaction of PBEC with MQ in response to DEP plays a pivotal role for MQ phenotypic alteration towards M2-subtypes, thereby promoting an efficient resolution of the inflammation. Furthermore, this study highlighted the fact that cell-cell interaction using multicellular ALI-models combined with an in vivo-like inhalation exposure system is critical in better mimicking the airway physiology compared with traditional cell culture systems.

Deciphering the details of RNA aminoglycoside interactions: from atomistic models to biotechnological applications

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ilgu, Muslum

A detailed study was done of the neomycin-B RNA aptamer for determining its selectivity and binding ability to both neomycin– and kanamycin-class aminoglycosides. A novel method to increase drug concentrations in cells for more efficiently killing is described. To test the method, a bacterial model system was adopted and several small RNA molecules interacting with aminoglycosides were cloned downstream of T7 RNA polymerase promoter in an expression vector. Then, the growth analysis of E. coli expressing aptamers was observed for 12-hour period. Our analysis indicated that aptamers helped to increase the intracellular concentration of aminoglycosides thereby increasing their efficacy.

Curcumin modulates the effect of histone modification on the expression of chemokines by type II alveolar epithelial cells in a rat COPD model.

PubMed

Gan, Lixing; Li, Chengye; Wang, Jian; Guo, Xuejun

2016-01-01

Studies have suggested that histone modification has a positive impact on various aspects associated with the progression of COPD. Histone deacetylase 2 (HDAC2) suppresses proinflammatory gene expression through deacetylation of core histones. To investigate the effect of histone modification on the expression of chemokines in type II alveolar epithelial cells (AEC II) in a rat COPD model and regulation of HDAC2 expression by curcumin in comparison with corticosteroid. The rat COPD model was established by cigarette smoke exposure and confirmed by histology and pathophysioloy. AEC II were isolated and cultured in vitro from the COPD models and control animals. The cells were treated with curcumin, corticosteroid, or trichostatin A, and messenger RNA (mRNA) expression of interleukin-8 (IL-8), monocyte chemoattractant protein-1 (MCP-1), and macrophage inflammatory protein-2α (MIP-2α) was assessed by quantitative real-time polymerase chain reaction (RT-PCR). The expression of HDAC2 was measured by Western blot. Chromatin immunoprecipitation was used to detect H3/H4 acetylation and H3K9 methylation in the promoter region of three kinds of chemokine genes (IL-8, MCP-1, and MIP-2α). Compared to the control group, the mRNAs of MCP-1, IL-8, and MIP-2α were upregulated 4.48-fold, 3.14-fold, and 2.83-fold, respectively, in the AEC II from COPD model. The protein expression of HDAC2 in the AEC II from COPD model was significantly lower than from the control group ( P <0.05). The decreased expression of HDAC2 was negatively correlated with the increased expression of IL-8, MCP-1, and MIP-2α mRNAs (all P <0.05). The level of H3/H4 acetylation was higher but H3K9 methylation in the promoter region of chemokine genes was lower in the cells from COPD model than from the control group (all P <0.05). Curcumin downregulated the expression of MCP-1, IL-8, and MIP-2α, and the expression was further enhanced in the presence of corticosteroid. Moreover, curcumin restored HDAC2 expression, decreased the levels of H3/H4 acetylation, and increased H3K9 methylation in the promoter region of chemokine in the presence or absence of dexamethasone (all P <0.05). Curcumin may suppress chemokines and restore corticosteroid resistance in COPD through modulating HDAC2 expression and its effect on histone modification.

Effects of "nourishing liver and kidney" acupuncture therapy on expression of brain derived neurotrophic factor and synaptophysin after cerebral ischemia reperfusion in rats.

PubMed

Xia, Wen-Guang; Zheng, Chan-Juan; Zhang, Xuan; Wang, Juan

2017-04-01

The aim of the present study was to investigate the effect of "nourishing liver and kidney" acupuncture therapy on motor and cognitive deficits, and the underlying mechanism following cerebral ischemia-reperfusion (I/R) via increasing the expression of brain derived neurotrophic factor (BDNF) and synaptophysin (SYN) in the hippocampus. Healthy adult male SD rats were randomly divided into sham operation group (n=51), model group (n=51), acupuncture group (n=51) and acupuncture control group (n=51). The middle cerebral I/R model was established. Acupunctures were performed in the acupuncture group and acupuncture control group at acupoints of Taixi (K103), Taichong (ST09) of both sides, for 30 min once daily every morning. The animals in the sham operation group and model group were conventionally fed in the cage, without any intervention therapy. The rats of each group were assessed with modified neurological severity scores (mNSS). The expression of BDNF and SYN in the hippocampus was detected by immunohistochemical SP method and the synaptic structure in hippocampus area was assessed morphologically and quantitatively at the 3rd, 7th and 14th day. The Morris water Maze (MWM) test was used to evaluate the rats' learning and memory abilities on the 15th day after acupuncture. The animals in the acupuncture control group and sham operation group presented no neurological deficit. In the acupuncture group, the nerve functional recovery was significantly better than that in the model group at the 7th and 14th day after modeling. The average MWM escape latency in the acupuncture group was shorter than that in the model group at the 3rd, 4th and 5th day. The number of crossings of the platform quadrant in the acupuncture group was significantly more than that in the model group. At the each time point, the expression levels of BDNF and SYN in the hippocampal regions increased significantly in the model group as compared with the sham operation group and the acupuncture control group. In the acupuncture group, the expression levels of BDNF at the 7th and 14th day increased more significantly than those in the model group. In the acupuncture group, the expression levels of SYN at the each time point increased more significantly than those in the model group. The post-synaptic density (PSD) was significantly increased and the synapse cleft width was narrowed in the acupuncture group as compared with other groups. The synaptic curvatures were improved obviously in the acupuncture group in contrast to the model group. It was concluded that the "nourishing liver and kidney" acupuncture therapy has positive effects on behavioral recovery, as well as learning and memory abilities, probably by promoting the expression of BDNF and SYN, and synaptic structure reconstruction in the ipsilateral hippocampus after I/R in rats. The "nourishing liver and kidney" acupuncture therapy can promote the functional recovery in rats after cerebral ischemia injury.

Increased cholecystokinin labeling in the hippocampus of a mouse model of epilepsy maps to spines and glutamatergic terminals

PubMed Central

Wyeth, Megan S.; Zhang, Nianhui; Houser, Carolyn R.

2011-01-01

The neuropeptide cholecystokinin (CCK) is abundant in the central nervous system and expressed in a subset of inhibitory interneurons, particularly in their axon terminals. The expression profile of CCK undergoes numerous changes in several models of temporal lobe epilepsy. Previous studies in the pilocarpine model of epilepsy have shown that CCK immunohistochemical labeling is substantially reduced in several regions of the hippocampal formation, consistent with decreased CCK expression as well as selective neuronal degeneration. However, in a mouse pilocarpine model of recurrent seizures, increases in CCK-labeling also occur and are especially striking in the hippocampal dendritic layers of strata oriens and radiatum. Characterizing these changes and determining the cellular basis of the increased labeling were the major goals of the current study. One possibility was that the enhanced CCK labeling could be associated with an increase in GABAergic terminals within these regions. However, in contrast to the marked increase in CCK-labeled structures, labeling of GABAergic axon terminals was decreased in the dendritic layers. Likewise, cannabinoid receptor 1-labeled axon terminals, many of which are CCK-containing GABAergic terminals, were also decreased. These findings suggested that the enhanced CCK labeling was not due to an increase in GABAergic axon terminals. The subcellular localization of CCK immunoreactivity was then examined using electron microscopy, and the identities of the structures that formed synaptic contacts were determined. In pilocarpine-treated mice, CCK was observed in dendritic spines and these were proportionally increased relative to controls, whereas the proportion of CCK-labeled terminals forming symmetric synapses was decreased. In addition, CCK-positive axon terminals forming asymmetric synapses were readily observed in these mice. Double labeling with vesicular glutamate transporter 1 and CCK revealed co-localization in numerous terminals forming asymmetric synapses, confirming the glutamatergic identity of these terminals. These data raise the possibility that expression of CCK is increased in hippocampal pyramidal cells in mice with recurrent, spontaneous seizures. PMID:22155653

Hypoxia/Aglycemia-Induced Endothelial Barrier Dysfunction and Tight Junction Protein Downregulation Can Be Ameliorated by Citicoline

PubMed Central

Pan, Qunwen; Zhao, Yuhui; Chen, Ji; Zhao, Bin; Chen, Yanfang

2013-01-01

This study explores the effect of citicoline on the permeability and expression of tight junction proteins (TJPs) in endothelial cells under hypoxia/aglycemia conditions. Hypoxia or oxygen and glucose deprivation (OGD) was utilized to induce endothelial barrier breakdown model on human umbilical vein endothelial cells (HUVECs) and mouse brain microvascular endothelial cells (bEnd.3s). The effect of citicoline on endothelial barrier breakdown models was determined at either low or high concentrations. FITC-Dextran flux was used to examine the endothelial permeability. The expression of TJPs was measured by immunofluorescence, Real-time PCR and Western Blot methods. Results showed that hypoxia or OGD increased the permeability of HUVECs accompanied with down-regulation of occludens-1 (ZO-1) and occludin at both mRNA and protein levels. Similarly in bEnd.3s, hypoxia increased the permeability and decreased the expression of ZO-1 and claudin-5. Citicoline treatment dose-dependently decreased the permeability in these two models, which paralleled with elevated expression of TJPs. The data demonstrate that citicoline restores the barrier function of endothelial cells compromised by hypoxia/aglycemia probably via up-regulating the expression of TJPs. PMID:24358213

Dietary diindolylmethane suppresses inflammation-driven lung squamous cell carcinoma in mice

PubMed Central

Song, Jung Min; Qian, Xuemin; Teferi, Fitsum; Pan, Jing; Wang, Yian; Kassie, Fekadu

2014-01-01

Inflammatory conditions of the lung such as chronic obstructive pulmonary disease (COPD) are known to increase lung cancer risk, particularly lung squamous cell carcinoma (LSCC). In the present study, we developed a mouse model of inflammation-driven LSCC that was induced by N-nitroso-trischloroethylurea (NTCU) and enhanced by lipopolysaccharide (LPS), a potent proinflammatory agent contained in tobacco and tobacco smoke, and determined the chemopreventive effects of BioResponse diindolylmethane (DIM) in the same model. Compared to mice treated with NTCU alone, mice treated with the combination of NTCU and LPS had a 9-fold increase in the number of bronchioles with LSCC. Also, compared to mice treated with LPS alone, mice treated with NTCU plus LPS showed significantly increased expression of the inflammatory cytokines IL-1α, IL-6, and TNFα (all three increased about 7-fold). Parallel to the increased cytokine gene expression, the NTCU plus LPS-treated group exhibited significantly enhanced activation of NF-κB, STAT3, ERK, p-38, and Akt, expression of p53, COX-2, and Mcl-1, and NF-κB- and STAT3-DNA binding in the lung. Dietary administration of DIM (10 µmol/g diet or 2460 ppm) to mice treated with NTCU plus LPS reduced the incidence of LSCC by 2-fold, suppressed activation/expression of proinflammatory and procarcinogenic proteins and NF-κB- and STAT3-DNA binding, but not the expression of cytokines and p53. This study highlights the potential significance of our mouse model to identify promising drugs or dietary agents for the chemoprevention of human LSCC and that DIM is a very good candidate for clinical lung cancer chemoprevention trials. PMID:25403850

Sex differences in pain-related behavior and expression of calcium/calmodulin-dependent protein kinase II in dorsal root ganglia of rats with diabetes type 1 and type 2.

PubMed

Ferhatovic, Lejla; Banozic, Adriana; Kostic, Sandra; Sapunar, Damir; Puljak, Livia

2013-06-01

Sex differences in pain-related behavior and expression of calcium/calmodulin dependent protein kinase II (CaMKII) in dorsal root ganglia were studied in rat models of Diabetes mellitus type 1 (DM1) and type 2 (DM2). DM1 was induced with 55mg/kg streptozotocin, and DM2 with a combination of high-fat diet and 35mg/kg of streptozotocin. Pain-related behavior was analyzed using thermal and mechanical stimuli. The expression of CaMKII was analyzed with immunofluorescence. Sexual dimorphism in glycemia, and expression of CaMKII was observed in the rat model of DM1, but not in DM2 animals. Increased expression of total CaMKII (tCaMKII) in small-diameter dorsal root ganglia neurons, which are associated with nociception, was found only in male DM1 rats. None of the animals showed increased expression of the phosphorylated alpha CaMKII isoform in small-diameter neurons. The expression of gamma and delta isoforms of CaMKII remained unchanged in all analyzed animal groups. Different patterns of glycemia and tCaMKII expression in male and female model of DM1 were not associated with sexual dimorphism in pain-related behavior. The present findings do not suggest sex-related differences in diabetic painful peripheral neuropathy in male and female diabetic rats. Copyright © 2012 Elsevier GmbH. All rights reserved.

Effects of Exercise on AMPK Signaling and Downstream Components to PI3K in Rat with Type 2 Diabetes

PubMed Central

Cao, Shicheng; Li, Bowen; Yi, Xuejie; Chang, Bo; Zhu, Beibei; Lian, Zhenzhen; Zhang, Zhaoran; Zhao, Gang; Liu, Huili; Zhang, He

2012-01-01

Exercise can increase skeletal muscle sensitivity to insulin, improve insulin resistance and regulate glucose homeostasis in rat models of type 2 diabetes. However, the potential mechanism remains poorly understood. In this study, we established a male Sprague–Dawley rat model of type 2 diabetes, with insulin resistance and β cell dysfunction, which was induced by a high-fat diet and low-dose streptozotocin to replicate the pathogenesis and metabolic characteristics of type 2 diabetes in humans. We also investigated the possible mechanism by which chronic and acute exercise improves metabolism, and the phosphorylation and expression of components of AMP-activated protein kinase (AMPK) and downstream components of phosphatidylinositol 3-kinase (PI3K) signaling pathways in the soleus. As a result, blood glucose, triglyceride, total cholesterol, and free fatty acid were significantly increased, whereas insulin level progressively declined in diabetic rats. Interestingly, chronic and acute exercise reduced blood glucose, increased phosphorylation and expression of AMPKα1/2 and the isoforms AMPKα1 and AMPKα2, and decreased phosphorylation and expression of AMPK substrate, acetyl CoA carboxylase (ACC). Chronic exercise upregulated phosphorylation and expression of AMPK upstream kinase, LKB1. But acute exercise only increased LKB1 expression. In particular, exercise reversed the changes in protein kinase C (PKC)ζ/λ phosphorylation, and PKCζ phosphorylation and expression. Additionally, exercise also increased protein kinase B (PKB)/Akt1, Akt2 and GLUT4 expression, but AS160 protein expression was unchanged. Chronic exercise elevated Akt (Thr308) and (Ser473) and AS160 phosphorylation. Finally, we found that exercise increased peroxisome proliferator-activated receptor-γ coactivator 1 (PGC1) mRNA expression in the soleus of diabetic rats. These results indicate that both chronic and acute exercise influence the phosphorylation and expression of components of the AMPK and downstream to PIK3 (aPKC, Akt), and improve GLUT4 trafficking in skeletal muscle. These data help explain the mechanism how exercise regulates glucose homeostasis in diabetic rats. PMID:23272147

Induction of autophagy by ARHI (DIRAS3) alters fundamental metabolic pathways in ovarian cancer models.

PubMed

Ornelas, Argentina; McCullough, Christopher R; Lu, Zhen; Zacharias, Niki M; Kelderhouse, Lindsay E; Gray, Joshua; Yang, Hailing; Engel, Brian J; Wang, Yan; Mao, Weiqun; Sutton, Margie N; Bhattacharya, Pratip K; Bast, Robert C; Millward, Steven W

2016-10-26

Autophagy is a bulk catabolic process that modulates tumorigenesis, therapeutic resistance, and dormancy. The tumor suppressor ARHI (DIRAS3) is a potent inducer of autophagy and its expression results in necroptotic cell death in vitro and tumor dormancy in vivo. ARHI is down-regulated or lost in over 60 % of primary ovarian tumors yet is dramatically up-regulated in metastatic disease. The metabolic changes that occur during ARHI induction and their role in modulating death and dormancy are unknown. We employed Nuclear Magnetic Resonance (NMR)-based metabolomic strategies to characterize changes in key metabolic pathways in both cell culture and xenograft models of ARHI expression and autophagy. These pathways were further interrogated by cell-based immunofluorescence imaging, tracer uptake studies, targeted metabolic inhibition, and in vivo PET/CT imaging. Induction of ARHI in cell culture models resulted in an autophagy-dependent increase in lactate production along with increased glucose uptake and enhanced sensitivity to glycolytic inhibitors. Increased uptake of glutamine was also dependent on autophagy and dramatically sensitized cultured ARHI-expressing ovarian cancer cell lines to glutaminase inhibition. Induction of ARHI resulted in a reduction in mitochondrial respiration, decreased mitochondrial membrane potential, and decreased Tom20 staining suggesting an ARHI-dependent loss of mitochondrial function. ARHI induction in mouse xenograft models resulted in an increase in free amino acids, a transient increase in [ 18 F]-FDG uptake, and significantly altered choline metabolism. ARHI expression has previously been shown to trigger autophagy-associated necroptosis in cell culture. In this study, we have demonstrated that ARHI expression results in decreased cellular ATP/ADP, increased oxidative stress, and decreased mitochondrial function. While this bioenergetic shock is consistent with programmed necrosis, our data indicates that the accompanying up-regulation of glycolysis and glutaminolysis is autophagy-dependent and serves to support cell viability rather than facilitate necroptotic cell death. While the mechanistic basis for metabolic up-regulation following ARHI induction is unknown, our preliminary data suggest that decreased mitochondrial function and increased metabolic demand may play a role. These alterations in fundamental metabolic pathways during autophagy-associated necroptosis may provide the basis for new therapeutic strategies for the treatment of dormant ovarian tumors.

Constitutive TL1A expression under colitogenic conditions modulates the severity and location of gut mucosal inflammation and induces fibrostenosis.

PubMed

Barrett, Robert; Zhang, Xiaolan; Koon, Hon Wai; Vu, Michelle; Chang, Jyh-Yau; Yeager, Nicole; Nguyen, Mary Ann; Michelsen, Kathrin S; Berel, Dror; Pothoulakis, Charalabos; Targan, Stephan R; Shih, David Q

2012-02-01

Intestinal fibrostenosis is a hallmark of severe Crohn's disease and can lead to multiple surgeries. Patients with certain TNFSF15 variants overexpress TL1A. The aim of this study was to determine the effect of TL1A overexpression on intestinal inflammation and the development of fibrostenosis. We assessed the in vivo consequences of constitutive TL1A expression on gut mucosal inflammation and fibrostenosis using two murine models of chronic colitis. In the dextran sodium sulfate (DSS) and adoptive T-cell transfer models, there was proximal migration of colonic inflammation, worsened patchy intestinal inflammation, and long gross intestinal strictures in Tl1a transgenic compared to wild-type littermates. In the DSS model, myeloid- and T-cell-expressing Tl1a transgenic mice had increased T-cell activation markers and interleukin-17 expression compared to wild-type mice. In the T-cell transfer model, Rag1(-/-) mice receiving Tl1a transgenic T cells had increased interferon-γ expression but reduced T-helper 17 cells and IL-17 production. Narrowed ureters with hydronephrosis were found only in the Tl1a transgenic mice in all chronic colitis models. In human translational studies, Crohn's disease patients with higher peripheral TL1A expression also exhibited intestinal fibrostenosis and worsened ileocecal inflammation with relative sparing of rectosigmoid inflammation. These data show that TL1A is an important cytokine that not only modulates the location and severity of mucosal inflammation, but also induces fibrostenosis. Copyright © 2012 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

PARP10 (ARTD10) modulates mitochondrial function

PubMed Central

Nagy, Lilla; Vida, András; Kis, Gréta; Brunyánszki, Attila; Antal, Miklós; Lüscher, Bernhard; Bai, Péter

2018-01-01

Poly(ADP-ribose) polymerase (PARP)10 is a PARP family member that performs mono-ADP-ribosylation of target proteins. Recent studies have linked PARP10 to metabolic processes and metabolic regulators that prompted us to assess whether PARP10 influences mitochondrial oxidative metabolism. The depletion of PARP10 by specific shRNAs increased mitochondrial oxidative capacity in cellular models of breast, cervical, colorectal and exocrine pancreas cancer. Upon silencing of PARP10, mitochondrial superoxide production decreased in line with increased expression of antioxidant genes pointing out lower oxidative stress upon PARP10 silencing. Improved mitochondrial oxidative capacity coincided with increased AMPK activation. The silencing of PARP10 in MCF7 and CaCo2 cells decreased the proliferation rate that correlated with increased expression of anti-Warburg enzymes (Foxo1, PGC-1α, IDH2 and fumarase). By analyzing an online database we showed that lower PARP10 expression increases survival in gastric cancer. Furthermore, PARP10 expression decreased upon fasting, a condition that is characterized by increases in mitochondrial biogenesis. Finally, lower PARP10 expression is associated with increased fatty acid oxidation. PMID:29293500

Altered Left Ventricular Ion Channel Transcriptome in a High-Fat-Fed Rat Model of Obesity: Insight into Obesity-Induced Arrhythmogenesis

PubMed Central

Yon, Marianne; Pickavance, Lucy; Yanni Gerges, Joseph; Davis, Gershan; Wilding, John; Jian, Kun; Hart, George; Boyett, Mark

2016-01-01

Introduction. Obesity is increasingly common and is associated with an increased prevalence of cardiac arrhythmias. The aim of this study was to see whether in obesity there is proarrhythmic gene expression of ventricular ion channels and related molecules. Methods and Results. Rats were fed on a high-fat diet and compared to control rats on a normal diet (n = 8). After 8 weeks, rats on the high-fat diet showed significantly greater weight gain and higher adiposity. Left ventricle samples were removed at 8 weeks and mRNA expression of ion channels and other molecules was measured using qPCR. Obese rats had significant upregulation of Cav1.2, HCN4, Kir2.1, RYR2, NCX1, SERCA2a, and RYR2 mRNA and downregulation of ERG mRNA. In the case of HCN4, it was confirmed that there was a significant increase in protein expression. The potential effects of the mRNA changes on the ventricular action potential and intracellular Ca2+ transient were predicted using computer modelling. Modelling predicted prolongation of the ventricular action potential and an increase in the intracellular Ca2+ transient, both of which would be expected to be arrhythmogenic. Conclusion. High-fat diet causing obesity results in arrhythmogenic cardiac gene expression of ion channels and related molecules. PMID:27747100

[Effects of electroacupuncture at "Jiaji" (EX-B 2) on myocardial apoptosis and the expression of Bcl-2 and Bax in rats with ischemia-reperfusion injury].

PubMed

Kong, Su-Ping; Zhang, Xin; Tan, Qi-Wen

2013-06-01

To explore the protective mechanism of acupuncture on myocardial ischemia-reperfusion injury. Fifty Wistar rats were randomly divided into a sham-operation group, a model group, a Jiaji group , a Neiguan group and a Yanglingquan group, 10 rats in each group. The model of myocardial ischemia-reperfusion injury was duplicated by ligating the left anterior descending coronary artery (LADCA) in the later four groups, and the LADCA was not ligated in the sham-operation group. The rats in the treatment groups were treated with electroacupuncture at "Jiaji" (EX-B 2), "Neiguan" (PC 6) and "Yanglingquan" (GB 34) on both sides, respectively, once a day for 7 days. No interventions were produced in the sham-operation group and model group. Myocardial apoptosis were examined by the TUNEL method. The expressions of Bcl-2 and Bax proteins were measured by immunohistochemical method. Apoptosis index(AI) was significantly lower in the Jiaji group, Neiguan group and sham-operation group compared with model group (P < 0.01). Compared with the sham-operation group, AI were significantly increased in other groups (all P < 0.01). There was no significant difference in AI between Jiaji and Neiguan group (P > 0.05). Compared with the model group, the expression of Bcl-2 was significantly increased and Bax protein was significantly decreased in Jiaji group and Neiguan group (both P < 0.01). The expressions of Bcl-2 and Bax proteins were significantly increased in other groups compared with the sham-operation group (all P < 0.01). There was no significant difference in the expression of Bcl-2 and Bax proteins between Jiaji and Neiguan group (P > 0.05). Electroacupuncture at both Jiaji (EX-B 2) and Neiguan (PC 6) has protective effects on myocardial ischemia-reperfusion injury, and the mechanism is closely related to inhibiting myocardial apoptosis by adjusting the expression of Bcl-2 and Bax.

Genetic control of ColE1 plasmid stability that is independent of plasmid copy number regulation.

PubMed

Standley, Melissa S; Million-Weaver, Samuel; Alexander, David L; Hu, Shuai; Camps, Manel

2018-06-16

ColE1-like plasmid vectors are widely used for expression of recombinant genes in E. coli. For these vectors, segregation of individual plasmids into daughter cells during cell division appears to be random, making them susceptible to loss over time when no mechanisms ensuring their maintenance are present. Here we use the plasmid pGFPuv in a recA relA strain as a sensitized model to study factors affecting plasmid stability in the context of recombinant gene expression. We find that in this model, plasmid stability can be restored by two types of genetic modifications to the plasmid origin of replication (ori) sequence: point mutations and a novel 269 nt duplication at the 5' end of the plasmid ori, which we named DAS (duplicated anti-sense) ori. Combinations of these modifications produce a range of copy numbers and of levels of recombinant expression. In direct contradiction with the classic random distribution model, we find no correlation between increased plasmid copy number and increased plasmid stability. Increased stability cannot be explained by reduced levels of recombinant gene expression either. Our observations would be more compatible with a hybrid clustered and free-distribution model, which has been recently proposed based on detection of individual plasmids in vivo using super-resolution fluorescence microscopy. This work suggests a role for the plasmid ori in the control of segregation of ColE1 plasmids that is distinct from replication initiation, opening the door for the genetic regulation of plasmid stability as a strategy aimed at enhancing large-scale recombinant gene expression or bioremediation.

ACE Over Expression in Myelomonocytic Cells: Effect on a Mouse Model of Alzheimer's Disease

PubMed Central

Koronyo-Hamaoui, Maya; Shah, Kandarp; Koronyo, Yosef; Bernstein, Ellen; Giani, Jorge F.; Janjulia, Tea; Black, Keith L.; Shi, Peng D.; Gonzalez-Villalobos, Romer A.; Fuchs, Sebastien; Shen, Xiao Z.; Bernstein, Kenneth E.

2014-01-01

While it is well known that angiotensin converting enzyme (ACE) plays an important role in blood pressure control, ACE also has effects on renal function, hematopoiesis, reproduction, and aspects of the immune response. ACE 10/10 mice over express ACE in myelomonocytic cells. Macrophages from these mice have an increased polarization towards a pro-inflammatory phenotype that results in a very effective immune response to challenge by tumors or bacterial infection. In a mouse model of Alzheimer's disease (AD), the ACE 10/10 phenotype provides significant protection against AD pathology, including reduced inflammation, reduced burden of the neurotoxic amyloid-β protein and preserved cognitive function. Taken together, these studies show that increased myelomonocytic ACE expression in mice alters the immune response to better defend against many different types of pathologic insult, including the cognitive decline observed in an animal model of AD. PMID:24792094

Withaferin A effectively targets soluble vimentin in the glaucoma filtration surgical model of fibrosis.

PubMed

Bargagna-Mohan, Paola; Deokule, Sunil P; Thompson, Kyle; Wizeman, John; Srinivasan, Cidambi; Vooturi, Sunil; Kompella, Uday B; Mohan, Royce

2013-01-01

Withaferin A (WFA) is a natural product that binds to soluble forms of the type III intermediate filament (IF) vimentin. Currently, it is unknown under what pathophysiological contexts vimentin is druggable, as cytoskeltal vimentin-IFs are abundantly expressed. To investigate druggability of vimentin, we exploited rabbit Tenon's capsule fibroblast (RbTCF) cell cultures and the rabbit glaucoma filtration surgical (GFS) model of fibrosis. WFA potently caused G₀/G₁ cell cycle inhibition (IC₅₀ 25 nM) in RbTCFs, downregulating ubiquitin E3 ligase skp2 and inducing p27(Kip1) expression. Transforming growth factor (TGF)-ß-induced myofibroblast transformation caused development of cell spheroids with numerous elongated invadopodia, which WFA blocked potently by downregulating soluble vimentin and α-smooth muscle actin (SMA) expression. In the pilot proof-of-concept study using the GFS model, subconjunctival injections of a low WFA dose reduced skp2 expression in Tenon's capsule and increased p27(Kip1) expression without significant alteration to vimentin-IFs. This treatment maintains significant nanomolar WFA concentrations in anterior segment tissues that correspond to WFA's cell cycle targeting activity. A ten-fold higher WFA dose caused potent downregulation of soluble vimentin and skp2 expression, but as found in cell cultures, no further increase in p27(Kip1) expression was observed. Instead, this high WFA dose potently induced vimentin-IF disruption and downregulated α-SMA expression that mimicked WFA activity in TGF-ß-treated RbTCFs that blocked cell contractile activity at submicromolar concentrations. These findings illuminate that localized WFA injection to ocular tissues exerts pharmacological control over the skp2-p27(Kip1) pathway by targeting of soluble vimentin in a model of surgical fibrosis.

Mechanisms altering airway smooth muscle cell Ca+ homeostasis in two asthma models.

PubMed

Kellner, Julia; Tantzscher, Juliane; Oelmez, Hamza; Edelmann, Martin; Fischer, Rainald; Huber, Rudolf Maria; Bergner, Albrecht

2008-01-01

Asthma is characterized by airway remodeling, altered mucus production and airway smooth muscle cell (ASMC) contraction causing extensive airway narrowing. In particular, alterations of ASMC contractility seem to be of crucial importance. The elevation of the cytoplasmic Ca(2+) concentration is a key event leading to ASMC contraction and changes in the agonist-induced Ca(2+) increase in ASMC have been reported in asthma. The aim of this study was to investigate mechanisms underlying these changes. Murine tracheal smooth muscle cells (MTSMC) from T-bet KO mice and human bronchial smooth muscle cells (HBSMC) incubated with IL-13 and IL-4 served as asthma models. Acetylcholine-induced changes in the cytoplasmic Ca(2+) concentration were recorded using fluorescence microscopy and the expression of Ca(2+) homeostasis regulating proteins was investigated with Western blot analysis. Acetylcholine-induced Ca(2+) transients were elevated in both asthma models. This correlated with an increased Ca(2+) content of the sarcoplasmic reticulum (SR). In MTSMC from T-bet KO mice, the expression of the SR Ca(2+) buffers calreticulin and calsequestrin was higher compared to wild-type mice. In HBSMC incubated with IL-13 or IL-4, the expression of ryanodine receptors, inositol-3-phosphate receptors and sarcoplasmic/endoplasmic reticulum Ca(2+) ATPases 2 was increased compared to HBSMC without incubation with interleukins. The enlarged acetylcholine-induced Ca(2+) transients could be reversed by blocking inositol-3-phosphate receptors. We conclude that in the murine asthma model the SR Ca(2+) buffer capacity is increased, while in the human asthma model the expression of SR Ca(2+) channels is altered. The investigation of the Ca(2+) homeostasis of ASMC has the potential to provide new therapeutical options in asthma. Copyright 2008 S. Karger AG, Basel.

Melatonin attenuates scopolamine-induced cognitive impairment via protecting against demyelination through BDNF-TrkB signaling in the mouse dentate gyrus.

PubMed

Chen, Bai Hui; Park, Joon Ha; Lee, Tae-Kyeong; Song, Minah; Kim, Hyunjung; Lee, Jae Chul; Kim, Young-Myeong; Lee, Choong-Hyun; Hwang, In Koo; Kang, Il Jun; Yan, Bing Chun; Won, Moo-Ho; Ahn, Ji Hyeon

2018-04-01

Animal models of scopolamine-induced amnesia are widely used to study underlying mechanisms and treatment of cognitive impairment in neurodegenerative diseases such as Alzheimer's disease (AD). Previous studies have identified that melatonin improves cognitive dysfunction in animal models. In this study, using a mouse model of scopolamine-induced amnesia, we assessed spatial and short-term memory functions for 4 weeks, investigated the expression of myelin-basic protein (MBP) in the dentate gyrus, and examined whether melatonin and scopolamine cotreatment could keep cognitive function and MBP expression. In addition, to study functions of melatonin for keeping cognitive function and MBP expression, we examined expressions of brain-derived neurotrophic factor (BDNF) and tropomycin receptor kinase B (TrkB) in the mouse dentate gyrus. Scopolamine (1 mg/kg) and melatonin (10 mg/kg) were intraperitoneally treated for 2 and 4 weeks. Two and 4 weeks after scopolamine treatment, mice showed significant cognitive impairment; however, melatonin and scopolamine cotreatment recovered cognitive impairment. Two and 4 weeks of scopolamine treatment, the density of MBP immunoreactive myelinated nerve fibers was significantly decreased in the dentate gyrus; however, scopolamine and melatonin cotreatment significantly increased the scopolamine-induced reduction of MBP expression in the dentate gyrus. Furthermore, the cotreatment of scopolamine and melatonin significantly increased the scopolamine-induced decrease of BDNF and TrKB immunoreactivity in the dentate gyrus. Taken together, our results indicate that melatonin treatment exerts anti-amnesic effect and restores the scopolamine-induced reduction of MBP expression through increasing BDNF and TrkB expressions in the mouse dentate gyrus. Copyright © 2018 Elsevier B.V. All rights reserved.

Dynamics and reproductive effects of complement factors in the spontaneous abortion model of CBA/J×DBA/2 mice.

PubMed

Takeshita, Ai; Kusakabe, Ken Takeshi; Hiyama, Masato; Kuniyoshi, Nobue; Kondo, Tomohiro; Kano, Kiyoshi; Kiso, Yasuo; Okada, Toshiya

2014-05-01

The complement system is one component of innate immunity that could participate in fetal loss. We have already reported that adipsin, a complement activator in the alternative pathway, is stably expressed in the placenta and that an increase in this expression is related to spontaneous abortion. However, complement inhibitor Crry was concurrently expressed in the placenta, and the role of complement factors during pregnancy was not clear. In the present study, we examined the endogenous regulation of complement factors in placenta and serum by using another model mouse for spontaneous abortion and studied the effect of exogenous complement disruption on pregnancy. Compared to control mice, the CBA/J×DBA/2 model mice had higher expression levels of adipsin in the placenta and serum. Adipsin and complement C3 were localized in the metrial gland and labyrinth regions, and both positive reactive ranges were limited in the maternal blood current in normal implantation sites. These results suggest that extrauterine adipsin hematogenously reaches the placenta, activates complement C3, and promotes destruction of the feto-maternal barrier in aborted implantation sites. Crry was consistently expressed in the placenta and serum and reduced in the resorption sites of CBA/J×DBA/2 mice as compared to normal sites. Injection of recombinant adipsin increased the resorption rate and changed the expression of Th-type cytokines toward a Th1 bias. The present study indicates that adipsin could induce the fetal loss that accompanies the Th1 bias and may be a crucial cause of spontaneous abortion. In addition, the local expression of Crry prevents complement activation in placenta in response to a systemic increase of adipsin. Copyright © 2014 Elsevier GmbH. All rights reserved.

Increased expression of deleted in malignant brain tumors (DMBT1) gene in precancerous gastric lesions: Findings from human and animal studies.

PubMed

Garay, Jone; Piazuelo, M Blanca; Lopez-Carrillo, Lizbeth; Leal, Yelda A; Majumdar, Sumana; Li, Li; Cruz-Rodriguez, Nataly; Serrano-Gomez, Silvia J; Busso, Carlos S; Schneider, Barbara G; Delgado, Alberto G; Bravo, Luis E; Crist, Angela M; Meadows, Stryder M; Camargo, M Constanza; Wilson, Keith T; Correa, Pelayo; Zabaleta, Jovanny

2017-07-18

Helicobacter pylori infection triggers a cascade of inflammatory stages that may lead to the appearance of non-atrophic gastritis, multifocal atrophic, intestinal metaplasia, dysplasia, and cancer. Deleted in malignant brain tumors 1 (DMBT1) belongs to the group of secreted scavenger receptor cysteine-rich proteins and is considered to be involved in host defense by binding to pathogens. Initial studies showed its deletion and loss of expression in a variety of tumors but the role of this gene in tumor development is not completely understood. Here, we examined the role of DMBT1 in gastric precancerous lesions in Caucasian, African American and Hispanic individuals as well as in the development of gastric pathology in a mouse model of H. pylori infection. We found that in 3 different populations, mucosal DMBT1 expression was significantly increased (2.5 fold) in individuals with dysplasia compared to multifocal atrophic gastritis without intestinal metaplasia; the increase was also observed in individuals with advanced gastritis and positive H. pylori infection. In our animal model, H. pylori infection of Dmbt1-/- mice resulted in significantly higher levels of gastritis, more extensive mucous metaplasia and reduced Il33 expression levels in the gastric mucosa compared to H. pylori-infected wild type mice. Our data in the animal model suggest that in response to H. pylori infection DMBT1 may mediate mucosal protection reducing the risk of developing gastric precancerous lesions. However, the increased expression in human gastric precancerous lesions points to a more complex role of DMBT1 in gastric carcinogenesis.

Increased expression of deleted in malignant brain tumors (DMBT1) gene in precancerous gastric lesions: Findings from human and animal studies

PubMed Central

Garay, Jone; Piazuelo, M. Blanca; Lopez-Carrillo, Lizbeth; Leal, Yelda A; Majumdar, Sumana; Li, Li; Cruz-Rodriguez, Nataly; Serrano-Gomez, Silvia J; Busso, Carlos S; Schneider, Barbara G; Delgado, Alberto G; Bravo, Luis E; Crist, Angela M; Meadows, Stryder M; Camargo, M. Constanza; Wilson, Keith T; Correa, Pelayo; Zabaleta, Jovanny

2017-01-01

Helicobacter pylori infection triggers a cascade of inflammatory stages that may lead to the appearance of non-atrophic gastritis, multifocal atrophic, intestinal metaplasia, dysplasia, and cancer. Deleted in malignant brain tumors 1 (DMBT1) belongs to the group of secreted scavenger receptor cysteine-rich proteins and is considered to be involved in host defense by binding to pathogens. Initial studies showed its deletion and loss of expression in a variety of tumors but the role of this gene in tumor development is not completely understood. Here, we examined the role of DMBT1 in gastric precancerous lesions in Caucasian, African American and Hispanic individuals as well as in the development of gastric pathology in a mouse model of H. pylori infection. We found that in 3 different populations, mucosal DMBT1 expression was significantly increased (2.5 fold) in individuals with dysplasia compared to multifocal atrophic gastritis without intestinal metaplasia; the increase was also observed in individuals with advanced gastritis and positive H. pylori infection. In our animal model, H. pylori infection of Dmbt1−/− mice resulted in significantly higher levels of gastritis, more extensive mucous metaplasia and reduced Il33 expression levels in the gastric mucosa compared to H. pylori-infected wild type mice. Our data in the animal model suggest that in response to H. pylori infection DMBT1 may mediate mucosal protection reducing the risk of developing gastric precancerous lesions. However, the increased expression in human gastric precancerous lesions points to a more complex role of DMBT1 in gastric carcinogenesis. PMID:28423364

Increased CD147 (EMMPRIN) expression in the rat brain following traumatic brain injury.

PubMed

Wei, Ming; Li, Hong; Shang, Yanguo; Zhou, Ziwei; Zhang, Jianning

2014-10-17

The extracellular matrix metalloproteinase inducer (EMMPRIN), or CD147, has been known to play a key regulatory role in vascular permeability and leukocyte activation by inducing the expression of matrix metalloproteinases (MMPs). The effects of traumatic brain injury on the expression of EMMPRIN remain poorly understood. In this study, we investigated changes in EMMPRIN expression in a rat model of fluid percussion injury (FPI) and examined the potential association between EMMPRIN and MMP-9 expression. Adult male rats were subjected to FPI. EMMPRIN expression was markedly up-regulated in the brain tissue surrounding the injured region 6-48 h after TBI, as measured by immunoblot and immunohistochemistry. EMMPRIN expression was localized to inflammatory cells. The increase in EMMPRIN expression was temporally correlated with an increase in MMP-9 levels. These data demonstrate, for the first time, changes in CD147 and MMP-9 expression following TBI. These data also suggest that CD147 and MMP-9 may play a role in vascular injuries after TBI. Copyright © 2014 Elsevier B.V. All rights reserved.

Prenatal administration of retinoic acid increases the trophoblastic insulin-like growth factor 2 protein expression in the nitrofen model of congenital diaphragmatic hernia.

PubMed

Kutasy, Balazs; Friedmacher, Florian; Duess, Johannes W; Puri, Prem

2014-02-01

The high mortality rate in congenital diaphragmatic hernia (CDH) is attributed to pulmonary hypoplasia (PH). Insulin-like growth factor 2 (IGF2) is an important regulator of fetal growth. The highest levels of IGF2 expression are found in the placenta, which are negatively regulated by decidual retinoid acid receptor alpha (RARα). It has been demonstrated that prenatal administration of retinoic acid (RA) suppresses decidual RARα expression. Previous studies have further shown that prenatal administration of RA can reverse PH in nitrofen-induced CDH model. In IGF2 knockout animals, low levels of IGF2 are associated with decreased placental growth and PH. We therefore hypothesized that nitrofen decreases trophoblastic IGF2 expression and prenatal administration of RA increases it through decidual RARα in the nitrofen-induced CDH model. Pregnant rats were exposed to either olive oil or nitrofen on day 9 of gestation (D9). RA was given intraperitoneally on D18, D19 and D20. Fetuses were harvested on D21 and divided into three groups: control, CDH and nitrofen+RA. Immunohistochemistry was performed to evaluate decidual RARα and trophoblastic IGF2 expression. Protein levels of IGF2 in serum, intra-amniotic fluid and left lungs were measured by enzyme-linked immunosorbent assay. Significant growth retardation of placenta and left lungs was observed in the CDH group compared to control and nitrofen+RA group. Markedly increased decidual RARα and decreased IGF2 immunoreactivity were found in the CDH group compared to control and nitrofen+RA group. Significantly decreased IGF2 protein levels were detected in serum, intra-amniotic fluid and left lungs in the CDH group compared to control and nitrofen+RA group. Our findings suggest that nitrofen may disturb trophoblastic IGF2 expression through decidual RARα resulting in retarded placental growth and PH in the nitrofen-induced CDH. Prenatal administration of RA may promote lung and placental growth by increasing trophoblastic IGF2 expression.

Ethanolic extract Ocimum sanctum. Enhances cognitive ability from young adulthood to middle aged mediated by increasing choline acetyl transferase activity in rat model.

PubMed

Kusindarta, Dwi Liliek; Wihadmadyatami, Hevi; Jadi, Arvendi R; Karnati, Srikanth; Lochnit, Guenter; Hening, Puspa; Haryanto, Aris; Auriva, Made B; Purwaningrum, Medania

2018-06-01

Patients with dementia are increasing steadily, cognitive impairment by dementia not only exclusively suffers by old people but also young to middle aged individuals. However, the mechanism of cognitive impairment occurs in young people is not understood. Further, current medication to impairment did not provide satisfactory results. Therefore, we investigated the potential role of Ocimum sanctum ethanolic extract to enhance cognitive ability in the rat in vivo model. Young to middle aged rats were divided into 3 groups (3, 6, 9 months old) were treated with (0, 50 and 100 mg/kg b.w.) O. sanctum for 45 days. We employed a behavioral assay to assess cognitive ability. Further, Nissl staining was performed to analyze hippocampus formation in dentate gyrus (DG), cornu ammonis 1 (CA1), cornu ammonis 3 (CA3). The expression and activity of ChAT in brain was analyzed by RT-PCR and ELISA. Our results showed that treatment of O. sanctum with a dosage of 100 mg/kg b.w. for 45 days induced the cognitive ability in nine months old rats. Further, we observed a significant increase in density of granular and pyramidal cells in DG, CA1, and CA3. These results were corroborated by an increase in the ChAT activity and gene expression in the rat model as well as HEK 293 cell culture model. Taken together, the administration of 100 mg/kg b.w. O.sanctum induced the expression of ChAT. The increased ChAT expression and activity may enhance the cognitive ability in 9 months old rats mimicking young and middle aged condition in humans. Copyright © 2018 Elsevier Ltd. All rights reserved.

Cardiovascular-renal and metabolic characterization of a rat model of polycystic ovary syndrome.

PubMed

Yanes, Licy L; Romero, Damian G; Moulana, Mohaddetheh; Lima, Roberta; Davis, Deborah D; Zhang, Huimin; Lockhart, Rachel; Racusen, Lorraine C; Reckelhoff, Jane F

2011-04-01

Polycystic ovary syndrome (PCOS) is the most common reproductive dysfunction in premenopausal women. PCOS is also associated with increased risk of cardiovascular disease when PCOS first occurs and later in life. Hypertension, a common finding in women with PCOS, is a leading risk factor for cardiovascular disease. The mechanisms responsible for hypertension in women with PCOS have not been elucidated. This study characterized the cardiovascular-renal consequences of hyperandrogenemia in a female rat model. Female Sprague-Dawley rats (aged 4-6 weeks) were implanted with dihydrotestosterone or placebo pellets lasting 90 days. After 10 to 12 weeks, blood pressure (by radiotelemetry), renal function (glomerular filtration rate, morphology, protein, and albumin excretion), metabolic parameters (plasma insulin, glucose, leptin, cholesterol, and oral glucose tolerance test), inflammation (plasma tumor necrosis factor-α), oxidative stress (mRNA expression of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase subunits, p22(phox), p47(phox), gp91(phox), and NOX4), nitrate/nitrite excretion and mRNA expression of components of the renin-angiotensin system (angiotensinogen, angiotensin-I-converting enzyme [ACE], and AT1 receptor) were determined. Plasma dihydrotestosterone increased 3-fold in hyperandrogenemic female (HAF) rats, whereas plasma estradiol levels did not differ compared with control females. HAF rats exhibited estrus cycle dysfunction. They also had increased food intake and body weight, increased visceral fat, glomerular filtration rate, renal injury, insulin resistance and metabolic dysfunction, oxidative stress, and increased expression of angiotensinogen and ACE and reduced AT1 receptor expression. The HAF rat is a unique model that exhibits many of the characteristics of PCOS in women and is a useful model to study the mechanisms responsible for PCOS-mediated hypertension. Copyright © 2011 Elsevier HS Journals, Inc. All rights reserved.

Colorectal cancer cells suppress CD4+ T cells immunity through canonical Wnt signaling.

PubMed

Sun, Xuan; Liu, Suoning; Wang, Daguang; Zhang, Yang; Li, Wei; Guo, Yuchen; Zhang, Hua; Suo, Jian

2017-02-28

Understanding how colorectal cancer escapes from immunosurveillance and immune attack is important for developing novel immunotherapies for colorectal cancer. In this study we evaluated the role of canonical Wnt signaling in the regulation of T cell function in a mouse colorectal cancer model. We found that colorectal cancer cells expressed abundant Wnt ligands, and intratumoral T cells expressed various Frizzled proteins. Meanwhile, both active β-catenin and total β-catenin were elevated in intratumoral T cells. In vitro study indicated that colorectal cancer cells suppressed IFN-γ expression and increased IL-17a expression in activated CD4+ T cells. However, the cytotoxic activity of CD8+ T cells was not altered by colorectal cancer cells. To further evaluate the importance of Wnt signaling for CD4+ T cell-mediated cancer immunity, β-catenin expression was enforced in CD4+ T cells using lentiviral transduction. In an adoptive transfer model, enforced expression of β-catenin in intratumoral CD4+ T cells increased IL-17a expression, enhanced proliferation and inhibited apoptosis of colorectal cancer cells. Taken together, our study disclosed a new mechanism by which colorectal cancer impairs T cell immunity.

Carbon Monoxide Releasing Molecule-A1 (CORM-A1) Improves Neurogenesis: Increase of Neuronal Differentiation Yield by Preventing Cell Death.

PubMed

Almeida, Ana S; Soares, Nuno L; Vieira, Melissa; Gramsbergen, Jan Bert; Vieira, Helena L A

2016-01-01

Cerebral ischemia and neurodegenerative diseases lead to impairment or death of neurons in the central nervous system. Stem cell based therapies are promising strategies currently under investigation. Carbon monoxide (CO) is an endogenous product of heme degradation by heme oxygenase (HO) activity. Administration of CO at low concentrations produces several beneficial effects in distinct tissues, namely anti-apoptotic and anti-inflammatory. Herein the CO role on modulation of neuronal differentiation was assessed. Three different models with increasing complexity were used: human neuroblastoma SH-S5Y5 cell line, human teratocarcinoma NT2 cell line and organotypic hippocampal slice cultures (OHSC). Cell lines were differentiated into post-mitotic neurons by treatment with retinoic acid (RA) supplemented with CO-releasing molecule A1 (CORM-A1). CORM-A1 positively modulated neuronal differentiation, since it increased final neuronal production and enhanced the expression of specific neuronal genes: Nestin, Tuj1 and MAP2. Furthermore, during neuronal differentiation process, there was an increase in proliferative cell number (ki67 mRNA expressing cells) and a decrease in cell death (lower propidium iodide (PI) uptake, limitation of caspase-3 activation and higher Bcl-2 expressing cells). CO supplementation did not increase the expression of RA receptors. In the case of SH-S5Y5 model, small amounts of reactive oxygen species (ROS) generation emerges as important signaling molecules during CO-promoted neuronal differentiation. CO's improvement of neuronal differentiation yield was validated using OHSC as ex vivo model. CORM-A1 treatment of OHSC promoted higher levels of cells expressing the neuronal marker Tuj1. Still, CORM-A1 increased cell proliferation assessed by ki67 expression and also prevented cell death, which was followed by increased Bcl-2 expression, decreased levels of active caspase-3 and PI uptake. Likewise, ROS signaling emerged as key factors in CO's increasing number of differentiated neurons in OHSC. In conclusion, CO's increasing number of differentiated neurons is a novel biological role disclosed herein. CO improves neuronal yield due to its capacity to reduce cell death, promoting an increase in proliferative population. However, one cannot disregard a direct CO's effect on specific cellular processes of neuronal differentiation. Further studies are needed to evaluate how CO can potentially modulate cell mechanisms involved in neuronal differentiation. In summary, CO appears as a promising therapeutic molecule to stimulate endogenous neurogenesis or to improve in vitro neuronal production for cell therapy strategies.

Carbon Monoxide Releasing Molecule-A1 (CORM-A1) Improves Neurogenesis: Increase of Neuronal Differentiation Yield by Preventing Cell Death

PubMed Central

Almeida, Ana S.; Soares, Nuno L.; Vieira, Melissa; Gramsbergen, Jan Bert

2016-01-01

Cerebral ischemia and neurodegenerative diseases lead to impairment or death of neurons in the central nervous system. Stem cell based therapies are promising strategies currently under investigation. Carbon monoxide (CO) is an endogenous product of heme degradation by heme oxygenase (HO) activity. Administration of CO at low concentrations produces several beneficial effects in distinct tissues, namely anti-apoptotic and anti-inflammatory. Herein the CO role on modulation of neuronal differentiation was assessed. Three different models with increasing complexity were used: human neuroblastoma SH-S5Y5 cell line, human teratocarcinoma NT2 cell line and organotypic hippocampal slice cultures (OHSC). Cell lines were differentiated into post-mitotic neurons by treatment with retinoic acid (RA) supplemented with CO-releasing molecule A1 (CORM-A1). CORM-A1 positively modulated neuronal differentiation, since it increased final neuronal production and enhanced the expression of specific neuronal genes: Nestin, Tuj1 and MAP2. Furthermore, during neuronal differentiation process, there was an increase in proliferative cell number (ki67 mRNA expressing cells) and a decrease in cell death (lower propidium iodide (PI) uptake, limitation of caspase-3 activation and higher Bcl-2 expressing cells). CO supplementation did not increase the expression of RA receptors. In the case of SH-S5Y5 model, small amounts of reactive oxygen species (ROS) generation emerges as important signaling molecules during CO-promoted neuronal differentiation. CO’s improvement of neuronal differentiation yield was validated using OHSC as ex vivo model. CORM-A1 treatment of OHSC promoted higher levels of cells expressing the neuronal marker Tuj1. Still, CORM-A1 increased cell proliferation assessed by ki67 expression and also prevented cell death, which was followed by increased Bcl-2 expression, decreased levels of active caspase-3 and PI uptake. Likewise, ROS signaling emerged as key factors in CO’s increasing number of differentiated neurons in OHSC. In conclusion, CO’s increasing number of differentiated neurons is a novel biological role disclosed herein. CO improves neuronal yield due to its capacity to reduce cell death, promoting an increase in proliferative population. However, one cannot disregard a direct CO’s effect on specific cellular processes of neuronal differentiation. Further studies are needed to evaluate how CO can potentially modulate cell mechanisms involved in neuronal differentiation. In summary, CO appears as a promising therapeutic molecule to stimulate endogenous neurogenesis or to improve in vitro neuronal production for cell therapy strategies. PMID:27144388

Effects of an Antimutagenic 1,4-Dihydropyridine AV-153 on Expression of Nitric Oxide Synthases and DNA Repair-related Enzymes and Genes in Kidneys of Rats with a Streptozotocin Model of Diabetes Mellitus.

PubMed

Ošiņa, Kristīne; Rostoka, Evita; Isajevs, Sergejs; Sokolovska, Jelizaveta; Sjakste, Tatjana; Sjakste, Nikolajs

2016-11-01

Development of complications of diabetes mellitus (DM), including diabetic nephropathy, is a complex multi-stage process, dependent on many factors including the modification of nitric oxide (NO) production and an impaired DNA repair. The goal of this work was to study in vivo effects of 1,4-dihydropyridine AV-153, known as antimutagen and DNA binder, on the expression of several genes and proteins involved in NO metabolism and DNA repair in the kidneys of rats with a streptozotocin (STZ)-induced model of DM. Transcription intensity was monitored by means of real-time RT-PCR and the expression of proteins by immunohistochemistry. Development of DM significantly induced PARP1 protein expression, while AV-153 (0.5 mg/kg) administration decreased it. AV-153 increased the expression of Parp1 gene in the kidneys of both intact and diabetic animals. Expression of H2afx mRNA and γH2AX histone protein, a marker of DNA breakage, was not changed in diabetic animals, but AV-153 up-regulated the expression of the gene without any impact on the protein expression. Development of DM was followed by a significant increase in iNOS enzyme expression, while AV-153 down-regulated the enzyme expression up to normal levels. iNos gene expression was also found to be increased in diabetic animals, but unlike the protein, the expression of mRNA was found to be enhanced by AV-153 administration. Expression of both eNOS protein and eNos gene in the kidneys was down-regulated, and the administration of AV-153 normalized the expression level. The effects of the compound in the kidneys of diabetic animals appear to be beneficial, as a trend for the normalization of expression of NO synthases is observed. © 2016 Nordic Association for the Publication of BCPT (former Nordic Pharmacological Society).

Enhancement of SMN protein levels in a mouse model of spinal muscular atrophy using novel drug-like compounds

PubMed Central

Cherry, Jonathan J; Osman, Erkan Y; Evans, Matthew C; Choi, Sungwoon; Xing, Xuechao; Cuny, Gregory D; Glicksman, Marcie A; Lorson, Christian L; Androphy, Elliot J

2013-01-01

Spinal muscular atrophy (SMA) is a neurodegenerative disease that causes progressive muscle weakness, which primarily targets proximal muscles. About 95% of SMA cases are caused by the loss of both copies of the SMN1 gene. SMN2 is a nearly identical copy of SMN1, which expresses much less functional SMN protein. SMN2 is unable to fully compensate for the loss of SMN1 in motor neurons but does provide an excellent target for therapeutic intervention. Increased expression of functional full-length SMN protein from the endogenous SMN2 gene should lessen disease severity. We have developed and implemented a new high-throughput screening assay to identify small molecules that increase the expression of full-length SMN from a SMN2 reporter gene. Here, we characterize two novel compounds that increased SMN protein levels in both reporter cells and SMA fibroblasts and show that one increases lifespan, motor function, and SMN protein levels in a severe mouse model of SMA. PMID:23740718

Epistemological beliefs of physics undergraduate and graduate students and faculty in the context of a well-structured and an ill-structured problem

NASA Astrophysics Data System (ADS)

Mercan, Fatih C.

This study examines epistemological beliefs of physics undergraduate and graduate students and faculty in the context of solving a well-structured and an ill-structured problem. The data collection consisted of a think aloud problem solving session followed by a semi-structured interview conducted with 50 participants, 10 participants at freshmen, seniors, masters, PhD, and faculty levels. The data analysis involved (a) identification of the range of beliefs about knowledge in the context of the well-structured and the ill-structured problem solving, (b) construction of a framework that unites the individual beliefs identified in each problem context under the same conceptual base, and (c) comparisons of the problem contexts and expertise level groups using the framework. The results of the comparison of the contexts of the well-structured and the ill-structured problem showed that (a) authoritative beliefs about knowledge were expressed in the well-structured problem context, (b) relativistic and religious beliefs about knowledge were expressed in the ill-structured problem context, and (c) rational, empirical, modeling beliefs about knowledge were expressed in both problem contexts. The results of the comparison of the expertise level groups showed that (a) undergraduates expressed authoritative beliefs about knowledge more than graduate students and faculty did not express authoritative beliefs, (b) faculty expressed modeling beliefs about knowledge more than graduate students and undergraduates did not express modeling beliefs, and (c) there were no differences in rational, empirical, experiential, relativistic, and religious beliefs about knowledge among the expertise level groups. As the expertise level increased the number of participants who expressed authoritative beliefs about knowledge decreased and the number of participants who expressed modeling based beliefs about knowledge increased. The results of this study implied that existing developmental and cognitive models of personal epistemology can explain personal epistemology in physics to a limited extent, however, these models cannot adequately account for the variation of epistemological beliefs across problem contexts. Modeling beliefs about knowledge emerged as a part of personal epistemology and an indicator of epistemological sophistication, which do not develop until extensive experience in the field. Based on these findings, the researcher recommended providing opportunities for practicing model construction for students.

Irradiated fibroblasts promote epithelial–mesenchymal transition and HDGF expression of esophageal squamous cell carcinoma

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bao, Ci-Hang; Wang, Xin-Tong; Ma, Wei

2015-03-06

Recent evidence suggested that nonirradiated cancer-associated fibroblasts (CAFs) promoted aggressive phenotypes of cancer cells through epithelial–mesenchymal transition (EMT). Hepatoma-derived growth factor (HDGF) is a radiosensitive gene of esophageal squamous cell carcinoma (ESCC). This study aimed to investigate the effect of irradiated fibroblasts on EMT and HDGF expression of ESCC. Our study demonstrated that coculture with nonirradiated fibroblasts significantly increased the invasive ability of ESCC cells and the increased invasiveness was further accelerated when they were cocultured with irradiated fibroblasts. Scattering of ESCC cells was also accelerated by the supernatant from irradiated fibroblasts. Exposure of ESCC cells to supernatant from irradiatedmore » fibroblasts resulted in decreased E-cadherin, increased vimentin in vitro and β-catenin was demonstrated to localize to the nucleus in tumor cells with irradiated fibroblasts in vivo models. The expression of HDGF and β-catenin were increased in both fibroblasts and ESCC cells of irradiated group in vitro and in vivo models. Interestingly, the tumor cells adjoining the stromal fibroblasts displayed strong nuclear HDGF immunoreactivity, which suggested the occurrence of a paracrine effect of fibroblasts on HDGF expression. These data suggested that irradiated fibroblasts promoted invasion, growth, EMT and HDGF expression of ESCC. - Highlights: • Irradiated CAFs accelerated invasiveness and scattering of ESCC cell lines. • Irradiated CAFs promoted EMT of ESCC cells. • Irradiated fibroblasts induced nuclear β-catenin relocalization in ESCC cells. • Irradiated fibroblasts increased HDGF expression in vitro and in vivo.« less

[Effect of electroacupuncture on expression of ionotropic glutamate receptor subunits and their genes in lumbar segments of spinal cord in rats with neuropathic pain].

PubMed

Ma, Cheng; Yu, Li; Yan, Li-ping

2010-12-01

To observe the effect of electroacupuncture (EA) on the expression of ionotropic glutamate receptor (iGluR) subunits and their mRNAs in the lumbar segments of spinal cord in rats with neuropathic pain, so as to explore its underlying mechanism in relieving spinal hyperalgesia. Thirty SD rats were randomly divided into control, model, and EA groups, with 10 rats in each. The spared nerve injury (SNI) model was established by ligature of the sural nerve after cutting off the common peroneal nerve and anterior tibial nerve. EA (2 Hz, 1 mA) was applied to "Huantiao" (GB 30) and "Weizhong" (BL 40) for 30 min, once daily for 7 days. Mechanical pain threshold was detected before and after modeling and before and after EA treatment. The expression levels of N-methyl-d-aspartic acid (NMDA) receptor subunits NR1 and NR 2 B,and AMPA receptor subunit GluR 1 of iGluR and their genes were assayed by Western blot and reverse transcription polymerase chain reaction (RT-PCR) separately. In comparison with control group, the mechanical pain thresholds were decreased significantly on day 2, 7 and day 14 following modeling in the model group (P < 0.05, P < 0.01). While compared with the model group, the pain threshold was increased considerably on day 14 in the EA group (P < 0.01). Compared with the control group, the expression levels of lumbar spinal cord NR 2 B and NR 2 B mRNA in the model group were increased significantly (P < 0.05), and those of lumbar spinal cord NR 1 and NR 1 mRNA, GluR 1 and GluR 1 mRNA in the model group increased slightly (P > 0.05). In comparison with the model group, the expression levels of lumbar spinal cord NR 2 B and NR 2 B mRNA in the EA group were downregulated remarkably (P < 0.05), and those of lumbar spinal cord NR 1 and NR 1 mRNA, GluR 1 and GluR 1 mRNA in the EA group down-regulated slightly (P > 0.05). EA can significantly suppress pain reaction in rats with neuropathic pain probably through down-regulating the expression of lumbar spinal cord NR 2 B protein and NR 2 B mRNA.

Mimicking emotions: how 3-12-month-old infants use the facial expressions and eyes of a model.

PubMed

Soussignan, Robert; Dollion, Nicolas; Schaal, Benoist; Durand, Karine; Reissland, Nadja; Baudouin, Jean-Yves

2018-06-01

While there is an extensive literature on the tendency to mimic emotional expressions in adults, it is unclear how this skill emerges and develops over time. Specifically, it is unclear whether infants mimic discrete emotion-related facial actions, whether their facial displays are moderated by contextual cues and whether infants' emotional mimicry is constrained by developmental changes in the ability to discriminate emotions. We therefore investigate these questions using Baby-FACS to code infants' facial displays and eye-movement tracking to examine infants' looking times at facial expressions. Three-, 7-, and 12-month-old participants were exposed to dynamic facial expressions (joy, anger, fear, disgust, sadness) of a virtual model which either looked at the infant or had an averted gaze. Infants did not match emotion-specific facial actions shown by the model, but they produced valence-congruent facial responses to the distinct expressions. Furthermore, only the 7- and 12-month-olds displayed negative responses to the model's negative expressions and they looked more at areas of the face recruiting facial actions involved in specific expressions. Our results suggest that valence-congruent expressions emerge in infancy during a period where the decoding of facial expressions becomes increasingly sensitive to the social signal value of emotions.

Electroacupuncture improves cerebral blood flow and attenuates moderate ischemic injury via Angiotensin II its receptors-mediated mechanism in rats.

PubMed

Li, Jing; He, Jiaojun; Du, Yuanhao; Cui, Jingjun; Ma, Ying; Zhang, Xuezhu

2014-11-11

To investigate the effects and potential mechanism of electroacupuncture intervention on expressions of Angiotensin II and its receptors-mediated signaling pathway in experimentally induced cerebral ischemia. Totally 126 male Wistar rats were randomly divided into control group, model group and EA group. The latter two were further divided into ten subgroups (n = 6) following Middle Cerebral Artery Occlusion (MCAO). Changes in regional cerebral blood flow (rCBF) and expressions of Angiotensin II and its receptors (AT1R, AT2R), as well as effector proteins in phosphatidyl inositol signal pathway were monitored before and at different times after MCAO. MCAO-induced decline of ipsilateral rCBF was partially suppressed by electroacupuncture, and contralateral blood flow was also superior to that of model group. Angiotensin II level was remarkably elevated immediately after MCAO, while electroacupuncture group exhibited significantly lower levels at 1 to 3 h and the value was significantly increased thereafter. The enhanced expression of AT1R was partially inhibited by electroacupuncture, while increased AT2R level was further induced. Electroacupuncture stimulation attenuated and postponed the upregulated-expressions of Gq and CaM these upregulations. ELISA results showed sharply increased expressions of DAG and IP3, which were remarkably neutralized by electroacupuncture. MCAO induced significant increases in expression of Angiotensin II and its receptor-mediated signal pathway. These enhanced expressions were significantly attenuated by electroacupuncture intervention, followed by reduced vasoconstriction and improved blood supply in ischemic region, and ultimately conferred beneficial effects on cerebral ischemia.

Gallic acid inhibits gastric cancer cells metastasis and invasive growth via increased expression of RhoB, downregulation of AKT/small GTPase signals and inhibition of NF-κB activity

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ho, Hsieh-Hsun; Chang, Chi-Sen; Division of Gastroenterology, Taichung Veterans General Hospital, Taichung 402, Taiwan

2013-01-01

Our previous study demonstrated the therapeutic potential of gallic acid (GA) for controlling tumor metastasis through its inhibitory effect on the motility of AGS cells. A noteworthy finding in our previous experiment was increased RhoB expression in GA-treated cells. The aim of this study was to evaluate the role of RhoB expression on the inhibitory effects of GA on AGS cells. By applying the transfection of RhoB siRNA into AGS cells and an animal model, we tested the effect of GA on inhibition of tumor growth and RhoB expression. The results confirmed that RhoB-siRNA transfection induced GA to inhibit AGSmore » cells’ invasive growth involving blocking the AKT/small GTPase signals pathway and inhibition of NF-κB activity. Finally, we evaluated the effect of GA on AGS cell metastasis by colonization of tumor cells in nude mice. It showed GA inhibited tumor cells growth via the expression of RhoB. These data support the inhibitory effect of GA which was shown to inhibit gastric cancer cell metastasis and invasive growth via increased expression of RhoB, downregulation of AKT/small GTPase signals and inhibition of NF-κB activity. Thus, GA might be a potential agent in treating gastric cancer. Highlights: ► GA could downregulate AKT signal via increased expression of RhoB. ► GA inhibits metastasis in vitro in gastric carcinoma. ► GA inhibits tumor growth in nude mice model.« less

Increased tenascin C and Toll-like receptor 4 levels in visceral adipose tissue as a link between inflammation and extracellular matrix remodeling in obesity.

PubMed

Catalán, Victoria; Gómez-Ambrosi, Javier; Rodríguez, Amaia; Ramírez, Beatriz; Rotellar, Fernando; Valentí, Victor; Silva, Camilo; Gil, María J; Salvador, Javier; Frühbeck, Gema

2012-10-01

Obesity is associated with an altered inflammatory and extracellular matrix (ECM) profile. Tenascin C (TNC) is an ECM glycoprotein with proinflammatory effects. We aimed to explore the expression levels of TNC in adipose tissue analyzing the contribution of adipocytes and stromovascular fraction cells (SVFC) as well as its impact on inflammation and ECM regulation. We also analyzed the effect of the stimulation with TNF-α and lipopolysaccharide (LPS) on both SVFC and adipocytes. Samples obtained from 75 subjects were used in the study. Expression levels of TNC, TLR4, MMP2, and MMP9 were analyzed in visceral adipose tissue (VAT) as well as in both adipocytes and SVFC. In addition, Tnc expression was measured in two mice models of obesity. We show, for the first time, that VAT expression levels of TNC are increased in normoglycemic and type 2 diabetic obese patients (P<0.01) as well as in obese patients with nonalcoholic steatohepatitis (P<0.01). Furthermore, expression levels of Tnc in epididymal adipose tissue from two different mice models of obesity were significantly increased (P<0.01). TNC and TLR4 were mainly expressed by SVFC, and its expression was significantly enhanced (P<0.01) by TNF-α treatment. LPS treatment also increased mRNA levels of TNC. Moreover, the addition of exogenous TNC induced (P<0.05) TLR4 and CCL2 mRNA expression in human adipocyte cultures. These findings indicate that TNC is involved in the etiopathology of obesity via visceral adipose tissue inflammation representing a link with ECM remodeling.

Muscle wasting in osteoarthritis model induced by anterior cruciate ligament transection.

PubMed

Silva, Jordana Miranda de Souza; Alabarse, Paulo Vinicius Gil; Teixeira, Vivian de Oliveira Nunes; Freitas, Eduarda Correa; de Oliveira, Francine Hehn; Chakr, Rafael Mendonça da Silva; Xavier, Ricardo Machado

2018-01-01

This study aimed to investigate the molecular pathways involved in muscle wasting in an animal model of osteoarthritis (OA) induced by anterior cruciate ligament transection (ACLT) in rats. Reduction of protein syntheses, increased proteolysis and impaired muscle regeneration are important pathways related to muscle wasting, and myogenin, MyoD, myostatin and MuRF-1 are some of their markers. Female Wistar rats were allocated into two groups: OA (submitted to the ACLT) and SHAM (submitted to surgery without ACLT). Nociception, spontaneous exploratory locomotion and body weight of animals were evaluated weekly. Twelve weeks after the disease induction, animals were euthanized, and the right knee joints were collected. Gastrocnemius muscle of the right hind paw were dissected and weighed. Gastrocnemius was used for evaluation of muscle atrophy and expression of IL-1β, TNF-α, Pax7, myogenin, MyoD, myostatin and MuRF-1. Histopathology of the knee confirmed the development of the disease in animals of OA group. Gastrocnemius of OA animals showed a reduction of about 10% in area and an increased IL-1β expression compared to animals of SHAM group. Expression of myostatin was increased in OA group, while myogenin expression was decreased. TNF-α, Pax7, MuRF-1 and MyoD expression was similar in both OA and SHAM groups. Nociception was significantly elevated in OA animals in the last two weeks of experimental period. Spontaneous exploratory locomotion, body weight and weight of gastrocnemius showed no difference between OA and SHAM groups. Gastrocnemius atrophy in OA induced by ACLT involves elevated expression of IL-1β within the muscle, as well as increased expression of myostatin and decreased expression of myogenin. Therefore, muscle wasting may be linked to impaired muscle regeneration.

Decidual β-carotene-15,15'-oxygenase-1 and 2 (BCMO1,2) expression is increased in nitrofen model of congenital diaphragmatic hernia.

PubMed

Takahashi, Hiromizu; Kutasy, Balazs; Pes, Lara; Paradisi, Francesca; Puri, Prem

2015-01-01

Retinoids are essential for fetal and lung development. Beta-carotene(BC) is the main dietary retinoid source and beta-carotene-15,15'-oxygenase-1 and 2 (Bcmo1,2) is the primary enzyme generating retinoid from BC in adult mammalian tissues. Placenta has a major role in the retinol homeostasis in fetal life: Since there is no fetal retinol synthesis, maternal retinol has to cross the placenta. It has been recently shown that BC can be converted to retinol by Bcmo1,2 in placenta for retinol transfer and moreover, BC can cross the placenta intact. The placental Bcmo1,2 expression is tightly controlled by placental retinol level. In severe retinol deficiency it has been shown that placental Bcmo1,2 expression are increased for generating retinol from dietary maternal BC even when the main retinol transfer is blocked. In recent years, low pulmonary retinol levels and disrupted retinoid signaling pathway have been implicated in the pathogenesis of pulmonary hypoplasia and congenital diaphragmatic hernia (CDH) in the nitrofen model of CDH. Recently, it has been demonstrated that the main retinol transfer in the placenta is blocked in the nitrofen model of CDH causing increased placental and decreased serum retinol level. The aim of our study was to determine maternal and fetal β-carotene levels and to investigate the hypothesis that placental expression of BCMO1 and BCMO2 is altered in nitrofen-exposed rat fetuses with CDH. Pregnant rats were exposed to either olive oil or nitrofen on day 9 of gestation (D9). Maternal and fetal serum, placenta, liver and left lungs were harvested on D21 and divided into two groups: control (n = 8) and nitrofen with CDH (n = 8). Immunochistochemistry was performed to evaluate trophoblasts by cytokeratin expression and placental Bcmo1,2 expression. Expression levels of Bcmo1,2 genes in fetal lungs and liver were determined using RT-PCR and immunohistochemistry. BC level was measured using HPLC. Markedly increased decidual Bcmo1,2 immunoreactivity was observed in CDH group compared to controls. There was no difference neither in the trophoblastic Bcmo1,2 immunoreactivity nor in the pulmonary and liver Bcmo1,2 expression compared to controls. There was no significant difference in maternal serum BC levels between control and CDH mothers (2.14 ± 0.55 vs 2.56 ± 1.6 μM/g, p = 0.8). BC was not detectable neither in the fetal serum nor liver or lungs. Our data show that nitrofen increases maternal but not fetal Bcmo1,2 expression in the placenta in nitrofen-induced CDH group. The markedly increased decidual Bcmo1,2 expression suggests that nitrofen may trigger local, decidual retinol synthesis in the nitrofen model of CDH.

Cooperation in the dark: signalling and collective action in quorum-sensing bacteria.

PubMed

Brown, S P; Johnstone, R A

2001-05-07

The study of quorum-sensing bacteria has revealed a widespread mechanism of coordinating bacterial gene expression with cell density. By monitoring a constitutively produced signal molecule, individual bacteria can limit their expression of group-beneficial phenotypes to cell densities that guarantee an effective group outcome. In this paper, we attempt to move away from a commonly expressed view that these impressive feats of coordination are examples of multicellularity in prokaryotic populations. Here, we look more closely at the individual conflict underlying this cooperation, illustrating that, even under significant levels of genetic conflict, signalling and resultant cooperative behaviour can stably exist. A predictive two-trait model of signal strength and of the extent of cooperation is developed as a function of relatedness (reflecting multiplicity of infection) and basic population demographic parameters. The model predicts that the strength of quorum signalling will increase as conflict (multiplicity of infecting strains) increases, as individuals attempt to coax more cooperative contributions from their competitors, leading to a devaluation of the signal as an indicator of density. Conversely, as genetic conflict increases, the model predicts that the threshold density for cooperation will increase and the subsequent strength of group cooperation will be depressed.

Short-Lived Human Umbilical Cord-Blood-Derived Neural Stem Cells Influence the Endogenous Secretome and Increase the Number of Endogenous Neural Progenitors in a Rat Model of Lacunar Stroke.

PubMed

Jablonska, Anna; Drela, Katarzyna; Wojcik-Stanaszek, Luiza; Janowski, Miroslaw; Zalewska, Teresa; Lukomska, Barbara

2016-11-01

Stroke is the leading cause of severe disability, and lacunar stroke is related to cognitive decline and hemiparesis. There is no effective treatment for the majority of patients with stroke. Thus, stem cell-based regenerative medicine has drawn a growing body of attention due to the capabilities for trophic factor expression and neurogenesis enhancement. Moreover, it was shown in an experimental autoimmune encephalomyelitis (EAE) model that even short-lived stem cells can be therapeutic, and we have previously observed that phenomenon indirectly. Here, in a rat model of lacunar stroke, we investigated the molecular mechanisms underlying the positive therapeutic effects of short-lived human umbilical cord-blood-derived neural stem cells (HUCB-NSCs) through the distinct measurement of exogenous human and endogenous rat trophic factors. We have also evaluated neurogenesis and metalloproteinase activity as cellular components of therapeutic activity. As expected, we observed an increased proliferation and migration of progenitors, as well as metalloproteinase activity up to 14 days post transplantation. These changes were most prominent at the 7-day time point when we observed 30 % increases in the number of bromodeoxyuridine (BrdU)-positive cells in HUCB-NSC transplanted animals. The expression of human trophic factors was present until 7 days post transplantation, which correlated well with the survival of the human graft. For these 7 days, the level of messenger RNA (mRNA) in the analyzed trophic factors was from 300-fold for CNTF to 10,000-fold for IGF, much higher compared to constitutive expression in HUCB-NSCs in vitro. What is interesting is that there was no increase in the expression of rat trophic factors during the human graft survival, compared to that in non-transplanted animals. However, there was a prolongation of a period of increased trophic expression until 14 days post transplantation, while, in non-transplanted animals, there was a significant drop in rat trophic expression at that time point. We conclude that the positive therapeutic effect of short-lived stem cells may be related to the net increase in the amount of trophic factors (rat + human) until graft death and to the prolonged increase in rat trophic factor expression subsequently.

Evaluation of the reversal of multidrug resistance by MDR1 ribonucleic acid interference in a human colon cancer model using a Renilla luciferase reporter gene and coelenterazine.

PubMed

Jeon, Yong Hyun; Bae, Seon-ae; Lee, Yong Jin; Lee, You La; Lee, Sang-Woo; Yoon, Ghil-Suk; Ahn, Byeong-Cheol; Ha, Jeoung-Hee; Lee, Jaetae

2010-12-01

The reversal effect of multidrug resistance (MDR1) gene expression by adenoviral vector-mediated MDR1 ribonucleic acid interference was assessed in a human colon cancer animal model using bioluminescent imaging with Renilla luciferase (Rluc) gene and coelenterazine, a substrate for Rluc or MDR1 gene expression. A fluorescent microscopic examination demonstrated an increased green fluorescent protein signal in Ad-shMDR1- (recombinant adenovirus that coexpressed MDR1 small hairpin ribonucleic acid [shRNA] and green fluorescent protein) infected HCT-15/Rluc cells in a virus dose-dependent manner. Concurrently, with an increasing administered virus dose (0, 15, 30, 60, and 120 multiplicity of infection), Rluc activity was significantly increased in Ad-shMDR1-infected HCT-15/Rluc cells in a virus dose-dependent manner. In vivo bioluminescent imaging showed about 7.5-fold higher signal intensity in Ad-shMDR1-infected tumors than in control tumors (p < .05). Immunohistologic analysis demonstrated marked reduction of P-glycoprotein expression in infected tumor but not in control tumor. In conclusion, the reversal of MDR1 gene expression by MDR1 shRNA was successfully evaluated by bioluminescence imaging with Rluc activity using an in vivo animal model with a multidrug resistance cancer xenograft.

Tissue responses to hexyl 5-aminolevulinate-induced photodynamic treatment in syngeneic orthotopic rat bladder cancer model: possible pathways of action

NASA Astrophysics Data System (ADS)

Arum, Carl-Jørgen; Gederaas, Odrun A.; Larsen, Eivind L. P.; Randeberg, Lise L.; Hjelde, Astrid; Krokan, Hans E.; Svaasand, Lars O.; Chen, Duan; Zhao, Chun-Mei

2011-02-01

Orthotopic bladder cancer model in rats mimics human bladder cancer with respect to urothelial tumorigenesis and progression. Utilizing this model at pT1 (superficial stage), we analyze the tissue responses to hexyl 5-aminolevulinate-induced photodynamic therapy (HAL-PDT). In comparison to untreated rats, HAL-PDT causes little change in tumor-free rat bladder but induces inflammatory changes with increased lymphocytes and mononuclear cell infiltration in rat bladders with tumor. Immunohistochemistry reveals that HAL-PDT is without effect on proliferating cell nuclear antigen expression within the tumor and increases caspase-3 expression in both normal urothelium and the tumor. Transmission electron microscopy reveals severe mitochondrial damage, formations of apoptotic bodies, vacuoles, and lipofuscin bodies, but no microvillus-formed niches in HAL-PDT-treated bladder cancer rats. Bioinformatics analysis of the gene expression profile indicates an activation of T-cell receptor signaling pathway in bladder cancer rats without PDT. HAL-PDT increases the expression of CD3 and CD45RA in the tumor (determined by immunohistochemistry). We suggest that pathways of action of HAL-PDT may include, at least, activations of mitochondrial apoptosis and autophagy, breakdown of cancer stem cell niches, and importantly, enhancement of T-cell activation.

Modeled Microgravity Disrupts Collagen I/Integrin Signaling During Osteoblastic Differentiation of Human Mesenchymal Stem Cells

NASA Technical Reports Server (NTRS)

Meyers, Valerie E.; Zayzafoon, Majd; Gonda, Steven R.; Gathings, William E.; McDonald, Jay M.

2004-01-01

Spaceflight leads to reduced bone mineral density in weight bearing bones that is primarily attributed to a reduction in bone formation. We have previously demonstrated severely reduced osteoblastogenesis of human mesenchymal stem cells (hMSC) following seven days culture in modeled microgravity. One potential mechanism for reduced osteoblastic differentiation is disruption of type I collagen-integrin interactions and reduced integrin signaling. Integrins are heterodimeric transmembrane receptors that bind extracellular matrix proteins and produce signals essential for proper cellular function, survival, and differentiation. Therefore, we investigated the effects of modeled microgravity on integrin expression and function in hMSC. We demonstrate that seven days of culture in modeled microgravity leads to reduced expression of the extracellular matrix protein, type I collagen (Col I). Conversely, modeled microgravity consistently increases Col I-specific alpha2 and beta1 integrin protein expression. Despite this increase in integrin sub-unit expression, autophosphorylation of adhesion-dependent kinases, focal adhesion kinase (FAK) and proline-rich tyrosine kinase 2 (PYK2), is significantly reduced. Activation of Akt is unaffected by the reduction in FAK activation. However, reduced downstream signaling via the Ras-MAPK pathway is evidenced by a reduction in Ras and ERK activation. Taken together, our findings indicate that modeled microgravity decreases integrin/MAPK signaling, which likely contributes to the observed reduction in osteoblastogenesis.

Polysaccharides from Enteromorpha prolifera Improve Glucose Metabolism in Diabetic Rats

PubMed Central

Lin, Wenting; Wang, Wenxiang; Liao, Dongdong; Chen, Damiao; Zhu, Pingping; Cai, Guoxi; Kiyoshi, Aoyagi

2015-01-01

This study investigated the effects of polysaccharides from Enteromorpha prolifera (PEP) on glucose metabolism in a rat model of diabetes mellitus (DM). PEP (0, 150, 300, and 600 mg/kg) was administered intragastrically to rats for four weeks. After treatment, fasting blood glucose (FBG) and insulin (INS) levels were measured, and the insulin sensitivity index (ISI) was calculated. The morphopathological changes in the pancreas were observed. Serum samples were collected to measure the oxidant-antioxidant status. The mRNA expression levels of glucokinase (GCK) and insulin receptor (InsR) in liver tissue and glucose transporter type 4 (GLUT-4) and adiponectin (APN) in adipose tissue were determined. Compared with the model group, the FBG and INS levels were lower, the ISI was higher, and the number of islet β-cells was significantly increased in all the PEP groups. In the medium- and high-dose PEP groups, MDA levels decreased, and the enzymatic activities of SOD and GSH-Px increased. The mRNA expression of InsR and GCK increased in all the PEP groups; APN mRNA expression increased in the high-dose PEP group, and GLUT-4 mRNA expression increased in adipose tissue. These findings suggest that PEP is a potential therapeutic agent that can be utilized to treat DM. PMID:26347892

[Effects of Salvianolate on Myosin Heavy Chain in Cardiomyocytes of Congestive Heart Failure Rats].

PubMed

Chen, Cheng; Zou, Xiang-gu; Qiu, Shan-dong; Chen, Hui; Chen, Yong-zhong; Lin, Xiu-ming

2015-07-01

To explore the effect of Salvianolate on myosin heavy chain (MHC) in cardiomyocytes of congestive heart failure (CHF) rats. Sixty male SD rats were divided into 6 groups according to random digit table, i.e., the normal control group (NCG), the model group, the Captopril group (CAG), the low dose Salvianolate group (LSG), the high dose Salvianolate group (HSG), the Captopril and high dose Salvianolate group (CSG), 10 in each group. CHF rat model was established with peritoneal injection of adriamycin in all rats except those in the NCG. Equal volume of normal saline was peritoneally injected to rats in the NCG, once per week for 6 successive weeks. Corresponding medication was started from the 5th week of injecting adriamycin. Rats in the CAG were administered with Captopril solution at the daily dose of 10 mg/kg by gastrogavage. Rats in the LSG and the HSG were administered with Salvianolate solution at the daily dose of 24.219 mg/kg and 48.438 mg/kg respectively by gastrogavage. Salvianolate was dissolved in 2 mL 5% glucose solution and administered by peritoneal injection. Rats in the CSG were peritoneally injected with high dose Salvianolate solution and administered with Captopril solution by gastrogavage. Two mL normal saline was peritoneally injected to rats in the model group, once per day for 8 successive weeks. Eight weeks later, the cardiac function and myocardial hypertrophy indices were detected by biological signal collecting and processing system. mRNA expression levels of alpha-MHC and beta-MHC in cardiac muscle were detected by fluorescence quantitative PCR. Expressions of protein kinase C (PKC) in cardiac muscle were detected by Western blot. Compared with the normal control group, heart mass index (HMI) and left ventricular mass index (LVMI) obviously increased in the model group (P < 0.01). Compared with the model group, HMI and LVMI decreased in HSG, CAG, and CSG groups (P < 0.05, P < 0.01). It was more obviously lowered in the CSG group than in the CAG group (P < 0.05). Compared with the NCG, the mRNA expression level of alpha-MHC in cardiac muscle decreased, the mRNA expression level of p-MHC and the expression of PKC in cardiac muscle increased in the model group (P < 0.01). Compared with the model group, the mRNA expression level of alpha-MHC in cardiac muscle was increased, and the mRNA expression level of beta-MHC and the expression of PKC in cardiac muscle were decreased in HSG, CAG, and CSG groups (P < 0.05, P < 0.01). There was statistical difference between the CSG group and the CAG group (P < 0.05). Salvianolate could up-regulate the mRNA expression level of alpha-MHC, and down-regulate the mRNA expression level of beta-MHC in cardiac muscle. Its mechanism might be related to decreasing the expression of PKC.

DNM3, p65 and p53 from exosomes represent potential clinical diagnosis markers for glioblastoma multiforme

PubMed Central

Yang, Jian-kai; Song, Jian; Huo, Hao-ran; Zhao, Yin-long; Zhang, Guang-yu; Zhao, Zong-mao; Sun, Guo-zhu; Jiao, Bao-hua

2017-01-01

Background: Glioblastoma multiforme (GBM) is the most aggressive and deadly primary brain cancer that arises from astrocytes and classified as grade IV. Recently, exosomes have been reported as an essential mediator in diverse cancer carcinogenesis and metastasis. However, their role in GBM is still unclear. In this study, we aimed to investigate whether blood exosomes can be potential clinical diagnostic markers for GBM. Methods: We used a xenograft orthotopic mouse model to detect the differentially expressed genes in the brain and blood exosomes of original/recurrent GBM. Results: We found that recurrent GBM had stronger growth capacity and lethality than original GBM in the mouse model. A gene microarray of original tumors and blood exosomes from GBM orthotopic xenografts results showed that DNM3, p65 and CD117 expressions increased, whereas PTEN and p53 expressions decreased in both original tumors and blood exosomes. In the recurrent GBM tumor model, DNM3 and p65 showed increased expressions, whereas ST14 and p53 showed decreased expressions in tumor and blood exosomes of the recurrent GBM mouse model. Conclusion: In summary, we found that DNM3, p65 and p53 had a similar trend in brain and blood exosomes both for original and recurrent GBM, and could serve as potential clinical diagnostic markers for GBM. PMID:29449895

TBX1 Represses Vegfr2 Gene Expression and Enhances the Cardiac Fate of VEGFR2+ Cells

PubMed Central

Lania, Gabriella; Ferrentino, Rosa; Baldini, Antonio

2015-01-01

The T-box transcription factor TBX1 has critical roles in maintaining proliferation and inhibiting differentiation of cardiac progenitor cells of the second heart field (SHF). Haploinsufficiency of the gene that encodes it is a cause of congenital heart disease. Here, we developed an embryonic stem (ES) cell-based model in which Tbx1 expression can be modulated by tetracycline. Using this model, we found that TBX1 down regulates the expression of VEGFR2, and we confirmed this finding in vivo during embryonic development. In addition, we found a Vegfr2 domain of expression, not previously described, in the posterior SHF and this expression is extended by loss of Tbx1. VEGFR2 has been previously described as a marker of a subpopulation of cardiac progenitors. Clonal analysis of ES-derived VEGFR2+ cells indicated that 12.5% of clones expressed three markers of cardiac lineage (cardiomyocyte, smooth muscle and endothelium). However, a pulse of Tbx1 expression was sufficient to increase the percentage to 20.8%. In addition, the percentage of clones expressing markers of multiple cardiac lineages increased from 41.6% to 79.1% after Tbx1 pulse. These results suggest that TBX1 plays a role in maintaining a progenitor state in VEGFR2+ cells. PMID:26382615

Skeletal muscle deiodinase type 2 regulation during illness in mice.

PubMed

Kwakkel, J; van Beeren, H C; Ackermans, M T; Platvoet-Ter Schiphorst, M C; Fliers, E; Wiersinga, W M; Boelen, A

2009-11-01

We have previously shown that skeletal muscle deiodinase type 2 (D2) mRNA (listed as Dio2 in MGI Database) is upregulated in an animal model of acute illness. However, human studies on the expression of muscle D2 during illness report conflicting data. Therefore, we evaluated the expression of skeletal muscle D2 and D2-regulating factors in two mouse models of illness that differ in timing and severity of illness: 1) turpentine-induced inflammation, and 2) Streptococcus pneumoniae infection. During turpentine-induced inflammation, D2 mRNA and activity increased compared to pair-fed controls, most prominently at day 1 and 2, whereas after S. pneumoniae infection D2 mRNA decreased. We evaluated the association of D2 expression with serum thyroid hormones, (de-)ubiquitinating enzymes ubiquitin-specific peptidase 33 and WD repeat and SOCS box-containing 1 (Wsb1), cytokine expression and activation of inflammatory pathways and cAMP pathway. During chronic inflammation the increased muscle D2 expression is associated with the activation of the cAMP pathway. The normalization of D2 5 days after turpentine injection coincides with increased Wsb1 and tumor necrosis factor alpha expression. Muscle interleukin-1beta (Il1b) expression correlated with decreased D2 mRNA expression after S. pneumoniae infection. In conclusion, muscle D2 expression is differentially regulated during illness, probably related to differences in the inflammatory response and type of pathology. D2 mRNA and activity increases in skeletal muscle during the acute phase of chronic inflammation compared to pair-fed controls probably due to activation of the cAMP pathway. In contrast, muscle D2 mRNA decreases 48 h after a severe bacterial infection, which is associated with local Il1b mRNA expression and might also be due to diminished food-intake.

Heterogeneity of leukemia-initiating capacity of chronic myelogenous leukemia stem cells

PubMed Central

Zhang, Bin; Li, Ling; Ho, Yinwei; Li, Min; Marcucci, Guido

2016-01-01

Chronic myelogenous leukemia (CML) results from transformation of a long-term hematopoietic stem cell (LTHSC) by expression of the BCR-ABL fusion gene. However, BCR-ABL–expressing LTHSCs are heterogeneous in their capacity as leukemic stem cells (LSCs). Although discrepancies in proliferative, self-renewal, and differentiation properties of normal LTHSCs are being increasingly recognized, the mechanisms underlying heterogeneity of leukemic LTHSCs are poorly understood. Using a CML mouse model, we identified gene expression differences between leukemic and nonleukemic LTHSCs. Expression of the thrombopoietin (THPO) receptor MPL was elevated in leukemic LTHSC populations. Compared with LTHSCs with low MPL expression, LTHSCs with high MPL expression showed enhanced JAK/STAT signaling and proliferation in response to THPO in vitro and increased leukemogenic capacity in vivo. Although both G0 and S phase subpopulations were increased in LTHSCs with high MPL expression, LSC capacity was restricted to quiescent cells. Inhibition of MPL expression in CML LTHSCs reduced THPO-induced JAK/STAT signaling and leukemogenic potential. These same phenotypes were also present in LTHSCs from patients with CML, and patient LTHSCs with high MPL expression had reduced sensitivity to BCR-ABL tyrosine kinase inhibitor treatment but increased sensitivity to JAK inhibitors. Together, our studies identify MPL expression levels as a key determinant of heterogeneous leukemia-initiating capacity and drug sensitivity of CML LTHSCs and suggest that high MPL–expressing CML stem cells are potential targets for therapy. PMID:26878174

The Roles of Phasic and Tonic Dopamine in Tic Learning and Expression.

PubMed

Maia, Tiago V; Conceição, Vasco A

2017-09-15

Tourette syndrome (TS) prominently involves dopaminergic disturbances, but the precise nature of those disturbances has remained elusive. A substantial body of empirical work and recent computational models have characterized the specific roles of phasic and tonic dopamine (DA) in action learning and selection, respectively. Using insights from this work and models, we suggest that TS involves increases in both phasic and tonic DA, which produce increased propensities for tic learning and expression, respectively. We review the evidence from reinforcement-learning and habit-learning studies in TS, which supports the idea that TS involves increased phasic DA responses; we also review the evidence that tics engage the habit-learning circuitry. On the basis of these findings, we suggest that tics are exaggerated, maladaptive, and persistent motor habits reinforced by aberrant, increased phasic DA responses. Increased tonic DA amplifies the tendency to execute learned tics and also provides a fertile ground of motor hyperactivity for tic learning. We review evidence suggesting that antipsychotics may counter both the increased propensity for tic expression, by increasing excitability in the indirect pathway, and the increased propensity for tic learning, by shifting plasticity in the indirect pathway toward long-term potentiation (and possibly also through more complex mechanisms). Finally, we review evidence suggesting that low doses of DA agonists that effectively treat TS decrease both phasic and tonic DA, thereby also reducing the propensity for both tic learning and tic expression, respectively. Copyright © 2017 Society of Biological Psychiatry. Published by Elsevier Inc. All rights reserved.

Adverse early life environment increases hippocampal microglia abundance in conjunction with decreased neural stem cells in juvenile mice.

PubMed

Cohen, Susan; Ke, Xingrao; Liu, Qiuli; Fu, Qi; Majnik, Amber; Lane, Robert

2016-12-01

Adverse maternal lifestyle resulting in adverse early life environment (AELE) increases risks for neuropsychiatric disorders in offspring. Neuropsychiatric disorders are associated with impaired neurogenesis and neuro-inflammation in the hippocampus (HP). Microglia are neuro-inflammatory cells in the brain that regulate neurogenesis via toll-like receptors (TLR). TLR-9 is implicated in neurogenesis inhibition and is responsible for stress-related inflammatory responses. We hypothesized that AELE would increase microglia cell count and increase TLR-9 expression in juvenile mouse HP. These increases in microglia cell count and TLR-9 expression would be associated with decrease neural stem cell count and neuronal cell count. We developed a mouse model of AELE combining Western diet and a stress environment. Stress environment consisted of random change from embryonic day 13 (E13) to E17 as well as static change in maternal environment from E13 to postnatal day 21(P21). At P21, we measured hippocampal cell numbers of microglia, neural stem cell and neuron, as well as hippocampal TLR-9 expression. AELE significantly increased total microglia number and TLR-9 expression in the hippocampus. Concurrently, AELE significantly decreased neural stem cell and neuronal numbers. AELE increased the neuro-inflammatory cellular response in the juvenile HP. We speculate that increased neuro-inflammatory responses may contribute to impaired neurogenesis seen in this model. Copyright © 2016 ISDN. Published by Elsevier Ltd. All rights reserved.

Changes in mRNA expression precede changes in microRNA expression in lesional psoriatic skin during treatment with adalimumab.

PubMed

Raaby, L; Langkilde, A; Kjellerup, R B; Vinter, H; Khatib, S H; Hjuler, K F; Johansen, C; Iversen, L

2015-08-01

Tumour necrosis factor (TNF)-α inhibition is an effective treatment for moderate to severe plaque-type psoriasis. A change in the cytokine expression profile occurs in the skin after 4 days of treatment, preceding any clinical or histological improvements. MicroRNAs (miRNAs) are important post-transcriptional regulators of gene expression, but miRNA expression has never been studied in psoriatic skin during treatment. To investigate changes in miRNA expression in psoriatic skin during adalimumab treatment and to compare results with changes in miRNA expression in a mouse model of Aldara-induced psoriasis-like skin inflammation. Punch biopsies were obtained from nonlesional and lesional psoriatic skin during adalimumab treatment. In the mouse model of Aldara-induced skin inflammation, biopsies were obtained from TNF-α knockout (KO), IL-17A KO and wild-type mice. miRNA expression levels were analysed with microarray, reverse transcriptase quantitative polymerase chain reaction and in situ hybridization. In psoriatic skin, no changes in miRNA expression were seen 4 days after treatment initiation. After 14 days of treatment, the expression of several miRNAs was normalized towards the level seen in nonlesional skin before treatment. miR-23b expression increased after 14 days of treatment and remained high for 84 days, despite unaltered levels at baseline. In the mouse model of Aldara-induced skin inflammation, the level of miR-146a increased, whereas no regulation was seen for miR-203, miR-214-3p, miR-125a, miR-23b or let-7d-5p. This study demonstrates that the changes seen in the cytokine expression levels after 4 days of treatment with adalimumab are not facilitated by early changes in miRNA expression. © 2015 British Association of Dermatologists.

Saponins extracted from Dioscorea collettii rhizomes regulate the expression of urate transporters in chronic hyperuricemia rats.

PubMed

Zhu, Liran; Dong, Yifan; Na, Sha; Han, Ru; Wei, Chengyin; Chen, Guangliang

2017-09-01

The current study aimed to investigate whether the saponins, bioactive component of effects of D. collettii, could reduce the serum uric acid level in a hyperuricemic mouse via regulation of urate transporters. Chronic hyperuricemia model was established by combine administration of adenine (100mg/kg) and ethambutol (250mg/kg). In the model group, the serum uric acid (SUA), urine uric acid (UUA) volume, and 24-h UUA values increased significantly, while the uric acid clearance rate (CUr) and creatinine clearance rate (CCr) values decreased. Further, the model groups showed significantly lower expression of organic anion transporter 1 (OAT1) and organic anion transporter 3 (OAT3) and significantly higher expression of renal tubular urate transporter 1 (URAT1), glucose transporter 9 (GLUT9) and URAT1 mRNA than the normal control group. Saponins administration was found to have a dose-dependent effect, as evidenced by the increase in the 24-h UUA, CUr and CCr values; the decrease in SUA; the decrease in the renal expression of URAT1 mRNA and URAT1 and GLUT9 proteins; and the increase in the renal expression of the OAT1 and OAT3 proteins. The saponins extracted from D. collettii rhizomes had an obvious anti-hyperuricemic effect through downregulation of the URAT1 mRNA and the URAT1 and GLUT9 proteins and upregulation of the OAT1 and OAT3 proteins. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

[The expression and significance of chemokines eotaxin and RANTES in the rat model of allergic rhinitis].

PubMed

Tian, Cuiling; Lei, Xiaoping; Shui, Minhong; Zhang, Yanhong; Jia, Qianwei; Tu, Jing; Lian, Gang; Tang, Siquan

2014-07-01

To explore the expression and significance of Eotaxin and RANTES in the rat model of allergic rhinitis (AR). 20 female SD rats in 6-7 weeks were randomly divided into control group and AR group (n = 10, respectively). AR rat model was made with ovalbumin stimulation. To detect pathological changes in mucosa and chemokine Eotaxin, RANTES in their nasal and lung tissues after execution. Compared with the control group, Lung EOS cell counted higher in AR group and the difference was significant (P < 0.01); the AR rats nasal mucosa and lung tissue of Eotaxin, RANTES expression was significantly increased (P < 0.01). There exist high expression of Eotaxin, RANTES, infiltration of eosinophils in nasal and lung tissue of model rats with allergic rhinitis, inferring that the upper and lower respiratory tract inflammatory response has obvious consistency.

Local expression of vaginal Th1 and Th2 cytokines in murine vaginal candidiasis under different immunity conditions.

PubMed

Chen, Shanjuan; Li, Shaohua; Wu, Yan; Liu, Zhixiang; Li, Jiawen

2008-08-01

To investigate the expression of vaginal Th1 and Th2 cytokines in rats with experimental vaginal candidiasis under different immune conditions, ICR murine vaginal candidiasis model was established and immno-suppressed murine models of vaginal cadidiasis were established in estrogen-treated mice. Non-estrogen-treated mice were used as controls. The mRNA level of Th1 (IL-2)/Th2 (IL-4, IL-10, TGF-beta1) cytokines in murine vaginal tissues was determined by RT-PCR. The cykotine in local tissues was increased to different extent under normal immune condition. IL-2 mRNA was increased during early stage of infection, while IL-10 was increased transiently during late stage of infection. TGF-beta1 production was found to be increased persistently. At same time, the expression of IL-2 mRNA was suppressed in immno-suppressed group, and the level of IL-4, IL-10, and TGF-beta1 were higher than the normal immunity group to different degree during infection. The high level of IL-2 mRNA during early stage of infection was associated with clearance of mucosal Candidia albicans (C. albicans), and its expression suppressed leading to decreased clearance of mucosal C. albican in immuno-suppression. The over-expression of IL-4 and IL-10 could significantly enhance the susceptibility to C. albicans infection in mice.

Ursolic acid induces neural regeneration after sciatic nerve injury

PubMed Central

Liu, Biao; Liu, Yan; Yang, Guang; Xu, Zemin; Chen, Jiajun

2013-01-01

In this study, we aimed to explore the role of ursolic acid in the neural regeneration of the injured sciatic nerve. BALB/c mice were used to establish models of sciatic nerve injury through unilateral sciatic nerve complete transection and microscopic anastomosis at 0.5 cm below the ischial tube-rosity. The successfully generated model mice were treated with 10, 5, or 2.5 mg/kg ursolic acid via intraperitoneal injection. Enzyme-linked immunosorbent assay results showed that serum S100 protein expression level gradually increased at 1–4 weeks after sciatic nerve injury, and significantly decreased at 8 weeks. As such, ursolic acid has the capacity to significantly increase S100 protein expression levels. Real-time quantitative PCR showed that S100 mRNA expression in the L4–6 segments on the injury side was increased after ursolic acid treatment. In addition, the muscular mass index in the soleus muscle was also increased in mice treated with ursolic acid. Toluidine blue staining revealed that the quantity and average diameter of myelinated nerve fibers in the injured sciatic nerve were significantly increased after treatment with ursolic acid. 10 and 5 mg/kg of ursolic acid produced stronger effects than 2.5 mg/kg of ursolic acid. Our findings indicate that ursolic acid can dose-dependently increase S100 expression and promote neural regeneration in BALB/c mice following sciatic nerve injury. PMID:25206561

A predictive biophysical model of translational coupling to coordinate and control protein expression in bacterial operons

PubMed Central

Tian, Tian; Salis, Howard M.

2015-01-01

Natural and engineered genetic systems require the coordinated expression of proteins. In bacteria, translational coupling provides a genetically encoded mechanism to control expression level ratios within multi-cistronic operons. We have developed a sequence-to-function biophysical model of translational coupling to predict expression level ratios in natural operons and to design synthetic operons with desired expression level ratios. To quantitatively measure ribosome re-initiation rates, we designed and characterized 22 bi-cistronic operon variants with systematically modified intergenic distances and upstream translation rates. We then derived a thermodynamic free energy model to calculate de novo initiation rates as a result of ribosome-assisted unfolding of intergenic RNA structures. The complete biophysical model has only five free parameters, but was able to accurately predict downstream translation rates for 120 synthetic bi-cistronic and tri-cistronic operons with rationally designed intergenic regions and systematically increased upstream translation rates. The biophysical model also accurately predicted the translation rates of the nine protein atp operon, compared to ribosome profiling measurements. Altogether, the biophysical model quantitatively predicts how translational coupling controls protein expression levels in synthetic and natural bacterial operons, providing a deeper understanding of an important post-transcriptional regulatory mechanism and offering the ability to rationally engineer operons with desired behaviors. PMID:26117546

Expression, regulation, and function of drug transporters in cervicovaginal tissues of a mouse model used for microbicide testing

PubMed Central

Zhou, Tian; Hu, Minlu; Pearlman, Andrew; Rohan, Lisa C.

2016-01-01

P-glycoprotein (P-gp), breast cancer resistance protein (BCRP), and multidrug resistance protein 4 (MRP4) are three efflux transporters that play key roles in the pharmacokinetics of antiretroviral drugs used in the pre-exposure prophylaxis of HIV sexual transmission. In this study, we investigated the expression, regulation, and function of these transporters in cervicovaginal tissues of a mouse model. Expression and regulation were examined using real-time RT-PCR and immunohistochemical staining, in the mouse tissues harvested at estrus and diestrus stages under natural cycling or after hormone synchronization. The three transporters were expressed at moderate to high levels compared to the liver. Transporter proteins were localized in various cell types in different tissue segments. Estrous cycle and exogenous hormone treatment affected transporter mRNA and protein expression, in a tissue- and transporter-dependent manner. Depo-Provera-synchronized mice were dosed vaginally or intraperitoneally with 3H-TFV, with or without MK571 co-administration, to delineate the function of cervicovaginal Mrp4. Co-administration of MK571 significantly increased the concentration of vaginally-administered TFV in endocervix and vagina. MK571 increased the concentration of intraperitoneally-administered TFV in the cervicovaginal lavage and vagina by several fold. Overall, P-gp, Bcrp, and Mrp4 were positively expressed in mouse cervicovaginal tissues, and their expression can be regulated by the estrous cycle or by exogenous hormones. In this model, the Mrp4 transporter impacted TFV distribution in cervicovaginal tissues. PMID:27453435

Higher thyroid hormone receptor expression correlates with short larval periods in spadefoot toads and increases metamorphic rate

PubMed Central

Hollar, Amy R.; Choi, Jinyoung; Grimm, Adam T.; Buchholz, Daniel R.

2011-01-01

Spadefoot toad species display extreme variation in larval period duration, due in part to evolution of thyroid hormone (TH) physiology. Specifically, desert species with short larval periods have higher tail tissue content of TH and exhibit increased responsiveness to TH. To address the molecular basis of larval period differences, we examined TH receptor (TR) expression across species. Based on the dual function model for the role of TR in development, we hypothesized that desert spadefoot species with short larval periods would have 1) late onset of TR expression prior to the production of endogenous TH and 2) higher TR levels when endogenous TH becomes available. To test these hypotheses, we cloned fragments of TRα and TRβ genes from the desert spadefoot toads Scaphiopus couchii and Spea multiplicata and their non-desert relative Pelobates cultripes and measured their mRNA levels in tails using quantitative PCR in the absence (premetamorphosis) or presence (natural metamorphosis) of TH. All species express TRα and TRβ from the earliest stages measured (from just after hatching), but S. couchii, which has the shortest larval period, had more TRα throughout development compared to P. cultripes, which has the longest larval period. TRβ mRNA levels were similar across species. Exogenous T3 treatment induced faster TH-response gene expression kinetics in S. couchii compared to the other species, consistent with its increased TRα mRNA expression and indicative of a functional consequence of more TRα activity at the molecular level. To directly test whether higher TRα expression may contribute to shorter larval periods, we overexpressed TRα via plasmid injection into tail muscle cells of the model frog Xenopus laevis and found an increased rate of muscle cell death in response to TH. These results suggest that increased TRα expression evolved in S. couchii and contribute to its higher metamorphic rates. PMID:21651912

Novel antidepressant candidate RO-05 modulated glucocorticoid receptors activation and FKBP5 expression in chronic mild stress model in rats.

PubMed

Xing, Y; Hou, J; Meng, Q; Yang, M; Kurihara, H; Tian, J

2015-04-02

In this study, a novel TRI (triple reuptake inhibitors) antidepressant candidate RO-05 (4-[1-[1-(benzoyloxy)cyclohexyl]-2-(dimethylamino)ethyl]-phenyl benzoate) was investigated in TST (tail suspension test), FST (forced swimming test) and CMS (chronic mild stress) model. Results showed RO-05 significantly decreased the immobility time in FST and TST at 4.5-, 9-, 18-mg/kg in rats and 9-, 18-, 36-mg/kg in mice. Chronic administration of 18-mg/kg RO-05 improved the behavioral index, anhedonia and normalized the hyperactivity of HPA (hypothalamic-pituitary-adrenal axis) of CMS rats. We further investigated the possible mechanisms of RO-05 in the CMS model. Eighteen milligrams per kilogram of RO-05 chronic administration significantly reversed the increase of mRNA and protein expression of FKBP5 in the CMS rat hippocampus, which facilitated the activation of GR- (glucocorticoid receptor) and GR-responsive gene Foxo1 expression. RO-05 also elevated the expression of BDNF (brain-derived neurotrophic factor) in CMS rat hippocampus. In summary, our results indicated that RO-05 is a promising antidepressant candidate. The possible antidepressant mechanisms of RO-05 were the modulation of FKBP5 expression, GR activation, corresponding inhibition of HPA axis hyperactivity, and the increase of BDNF expression. Copyright © 2015 IBRO. Published by Elsevier Ltd. All rights reserved.

Effect of Retinol Palmitate on Corneal and Conjunctival Mucin Gene Expression in a Rat Dry Eye Model After Injury.

PubMed

Tabuchi, Nobuhito; Toshida, Hiroshi; Koike, Daisuke; Odaka, Akito; Suto, Chikako; Ohta, Toshihiko; Murakami, Akira

We examined the wound-healing effect of retinol palmitate (VApal) on mucin gene and protein expressions in a rat dry eye model based on lacrimal gland (LG) resection after injury. The rat dry eye model was prepared by surgical resection of the main LG in male Long-Evans rats. After alkaline injury of the central part of the lower palpebral conjunctiva bilaterally, VApal eye drops at 1,500 IU/mL in one eye and a vehicle in the fellow eye were both administered 6 times a day for 7 days. The expression of mucin gene and protein was analyzed by real-time polymerase chain reaction or enzyme-linked immunosorbent assay in the cornea and conjunctiva of MUC1, MUC4, MUC16, and MUC5AC after 1, 3, (5), and 7 days of treatment with VApal. Significant decreases in fluorescein-stained areas and rose bengal scores were observed in VApal-treated dry eyes compared with vehicle-treated dry eyes at both 3 (P < 0.05) and 7 days (P < 0.01). Significant increases in corneal rMuc4 and conjunctival rMuc5AC after 1 day (P < 0.01) and conjunctival rMuc16 gene expression after 3 days were observed with VApal treatment (P < 0.05). Furthermore, conjunctival MUC16 expression significantly increased after 3 days of VApal treatment (P < 0.05). VApal promoted corneal rMuc4, conjunctival rMuc5AC, and conjunctival rMuc16 gene expression in a rat dry eye model after injury. VApal also promoted conjunctival MUC16 expression. These results indicate that VApal has efficacy in improving keratoconjunctival epithelial damage associated with decreased tear production.

Gene Expression Analysis to Assess the Relevance of Rodent Models to Human Lung Injury.

PubMed

Sweeney, Timothy E; Lofgren, Shane; Khatri, Purvesh; Rogers, Angela J

2017-08-01

The relevance of animal models to human diseases is an area of intense scientific debate. The degree to which mouse models of lung injury recapitulate human lung injury has never been assessed. Integrating data from both human and animal expression studies allows for increased statistical power and identification of conserved differential gene expression across organisms and conditions. We sought comprehensive integration of gene expression data in experimental acute lung injury (ALI) in rodents compared with humans. We performed two separate gene expression multicohort analyses to determine differential gene expression in experimental animal and human lung injury. We used correlational and pathway analyses combined with external in vitro gene expression data to identify both potential drivers of underlying inflammation and therapeutic drug candidates. We identified 21 animal lung tissue datasets and three human lung injury bronchoalveolar lavage datasets. We show that the metasignatures of animal and human experimental ALI are significantly correlated despite these widely varying experimental conditions. The gene expression changes among mice and rats across diverse injury models (ozone, ventilator-induced lung injury, LPS) are significantly correlated with human models of lung injury (Pearson r = 0.33-0.45, P < 1E -16 ). Neutrophil signatures are enriched in both animal and human lung injury. Predicted therapeutic targets, peptide ligand signatures, and pathway analyses are also all highly overlapping. Gene expression changes are similar in animal and human experimental ALI, and provide several physiologic and therapeutic insights to the disease.

Changes in the Brain Endocannabinoid System in Rat Models of Depression.

PubMed

Smaga, Irena; Jastrzębska, Joanna; Zaniewska, Magdalena; Bystrowska, Beata; Gawliński, Dawid; Faron-Górecka, Agata; Broniowska, Żaneta; Miszkiel, Joanna; Filip, Małgorzata

2017-04-01

A growing body of evidence implicates the endocannabinoid (eCB) system in the pathophysiology of depression. The aim of this study was to investigate the influence of changes in the eCB system, such as levels of neuromodulators, eCB synthesizing and degrading enzymes, and cannabinoid (CB) receptors, in different brain structures in animal models of depression using behavioral and biochemical analyses. Both models used, i.e., bulbectomized (OBX) and Wistar Kyoto (WKY) rats, were characterized at the behavioral level by increased immobility time. In the OBX rats, anandamide (AEA) levels were decreased in the prefrontal cortex, hippocampus, and striatum and increased in the nucleus accumbens, while 2-arachidonoylglycerol (2-AG) levels were increased in the prefrontal cortex and decreased in the nucleus accumbens with parallel changes in the expression of eCB metabolizing enzymes in several structures. It was also observed that CB 1 receptor expression decreased in the hippocampus, dorsal striatum, and nucleus accumbens, and CB 2 receptor expression decreased in the prefrontal cortex and hippocampus. In WKY rats, the levels of eCBs were reduced in the prefrontal cortex (2-AG) and dorsal striatum (AEA) and increased in the prefrontal cortex (AEA) with different changes in the expression of eCB metabolizing enzymes, while the CB 1 receptor density was increased in several brain regions. These findings suggest that dysregulation in the eCB system is implicated in the pathogenesis of depression, although neurochemical changes were linked to the particular brain structure and the factor inducing depression (surgical removal of the olfactory bulbs vs. genetic modulation).

Ethanol-induced hyponatremia augments brain edema after traumatic brain injury.

PubMed

Katada, Ryuichi; Watanabe, Satoshi; Ishizaka, Atsushi; Mizuo, Keisuke; Okazaki, Shunichiro; Matsumoto, Hiroshi

2012-04-01

Alcohol consumption augments brain edema by expression of brain aquaporin-4 after traumatic brain injury. However, how ethanol induces brain aquaporin-4 expression remains unclear. Aquaporin-4 can operate with some of ion channels and transporters. Therefore, we hypothesized that ethanol may affect electrolytes through regulating ion channels, leading to express aquaporin-4. To clarify the hypothesis, we examined role of AQP4 expression in ethanol-induced brain edema and changes of electrolyte levels after traumatic brain injury in the rat. In the rat traumatic brain injury model, ethanol administration reduced sodium ion concentration in blood significantly 24 hr after injury. An aquaporin-4 inhibitor recovered sodium ion concentration in blood to normal. We observed low sodium ion concentration in blood and the increase of brain aquaporin-4 in cadaver with traumatic brain injury. Therefore, ethanol increases brain edema by the increase of aquaporin-4 expression with hyponatremia after traumatic brain injury.

Alogliptin alleviates hepatic steatosis in a mouse model of nonalcoholic fatty liver disease by promoting CPT1a expression via Thr172 phosphorylation of AMPKα in the liver.

PubMed

Tobita, Hiroshi; Sato, Shuichi; Yazaki, Tomotaka; Mishiro, Tsuyoshi; Ishimura, Norihisa; Ishihara, Shunnji; Kinoshita, Yoshikazu

2018-05-01

Pioglitazone (PIO) has been reported to be effective for nonalcoholic fatty liver disease (NAFLD) and alogliptin (ALO) may have efficacy against NAFLD progression in patients with type 2 diabetes mellitus (T2DM). The present study examined the effectiveness of ALO in a rodent model of NAFLD and diabetes mellitus. KK‑Ay mice were used to produce an NAFLD model via administration of a choline‑deficient (CD) diet. To examine the effects of alogliptin, KK‑Ay mice were provided with a CD diet with 0.03% ALO and/or 0.02% PIO orally for 8 weeks. Biochemical parameters, pathological alterations and hepatic mRNA levels associated with fatty acid metabolism were assessed. Severe hepatic steatosis was observed in KK‑Ay mice fed with a CD diet, which was alleviated by the administration of ALO and/or PIO. ALO administration increased the hepatic carnitine palmitoyltransferase 1a (CPT1a) mRNA expression level and enhanced the Thr172 phosphorylation of AMP‑activated protein kinase α (AMPKα) in the liver. PIO administration tended to decrease the hepatic fatty acid synthase mRNA expression level and increase the serum adiponectin level. Homeostasis model of assessment‑insulin resistance values tended to improve with ALO and PIO administration. ALO and PIO alleviated hepatic steatosis in KK‑Ay mice fed with a CD diet. ALO increased hepatic mRNA expression levels associated with fatty acid oxidation. In addition, the results of the present study suggested that ALO promotes CPT1a expression via Thr172 phosphorylation of AMPKα.

Effect of electroacupuncture on brain-derived neurotrophic factor mRNA expression in mouse hippocampus following cerebral ischemia-reperfusion injury.

PubMed

Zhao, Jianxin; Xu, Huazhou; Tian, Yuanxiang; Hu, Manxiang; Xiao, Hongling

2013-04-01

This work aims to observe the effects of electroacupuncture on brain-derived neurotrophic factor (BDNF) mRNA expression in mouse hippocampus following cerebral ischemia-reperfusion injury. The models of mouse cerebral ischemia-reperfusion injury were established. A total of 96 healthy mice were randomly assigned into 4 groups, namely, the sham surgery, model, model + electroacupuncture, and mode + hydergine groups. Mice in the model + electroacupuncture group were treated through electroacupuncture at the Shenshu (BL 23), Geshu (BL 17), and Baihui (GV 20) acupoints. Mice in the model+hydergine group were intragastrically administered with hydergine (0.77 mg/kg(-1) x day(-1)). The levels of BDNF mRNA expressions in the hippocampus were ana lyzed through a semi-quantitative reverse transcription-polymerase chain reaction assay on days 1 and 7 after the surgeries. BDNF mRNA expressions in the mouse hippocampus of the model group on days 1 and 7 after the surgery were higher than those of the sham surgery group (both P < 0.01). On days 1 and 7 of the electroacupuncture treatment, BDNF mRNA expression in the mouse hippocampus of the model + electroacupuncture group was significantly elevated compared with the model group (both P < 0.01) or the model + hydergine group (both P < 0.01). On days 1 and 7 of the hydergine treatment, BDNF mRNA expression in the mouse hippocampus of the model + hydergine group tended to increase compared with the model group; however, statistical significance was not achieved (both P > 0.05). Electroacupuncture treatment enhances endogenous BDNF expression, which may improve the survival environment for intracerebral neurons and inhibit the apoptosis of hippocampal cells.

HPV16-E7 Expression in skin induces TSLP secretion, type 2 ILC infiltration and atopic dermatitis-like lesions

PubMed Central

Bergot, Anne-Sophie; Monnet, Nastasia; Tran, Le Son; Mittal, Deepak; Al-Kouba, Jane; Steptoe, Raymond J.; Grimbaldeston, Michele A.; Frazer, Ian H.; Wells, James W.

2014-01-01

Atopic dermatitis is a common pruritic and inflammatory skin disorder with unknown etiology. Most commonly occurring during early childhood, atopic dermatitis is associated with eczematous lesions and lichenification, in which the epidermis becomes hypertrophied resulting in thickening of the skin. In this study, we report an atopic dermatitis-like pathophysiology results in a murine model following the expression of the high-risk Human Papillomavirus (HPV) 16 oncoprotein E7 in keratinocytes under the Keratin 14 promoter. We show that HPV 16 E7 expression in the skin is associated with skin thickening, acanthosis and light spongiosis. Locally, HPV 16 E7 expressing skin secreted high levels of TSLP and contained increased numbers of ILCs. High levels of circulating IgE were associated with increased susceptibility to skin allergy in a model of cutaneous challenge, and to airway bronchiolar inflammation, enhanced airway goblet cell metaplasia and mucus production in a model of atopic march. Surprisingly, skin pathology occurred independently of T-cells and mast cells. Thus, our findings suggest that the expression of a single HPV oncogene in the skin can drive the onset of atopic dermatitis-like pathology through the induction of TSLP and type 2 ILC infiltration. PMID:25601274

Mushroom β-Glucan May Immunomodulate the Tumor-Associated Macrophages in the Lewis Lung Carcinoma

PubMed Central

Wang, Wan-Jhen; Wu, Yu-Sheng; Chen, Sherwin; Liu, Chi-Feng

2015-01-01

The present study showed that oral mushroom beta-glucan treatment significantly increased IFN-γ mRNA expression but significantly reduced COX-2 mRNA expression within the lung. For LLC tumor model, oral Ganoderma lucidum or Antrodia camphorata polysaccharides treatments significantly reduced TGF-β production in serum. In addition, IL-12 and IFN-γ mRNA expression were significantly increased, but IL-6, IL-10, COX-2, and TGF-β mRNA expression were substantially following oral mushroom polysaccharides treatments. The study highlights the efficacious effect of mushroom polysaccharides for ameliorating the immune suppression in the tumor microenvironment. Increased M1 phenotype of tumor-associated macrophages and attenuated M2 phenotype of tumor-associated macrophages could be achieved by ingesting mushroom polysaccharides. PMID:26167490

EWS Knockdown and Taxifolin Treatment Induced Differentiation and Removed DNA Methylation from p53 Promoter to Promote Expression of Puma and Noxa for Apoptosis in Ewing’s Sarcoma

PubMed Central

Hossain, Mohammad Motarab; Ray, Swapan Kumar

2016-01-01

Ewing’s sarcoma is a pediatric tumor that mainly occurs in soft tissues and bones. Malignant characteristics of Ewing’s sarcoma are correlated with expression of EWS oncogene. We achieved knockdown of EWS expression using a plasmid vector encoding EWS short hairpin RNA (shRNA) to increase anti-tumor mechanisms of taxifolin (TFL), a new flavonoid, in human Ewing’s sarcoma cells in culture and animal models. Immunofluorescence microscopy and flow cytometric analysis showed high expression of EWS in human Ewing’s sarcoma SK-N-MC and RD-ES cell lines. EWS shRNA plus TFL inhibited 80% cell viability and caused the highest decreases in EWS expression at mRNA and protein levels in both cell lines. Knockdown of EWS expression induced morphological features of differentiation. EWS shRNA plus TFL caused more alterations in molecular markers of differentiation than either agent alone. EWS shRNA plus TFL caused the highest decreases in cell migration with inhibition of survival, angiogenic and invasive factors. Knockdown of EWS expression was associated with removal of DNA methylation from p53 promoter, promoting expression of p53, Puma, and Noxa. EWS shRNA plus TFL induced the highest amounts of apoptosis with activation of extrinsic and intrinsic pathways in both cell lines in culture. EWS shRNA plus TFL also inhibited growth of Ewing’s sarcoma tumors in animal models due to inhibition of differentiation inhibitors and angiogenic and invasive factors and also induction of activation of caspase-3 for apoptosis. Collectively, knockdown of EWS expression increased various anti-tumor mechanisms of TFL in human Ewing’s sarcoma in cell culture and animal models. PMID:27547487

EWS Knockdown and Taxifolin Treatment Induced Differentiation and Removed DNA Methylation from p53 Promoter to Promote Expression of Puma and Noxa for Apoptosis in Ewing's Sarcoma.

PubMed

Hossain, Mohammad Motarab; Ray, Swapan Kumar

2014-10-01

Ewing's sarcoma is a pediatric tumor that mainly occurs in soft tissues and bones. Malignant characteristics of Ewing's sarcoma are correlated with expression of EWS oncogene. We achieved knockdown of EWS expression using a plasmid vector encoding EWS short hairpin RNA (shRNA) to increase anti-tumor mechanisms of taxifolin (TFL), a new flavonoid, in human Ewing's sarcoma cells in culture and animal models. Immunofluorescence microscopy and flow cytometric analysis showed high expression of EWS in human Ewing's sarcoma SK-N-MC and RD-ES cell lines. EWS shRNA plus TFL inhibited 80% cell viability and caused the highest decreases in EWS expression at mRNA and protein levels in both cell lines. Knockdown of EWS expression induced morphological features of differentiation. EWS shRNA plus TFL caused more alterations in molecular markers of differentiation than either agent alone. EWS shRNA plus TFL caused the highest decreases in cell migration with inhibition of survival, angiogenic and invasive factors. Knockdown of EWS expression was associated with removal of DNA methylation from p53 promoter, promoting expression of p53, Puma, and Noxa. EWS shRNA plus TFL induced the highest amounts of apoptosis with activation of extrinsic and intrinsic pathways in both cell lines in culture. EWS shRNA plus TFL also inhibited growth of Ewing's sarcoma tumors in animal models due to inhibition of differentiation inhibitors and angiogenic and invasive factors and also induction of activation of caspase-3 for apoptosis. Collectively, knockdown of EWS expression increased various anti-tumor mechanisms of TFL in human Ewing's sarcoma in cell culture and animal models.

Caricaturing facial expressions.

PubMed

Calder, A J; Rowland, D; Young, A W; Nimmo-Smith, I; Keane, J; Perrett, D I

2000-08-14

The physical differences between facial expressions (e.g. fear) and a reference norm (e.g. a neutral expression) were altered to produce photographic-quality caricatures. In Experiment 1, participants rated caricatures of fear, happiness and sadness for their intensity of these three emotions; a second group of participants rated how 'face-like' the caricatures appeared. With increasing levels of exaggeration the caricatures were rated as more emotionally intense, but less 'face-like'. Experiment 2 demonstrated a similar relationship between emotional intensity and level of caricature for six different facial expressions. Experiments 3 and 4 compared intensity ratings of facial expression caricatures prepared relative to a selection of reference norms - a neutral expression, an average expression, or a different facial expression (e.g. anger caricatured relative to fear). Each norm produced a linear relationship between caricature and rated intensity of emotion; this finding is inconsistent with two-dimensional models of the perceptual representation of facial expression. An exemplar-based multidimensional model is proposed as an alternative account.

The effects of sildenafil citrate on urinary podocin and nephrin mRNA expression in an L-NAME model of pre-eclampsia.

PubMed

Baijnath, Sooraj; Murugesan, Saravanakumar; Mackraj, Irene; Gathiram, Prem; Moodley, Jagidesa

2017-03-01

We investigated the effects of sildenafil citrate (SC) on podocyturia in N ω -nitro-L-arginine methyl ester hydrochloride (L-NAME) model of pre-eclampsia (PE). One hundred and twenty Sprague-Dawley rats (SDR) were divided into five groups like pregnant control (PC), early-onset PE (EOPE), late-onset PE(LOPE), early and late-onset PE with SC-treated groups [EOPE (SC); LOPE (SC)]. PE was induced in SDR by oral administration of L-NAME in drinking water for 4-8 days for EOPE and 8-14 day for LOPE. The blood pressure, urine volume and total urine protein were increased in EOPE and LOPE groups when compared to PC, and all the above parameters decreased in EOPE (SC) and LOPE (SC) groups when compared to EOPE and LOPE groups, respectively. The EOPE and LOPE groups showed an increase in urinary nephrin mRNA and podocin mRNA levels compared to PC group. Increases in serum and renal soluble fms-like tyrosine kinase-1 (sFlt-1) expression levels and decreases in renal vascular endothelial growth factor (VEGF) expression and serum placenta growth factor (PlGF) levels were observed in EOPE and LOPE groups when compared to PC group. In addition, decreases in serum and renal sFlt-1 expression levels and increases in renal VEGF expression and serum PlGF levels were observed in EOPE (SC) and LOPE (SC) groups when compared to EOPE and LOPE groups, respectively. The light microscopy showed that the renal tissue of L-NAME-treated rats had extensive glomerular damage, tubular damage and infiltration by mononuclear cells when compared to PC group. Therefore, SC ameliorated podocyturia through its effects on the antiangiogenic/angiogenic status in this animal model.

Effects of Exposure to Ozone on the Ocular Surface in an Experimental Model of Allergic Conjunctivitis

PubMed Central

Lee, Hun; Kim, Eung Kweon; Kim, Hee Young; Kim, Tae-im

2017-01-01

Based on previous findings that ozone can induce an inflammatory response in the ocular surface of an animal model and in cultured human conjunctival epithelial cells, we investigated whether exposure to ozone exacerbates symptoms of allergic conjunctivitis. We evaluated the effects of exposure to ozone on conjunctival chemosis, conjunctival injection, corneal and conjunctival fluorescein staining scores, production of inflammatory cytokines in tears, and aqueous tear production in a mouse model of allergic conjunctivitis. To validate our in vivo results, we used interleukin (IL)-1α-pretreated conjunctival epithelial cells as an in vitro substitute for the mouse model. We evaluated whether exposure to ozone increased the inflammatory response and altered oxidative status and mitochondrial function in IL-1α-pretreated conjunctival epithelial cells. In the in vivo study, ozone induced increases in conjunctival chemosis, conjunctival injection, corneal and conjunctival fluorescein staining scores, and production of inflammatory cytokines, accompanied by a decrease in tear volume. In the in vitro study, exposure to ozone led to additional increases in IL-6 and tumor necrosis factor-α mRNA levels, which were already induced by treatment with IL-1α. Ozone did not induce any changes in cell viability. Pretreatment with IL-1α increased the expression of manganese superoxide dismutase, and exposure to ozone led to additional increments in the expression of this antioxidant enzyme. Ozone did not induce any changes in mitochondrial activity or expression of mitochondrial enzymes and proteins related to mitochondrial function, with the exception of phosphor-mammalian target of rapamycin. Treatment with butylated hydroxyanisole, a free radical scavenger, attenuated the ozone-induced increases in IL-6 expression in IL-1α-pretreated conjunctival epithelial cells. Therefore, we conclude that exposure to ozone exacerbates the detrimental effects on the integrity of the ocular surface caused by conjunctival allergic reactions, and further increases the inflammatory response in IL-1α-pretreated conjunctival epithelial cells. PMID:28046113

Exercise Training and PI3Kα-Induced Electrical Remodeling Is Independent of Cellular Hypertrophy and Akt Signaling

PubMed Central

Yang, Kai-Chien; Tseng, Yi-Tang; Nerbonne, Jeanne M.

2012-01-01

In contrast with pathological hypertrophy, exercise-induced physiological hypertrophy is not associated with electrical abnormalities or increased arrhythmia risk. Recent studies have shown that increased cardiac-specific expression of phosphoinositide-3-kinase-α (PI3Kα), the key mediator of physiological hypertrophy, results in transcriptional upregulation of ion channel subunits in parallel with the increase in myocyte size (cellular hypertrophy) and the maintenance of myocardial excitability. The experiments here were undertaken to test the hypothesis that Akt1, which underlies PI3Kα-induced cellular hypertrophy, mediates the effects of augmented PI3Kα signaling on the transcriptional regulation of cardiac ion channels. In contrast to wild-type animals, chronic exercise (swim) training of mice (Akt1−/−) lacking Akt1 did not result in ventricular myocyte hypertrophy. Ventricular K+ current amplitudes and the expression of K+ channel subunits, however, were increased markedly in Akt1−/− animals with exercise training. Expression of the transcripts encoding inward (Na+ and Ca2+) channel subunits were also increased in Akt1−/− ventricles following swim training. Additional experiments in a transgenic mouse model of inducible cardiac-specific expression of constitutively active PI3Kα (icaPI3Kα) revealed that short-term activation of PI3Kα signaling in the myocardium also led to the transcriptional upregulation of ion channel subunits. Inhibition of cardiac Akt activation with triciribine in this (inducible caPI3Kα expression) model did not prevent the upregulation of myocardial ion channel subunits. These combined observations demonstrate that chronic exercise training and enhanced PI3Kα expression/activity result in transcriptional upregulation of myocardial ion channel subunits independent of cellular hypertrophy and Akt signaling. PMID:22824041

The two-component system GrvRS (EtaRS) regulates ace expression in Enterococcus faecalis OG1RF.

PubMed

Roh, Jung Hyeob; Singh, Kavindra V; La Rosa, Sabina Leanti; Cohen, Ana Luisa V; Murray, Barbara E

2015-01-01

Expression of ace (adhesin to collagen of Enterococcus faecalis), encoding a virulence factor in endocarditis and urinary tract infection models, has been shown to increase under certain conditions, such as in the presence of serum, bile salts, urine, and collagen and at 46 °C. However, the mechanism of ace/Ace regulation under different conditions is still unknown. In this study, we identified a two-component regulatory system GrvRS as the main regulator of ace expression under these stress conditions. Using Northern hybridization and β-galactosidase assays of an ace promoter-lacZ fusion, we found transcription of ace to be virtually absent in a grvR deletion mutant under the conditions that increase ace expression in wild-type OG1RF and in the complemented strain. Moreover, a grvR mutant revealed decreased collagen binding and biofilm formation as well as attenuation in a murine urinary tract infection model. Here we show that GrvR plays a major role in control of ace expression and E. faecalis virulence. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

The regulation of pituitary-thyroid abnormalities by peripheral administration of levothyroxine increased brain-derived neurotrophic factor and reelin protein expression in an animal model of Alzheimer's disease.

PubMed

Shabani, Sahreh; Farbood, Yaghoob; Mard, Seyyed Ali; Sarkaki, Alireza; Ahangarpour, Akram; Khorsandi, Layasadat

2018-03-01

Alzheimer's disease (AD) is associated with decreased serum levels of thyroid hormones (THs), increased levels of thyroid-stimulating hormone (TSH), and decreased protein expression of brain-derived neurotrophic factor (BDNF) and reelin in the hippocampus. In this study, we have evaluated the effect of subcutaneous administration of levothyroxine (L-T 4 ) on levels of THs and TSH as well as protein expression of BDNF and reelin in AD rats. To make an animal model of AD, amyloid-beta peptide (Aβ) plus ibotenic acid were infused intrahippocampally, and rats were treated with L-T 4 and (or) saline for 10 days. The levels of THs and TSH were measured by ELISA kits. Protein synthesis was detected by Western blotting method. Results have been shown that serum level of THs, BDNF, and reelin protein expression in the hippocampus were significantly decreased (P < 0.001) in AD animals and elevated significantly in AD rats treated with L-T 4 (P < 0.01). Data showed that TSH level significantly decreased in AD rats treated with L-T 4 (P < 0.05). These findings indicated that L-T 4 increased BDNF and reelin protein expression by regulation of serum THs and TSH level in Aβ-induced AD rats.

Induction of Tissue Factor Pathway Inhibitor 2 by hCG Regulates Periovulatory Gene Expression and Plasmin Activity.

PubMed

Puttabyatappa, Muraly; Al-Alem, Linah F; Zakerkish, Farnosh; Rosewell, Katherine L; Brännström, Mats; Curry, Thomas E

2017-01-01

Increased proteolytic activity is a key event that aids in breakdown of the follicular wall to permit oocyte release. How the protease activity is regulated is still unknown. We hypothesize that tissue factor pathway inhibitor 2 (TFPI2), a Kunitz-type serine protease inhibitor, plays a role in regulating periovulatory proteolytic activity as in other tissues. TFPI2 is secreted into the extracellular matrix (ECM) where it is postulated to regulate physiological ECM remodeling. The expression profile of TFPI2 during the periovulatory period was assessed utilizing a well-characterized human menstrual cycle model and a gonadotropin-primed rat model. Administration of an ovulatory dose of human chorionic gonadotropin (hCG) increased TFPI2 expression dramatically in human and rat granulosa and theca cells. This increase in Tfpi2 expression in rat granulosa cells required hCG-mediated epidermal growth factor, protein kinase A, mitogen-activated protein kinase (MAPK) 1/2, p38 MAPK and protease activated receptor 1-dependent cell signaling. A small interferingRNA-mediated knockdown of TFPI2 in rat granulosa cells resulted in increased plasmin activity in the granulosa cell conditioned media. Knockdown of TFPI2 also reduced expression of multiple genes including interleukin 6 (Il6) and amphiregulin (Areg). Overexpression of TFPI2 using an adenoviral vector partially restored the expression of Il6 and Areg in TFPI2 siRNA treated rat granulosa cells. These data support the hypothesis that TFPI2 is important for moderating plasmin activity and regulating granulosa cell gene expression during the periovulatory period. We, therefore, propose that through these actions, TFPI2 aids in the tissue remodeling taking place during follicular rupture and corpus luteum formation. Copyright © 2017 by the Endocrine Society.

Induction of Tissue Factor Pathway Inhibitor 2 by hCG Regulates Periovulatory Gene Expression and Plasmin Activity

PubMed Central

Puttabyatappa, Muraly; Al-Alem, Linah F.; Zakerkish, Farnosh; Rosewell, Katherine L.; Brännström, Mats

2017-01-01

Increased proteolytic activity is a key event that aids in breakdown of the follicular wall to permit oocyte release. How the protease activity is regulated is still unknown. We hypothesize that tissue factor pathway inhibitor 2 (TFPI2), a Kunitz-type serine protease inhibitor, plays a role in regulating periovulatory proteolytic activity as in other tissues. TFPI2 is secreted into the extracellular matrix (ECM) where it is postulated to regulate physiological ECM remodeling. The expression profile of TFPI2 during the periovulatory period was assessed utilizing a well-characterized human menstrual cycle model and a gonadotropin-primed rat model. Administration of an ovulatory dose of human chorionic gonadotropin (hCG) increased TFPI2 expression dramatically in human and rat granulosa and theca cells. This increase in Tfpi2 expression in rat granulosa cells required hCG-mediated epidermal growth factor, protein kinase A, mitogen-activated protein kinase (MAPK) 1/2, p38 MAPK and protease activated receptor 1-dependent cell signaling. A small interferingRNA-mediated knockdown of TFPI2 in rat granulosa cells resulted in increased plasmin activity in the granulosa cell conditioned media. Knockdown of TFPI2 also reduced expression of multiple genes including interleukin 6 (Il6) and amphiregulin (Areg). Overexpression of TFPI2 using an adenoviral vector partially restored the expression of Il6 and Areg in TFPI2 siRNA treated rat granulosa cells. These data support the hypothesis that TFPI2 is important for moderating plasmin activity and regulating granulosa cell gene expression during the periovulatory period. We, therefore, propose that through these actions, TFPI2 aids in the tissue remodeling taking place during follicular rupture and corpus luteum formation. PMID:27813674

Maternal docosahexaenoic acid increases adiponectin and normalizes IUGR-induced changes in rat adipose deposition.

PubMed

Bagley, Heidi N; Wang, Yan; Campbell, Michael S; Yu, Xing; Lane, Robert H; Joss-Moore, Lisa A

2013-01-01

Intrauterine growth restriction (IUGR) predisposes to obesity and adipose dysfunction. We previously demonstrated IUGR-induced increased visceral adipose deposition and dysregulated expression of peroxisome proliferator activated receptor- γ 2 (PPAR γ 2) in male adolescent rats, prior to the onset of obesity. In other studies, activation of PPAR γ increases subcutaneous adiponectin expression and normalizes visceral adipose deposition. We hypothesized that maternal supplementation with docosahexaenoic acid (DHA), a PPAR γ agonist, would normalize IUGR adipose deposition in association with increased PPAR γ , adiponectin, and adiponectin receptor expression in subcutaneous adipose. To test these hypotheses, we used a well-characterized model of uteroplacental-insufficiency-(UPI-) induced IUGR in the rat with maternal DHA supplementation. Our primary findings were that maternal DHA supplementation during rat pregnancy and lactation (1) normalizes IUGR-induced changes in adipose deposition and visceral PPAR γ expression in male rats and (2) increases serum adiponectin, as well as adipose expression of adiponectin and adiponectin receptors in former IUGR rats. Our novel findings suggest that maternal DHA supplementation may normalize adipose dysfunction and promote adiponectin-induced improvements in metabolic function in IUGR.

[Induction of hypoxia-inducible factor-1alpha in two kinds of rats asphyxiation death models].

PubMed

Zhang, Bei-lei; Yang, Zhi-hui; Ran, Peng; Liang, Wei-bo; Zhou, Bin; Zhang, Geng-qian; Lu, Mei-li; Zhang, Lin

2007-02-15

To investigate the expression of hypoxia-inducible factor 1-alpha (HIF1-alpha) in the heart, lung, liver and kidney in rats died of two typical models of asphyxia. Two asphyxia models were made and tissue samples of the dead rats were collected from different groups at various postmortem duration. The expression and the changes of HIF1-alpha in various tissues were examined by immunohistochemistry and image analysis techniques. Results Significant expression of HIF1-alpha was observed in the myocardial fibers, kidney cells, liver cells and lung cells in both asphyxia models, but not in the control group. The expression of HIF1-alpha in various tissues in the rat died of nitrogen gas breathing was found in the nuclei at 0 hour and the expression level decreased gradually thereafter. The HIF1-alpha expression level and duration in various tissues of the rat died of hanging were higher and longer than that of the former group, with a peak of the expression level observed 6 hours after death, and then started to decline in all tissues except the heart where the expression still showed an increase 24 hours after death. The control groups showed a steady expression in the cytoplasm but not in the nuclei. HIF1-alpha appears to be a valuable biomarker in the diagnosis of asphyxia within 24 hours after death.

Characterization of docosahexaenoic acid (DHA)-induced heme oxygenase-1 (HO-1) expression in human cancer cells: the importance of enhanced BTB and CNC homology 1 (Bach1) degradation.

PubMed

Wang, Shuai; Hannafon, Bethany N; Wolf, Roman F; Zhou, Jundong; Avery, Jori E; Wu, Jinchang; Lind, Stuart E; Ding, Wei-Qun

2014-05-01

The effect of docosahexaenoic acid (DHA) on heme oxygenase-1 (HO-1) expression in cancer cells has never been characterized. This study examines DHA-induced HO-1 expression in human cancer cell model systems. DHA enhanced HO-1 gene expression in a time- and concentration-dependent manner, with maximal induction at 21 h of treatment. This induction of HO-1 expression was confirmed in vivo using a xenograft nude mouse model fed a fish-oil-enriched diet. The increase in HO-1 gene transcription induced by DHA was significantly attenuated by the antioxidant N-acetyl cysteine, suggesting the involvement of oxidative stress. This was supported by direct measurement of lipid peroxide levels after DHA treatment. Using a human HO-1 gene promoter reporter construct, we identified two antioxidant response elements (AREs) that mediate the DHA-induced increase in HO-1 gene transcription. Knockdown of nuclear factor (erythroid-derived 2)-like 2 (Nrf2) expression compromised the DHA-induced increase in HO-1 gene transcription, indicating the importance of the Nrf2 pathway in this event. However, the nuclear protein levels of Nrf2 remained unchanged upon DHA treatment. Further studies demonstrated that DHA reduces nuclear Bach1 protein expression by promoting its degradation and attenuates Bach1 binding to the AREs in the HO-1 gene promoter. In contrast, DHA enhanced Nrf2 binding to the AREs without affecting nuclear Nrf2 expression levels, indicating a new cellular mechanism that mediates DHA's induction of HO-1 gene transcription. To our knowledge, this is the first characterization of DHA-induced HO-1 expression in human malignant cells. Copyright © 2014 Elsevier Inc. All rights reserved.

Mechanisms of Neuroprotection from Hypoxia-Ischemia (HI) Brain Injury by Up-regulation of Cytoglobin (CYGB) in a Neonatal Rat Model*

PubMed Central

Tian, Shu-Feng; Yang, Han-Hua; Xiao, Dan-Ping; Huang, Yue-Jun; He, Gu-Yu; Ma, Hai-Ran; Xia, Fang; Shi, Xue-Chuan

2013-01-01

This study was designed to investigate the expression profile of CYGB, its potential neuroprotective function, and underlying molecular mechanisms using a model of neonatal hypoxia-ischemia (HI) brain injury. Cygb mRNA and protein expression were evaluated within the first 36 h after the HI model was induced using RT-PCR and Western blotting. Cygb mRNA expression was increased at 18 h in a time-dependent manner, and its level of protein expression increased progressively in 24 h. To verify the neuroprotective effect of CYGB, a gene transfection technique was employed. Cygb cDNA and shRNA delivery adenovirus systems were established (Cygb-cDNA-ADV and Cygb-shRNA-ADV, respectively) and injected into the brains of 3-day-old rats 4 days before they were induced with HI treatment. Rats from different groups were euthanized 24 h post-HI, and brain samples were harvested. 2,3,5-Triphenyltetrazolium chloride, TUNEL, and Nissl staining indicated that an up-regulation of CYGB resulted in reduced acute brain injury. The superoxide dismutase level was found to be dependent on expression of CYGB. The Morris water maze test in 28-day-old rats demonstrated that CYGB expression was associated with improvement of long term cognitive impairment. Studies also demonstrated that CYGB can up-regulate mRNA and protein levels of VEGF and increase both the density and diameter of the microvessels but inhibits activation of caspase-2 and -3. Thus, this is the first in vivo study focusing on the neuroprotective role of CYGB. The reduction of neonatal HI injury by CYGB may be due in part to antioxidant and antiapoptotic mechanisms and by promoting angiogenesis. PMID:23585565

Effect of compound Maqin decoction on TGF-β1/Smad proteins and IL-10 and IL-17 content in lung tissue of asthmatic rats.

PubMed

Xie, Y H; Li, X P; Xu, Z X; Qian, P; Li, X L; Wang, Y Q

2016-09-02

In this research, compound Maqin decoction (CMD) has been shown to positively affect in airway inflammation of asthma models. We evaluated the effects of CMD on the expression of transforming growth factor (TGF)-β1/Smad proteins, interleukin (IL)-17, and IL-10 in lung tissue of asthmatic rats. Asthma was induced in a rat model using ovalbumin. After a 4-week treatment with CMD, rats were killed to evaluate the expression of TGF-β1 and Smad proteins in lung tissue. IL-10 and IL-17 levels in lung tissue homogenates were determined by ELISA. The expression of TGF-β1 and Smad3 protein increased, whereas expression of Smad7 protein decreased upon high-dose or low-dose treatment with CMD or by intervention with dexamethasone, compared to the control. There was a significant difference between treatment with a high dose CMD and the control treatment, but no significant difference was found between high-dose CMD treatment and dexamethasone intervention. The expression of TGF-β1 and Smad7 protein increased, whereas the expression of Smad3 protein decreased in the model group compared to other groups. In the CMD high-dose group, low-dose group, and dexamethasone intervention group, the IL-17 concentrations in lung tissue homogenates were decreased, while IL-10 levels were increased. Again, there was a significant difference between CMD high-dose and control treatment, but not between CMD high-dose treatment and dexamethasone intervention. Thus, positive effects of CMD against asthmatic airway remodeling may be due to its regulatory effect on TGF-β1, Smad3, and Smad7 protein levels and on cytokines such as IL-10 and IL-17.

Increased pulmonary RhoA expression in the nitrofen-induced congenital diaphragmatic hernia rat model.

PubMed

Takayasu, Hajime; Masumoto, Kouji; Hagiwara, Koki; Sasaki, Takato; Ono, Kentaro; Jimbo, Takahiro; Uesugi, Toru; Gotoh, Chikashi; Urita, Yasuhisa; Shinkai, Toko; Tanaka, Hideaki

2015-09-01

Persistent pulmonary hypertension remains a major cause of mortality and morbidity in cases of congenital diaphragmatic hernia (CDH). Recently, RhoA/Rho-kinase-mediated vasoconstriction has been reported to be important in the pathogenesis of pulmonary hypertension (PH). Several recent reports have described that fasudil, a potent Rho-kinase inhibitor and vasodilator, could represent a potential therapeutic option for PH. We designed this study to investigate the hypothesis that the expression level of RhoA is increased in the nitrofen-induced CDH rat model. The expression level of Wnt11, an activator of RhoA, was also evaluated. Pregnant rats were treated with or without nitrofen on gestational day 9 (D9). Fetuses were sacrificed on D17, D19 and D21 and were divided into control and CDH groups. Quantitative real-time polymerase chain reaction was performed to determine the pulmonary gene expression levels of both Wnt11 and RhoA. An immunofluorescence study was also performed to evaluate the expression and localization of RhoA. The relative mRNA expression levels of pulmonary Wnt11 and RhoA on D21 were significantly increased in the CDH group compared with the control group (p=0.016 and p=0.008, respectively). The immunofluorescence study confirmed the overexpression of RhoA in the pulmonary vessels of CDH rats on D21. Our results provide evidence that the RhoA/Rho-kinase-mediated pathway is involved in the pathogenesis of PH in the nitrofen-induced CDH rat model. Our data also suggest that the fasudil, a Rho-kinase inhibitor, could represent a therapeutic option for the treatment of PH in CDH. Copyright © 2015 Elsevier Inc. All rights reserved.

A novel porcine ex vivo retina culture model for oxidative stress induced by H₂O₂.

PubMed

Hurst, José; Kuehn, Sandra; Jashari, Adelina; Tsai, Teresa; Bartz-Schmidt, Karl Ulrich; Schnichels, Sven; Joachim, Stephanie C

2017-03-01

Oxidative stress is a key player in many ophthalmic diseases. However, the role of oxidative stress in most degenerative processes is not yet known. Therefore, accurate and practical models are required to efficiently screen for therapeutics. Porcine eyes are closely related to the human eye, and can be obtained from the abattoir as a by-product of the food industry. Therefore, they offer excellent opportunities for the development of culture models with which to pre-screen potential therapies, while reducing the use of laboratory animals. To induce oxidative stress, organotypic cultures of porcine retina were treated with different doses of hydrogen peroxide (H₂O₂; 100, 300 and 500μM) for three hours. On days 3 and 8, the retinas were conserved for histological and Western blotting analyses and for evaluation of gene expression, which determined the number of retinal ganglion cells (RGCs), the activation state of glial cells, and the expression levels of several oxidative stress markers. H₂O₂ treatment led to a reduction in the number of RGCs and to an increase in apoptotic RGCs. In addition, a dose-dependent increase of microglia and an elevation of CD11b expression was observed. On day 3, a reduction of IL-1β, and an increase of iNOS, as well as of HSP70 mRNA were found. On day 8, an increase in TNF-α and IL-1β mRNA expression was detected. In conclusion, this ex vivo model offers an opportunity to study the molecular mechanisms underlying certain eye disorders and to test new therapeutic approaches to diminish the effects of oxidative stress. 2017 FRAME.

Immunomodulators targeting MARCO expression improve resistance to postinfluenza bacterial pneumonia.

PubMed

Wu, Muzo; Gibbons, John G; DeLoid, Glen M; Bedugnis, Alice S; Thimmulappa, Rajesh K; Biswal, Shyam; Kobzik, Lester

2017-07-01

Downregulation of the alveolar macrophage (AM) receptor with collagenous structure (MARCO) leads to susceptibility to postinfluenza bacterial pneumonia, a major cause of morbidity and mortality. We sought to determine whether immunomodulation of MARCO could improve host defense and resistance to secondary bacterial pneumonia. RNAseq analysis identified a striking increase in MARCO expression between days 9 and 11 after influenza infection and indicated important roles for Akt and Nrf2 in MARCO recovery. In vitro, primary human AM-like monocyte-derived macrophages (AM-MDMs) and THP-1 macrophages were treated with IFNγ to model influenza effects. Activators of Nrf2 (sulforaphane) or Akt (SC79) caused increased MARCO expression and a MARCO-dependent improvement in phagocytosis in IFNγ-treated cells and improved survival in mice with postinfluenza pneumococcal pneumonia. Transcription factor analysis also indicated a role for transcription factor E-box (TFEB) in MARCO recovery. Overexpression of TFEB in THP-1 cells led to marked increases in MARCO. The ability of Akt activation to increase MARCO expression in IFNγ-treated AM-MDMs was abrogated in TFEB-knockdown cells, indicating Akt increases MARCO expression through TFEB. Increasing MARCO expression by targeting Nrf2 signaling or the Akt-TFEB-MARCO pathway are promising strategies to improve bacterial clearance and survival in postinfluenza bacterial pneumonia. Copyright © 2017 the American Physiological Society.

Synthesis and deposition of basement membrane proteins by primary brain capillary endothelial cells in a murine model of the blood-brain barrier.

PubMed

Thomsen, Maj Schneider; Birkelund, Svend; Burkhart, Annette; Stensballe, Allan; Moos, Torben

2017-03-01

The brain vascular basement membrane is important for both blood-brain barrier (BBB) development, stability, and barrier integrity and the contribution hereto from brain capillary endothelial cells (BCECs), pericytes, and astrocytes of the BBB is probably significant. The aim of this study was to analyse four different in vitro models of the murine BBB for expression and possible secretion of major basement membrane proteins from murine BCECs (mBCECs). mBCECs, pericytes and glial cells (mainly astrocytes and microglia) were prepared from brains of C57BL/6 mice. The mBCECs were grown as monoculture, in co-culture with pericytes or mixed glial cells, or as a triple-culture with both pericytes and mixed glial cells. The integrity of the BBB models was validated by measures of transendothelial electrical resistance (TEER) and passive permeability to mannitol. The expression of basement membrane proteins was analysed using RT-qPCR, mass spectrometry and immunocytochemistry. Co-culturing mBCECs with pericytes, mixed glial cells, or both significantly increased the TEER compared to the monoculture, and a low passive permeability was correlated with high TEER. The mBCECs expressed all major basement membrane proteins such as laminin-411, laminin-511, collagen [α1(IV)] 2 α2(IV), agrin, perlecan, and nidogen 1 and 2 in vitro. Increased expression of the laminin α5 subunit correlated with the addition of BBB-inducing factors (hydrocortisone, Ro 20-1724, and pCPT-cAMP), whereas increased expression of collagen IV α1 primarily correlated with increased levels of cAMP. In conclusion, BCECs cultured in vitro coherently form a BBB and express basement membrane proteins as a feature of maturation. Cover Image for this issue: doi: 10.1111/jnc.13789. © 2016 International Society for Neurochemistry.

Telomere length and telomere repeating factors: Cellular markers for post-traumatic stress disorder-like model.

PubMed

Dong, Yuanjun; Zhang, Guiqing; Yuan, Xiuyu; Zhang, Yueqi; Hu, Min

2016-05-01

The aim of the present study was to explore the telomere length of peripheral blood leukocytes from a rat model of post-traumatic stress disorder (PTSD), as well as the expression level of telomere-binding protein in the hippocampal CA1 region. The PTSD model was established with 42 adult male Wistar rats. The relative telomere length of the leukocytes was measured by real-time fluorescence quantitative polymerase chain reaction, and the expression levels of telomere repeating factor 1 (TRF1) and telomere repeating factor 2 (TRF2) in the hippocampal CA1 region of the PTSD rat model were determined by immunofluorescence technology. The covariance analysis of repeated measurements by the mixed model approach was used for the telomere length analysis. The comparison of averaged data among groups was performed using least significant difference and analysis of variance. The Student's t test or the Mann-Whitney U test was used for intragroup comparison. The association study among groups was conducted using the Spearman test. The shortening speed of telomere length significantly accelerated in rats after Single Prolonged Stress (SPS) stimulation (P<0.05). The expression levels of TRF1 and TRF2 increased with the progress of PTSD, and the expression peak was shown in day 14, which was significantly different from the control group (P<0.05). The shortening speed of the telomere length of peripheral blood leukocytes accelerated in PTSD rats, and the expression levels of TRF1 and TRF2 increased in hippocampus, both of which were closely associated with the pathological progress of the PTSD-like model and unfavorable prognosis. Copyright © 2016 Elsevier B.V. All rights reserved.

Comparison of protective effects of safflor injection and extract of Ginkgo biloba on lung ischemia/reperfusion injury in rabbits.

PubMed

Tian, Xiao-xi; Wang, Bo-liang; Cao, Yi-zhan; Zhong, Yue-xia; Tu, Yan-yang; Xiao, Jian-bo; He, Qian-feng; Zhai, Li-na

2015-03-01

To observe the protective effects of safflor Injection (SI) and extract of Ginkgo biloba (EGB) on lung ischemia-reperfusion injury (LIRI) and investigate its mechanism. In vivo rabbit model of LIRI was reconstructed. Forty rabbits were randomly and equally divided into four groups: sham-operation group (sham group), ischemia-reperfusion group (model group), ischemia-reperfusion plus SI group (safflor group) and ischemia-reperfusion plus EGB injection group (EGB group). Malondialdehyde (MDA) content, superoxide dismutase (SOD) and xanthine oxidase (XO) activity in serum were measured. The wet/dry weight ratio (W/D) of the lung tissue and activity of myeloperoxidase (MPO) were also tested. Ultrastructure change of the lung tissue was observed by the electron microscope. The expression of intercellular adhesion molecule-1 (ICAM-1) was measured by immunohistochemistry (IHC). In the model group, MDA and XO increased and SOD decreased in serum compared with the sham group (P<0.01). The values of W/D, MPO and ICAM-1 of the model group were higher than those of the sham group (P<0.01), but those of the safflor group and EGB group were significantly lower than those of the model group (P<0.01). The IHC demonstrated that ICAM-1 expression in lung tissue of the model group was significantly higher than those of the safflor group (P<0.01). Compared with safflor group, in the EGB group MDA, XO, MPO decreased, SOD and ICAM-1 expression increased (P<0.05), but the change of W/D was not statistically significant (P>0.05). SI and EGB may attenuate LIRI through antioxidation, inhibition of neutrophil aggregation and down-regulation of ICAM-1 expression. But EGB had more effect on the antioxidation, while SI did better on regulating ICAM-1 expression.

Intra-tubular hydrodynamic forces influence tubulo-interstitial fibrosis in the kidney

PubMed Central

Rohatgi, Rajeev; Flores, Daniel

2010-01-01

Purpose of review Renal epithelial cells respond to mechanical stimuli with immediate transduction events (e.g., activation of ion channels), intermediate biological responses (e.g., changes in gene expression), and long term cellular adaptation (e.g., protein expression). Progressive renal disease is characterized by disturbed glomerular hydrodynamics that contributes to glomerulosclerosis, but, how intra-tubular biomechanical forces contribute to tubulo-interstital inflammation and fibrosis is poorly understood. Recent findings In vivo and in vitro models of obstructive uropathy demonstrate that tubular stretch induces robust expression of transforming growth factor β-1 (TGFβ-1), activation of tubular apoptosis, and induction of NF-κB signaling which contribute to the inflammatory and fibrotic milieu. Non-obstructive structural kidney diseases associated with nephron loss follow a course characterized by compensatory increases of single nephron glomerular filtration rate and tubular flow rate. Resulting increases in tubular fluid shear stress (FSS) reduce tissue-plasminogen activator and urokinase enzymatic activity which diminishes breakdown of extracellular matrix. In models of high dietary Na intake, which increase tubular flow, urinary TGFβ-1 concentrations and renal mitogen activated protein kinase activity are increased. Summary In conclusion, intra-tubular biomechanical forces, stretch and FSS, generate changes in intracellular signaling and gene expression that contribute to the pathobiology of obstructive, and non-obstructive kidney disease. PMID:19851105

Effect of various classes of pesticides on expression of stress genes in transgenic C. elegans model of Parkinson's disease.

PubMed

Jadiya, Pooja; Mir, Snober S; Nazir, Aamir

2012-12-01

Neurodegenerative diseases are known to be associated with genetic and environmental factors. The multifactorial Parkinson's disease (PD) is triggered and/or further worsened by exposure to certain pesticides. Existing literature suggests a link between pesticide exposure and increased incidence of PD. We carried out the present study to look into the stress gene expression pattern of transgenic Caenorhabditis elegans (C. elegans) model of PD after exposure to pesticides from different classes. Expression level of sod-1, sod-2, sod-3, hsp-70, hsp-60, and hsp-16.2 stress responsive genes was determined using qPCR. Our findings demonstrate that the expression of stress related genes does not follow a generalized pattern to different toxicants; rather each pesticide class has a specific expression signature.

RON is not a prognostic marker for resectable pancreatic cancer.

PubMed

Tactacan, Carole M; Chang, David K; Cowley, Mark J; Humphrey, Emily S; Wu, Jianmin; Gill, Anthony J; Chou, Angela; Nones, Katia; Grimmond, Sean M; Sutherland, Robert L; Biankin, Andrew V; Daly, Roger J

2012-09-07

The receptor tyrosine kinase RON exhibits increased expression during pancreatic cancer progression and promotes migration, invasion and gemcitabine resistance of pancreatic cancer cells in experimental models. However, the prognostic significance of RON expression in pancreatic cancer is unknown. RON expression was characterized in several large cohorts, including a prospective study, totaling 492 pancreatic cancer patients and relationships with patient outcome and clinico-pathologic variables were assessed. RON expression was associated with outcome in a training set, but this was not recapitulated in the validation set, nor was there any association with therapeutic responsiveness in the validation set or the prospective study. Although RON is implicated in pancreatic cancer progression in experimental models, and may constitute a therapeutic target, RON expression is not associated with prognosis or therapeutic responsiveness in resected pancreatic cancer.

SDF-1 promotes endochondral bone repair during fracture healing at the traumatic brain injury condition.

PubMed

Liu, Xiaoqi; Zhou, Changlong; Li, Yanjing; Ji, Ye; Xu, Gongping; Wang, Xintao; Yan, Jinglong

2013-01-01

The objective of this study was to investigate the role of stromal cell-derived factor-1 (SDF-1) and its receptor, CXCR4, on bone healing and whether SDF-1 contributes to accelerating bone repair in traumatic brain injury (TBI)/fracture model. Real-time polymerase chain reaction and immunohistochemical analysis were used to detect the expression of SDF-1 during the repair of femoral bone in TBI/fracture model. The TBI/fracture model was treated with anti-SDF-1 neutralizing antibody or AMD3100, an antagonist for CXCR4, and evaluated by histomorphometry. In vitro and in vivo migration assays were used to evaluate the functional effect of SDF-1 on primary mesenchymal stem cells. The expression of SDF1 and CXCR4 messenger RNA was increased during the bone healing in TBI/fracture model but was less increased in fracture only model. High expression of SDF-1 protein was observed in the surrounding tissue of the damaged bone. Treated with anti-SDF-1 antibody or AMD3100 could inhibit new bone formation. SDF-1 increased mesenchymal stem cell chemotaxis in vitro in a dose-dependent manner. The in vivo migration study demonstrated that mesenchymal stem cells recruited by SDF-1 participate in endochondral bone repair. The SDF-1/CXCR4 axis plays a crucial role in the accelerating fracture healing under the condition of TBI and contributes to endochondral bone repair.

The transcriptional landscape of age in human peripheral blood

PubMed Central

Peters, Marjolein J.; Joehanes, Roby; Pilling, Luke C.; Schurmann, Claudia; Conneely, Karen N.; Powell, Joseph; Reinmaa, Eva; Sutphin, George L.; Zhernakova, Alexandra; Schramm, Katharina; Wilson, Yana A.; Kobes, Sayuko; Tukiainen, Taru; Nalls, Michael A.; Hernandez, Dena G.; Cookson, Mark R.; Gibbs, Raphael J.; Hardy, John; Ramasamy, Adaikalavan; Zonderman, Alan B.; Dillman, Allissa; Traynor, Bryan; Smith, Colin; Longo, Dan L.; Trabzuni, Daniah; Troncoso, Juan; van der Brug, Marcel; Weale, Michael E.; O'Brien, Richard; Johnson, Robert; Walker, Robert; Zielke, Ronald H.; Arepalli, Sampath; Ryten, Mina; Singleton, Andrew B.; Ramos, Yolande F.; Göring, Harald H. H.; Fornage, Myriam; Liu, Yongmei; Gharib, Sina A.; Stranger, Barbara E.; De Jager, Philip L.; Aviv, Abraham; Levy, Daniel; Murabito, Joanne M.; Munson, Peter J.; Huan, Tianxiao; Hofman, Albert; Uitterlinden, André G.; Rivadeneira, Fernando; van Rooij, Jeroen; Stolk, Lisette; Broer, Linda; Verbiest, Michael M. P. J.; Jhamai, Mila; Arp, Pascal; Metspalu, Andres; Tserel, Liina; Milani, Lili; Samani, Nilesh J.; Peterson, Pärt; Kasela, Silva; Codd, Veryan; Peters, Annette; Ward-Caviness, Cavin K.; Herder, Christian; Waldenberger, Melanie; Roden, Michael; Singmann, Paula; Zeilinger, Sonja; Illig, Thomas; Homuth, Georg; Grabe, Hans-Jörgen; Völzke, Henry; Steil, Leif; Kocher, Thomas; Murray, Anna; Melzer, David; Yaghootkar, Hanieh; Bandinelli, Stefania; Moses, Eric K.; Kent, Jack W.; Curran, Joanne E.; Johnson, Matthew P.; Williams-Blangero, Sarah; Westra, Harm-Jan; McRae, Allan F.; Smith, Jennifer A.; Kardia, Sharon L. R.; Hovatta, Iiris; Perola, Markus; Ripatti, Samuli; Salomaa, Veikko; Henders, Anjali K.; Martin, Nicholas G.; Smith, Alicia K.; Mehta, Divya; Binder, Elisabeth B.; Nylocks, K Maria; Kennedy, Elizabeth M.; Klengel, Torsten; Ding, Jingzhong; Suchy-Dicey, Astrid M.; Enquobahrie, Daniel A.; Brody, Jennifer; Rotter, Jerome I.; Chen, Yii-Der I.; Houwing-Duistermaat, Jeanine; Kloppenburg, Margreet; Slagboom, P. Eline; Helmer, Quinta; den Hollander, Wouter; Bean, Shannon; Raj, Towfique; Bakhshi, Noman; Wang, Qiao Ping; Oyston, Lisa J.; Psaty, Bruce M.; Tracy, Russell P.; Montgomery, Grant W.; Turner, Stephen T.; Blangero, John; Meulenbelt, Ingrid; Ressler, Kerry J.; Yang, Jian; Franke, Lude; Kettunen, Johannes; Visscher, Peter M.; Neely, G. Gregory; Korstanje, Ron; Hanson, Robert L.; Prokisch, Holger; Ferrucci, Luigi; Esko, Tonu; Teumer, Alexander; van Meurs, Joyce B. J.; Johnson, Andrew D.

2015-01-01

Disease incidences increase with age, but the molecular characteristics of ageing that lead to increased disease susceptibility remain inadequately understood. Here we perform a whole-blood gene expression meta-analysis in 14,983 individuals of European ancestry (including replication) and identify 1,497 genes that are differentially expressed with chronological age. The age-associated genes do not harbor more age-associated CpG-methylation sites than other genes, but are instead enriched for the presence of potentially functional CpG-methylation sites in enhancer and insulator regions that associate with both chronological age and gene expression levels. We further used the gene expression profiles to calculate the ‘transcriptomic age' of an individual, and show that differences between transcriptomic age and chronological age are associated with biological features linked to ageing, such as blood pressure, cholesterol levels, fasting glucose, and body mass index. The transcriptomic prediction model adds biological relevance and complements existing epigenetic prediction models, and can be used by others to calculate transcriptomic age in external cohorts. PMID:26490707

Generation of a mouse model with a reversible hypomorphic cytochrome P450 reductase gene: utility for tissue-specific rescue of the reductase expression, and insights from a resultant mouse model with global suppression of P450 reductase expression in extrahepatic tissues.

PubMed

Wei, Yuan; Zhou, Xin; Fang, Cheng; Li, Lei; Kluetzman, Kerri; Yang, Weizhu; Zhang, Qing-Yu; Ding, Xinxin

2010-07-01

A mouse model termed Cpr-low (CL) was recently generated, in which the expression of the cytochrome P450 reductase (Cpr) gene was globally down-regulated. The decreased CPR expression was accompanied by phenotypical changes, including reduced embryonic survival, decreases in circulating cholesterol, increases in hepatic P450 expression, and female infertility (accompanied by elevated serum testosterone and progesterone levels). In the present study, a complementary mouse model [named reversible-CL (r-CL)] was generated, in which the reduced CPR expression can be reversed in an organ-specific fashion. The neo cassette, which was inserted into the last Cpr intron in r-CL mice, can be deleted by Cre recombinase, thus returning the structure of the Cpr gene (and hence CPR expression) to normal in Cre-expressing cells. All previously identified phenotypes of the CL mice were preserved in the r-CL mice. As a first application of the r-CL model, we have generated an extrahepatic-CL (xh-CL) mouse for testing of the functions of CPR-dependent enzymes in all extrahepatic tissues. The xh-CL mice, generated by mating of r-CL mice with albumin-Cre mice, had normal CPR expression in hepatocytes but down-regulated CPR expression elsewhere. They were indistinguishable from wild-type mice in body and liver weights, circulating cholesterol levels, and hepatic microsomal P450 expression and activities; however, they still showed elevated serum testosterone and progesterone levels and sterility in females. Embryonic lethality was prevented in males, but apparently not in females, indicating a critical role for fetal hepatic CPR-dependent enzymes in embryonic development, at least in males.

Usefulness of acr expression for monitoring latent Mycobacterium tuberculosis bacilli in 'in vitro' and 'in vivo' experimental models.

PubMed

Gordillo, S; Guirado, E; Gil, O; Díaz, J; Amat, I; Molinos, S; Vilaplana, C; Ausina, V; Cardona, P-J

2006-07-01

Real-time RT-PCR was used to quantify the expression of genes possibly involved in Mycobacterium tuberculosis latency in in vitro and murine models. Exponential and stationary phase (EP and SP) bacilli were exposed to decreasing pH levels (from 6.5 to 4.5) in an unstirred culture, and mRNA levels for 16S rRNA, sigma factors sigA,B,E,F,G,H and M, Rv0834c, icl, nirA, narG, fpbB, acr, rpoA, recA and cysH were quantified. The expression of acr was the one that best correlated with the CFU decrease observed in SP bacilli. In the murine model, the expressions of icl, acr and sigF tended to decrease when bacillary counts increased and vice versa. Values from immunodepressed mice (e.g. alpha/beta T cells, TNF, IFN-gamma and iNOs knock out strains), with accelerated bacillary growth rate, confirmed this fact. Finally, the expression of acr was maintained in mice following long-term treatment with antibiotics. The quantification of acr expression could be useful for monitoring the presence of latent bacilli in some murine models of tuberculosis.

[Hepatocyte apoptosis and mitochondrial permeability transition pore opening in rats with nonalcoholic fatty liver].

PubMed

Kang, Min; Li, Sen; Zhong, Dejun; Yang, Zhimin; Li, Peng

2013-07-01

To investigate the role of hepatocyte apoptosis and mitochondrial permeability transition pore (MPTP) opening in the pathogenesis of nonalcoholic fatty liver disease (NAFLD). Thirty male SD rats were randomized into normal diet group and high-fat diet group. At 4, 8 and 12 week of feeding. The hepatocyte apoptosis index (AI) was measured using flow cytometry, and MPTP opening was evaluated with ultraviolet spectrophotometry. Immunohistochemistry was employed to detect hepatic expressions of Bcl-2 and Bax, and Western blotting was used to detect Bax protein expression changes. High-fat feeding resulted in significantly increased hepatocyte AI at 4-12 weeks and gradually increased MPTP opening. In the high-fat diet group, hepatic Bcl-2 expression was detected but the positive cell number remained stable, whereas Bax-positive cell number increased steadily with time with progressively increased intensity of Bax protein expression, resulting in gradually decreased Bcl-2/Bax ratio. Hepatocyte apoptosis occurs in the rat model of NAFLD in close correlation with mitochondrial damage. Increased MPTP opening as the result of increased Bax expression and aberrant Bcl-2/Bax ratio is an important mechanism of hepatocyte mitochondrial damage in NAFLD.

Novel stimuli are negative stimuli: evidence that negative affect is reduced in the mere exposure effect.

PubMed

Robinson, Brent M; Elias, Lorin J

2005-04-01

Repeated exposure of a nonreinforced stimulus results in an increased preference for that stimulus, the mere exposure effect. The present study repeatedly presented positive, negative, and neutrally affective faces to 48 participants while they made judgments about the emotional expression. Participants then rated the likeability of novel neutrally expressive faces and some of these previously presented faces, this time in their neutral expression. Faces originally presented as happy were rated as the most likeable, followed by faces originally presented as neutral. Negative and novel faces were not rated significantly differently from each other. These findings support the notion that the increase in preference towards repeatedly presented stimuli is the result of the reduction in negative affect, consistent with the modified two-factor uncertainty-reduction model and classical conditioning model of the mere exposure effect.

Effect of fenhexamid and cyprodinil on the expression of cell cycle- and metastasis-related genes via an estrogen receptor-dependent pathway in cellular and xenografted ovarian cancer models

DOE Office of Scientific and Technical Information (OSTI.GOV)

Go, Ryeo-Eun; Kim, Cho-Won; Choi, Kyung-Chul, E-mail: kchoi@cbu.ac.kr

ABSTRACT: Fenhexamid and cyprodinil are antifungal agents (pesticides) used for agriculture, and are present at measurable amounts in fruits and vegetables. In the current study, the effects of fenhexamid and cyprodinil on cancer cell proliferation and metastasis were examined. Additionally, the protein expression levels of cyclin D1 and cyclin E as well as cathepsin D were analyzed in BG-1 ovarian cancer cells that express estrogen receptors (ERs). The cells were cultured with 0.1% dimethyl sulfoxide (DMSO; control), 17β-estradiol (E2; 10{sup −9} M), and fenhexamid or cyprodinil (10{sup –5}–10{sup −7} M). Results of a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay showed that fenhexamidmore » and cyprodinil increased BG-1 cell proliferation about 1.5 to 2 times similar to E2 (5 times) compared to the control. When the cells were co-treated with ICI 182,780 (10{sup −8} M), an ER antagonist, the proliferation of pesticide-treated BG-1 cells was decreased to the level of the control. A wound healing assay revealed that the pesticides reduced the disrupted area in the BG-1 cell monolayer similar to E2. Protein levels of cyclin D1 and E as well as cathepsin D were increased by fenhexamid and cyprodinil. This effect was reversed by co-treatment with ICI 182,780. In a xenograft mouse model with transplanted BG-1 cells, cyprodinil significantly increased tumor mass formation about 2 times as did E2 (6 times) compared to the vehicle (0.1% DMSO) over an 80-day period. In contrast, fenhexamid did not promote ovarian tumor formation in this mouse model. Cyprodinil also induced cell proliferation along with the expression of proliferating cell nuclear antigen (PCNA) and cathepsin D in tumor tissues similar to E2. Taken together, these results imply that fenhexamid and cyprodinil may have disruptive effects on ER-expressing cancer by altering the cell cycle- and metastasis-related gene expression via an ER-dependent pathway. - Highlights: • Fenhexamid and cyprodinil increased BG-1 cell proliferation via ER. • Two pesticides reduced the disrupted area in the BG-1 cell monolayer. • Cyclin D1 and E and cathepsin D proteins were induced by fenhexamid and cyprodinil. • Cyprodinil significantly increased tumor mass formation in a xenografted model. • Fenhexamid and cyprodinil may have disruptive effects on ER-expressing cancer.« less

[Curcumin inhibited rat colorectal carcinogenesis by activating PPAR-γ: an experimental study].

PubMed

Liu, Liu-bin; Duan, Chang-nong; Ma, Zeng-yi; Xu, Gang

2015-04-01

To explore the chemopreventive effect of curcumin on DMH induced colorectal carcinogenesis and the underlining mechanism. Totally 40 Wistar rats were divided into the model group and the curcumin group by random digit table, 20 in each group. Meanwhile, a normal control group was set up (n =10). A colorectal cancer model was induced by subcutaneously injecting 20 mg/kg DMH. The tumor incidence and the inhibition rate were calculated. The effect of curcumin on the expression of peroxisome proliferator-activated receptor gamma (PPARγ) in rat colon mucosal tissues was observed using immunohistochemistry and Western blot. HT 29 cell line were cultured and divided into a control group, the curcumin + GW9662 (2-chloro-5-nitro-N-4-phenylbenzamide) intervention group, and the curcumin group. The inhibition of different concentrations curcumin on HT29 cell line was detected using MTT. The expression of curcumin on PPARy was also detected using Western blot. The tumor incidence was 80. 00% (12/15 cases) in the model group, obviously higher than that of the curcumin group (58. 82%, 10/17 cases, P <0. 05). The inhibition rate of curcumin on DMH induced colorected carcinoma reached 26. 46%. Compared with the normal control group, the expression of PPARγ protein was significantly increased in the curcumin group and the model group (P <0. 01). Compared with the model group at the same time point, the expression of PPARy protein was significantly enhanced in the curcumin group (P <0. 05). MTT analysis showed that curcumin could inhibit the proliferation of in vitro HT 29 cells in dose and time dependent manners. The expression of PPARy protein was significantly increased in the GW9662 group and the curcumin group, showing statistical difference when compared with the normal control group (P <0. 01). Compared with the GW9662 group, the expression of PPARγ protein was significantly increased in the curcumin group (P <0. 01). Curcumin could inhibit DMH-induced rat colorectal carcinogenesis and the growth of in vitro cultured HT 29 cell line, which might be achieved by activating PPARy signal transduction pathway.

Genome-Wide Gene Expression Analysis of Bordetella pertussis Isolates Associated with a Resurgence in Pertussis: Elucidation of Factors Involved in the Increased Fitness of Epidemic Strains

PubMed Central

King, Audrey J.; van der Lee, Saskia; Mohangoo, Archena; van Gent, Marjolein; van der Ark, Arno; van de Waterbeemd, Bas

2013-01-01

Bordetella pertussis (B. pertussis) is the causative agent of whooping cough, which is a highly contagious disease in the human respiratory tract. Despite vaccination since the 1950s, pertussis remains the most prevalent vaccine-preventable disease in developed countries. A recent resurgence pertussis is associated with the expansion of B. pertussis strains with a novel allele for the pertussis toxin (ptx) promoter ptxP3 in place of resident ptxP1 strains. The recent expansion of ptxP3 strains suggests that these strains carry mutations that have increased their fitness. Compared to the ptxP1 strains, ptxP3 strains produce more Ptx, which results in increased virulence and immune suppression. In this study, we investigated the contribution of gene expression changes of various genes on the increased fitness of the ptxP3 strains. Using genome-wide gene expression profiling, we show that several virulence genes had higher expression levels in the ptxP3 strains compared to the ptxP1 strains. We provide the first evidence that wildtype ptxP3 strains are better colonizers in an intranasal mouse infection model. This study shows that the ptxP3 mutation and the genetic background of ptxP3 strains affect fitness by contributing to the ability to colonize in a mouse infection model. These results show that the genetic background of ptxP3 strains with a higher expression of virulence genes contribute to increased fitness. PMID:23776625

Deletion of Galgt2 (B4Galnt2) Reduces Muscle Growth in Response to Acute Injury and Increases Muscle Inflammation and Pathology in Dystrophin-Deficient Mice

PubMed Central

Xu, Rui; Singhal, Neha; Serinagaoglu, Yelda; Chandrasekharan, Kumaran; Joshi, Mandar; Bauer, John A.; Janssen, Paulus M.L.; Martin, Paul T.

2016-01-01

Transgenic overexpression of Galgt2 (official name B4Galnt2) in skeletal muscle stimulates the glycosylation of α dystroglycan (αDG) and the up-regulation of laminin α2 and dystrophin surrogates known to inhibit muscle pathology in mouse models of congenital muscular dystrophy 1A and Duchenne muscular dystrophy. Skeletal muscle Galgt2 gene expression is also normally increased in the mdx mouse model of Duchenne muscular dystrophy compared with the wild-type mice. To assess whether this increased endogenous Galgt2 expression could affect disease, we quantified muscular dystrophy measures in mdx mice deleted for Galgt2 (Galgt2−/−mdx). Galgt2−/− mdx mice had increased heart and skeletal muscle pathology and inflammation, and also worsened cardiac function, relative to age-matched mdx mice. Deletion of Galgt2 in wild-type mice also slowed skeletal muscle growth in response to acute muscle injury. In each instance where Galgt2 expression was elevated (developing muscle, regenerating muscle, and dystrophic muscle), Galgt2-dependent glycosylation of αDG was also increased. Overexpression of Galgt2 failed to inhibit skeletal muscle pathology in dystroglycan-deficient muscles, in contrast to previous studies in dystrophin-deficient mdx muscles. This study demonstrates that Galgt2 gene expression and glycosylation of αDG are dynamically regulated in muscle and that endogenous Galgt2 gene expression can ameliorate the extent of muscle pathology, inflammation, and dysfunction in mdx mice. PMID:26435413

Hypoxia activates muscle-restricted coiled-coil protein (MURC) expression via transforming growth factor-β in cardiac myocytes.

PubMed

Shyu, Kou-Gi; Cheng, Wen-Pin; Wang, Bao-Wei; Chang, Hang

2014-03-01

The expression of MURC (muscle-restricted coiled-coil protein), a hypertrophy-regulated gene, increases during pressure overload. Hypoxia can cause myocardial hypertrophy; however, how hypoxia affects the regulation of MURC in cardiomyocytes undergoing hypertrophy is still unknown. The aim of the present study was to test the hypothesis that hypoxia induces MURC expression in cardiomyocytes during hypertrophy. The expression of MURC was evaluated in cultured rat neonatal cardiomyocytes subjected to hypoxia and in an in vivo model of AMI (acute myocardial infarction) to induce myocardial hypoxia in adult rats. MURC protein and mRNA expression were significantly enhanced by hypoxia. MURC proteins induced by hypoxia were significantly blocked after the addition of PD98059 or ERK (extracellular-signal-regulated kinase) siRNA 30 min before hypoxia. Gel-shift assay showed increased DNA-binding activity of SRF (serum response factor) after hypoxia. PD98059, ERK siRNA and an anti-TGF-β (transforming growth factor-β) antibody abolished the SRF-binding activity enhanced by hypoxia or exogenous administration of TGF-β. A luciferase promoter assay demonstrated increased transcriptional activity of SRF in cardiomyocytes by hypoxia. Increased βMHC (β-myosin heavy chain) and BNP (B-type natriuretic peptide) protein expression and increased protein synthesis was identified after hypoxia with the presence of MURC in hypertrophic cardiomyocytes. MURC siRNA inhibited the hypertrophic marker protein expression and protein synthesis induced by hypoxia. AMI in adult rats also demonstrated increased MURC protein expression in the left ventricular myocardium. In conclusion, hypoxia in cultured rat neonatal cardiomyocytes increased MURC expression via the induction of TGF-β, SRF and the ERK pathway. These findings suggest that MURC plays a role in hypoxia-induced hypertrophy in cardiomyocytes.

Influence of TRAIL gene on biomechanical properties of the human leukemic cell line Jurkat.

PubMed

Yao, Weijuan; Chen, Kai; Wang, Xinjuan; Xie, Lide; Wen, Zongyao; Yan, Zongyi; Chien, Shu

2002-12-01

We cloned the cDNA fragment of human TNF-related apoptosis inducing ligand (TRAIL) into RevTet-On, a Tet-regulated and high-level gene expression system. Making use of the TRAIL gene expression system in Jurkat as a cell model, we studied the influence of TRAIL gene on the biomechanics properties of Jurkat through measuring changes of cellular biomechanics properties before and after the TRAIL gene expression, which was induced by adding tetracycline derivative doxycycline (Dox). The results indicated that the TRAIL gene expression led to significant changes in cellular biomechanics properties. The osmotic fragility increased and the cell stiffness increased after the expression of TRAIL gene. Thus, the apoptosis-inducing TRAIL gene caused significant changes in the biomechanics properties of Jurkat cells.

Expression of Inflammation-related Intercellular Adhesion Molecules in Cardiomyocytes In Vitro and Modulation by Pro-inflammatory Agents.

PubMed

El-Battrawy, Ibrahim; Tülümen, Erol; Lang, Siegfried; Akin, Ibrahim; Behnes, Michael; Zhou, Xiabo; Mavany, Martin; Bugert, Peter; Bieback, Karen; Borggrefe, Martin; Elmas, Elif

2016-01-01

Cell-surface adhesion molecules regulate multiple intercellular and intracellular processes and play important roles in inflammation by facilitating leukocyte endothelial transmigration. Whether cardiomyocytes express surface-adhesion molecules related to inflammation and the effect of pro-inflammatory mediators remain unknown. In the present study, the expression of different cell-adhesion molecules (CD11a, CD11b, CD31, CD62P, CD162, F11 receptor and mucosal vascular addressin cell adhesion molecule 1 (MADCAM1)) and the effect of pro-inflammatory mediators were investigated in an in vitro model of human cardiomyocytes. Cells were supplied as a primary culture of cardiac alpha actin-positive cells from human heart tissue. The cells were incubated for 24 h with 1 U/ml thrombin or 700 ng/ml lipopolysaccharide (LPS) or with a combination of both. The expression of the cell adhesion molecules was measured by flow cytometry. In cultured human cardiomyocytes, 22.8% of cells expressed CD31, 7.1% MADCAM1 and 2.6% F11R. CD11a, CD11b, CD62P and CD162 were expressed by fewer than 2% of the cells at baseline. CD31 expression increased on incubation of cardiomyocytes with thrombin by 26% (p<0.05) and with LPS by 26% (p=0.06). The combination of thrombin and LPS did not result in increased levels of CD31 (p>0.10). The pro-inflammatory agents LPS and thrombin had no effect on the expression of MADCAM1 and F11R. Inflammation-related cell-adhesion molecules CD31, MADCAM1 and F11R were shown to be expressed on the surface of human cardiomyocytes in an in vitro model. Incubation with LPS or thrombin resulted in increased expression of CD31, however, it did not modify the expression of the cell adhesion molecules MADCAM1 and F11R. Copyright © 2016 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

GPR30 Regulates Glutamate Transporter GLT-1 Expression in Rat Primary Astrocytes*

PubMed Central

Lee, Eunsook; Sidoryk-Wêgrzynowicz, Marta; Wang, Ning; Webb, Anton; Son, Deok-Soo; Lee, Kyuwon; Aschner, Michael

2012-01-01

The G protein-coupled estrogen receptor GPR30 contributes to the neuroprotective effects of 17β-estradiol (E2); however, the mechanisms associated with this protection have yet to be elucidated. Given that E2 increases astrocytic expression of glutamate transporter-1 (GLT-1), which would prevent excitotoxic-induced neuronal death, we proposed that GPR30 mediates E2 action on GLT-1 expression. To investigate this hypothesis, we examined the effects of G1, a selective agonist of GPR30, and GPR30 siRNA on astrocytic GLT-1 expression, as well as glutamate uptake in rat primary astrocytes, and explored potential signaling pathways linking GPR30 to GLT-1. G1 increased GLT-1 protein and mRNA levels, subject to regulation by both MAPK and PI3K signaling. Inhibition of TGF-α receptor suppressed the G1-induced increase in GLT-1 expression. Silencing GPR30 reduced the expression of both GLT-1 and TGF-α and abrogated the G1-induced increase in GLT-1 expression. Moreover, the G1-induced increase in GLT-1 protein expression was abolished by a protein kinase A inhibitor and an NF-κB inhibitor. G1 also enhanced cAMP response element-binding protein (CREB), as well as both NF-κB p50 and NF-κB p65 binding to the GLT-1 promoter. Finally, to model dysfunction of glutamate transporters, manganese was used, and G1 was found to attenuate manganese-induced impairment in GLT-1 protein expression and glutamate uptake. Taken together, the present data demonstrate that activation of GPR30 increases GLT-1 expression via multiple pathways, suggesting that GPR30 is worthwhile as a potential target to be explored for developing therapeutics of excitotoxic neuronal injury. PMID:22645130

Expression, regulation, and function of drug transporters in cervicovaginal tissues of a mouse model used for microbicide testing.

PubMed

Zhou, Tian; Hu, Minlu; Pearlman, Andrew; Rohan, Lisa C

2016-09-15

P-glycoprotein (P-gp), breast cancer resistance protein (BCRP), and multidrug resistance protein 4 (MRP4) are three efflux transporters that play key roles in the pharmacokinetics of antiretroviral drugs used in the pre-exposure prophylaxis of HIV sexual transmission. In this study, we investigated the expression, regulation, and function of these transporters in cervicovaginal tissues of a mouse model. Expression and regulation were examined using real-time RT-PCR and immunohistochemical staining, in the mouse tissues harvested at estrus and diestrus stages under natural cycling or after hormone synchronization. The three transporters were expressed at moderate to high levels compared to the liver. Transporter proteins were localized in various cell types in different tissue segments. Estrous cycle and exogenous hormone treatment affected transporter mRNA and protein expression, in a tissue- and transporter-dependent manner. Depo-Provera-synchronized mice were dosed vaginally or intraperitoneally with (3)H-TFV, with or without MK571 co-administration, to delineate the function of cervicovaginal Mrp4. Co-administration of MK571 significantly increased the concentration of vaginally-administered TFV in endocervix and vagina. MK571 increased the concentration of intraperitoneally-administered TFV in the cervicovaginal lavage and vagina by several fold. Overall, P-gp, Bcrp, and Mrp4 were positively expressed in mouse cervicovaginal tissues, and their expression can be regulated by the estrous cycle or by exogenous hormones. In this model, the Mrp4 transporter impacted TFV distribution in cervicovaginal tissues. Copyright © 2016. Published by Elsevier Inc.

Acute uriticaria-like lesions in allergen-unexposed cutaneous tissues in a mouse model of late allergic rhinitis

PubMed Central

Hayashi, Toshiharu; Fujii, Taeko

2008-01-01

The mechanisms of distant manifestation after a local allergic reaction are largely unknown. This study examined the development of cutaneous lesions in a mouse model of late allergic rhinitis (LAR). BALB/c mice were sensitized by ovalbumin (OVA) intraperitoneally two times (on days 0 and 10) and challenged by OVA intranasally on day 14. Four days after OVA challenge, nasal and cutaneous lesions including helper T (Th) responses, expression of adhesion molecules and presence of OVA and IgE were examined, and compared with unsensitized and unchallenged (control) mice. Compared with the control group, the LAR group developed LAR characterized by infiltration of lymphocytes and eosinophils, increased IgE values and increased productions of IL-4 and IL-5, but not IFN-γ. A dominant infiltration of eosinophils and increase in mast cells, attachment of eosinophils to endothelium, intense expression of VCAM-1 on endothelium in venules and VLA-4 expression on eosinophils and mast cells were recognized in the cutaneous tissues. There were no differences in the expression of ICAM-1 on vascular endothelium and LFA-1 on infiltrated leucocytes between the two groups. CLA expression on lymphocytes was not detected, and the binding of OVA and IgE on mast cells and eosinophils was found in the cutaneous lesions in the LAR group, but not in the control group. This study suggests that acute uriticaria-like lesions in OVA-unexposed cutaneous tissues may be induced by immediate allergic reaction due to the systemic development of Th2-type response in a mouse model of LAR. PMID:18460071

C/EBPβ-LAP*/LAP Expression Is Mediated by RSK/eIF4B-Dependent Signalling and Boosted by Increased Protein Stability in Models of Monocytic Differentiation

PubMed Central

Christmann, Martin; Friesenhagen, Judith; Westphal, Andreas; Pietsch, Daniel; Brand, Korbinian

2015-01-01

The transcription factor C/EBPβ plays a key role in monocytic differentiation and inflammation. Its small isoform LIP is associated with proliferation at early premonocytic developmental stages and regulated via mTOR-dependent signalling. During later stages of (pre)monocytic differentiation there is a considerable increase in the large C/EBPβ isoforms LAP*/LAP which inhibit proliferation thus supporting terminal differentiation. Here, we showed in different models of monocytic differentiation that this dramatic increase in the LAP*/LAP protein and LAP/LIP ratio was accompanied by an only modest/retarded mRNA increase suggesting an important role for (post)translational mechanisms. We found that LAP*/LAP formation was induced via MEK/RSK-dependent cascades, whereas mTOR/S6K1 were not involved. Remarkably, LAP*/LAP expression was dependent on phosphorylated eIF4B, an acceleratory protein of RNA helicase eIF4A. PKR inhibition reduced the expression of eIF4B and C/EBPβ in an eIF2α-independent manner. Furthermore, under our conditions a marked stabilisation of LAP*/LAP protein occurred, accompanied by reduced chymotrypsin-like proteasome/calpain activities and increased calpastatin levels. Our study elucidates new signalling pathways inducing LAP*/LAP expression and indicates new alternative PKR functions in monocytes. The switch from mTOR- to RSK-mediated signalling to orchestrate eIF4B-dependent LAP*/LAP translation, accompanied by increased protein stability but only small mRNA changes, may be a prototypical example for the regulation of protein expression during selected processes of differentiation/proliferation. PMID:26646662

Post-junctional facilitation of substance P signaling in a tibia fracture rat model of complex regional pain syndrome type I

PubMed Central

Wei, Tzuping; Li, Wen-wu; Guo, Tian-Zhi; Zhao, Rong; Wang, Liping; Clark, David J; Oaklander, Ann Louise; Schmelz, Martin; Kingery, Wade S.

2009-01-01

Tibia fracture in rats evokes nociceptive, vascular, and bone changes resembling complex regional pain syndrome (CRPS). Substance P (SP) signaling contributes to the hindpaw warmth, increased vascular permeability, and edema observed in this model, suggesting that neurogenic inflammatory responses could be enhanced after fracture. Four weeks after tibia fracture we measured SP and calcitonin gene-related peptide (CGRP) protein levels in the sciatic nerve and serum. Hindpaw skin extravasation responses and SP receptor (NK1), CGRP receptor (calcitonin receptor-like receptor, CRLR) and neutral endopeptidase (NEP) protein levels were also determined. Gene expression levels of these peptides, receptors, and peptidase were examined in the DRG and skin. Spontaneous and intravenous SP-evoked extravasation responses were increased ipsilateral, but not contralateral to the fracture. Fracture increased SP and CGRP gene expression in the ipsilateral L4,L5 DRG and neuropeptide protein levels in the sciatic nerve and in serum, but had no effect on electrically-evoked SP and CGRP release. NK1 receptor expression was increased in the ipsilateral hindpaw skin keratinocytes and endothelial cells after injury, but CRLR and NEP expression were unchanged. Fracture also increased epidermal thickness, but had no effect on epidermal skin neurite counts. These results demonstrate that spontaneous and intravenous SP-evoked extravasation responses are enhanced in the ipsilateral hindlimb after fracture and that fracture chronically increases the expression of endothelial and keratinocyte NK1 receptors in the injured limb. We postulate that SP activation of these up-regulated NK1 receptors results in skin warmth, protein leakage, edema, and keratinocyte proliferation in the injured limb. PMID:19464118

Post-junctional facilitation of Substance P signaling in a tibia fracture rat model of complex regional pain syndrome type I.

PubMed

Wei, Tzuping; Li, Wen-Wu; Guo, Tian-Zhi; Zhao, Rong; Wang, Liping; Clark, David J; Oaklander, Anne Louise; Schmelz, Martin; Kingery, Wade S

2009-08-01

Tibia fracture in rats evokes nociceptive, vascular, and bone changes resembling complex regional pain syndrome (CRPS). Substance P (SP) signaling contributes to the hindpaw warmth, increased vascular permeability, and edema observed in this model, suggesting that neurogenic inflammatory responses could be enhanced after fracture. Four weeks after tibia fracture we measured SP and calcitonin gene-related peptide (CGRP) protein levels in the sciatic nerve and serum. Hindpaw skin extravasation responses and SP receptor (NK1), CGRP receptor (calcitonin receptor-like receptor, CRLR) and neutral endopeptidase (NEP) protein levels were also determined. Gene expression levels of these peptides, receptors, and peptidase were examined in the DRG and skin. Spontaneous and intravenous SP-evoked extravasation responses were increased ipsilateral, but not contralateral to the fracture. Fracture increased SP and CGRP gene expression in the ipsilateral L4,L5 DRG and neuropeptide protein levels in the sciatic nerve and in serum, but had no effect on electrically evoked SP and CGRP release. NK1 receptor expression was increased in the ipsilateral hindpaw skin keratinocytes and endothelial cells after injury, but CRLR and NEP expression were unchanged. Fracture also increased epidermal thickness, but had no effect on epidermal skin neurite counts. These results demonstrate that spontaneous and intravenous SP-evoked extravasation responses are enhanced in the ipsilateral hindlimb after fracture and that fracture chronically increases the expression of endothelial and keratinocyte NK1 receptors in the injured limb. We postulate that SP activation of these up-regulated NK1 receptors results in skin warmth, protein leakage, edema, and keratinocyte proliferation in the injured limb.

Timp3 loss accelerates tumour invasion and increases prostate inflammation in a mouse model of prostate cancer.

PubMed

Adissu, Hibret A; McKerlie, Colin; Di Grappa, Marco; Waterhouse, Paul; Xu, Qiang; Fang, Hui; Khokha, Rama; Wood, Geoffrey A

2015-12-01

Altered expression and activity of proteases is implicated in inflammation and cancer progression. An important negative regulator of protease activity is TIMP3 (tissue inhibitor of metalloproteinase 3). TIMP3 expression is lacking in many cancers including advanced prostate cancer, and this may facilitate invasion and metastasis by allowing unrestrained protease activity. To investigate the role of TIMP3 in prostate cancer progression, we crossed TIMP3-deficient mice (Timp3(-/-)) to mice with prostate-specific deletion of the tumor suppressor Pten (Pten(-/-)), a well-established mouse model of prostate cancer. Tumor growth and progression were compared between Pten(-/-), Timp3(-/-) and control (Pten(-/-), Timp3(+/+)) mice at 16 weeks of age by histopathology and markers of proliferation, vascularity, and tumor invasion. Metalloproteinase activity within the tumors was assessed by gelatin zymography. Inflammatory infiltrates were assessed by immunohistochemistry for macrophages and lymphocytes whereas expression of cytokines and other inflammatory mediators was assessed by quantitative real time PCR and multiplex ELISA. Increased tumor growth, proliferation index, increased microvascular density, and invasion was observed in Pten(-/-), Timp3(-/-) prostate tumors compared to Pten(-/-), Timp3(+/+) tumors. Tumor cell invasion in Pten(-/-), Timp3(-/-) mice was associated with increased expression of matrix metalloprotease (MMP)-9 and activation of MMP-2. There was markedly increased inflammatory cell infiltration into the TIMP3-deficient prostate tumors along with increased expression of monocyte chemoattractant protein-1, cyclooxygenase-2, TNF-α, and interleukin-1β; all of which are implicated in inflammation and cancer. This study provides important insights into the role of altered protease activity in promoting prostate cancer invasion and implicates prostate inflammation as an important promoting factor in prostate cancer progression. © 2015 Wiley Periodicals, Inc.

Inflammation, Impaired Motility, and Permeability in a Guinea Pig Model of Postoperative Ileus.

PubMed

Lee, Yoo Jin; Hussain, Zahid; Huh, Cheal Wung; Lee, Young Ju; Park, Hyojin

2018-01-30

Postoperative ileus (POI) is characterized by impaired propulsive function of the gastrointestinal tract after surgery. Although inflammation is considered to be an important pathogenesis of POI, significant data are lacking. We aim to correlate the recovery time of postoperative dysmotility with that of inflammation and mucosal permeability. An experimental POI model of guinea pig was used. Contractile activity of the circular muscle of the stomach, jejunum, ileum, and proximal colon was measured through a tissue bath study. Inflammatory cells were counted, and the expression of calprotectin and tryptase were analyzed. The expression of protease-activated receptor 2 (PAR-2), claudin-1, and claudin-2 were analyzed with immunofluorescence. The small bowel and colon showed decreased contractile amplitude in the POI groups compared to control. In contrast to the colon, the contractile amplitude of the small bowel significantly recovered in the POI group at 6 hours after the operation compared to the control group. Inflammation was highly significant in the POI groups compared to the control and sham groups, especially in the colon. Immunofluorescence showed increased PAR-2 expression in the POI groups compared to sham. The decreased claudin-1 expression and increased claudin-2 expression may suggest increased mucosal permeability of the small bowel and colon in the POI groups. Increased inflammation and mucosal permeability may play an important role in the differential recovery stages in POI. These data may provide further insights into the pathophysiology and potential new therapeutic prospects of POI.

Inflammation, Impaired Motility, and Permeability in a Guinea Pig Model of Postoperative Ileus

PubMed Central

Lee, Yoo Jin; Hussain, Zahid; Huh, Cheal Wung; Lee, Young Ju; Park, Hyojin

2018-01-01

Background/Aims Postoperative ileus (POI) is characterized by impaired propulsive function of the gastrointestinal tract after surgery. Although inflammation is considered to be an important pathogenesis of POI, significant data are lacking. We aim to correlate the recovery time of postoperative dysmotility with that of inflammation and mucosal permeability. Methods An experimental POI model of guinea pig was used. Contractile activity of the circular muscle of the stomach, jejunum, ileum, and proximal colon was measured through a tissue bath study. Inflammatory cells were counted, and the expression of calprotectin and tryptase were analyzed. The expression of protease-activated receptor 2 (PAR-2), claudin-1, and claudin-2 were analyzed with immunofluorescence. Results The small bowel and colon showed decreased contractile amplitude in the POI groups compared to control. In contrast to the colon, the contractile amplitude of the small bowel significantly recovered in the POI group at 6 hours after the operation compared to the control group. Inflammation was highly significant in the POI groups compared to the control and sham groups, especially in the colon. Immunofluorescence showed increased PAR-2 expression in the POI groups compared to sham. The decreased claudin-1 expression and increased claudin-2 expression may suggest increased mucosal permeability of the small bowel and colon in the POI groups. Conclusions Increased inflammation and mucosal permeability may play an important role in the differential recovery stages in POI. These data may provide further insights into the pathophysiology and potential new therapeutic prospects of POI. PMID:29291615

Heat Shock Protein B1-Deficient Mice Display Impaired Wound Healing

PubMed Central

McNamee, Kay; Przybycien, Paulina M.; Lu, Xin; Williams, Richard O.; Bou-Gharios, George; Saklatvala, Jeremy; Dean, Jonathan L. E.

2013-01-01

There is large literature describing in vitro experiments on heat shock protein (hsp)B1 but understanding of its function in vivo is limited to studies in mice overexpressing human hspB1 protein. Experiments in cells have shown that hspB1 has chaperone activity, a cytoprotective role, regulates inflammatory gene expression, and drives cell proliferation. To investigate the function of the protein in vivo we generated hspB1-deficient mice. HspB1-deficient fibroblasts display increased expression of the pro-inflammatory cytokine, interleukin-6, compared to wild-type cells, but reduced proliferation. HspB1-deficient fibroblasts exhibit reduced entry into S phase and increased expression of cyclin-dependent kinase inhibitors p27kip1 and p21waf1. The expression of hspB1 protein and mRNA is also controlled by the cell cycle. To investigate the physiological function of hspB1 in regulating inflammation and cell proliferation we used an excisional cutaneous wound healing model. There was a significant impairment in the rate of healing of wounds in hspB1-deficient mice, characterised by reduced re-epithelialisation and collagen deposition but also increased inflammation. HspB1 deficiency augments neutrophil infiltration in wounds, driven by increased chemokine (C-X-C motif) ligand 1 expression. This appears to be a general mechanism as similar results were obtained in the air-pouch and peritonitis models of acute inflammation. PMID:24143227

Additional increased effects of mannitol-temozolomide combined treatment on blood-brain barrier permeability.

PubMed

Choi, Chunggab; Kim, Hye Min; Shon, Jeeheun; Park, Jiae; Kim, Hyeong-Taek; Oh, Seung-Hun; Kim, Nam Keun; Kim, Ok Joon

2018-03-04

The blood-brain barrier (BBB) is major obstacle in drug or stem cell treatment in chronic stroke. We hypothesized that adding mannitol to temozolomide (TMZ) is a practically applicable method for resolving the low efficacy of intravenous mannitol therapy. In this study, we investigated whether BBB permeability could be increased by this combined treatment. First, we established a chronic ischemic stroke rat model and examined changes in leakage of Evans blue dye within a lesion site, and in expression of tight junction proteins (TJPs), by this combined treatment. Additionally, in an in vitro BBB model using trans-wells, we analyzed changes in diffusion of a fluorescent tracer and in expression of TJPs. Mannitol-TMZ combined treatment not only increased the amount of Evans blue dye within the stroke lesion site, but also reduced occludin expression in rat brain microvessels. The in vitro study also showed that combined treatment increased the permeability for two different-sized fluorescent tracers, especially large size, and decreased expression of TJPs, such as occludin and ZO-1. Increased BBB permeability effects were more prominent with combined than with single treatments. Mannitol-TMZ combined treatment induced a decrease of TJPs with a consequent increase in BBB permeability. This combined treatment is clinically useful and might provide new therapeutic options by enabling efficient intracerebral delivery of various drugs that could not otherwise be used to treat many CNS diseases due to their inability to penetrate the BBB. Copyright © 2018 Elsevier Inc. All rights reserved.

Apigenin: Selective CK2 inhibitor increases Ikaros expression and improves T cell homeostasis and function in murine pancreatic cancer

PubMed Central

Nelson, Nadine; Szekeres, Karoly; Iclozan, Cristina; Rivera, Ivannie Ortiz; McGill, Andrew; Johnson, Gbemisola; Nwogu, Onyekachi

2017-01-01

Pancreatic cancer (PC) evades immune destruction by favoring the development of regulatory T cells (Tregs) that inhibit effector T cells. The transcription factor Ikaros is critical for lymphocyte development, especially T cells. We have previously shown that downregulation of Ikaros occurs as a result of its protein degradation by the ubiquitin-proteasome system in our Panc02 tumor-bearing (TB) mouse model. Mechanistically, we observed a deregulation in the balance between Casein Kinase II (CK2) and protein phosphatase 1 (PP1), which suggested that increased CK2 activity is responsible for regulating Ikaros’ stability in our model. We also showed that this loss of Ikaros expression is associated with a significant decrease in CD4+ and CD8+ T cell percentages but increased CD4+CD25+ Tregs in TB mice. In this study, we evaluated the effects of the dietary flavonoid apigenin (API), on Ikaros expression and T cell immune responses. Treatment of splenocytes from naïve mice with (API) stabilized Ikaros expression and prevented Ikaros downregulation in the presence of murine Panc02 cells in vitro, similar to the proteasome inhibitor MG132. In vivo treatment of TB mice with apigenin (TB-API) improved survival, reduced tumor weights and prevented splenomegaly. API treatment also restored protein expression of some Ikaros isoforms, which may be attributed to its moderate inhibition of CK2 activity from splenocytes of TB-API mice. This partial restoration of Ikaros expression was accompanied by a significant increase in CD4+ and CD8+ T cell percentages and a reduction in Treg percentages in TB-API mice. In addition, CD8+ T cells from TB-API mice produced more IFN-γ and their splenocytes were better able to prime allogeneic CD8+ T cell responses compared to TB mice. These results provide further evidence that Ikaros is regulated by CK2 in our pancreatic cancer model. More importantly, our findings suggest that API may be a possible therapeutic agent for stabilizing Ikaros expression and function to maintain T cell homeostasis in murine PC. PMID:28152014

Muscle-Specific SIRT1 Gain-of-Function Increases Slow-Twitch Fibers and Ameliorates Pathophysiology in a Mouse Model of Duchenne Muscular Dystrophy

PubMed Central

Chalkiadaki, Angeliki; Igarashi, Masaki; Nasamu, Armiyaw Sebastian; Knezevic, Jovana; Guarente, Leonard

2014-01-01

SIRT1 is a metabolic sensor and regulator in various mammalian tissues and functions to counteract metabolic and age-related diseases. Here we generated and analyzed mice that express SIRT1 at high levels specifically in skeletal muscle. We show that SIRT1 transgenic muscle exhibits a fiber shift from fast-to-slow twitch, increased levels of PGC-1α, markers of oxidative metabolism and mitochondrial biogenesis, and decreased expression of the atrophy gene program. To examine whether increased activity of SIRT1 protects from muscular dystrophy, a muscle degenerative disease, we crossed SIRT1 muscle transgenic mice to mdx mice, a genetic model of Duchenne muscular dystrophy. SIRT1 overexpression in muscle reverses the phenotype of mdx mice, as determined by histology, creatine kinase release into the blood, and endurance in treadmill exercise. In addition, SIRT1 overexpression also results in increased levels of utrophin, a functional analogue of dystrophin, as well as increased expression of PGC-1α targets and neuromuscular junction genes. Based on these findings, we suggest that pharmacological interventions that activate SIRT1 in skeletal muscle might offer a new approach for treating muscle diseases. PMID:25032964

Functionally distinct roles for different miR-155 expression levels through contrasting effects on gene expression, in acute myeloid leukaemia.

PubMed

Narayan, N; Morenos, L; Phipson, B; Willis, S N; Brumatti, G; Eggers, S; Lalaoui, N; Brown, L M; Kosasih, H J; Bartolo, R C; Zhou, L; Catchpoole, D; Saffery, R; Oshlack, A; Goodall, G J; Ekert, P G

2017-04-01

Enforced expression of microRNA-155 (miR-155) in myeloid cells has been shown to have both oncogenic or tumour-suppressor functions in acute myeloid leukaemia (AML). We sought to resolve these contrasting effects of miR-155 overexpression using murine models of AML and human paediatric AML data sets. We show that the highest miR-155 expression levels inhibited proliferation in murine AML models. Over time, enforced miR-155 expression in AML in vitro and in vivo, however, favours selection of intermediate miR-155 expression levels that results in increased tumour burden in mice, without accelerating the onset of disease. Strikingly, we show that intermediate and high miR-155 expression also regulate very different subsets of miR-155 targets and have contrasting downstream effects on the transcriptional environments of AML cells, including genes involved in haematopoiesis and leukaemia. Furthermore, we show that elevated miR-155 expression detected in paediatric AML correlates with intermediate and not high miR-155 expression identified in our experimental models. These findings collectively describe a novel dose-dependent role for miR-155 in the regulation of AML, which may have important therapeutic implications.

Bmp6 Expression in Murine Liver Non Parenchymal Cells: A Mechanism to Control their High Iron Exporter Activity and Protect Hepatocytes from Iron Overload?

PubMed Central

Rausa, Marco; Pagani, Alessia; Nai, Antonella; Campanella, Alessandro; Gilberti, Maria Enrica; Apostoli, Pietro; Camaschella, Clara; Silvestri, Laura

2015-01-01

Bmp6 is the main activator of hepcidin, the liver hormone that negatively regulates plasma iron influx by degrading the sole iron exporter ferroportin in enterocytes and macrophages. Bmp6 expression is modulated by iron but the molecular mechanisms are unknown. Although hepcidin is expressed almost exclusively by hepatocytes (HCs), Bmp6 is produced also by non-parenchymal cells (NPCs), mainly sinusoidal endothelial cells (LSECs). To investigate the regulation of Bmp6 in HCs and NPCs, liver cells were isolated from adult wild type mice whose diet was modified in iron content in acute or chronic manner and in disease models of iron deficiency (Tmprss6 KO mouse) and overload (Hjv KO mouse). With manipulation of dietary iron in wild-type mice, Bmp6 and Tfr1 expression in both HCs and NPCs was inversely related, as expected. When hepcidin expression is abnormal in murine models of iron overload (Hjv KO mice) and deficiency (Tmprss6 KO mice), Bmp6 expression in NPCs was not related to Tfr1. Despite the low Bmp6 in NPCs from Tmprss6 KO mice, Tfr1 mRNA was also low. Conversely, despite body iron overload and high expression of Bmp6 in NPCs from Hjv KO mice, Tfr1 mRNA and protein were increased. However, in the same cells ferritin L was only slightly increased, but the iron content was not, suggesting that Bmp6 in these cells reflects the high intracellular iron import and export. We propose that NPCs, sensing the iron flux, not only increase hepcidin through Bmp6 with a paracrine mechanism to control systemic iron homeostasis but, controlling hepcidin, they regulate their own ferroportin, inducing iron retention or release and further modulating Bmp6 production in an autocrine manner. This mechanism, that contributes to protect HC from iron loading or deficiency, is lost in disease models of hepcidin production. PMID:25860887

Ankyrin G expression is associated with androgen receptor stability, invasiveness, and lethal outcome in prostate cancer patients.

PubMed

Wang, Tingting; Abou-Ouf, Hatem; Hegazy, Samar A; Alshalalfa, Mohammed; Stoletov, Konstantin; Lewis, John; Donnelly, Bryan; Bismar, Tarek A

2016-12-01

Ankyrin G (ANK3) is a member of the Ankyrin family, which functions to provide cellular stability by anchoring the cytoskeleton to the plasma membrane. Deregulation of ANK3 expression has been observed in multiple human cancers but its mechanism remains unknown. ANK3 expression in relation to disease progression and patients' outcome was investigated in two cohorts of prostate cancer (PCA). Mechanistic studies were carried out in vitro and in vivo using several PCA cell lines and the avian embryo model. Silencing ANK3 resulted in significant reduction of cell proliferation through an AR-independent mechanism. Decreased ANK3 expression delayed S phase to G2/M cell cycle transition and reduced the expression of cyclins A and B. However, cells with knocked-down ANK3 exhibited significant increase in cell invasion through an AR-dependent mechanism. Furthermore, we found that ANK3 is a regulator of AR protein stability. ANK3 knockdown also promoted cancer cell invasion and extravasations in vivo using the avian embryo model (p < 0.01). In human samples, ANK3 expression was dramatically upregulated in high grade intraepithelial neoplasia (HGPIN) and localized PCA (p < 0.0001). However, it was downregulated castration resistant stage (p < 0.0001) and showed inverse relation to Gleason score (p < 0.0001). In addition, increased expression of ANK3 in cancer tissues was correlated with better cancer-specific survival of PCA patients (p = 0.012). Silencing ANK3 results in significant reduction of cell proliferation through an AR-independent mechanism. ANK3 knockdown results in significant increase in cell invasion through an AR-dependent mechanism. ANK3 is a regulator of AR protein stability. ANK3 knockdown also promotes cancer cell invasion and extravasation in vivo using the avian embryo model.

Model-directed engineering of "difficult-to-express" monoclonal antibody production by Chinese hamster ovary cells.

PubMed

Pybus, Leon P; Dean, Greg; West, Nathan R; Smith, Andrew; Daramola, Olalekan; Field, Ray; Wilkinson, Stephen J; James, David C

2014-02-01

Despite improvements in volumetric titer for monoclonal antibody (MAb) production processes using Chinese hamster ovary (CHO) cells, some "difficult-to-express" (DTE) MAbs inexplicably reach much lower process titers. These DTE MAbs require intensive cell line and process development activity, rendering them more costly or even unsuitable to manufacture. To rapidly and rationally identify an optimal strategy to improve production of DTE MAbs, we have developed an engineering design platform combining high-yielding transient production, empirical modeling of MAb synthesis incorporating an unfolded protein response (UPR) regulatory loop with directed expression and cell engineering approaches. Utilizing a panel of eight IgG1 λ MAbs varying >4-fold in volumetric titer, we showed that MAb-specific limitations on folding and assembly rate functioned to induce a proportionate UPR in host CHO cells with a corresponding reduction in cell growth rate. Derived from comparative empirical modeling of cellular constraints on the production of each MAb we employed two strategies to increase production of DTE MAbs designed to avoid UPR induction through an improvement in the rate/cellular capacity for MAb folding and assembly reactions. Firstly, we altered the transfected LC:HC gene ratio and secondly, we co-expressed a variety of molecular chaperones, foldases or UPR transactivators (BiP, CypB, PDI, and active forms of ATF6 and XBP1) with recombinant MAbs. DTE MAb production was significantly improved by both strategies, although the mode of action was dependent upon the approach employed. Increased LC:HC ratio or CypB co-expression improved cell growth with no effect on qP. In contrast, BiP, ATF6c and XBP1s co-expression increased qP and reduced cell growth. This study demonstrates that expression-engineering strategies to improve production of DTE proteins in mammalian cells should be product specific, and based on rapid predictive tools to assess the relative impact of different engineering interventions. © 2013 Wiley Periodicals, Inc.

Hypothalamic expression of inflammatory mediators in an animal model of binge eating.

PubMed

Alboni, Silvia; Micioni Di Bonaventura, Maria Vittoria; Benatti, Cristina; Giusepponi, Maria Elena; Brunello, Nicoletta; Cifani, Carlo

2017-03-01

Binge eating episodes are characterized by uncontrollable, distressing eating of a large amount of highly palatable food and represent a central feature of bingeing related eating disorders. Research suggests that inflammation plays a role in the onset and maintenance of eating-related maladaptive behavior. Markers of inflammation can be selectively altered in discrete brain regions where they can directly or indirectly regulate food intake. In the present study, we measured expression levels of different components of cytokine systems (IL-1, IL-6, IL-18, TNF-α and IFN-ɣ) and related molecules (iNOS and COX2) in the preoptic and anterior-tuberal parts of the hypothalamus of a validated animal model of binge eating. In this animal model, based on the exposure to both food restriction and frustration stress, binge-like eating behavior for highly palatable food is not shown when animals are exposed to the frustration stress during the estrus phase. We found a characteristic down-regulation of the IL-18/IL-18 receptor system (with increased expression of the inhibitor of the pro-inflammatory cytokine IL-18, IL-18BP, together with a decreased expression of the binding chain of the IL-18 receptor) and a three-fold increase in the expression of iNOS specifically in the anterior-tuberal region of the hypothalamus of animals that develop a binge-like eating behavior. Differently, when food restricted animals were stressed during the estrus phase, IL-18 expression increased, while iNOS expression was not significantly affected. Considering the role of this region of the hypothalamus in controlling feeding related behavior, this can be relevant in eating disorders and obesity. Our data suggest that by targeting centrally selected inflammatory markers, we may prevent that disordered eating turns into a full blown eating disorder. Copyright © 2016 Elsevier B.V. All rights reserved.

The anabolic/androgenic steroid nandrolone exacerbates gene expression modifications induced by mutant SOD1 in muscles of mice models of amyotrophic lateral sclerosis

PubMed Central

Galbiati, Mariarita; Onesto, Elisa; Zito, Arianna; Crippa, Valeria; Rusmini, Paola; Mariotti, Raffaella; Bentivoglio, Marina; Bendotti, Caterina; Poletti, Angelo

2012-01-01

Anabolic/androgenic steroids (AAS) are drugs that enhance muscle mass, and are often illegally utilized in athletes to improve their performances. Recent data suggest that the increased risk for amyotrophic lateral sclerosis (ALS) in male soccer and football players could be linked to AAS abuse. ALS is a motor neuron disease mainly occurring in sporadic (sALS) forms, but some familial forms (fALS) exist and have been linked to mutations in different genes. Some of these, in their wild type (wt) form, have been proposed as risk factors for sALS, i.e. superoxide dismutase 1 (SOD1) gene, whose mutations are causative of about 20% of fALS. Notably, SOD1 toxicity might occur both in motor neurons and in muscle cells. Using gastrocnemius muscles of mice overexpressing human mutant SOD1 (mutSOD1) at different disease stages, we found that the expression of a selected set of genes associated to muscle atrophy, MyoD, myogenin, atrogin-1, and transforming growth factor (TGF)β1, is up-regulated already at the presymptomatic stage. Atrogin-1 gene expression was increased also in mice overexpressing human wtSOD1. Similar alterations were found in axotomized mouse muscles and in cultured ALS myoblast models. In these ALS models, we then evaluated the pharmacological effects of the synthetic AAS nandrolone on the expression of the genes modified in ALS muscle. Nandrolone administration had no effects on MyoD, myogenin, and atrogin-1 expression, but it significantly increased TGFβ1 expression at disease onset. Altogether, these data suggest that, in fALS, muscle gene expression is altered at early stages, and AAS may exacerbate some of the alterations induced by SOD1 possibly acting as a contributing factor also in sALS. PMID:22178654

The anabolic/androgenic steroid nandrolone exacerbates gene expression modifications induced by mutant SOD1 in muscles of mice models of amyotrophic lateral sclerosis.

PubMed

Galbiati, Mariarita; Onesto, Elisa; Zito, Arianna; Crippa, Valeria; Rusmini, Paola; Mariotti, Raffaella; Bentivoglio, Marina; Bendotti, Caterina; Poletti, Angelo

2012-02-01

Anabolic/androgenic steroids (AAS) are drugs that enhance muscle mass, and are often illegally utilized in athletes to improve their performances. Recent data suggest that the increased risk for amyotrophic lateral sclerosis (ALS) in male soccer and football players could be linked to AAS abuse. ALS is a motor neuron disease mainly occurring in sporadic (sALS) forms, but some familial forms (fALS) exist and have been linked to mutations in different genes. Some of these, in their wild type (wt) form, have been proposed as risk factors for sALS, i.e. superoxide dismutase 1 (SOD1) gene, whose mutations are causative of about 20% of fALS. Notably, SOD1 toxicity might occur both in motor neurons and in muscle cells. Using gastrocnemius muscles of mice overexpressing human mutant SOD1 (mutSOD1) at different disease stages, we found that the expression of a selected set of genes associated to muscle atrophy, MyoD, myogenin, atrogin-1, and transforming growth factor (TGF)β1, is up-regulated already at the presymptomatic stage. Atrogin-1 gene expression was increased also in mice overexpressing human wtSOD1. Similar alterations were found in axotomized mouse muscles and in cultured ALS myoblast models. In these ALS models, we then evaluated the pharmacological effects of the synthetic AAS nandrolone on the expression of the genes modified in ALS muscle. Nandrolone administration had no effects on MyoD, myogenin, and atrogin-1 expression, but it significantly increased TGFβ1 expression at disease onset. Altogether, these data suggest that, in fALS, muscle gene expression is altered at early stages, and AAS may exacerbate some of the alterations induced by SOD1 possibly acting as a contributing factor also in sALS. Copyright © 2011 Elsevier Ltd. All rights reserved.

Erythropoietin over-expression protects against diet-induced obesity in mice through increased fat oxidation in muscles.

PubMed

Hojman, Pernille; Brolin, Camilla; Gissel, Hanne; Brandt, Claus; Zerahn, Bo; Pedersen, Bente Klarlund; Gehl, Julie

2009-06-12

Erythropoietin can be over-expressed in skeletal muscles by gene electrotransfer, resulting in 100-fold increase in serum EPO and significant increases in haemoglobin levels. Earlier studies have suggested that EPO improves several metabolic parameters when administered to chronically ill kidney patients. Thus we applied the EPO over-expression model to investigate the metabolic effect of EPO in vivo.At 12 weeks, EPO expression resulted in a 23% weight reduction (P<0.01) in EPO transfected obese mice; thus the mice weighed 21.9+/-0.8 g (control, normal diet,) 21.9+/-1.4 g (EPO, normal diet), 35.3+/-3.3 g (control, high-fat diet) and 28.8+/-2.6 g (EPO, high-fat diet). Correspondingly, DXA scanning revealed that this was due to a 28% reduction in adipose tissue mass.The decrease in adipose tissue mass was accompanied by a complete normalisation of fasting insulin levels and glucose tolerance in the high-fat fed mice. EPO expression also induced a 14% increase in muscle volume and a 25% increase in vascularisation of the EPO transfected muscle. Muscle force and stamina were not affected by EPO expression. PCR array analysis revealed that genes involved in lipid metabolism, thermogenesis and inflammation were increased in muscles in response to EPO expression, while genes involved in glucose metabolism were down-regulated. In addition, muscular fat oxidation was increased 1.8-fold in both the EPO transfected and contralateral muscles.In conclusion, we have shown that EPO when expressed in supra-physiological levels has substantial metabolic effects including protection against diet-induced obesity and normalisation of glucose sensitivity associated with a shift to increased fat metabolism in the muscles.

Erythropoietin Over-Expression Protects against Diet-Induced Obesity in Mice through Increased Fat Oxidation in Muscles

PubMed Central

Hojman, Pernille; Brolin, Camilla; Gissel, Hanne; Brandt, Claus; Zerahn, Bo; Pedersen, Bente Klarlund; Gehl, Julie

2009-01-01

Erythropoietin can be over-expressed in skeletal muscles by gene electrotransfer, resulting in 100-fold increase in serum EPO and significant increases in haemoglobin levels. Earlier studies have suggested that EPO improves several metabolic parameters when administered to chronically ill kidney patients. Thus we applied the EPO over-expression model to investigate the metabolic effect of EPO in vivo. At 12 weeks, EPO expression resulted in a 23% weight reduction (P<0.01) in EPO transfected obese mice; thus the mice weighed 21.9±0.8 g (control, normal diet,) 21.9±1.4 g (EPO, normal diet), 35.3±3.3 g (control, high-fat diet) and 28.8±2.6 g (EPO, high-fat diet). Correspondingly, DXA scanning revealed that this was due to a 28% reduction in adipose tissue mass. The decrease in adipose tissue mass was accompanied by a complete normalisation of fasting insulin levels and glucose tolerance in the high-fat fed mice. EPO expression also induced a 14% increase in muscle volume and a 25% increase in vascularisation of the EPO transfected muscle. Muscle force and stamina were not affected by EPO expression. PCR array analysis revealed that genes involved in lipid metabolism, thermogenesis and inflammation were increased in muscles in response to EPO expression, while genes involved in glucose metabolism were down-regulated. In addition, muscular fat oxidation was increased 1.8-fold in both the EPO transfected and contralateral muscles. In conclusion, we have shown that EPO when expressed in supra-physiological levels has substantial metabolic effects including protection against diet-induced obesity and normalisation of glucose sensitivity associated with a shift to increased fat metabolism in the muscles. PMID:19521513

Increased Cerebral Tff1 Expression in Two Murine Models of Neuroinflammation.

PubMed

Znalesniak, Eva B; Fu, Ting; Guttek, Karina; Händel, Ulrike; Reinhold, Dirk; Hoffmann, Werner

2016-01-01

The trefoil factor family (TFF) peptide TFF1 is a typical secretory product of the gastric mucosa and a very low level of expression occurs in nearly all regions of the murine brain. TFF1 possesses a lectin activity and binding to a plethora of transmembrane glycoproteins could explain the diverse biological effects of TFF1 (e.g., anti-apoptotic effect). It was the aim to test whether TFF expression is changed during neuroinflammation. Expression profiling was performed using semi-quantitative RT-PCR analyses in two murine models of neuroinflammation, i.e. Toxoplasma gondii-induced encephalitis and experimental autoimmune encephalomyelitis (EAE), the latter being the most common animal model of multiple sclerosis. Tff1 expression was also localized using RNA in situ hybridization histochemistry. We report for the first time on a significant transcriptional induction in cerebral Tff1 expression in both T. gondii-induced encephalitis and EAE. In contrast, Tff2 and Tff3 expression were not altered. Tff1 transcripts were predominantly localized in the internal granular layer of the cerebellum indicating neuronal expression. Furthermore, also glial cells are expected to express Tff1. Characterization of both experimental models by expression profiling (e.g., inflammasome sensors, inflammatory cytokines, microglial marker Iba1, ependymin related protein 1) revealed differences concerning the expression of the inflammasome sensor Nlrp1 and interleukin 17a. The up-regulated expression of Tff1 is probably the result of a complex inflammatory process as its expression is induced by tumor necrosis factor α as well as interleukins 1β and 17. However on the transcript level, Tff1KO mice did not show any significant signs of an altered immune response after infection with T. gondii in comparison with the wild type animals. © 2016 The Author(s) Published by S. Karger AG, Basel.

Co-culture of 3D tumor spheroids with fibroblasts as a model for epithelial–mesenchymal transition in vitro

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kim, Sun-Ah, E-mail: j.sarah.k@gmail.com; Lee, Eun Kyung, E-mail: leeek@catholic.ac.kr; Cancer Evolution Research Center, College of Medicine, The Catholic University of Korea, Seoul 137-701

Epithelial–mesenchymal transition (EMT) acts as a facilitator of metastatic dissemination in the invasive margin of malignant tumors where active tumor–stromal crosstalks take place. Co-cultures of cancer cells with cancer-associated fibroblasts (CAFs) are often used as in vitro models of EMT. We established a tumor–fibroblast proximity co-culture using HT-29 tumor spheroids (TSs) with CCD-18co fibroblasts. When co-cultured with TSs, CCD-18co appeared activated, and proliferative activity as well as cell migration increased. Expression of fibronectin increased whereas laminin and type I collagen decreased in TSs co-cultured with fibroblasts compared to TSs alone, closely resembling the margin of in vivo xenograft tissue. Activemore » TGFβ1 in culture media significantly increased in TS co-cultures but not in 2D co-cultures of cancer cells–fibroblasts, indicating that 3D context-associated factors from TSs may be crucial to crosstalks between cancer cells and fibroblasts. We also observed in TSs co-cultured with fibroblasts increased expression of α-SMA, EGFR and CTGF; reduced expression of membranous β-catenin and E-cadherin, together suggesting an EMT-like changes similar to a marginal region of xenograft tissue in vivo. Overall, our in vitro TS–fibroblast proximity co-culture mimics the EMT-state of the invasive margin of in vivo tumors in early metastasis. - Highlights: • An adjacent co-culture of tumor spheroids and fibroblasts is presented as EMT model. • Activation of fibroblasts and increased cell migration were shown in co-culture. • Expression of EMT-related factors in co-culture was similar to that in tumor tissue. • Crosstalk between spheroids and fibroblasts was demonstrated by secretome analysis.« less

[Effects of electro-acupuncture on expression of triggering receptor expressed on myeloid cells 1 in ankle joint synovial tissue of acute gouty arthritis rats].

PubMed

Zhang, Chao-Nan; Huang, Xue-Kuan; Luo, Yan; Jiang, Juan; Wan, Lei; Wang, Ling

2015-01-01

To investigate the effects of electro-acupuncture (EA) on the expression of triggering receptor expressed on myeloid cell (TREM)l in ankle joint synovial tissue of acute gouty arthritis (AGA) rats. Forty male SD rats were randomly divided into 4 groups: normal, AGA, medication and EA group, 10 rats in each group. AGA model was established by induced monosodium urate (MSU) method, except the normal group. Tow days before AGA model was established, normal and AGA groups were lavaged with normal saline (20 ml/kg), medication group was lavaged with colchicine solution (20 ml/kg), EA(1.5-2 Hz, D.-D.wave, 9v; 1-3 rnA) was applied to "Sanyinjiao" (SP6), "jiexi" (ST41) and "Kunlun" (BL60) for 20 min, once daily;continuously for 9 days. Then observed the changes in dysfunction, and the content of TNF-α and IL-lβ detected by ELISA, the expression of TREM-l detected by immunohistochemistry and western blot. Compared to the normal group, the AGA group of the dysfunction index increased significantly (P<0.01), the content of TNF-α and IL-lβ increased significantly (P<0.05), the expression of TREM-l in synovial tissue increased significantly (P<0.05); the medication and EA groups compared to the AGA group, the dysfunction index decreased significantly (P<0.01), the content of TNF-α and IL-lβ decreased significantly (P<0.05), the expression of TREM-l in synovial tissue decreased significantly (P<0.05); there were not statistically significant between the medication and EA group (P>0.05). EA treating AGA may be through down-regulating the expression of TREM -1 in synovial tissue.

Synergistically increased ILC2 and Th9 cells in lung tissue jointly promote the pathological process of asthma in mice.

PubMed

Ying, Xinyu; Su, Zhaoliang; Bie, Qingli; Zhang, Pan; Yang, Huijian; Wu, Yumin; Xu, Yunyun; Wu, Jing; Zhang, Mengying; Wang, Shengjun; Xu, Huaxi

2016-06-01

In recent years, T helper (Th) 9 cells have been demonstrated to be key mediators in immune responses in asthmatic lungs, and innate lymphoid cells 2 (ILC2s) have been described as a novel type of innate immunocyte with the ability to enhance immunoglobulin E (IgE) production. However, the interaction between ILC2s and Th9 cells in the pulmonary system of a mouse model of asthma remains to be elucidated. In the present study, the response state of lung tissue with regards to Th9 and ILC2s in a mouse model of asthma was investigated by detecting Th9‑ and ILC2‑associated cytokine receptors. The present study also investigated the association between the expression levels of the cytokine receptors in lung tissue samples and the IgE levels in sera samples from mouse models of asthma. Results from the present study demonstrated that the frequency of ILC2s and Th9 cells was significantly increased in the lung tissue samples, indicating that a Th2-type immune response had occurred. In addition, high mRNA expression levels of RAR‑related orphan receptor α, interleukin 1 receptor‑like 1, transcription factor PU.1 and interleukin (IL)‑9 were observed. Furthermore, IL‑5Rα, IL‑13Rα2 and high‑affinity IgE receptor were increased in mouse models of asthma, and a positive association was observed between the expression levels of ILC2‑ or Th9‑associated receptors in tissue samples and IgE levels in the sera. This indicated that ILC2s and Th9 were in a state of polarization and may promote each other in the lung tissue of mouse models of asthma, and that the lung tissue was responding to the two types of cells via increased expression of receptors.

Effect of Acupotomy on FAK-PI3K Signaling Pathways in KOA Rabbit Articular Cartilages

PubMed Central

Xie, Zhan-guo; Guo, Yan; Yu, Jia-Ni; Lu, Juan; Zhang, Wei; Wang, Li-Juan; Xu, Jing; Zhao, Rui-Li; Zhou, Shuai

2017-01-01

Objective By observing the needle-knife of KOA rabbit morphology, knee joint cartilage p-FAK, p-PI3K, Aggrecan gene, and protein expression, to study the effect of needle-knife to promote cartilage cell synthesis metabolism mechanism. Method 49 male New Zealand rabbits, randomly divided into normal group (Z), model group (M), model-inhibitors (MP), needle-knife group (D), needle-knife inhibitors group (DP), electroacupuncture group (E), and electroacupuncture inhibitors (EP). RT-PCR and Western Blot were used to test each animal cartilage p-FAK, p-PI3K, and Aggrecan gene and protein expression level. Results Compared with N group, p-FAK and p-PI3K protein and mRNA expression of M group, D group, and E group increased (P < 0.05), while the protein and mRNA expression of Aggrecan reduced (P < 0.05). Compared with M group, p-FAK, p-PI3K, Aggrecan protein, and mRNA of E and D group increased (P < 0.05). Compared with E group, p-FAK, p-PI3K, Aggrecan protein, and mRNA expression of D group increased (P < 0.05); after adding inhibitors, p-FAK, p-PI3K, Aggrecan protein, and mRNA expression reduced (P < 0.05). Conclusion Needle-knife therapy can promote the repairment of cartilage cells by activating FAK-PI3K signaling pathways, promoting the synthesis of cartilage cell metabolism. PMID:29234400

TRPM5 mediates acidic extracellular pH signaling and TRPM5 inhibition reduces spontaneous metastasis in mouse B16-BL6 melanoma cells

PubMed Central

Maeda, Toyonobu; Suzuki, Atsuko; Koga, Kaori; Miyamoto, Chihiro; Maehata, Yojiro; Ozawa, Shigeyuki; Hata, Ryu-Ichiro; Nagashima, Yoji; Nabeshima, Kazuki; Miyazaki, Kaoru; Kato, Yasumasa

2017-01-01

Extracellular acidity is a hallmark of solid tumors and is associated with metastasis in the tumor microenvironment. Acidic extracellular pH (pHe) has been found to increase intracellular Ca2+ and matrix metalloproteinase-9 (MMP-9) expression by activating NF-κB in the mouse B16 melanoma model. The present study assessed whether TRPM5, an intracellular Ca2+-dependent monovalent cation channel, is associated with acidic pHe signaling and induction of MMP-9 expression in this mouse melanoma model. Treatment of B16 cells with Trpm5 siRNA reduced acidic pHe-induced MMP-9 expression. Enforced expression of Trpm5 increased the rate of acidic pHe-induced MMP-9 expression, as well as increasing experimental lung metastasis. This genetic manipulation did not alter the pHe critical for MMP-9 induction but simply amplified the percentage of inducible MMP-9 at each pHe. Treatment of tumor bearing mice with triphenylphosphine oxide (TPPO), an inhibitor of TRPM5, significantly reduced spontaneous lung metastasis. In silico analysis of clinical samples showed that high TRPM5 mRNA expression correlated with poor overall survival rate in patients with melanoma and gastric cancer but not in patients with cancers of the ovary, lung, breast, and rectum. These results showed that TRPM5 amplifies acidic pHe signaling and may be a promising target for preventing metastasis of some types of tumor. PMID:29108231

Symbiosis induces widespread changes in the proteome of the model cnidarian Aiptasia.

PubMed

Oakley, Clinton A; Ameismeier, Michael F; Peng, Lifeng; Weis, Virginia M; Grossman, Arthur R; Davy, Simon K

2016-07-01

Coral reef ecosystems are metabolically founded on the mutualism between corals and photosynthetic dinoflagellates of the genus Symbiodinium. The glass anemone Aiptasia sp. has become a tractable model for this symbiosis, and recent advances in genetic information have enabled the use of mass spectrometry-based proteomics in this model. We utilized label-free liquid chromatography electrospray-ionization tandem mass spectrometry to analyze the effects of symbiosis on the proteomes of symbiotic and aposymbiotic Aiptasia. We identified and obtained relative quantification of more than 3,300 proteins in 1,578 protein clusters, with 81 protein clusters showing significantly different expression between symbiotic states. Symbiotic anemones showed significantly higher expression of proteins involved in lipid storage and transport, nitrogen transport and cycling, intracellular trafficking, endocytosis and inorganic carbon transport. These changes reflect shifts in host metabolism and nutrient reserves due to increased nutritional exchange with the symbionts, as well as mechanisms for supplying inorganic nutrients to the algae. Aposymbiotic anemones exhibited increased expression of multiple systems responsible for mediating reactive oxygen stress, suggesting that the host derives direct or indirect protection from oxidative stress while in symbiosis. Aposymbiotic anemones also increased their expression of an array of proteases and chitinases, indicating a metabolic shift from autotrophy to heterotrophy. These results provide a comprehensive Aiptasia proteome with more direct relative quantification of protein abundance than transcriptomic methods. The extension of "omics" techniques to this model system will allow more powerful studies of coral physiology, ecosystem function, and the effects of biotic and abiotic stress on the coral-dinoflagellate mutualism. © 2015 John Wiley & Sons Ltd.

Receptor-interacting protein kinase 3-mediated programmed cell necrosis in rats subjected to focal cerebral ischemia-reperfusion injury.

PubMed

Dong, Yanru; Bao, Cuifen; Yu, Jingwei; Liu, Xia

2016-07-01

In the current study, the activation of tumor necrosis factor-α receptor 1 (TNFR1) and receptor-interacting protein kinase 3 (RIP3) were investigated following cerebral ischemia-reperfusion injury (CIRI). Healthy SD rats were randomly divided into 3 groups: Sham operation group, model group and inhibitor group. The model group and inhibitor group were further divided into 4 subgroups of 6, 12, 24 and 72 h following CIRI. Using right middle cerebral artery embolization, the CIRI model was generated. To confirm that the CIRI model was established, neurological scores, TTC staining and brain water content measurements were conducted. Immunohistochemistry and western blotting were conducted to investigate the expression of TNFR1 and RIP3 in the cerebral cortex. It was observed that nerve cell necrosis occurred following 6 h of CIRI. The appearance of necrotic cells was gradually increased with increasing CIRI duration. TNFR1 and RIP3 were positively expressed following 6 h of CIRI. With increasing durations of CIRI, the protein expression levels of TNFR1 and RIP3 were significantly increased. Pre‑administration with Z-VAD-FMK (zVAD) significantly increased the protein level of RIP3, however, had no effect on the levels of TNFR1, and was accompanied by a reduction in necrosis. In conclusion, RIP3‑mediated cell necrosis was enhanced by caspase blockade zVAD and the function of zVAD was independent of TNFR1 signaling following IR.

Inhibition of Excessive Monoamine Oxidase A/B Activity Protects Against Stress-induced Neuronal Death in Huntington Disease.

PubMed

Ooi, Jolene; Hayden, Michael R; Pouladi, Mahmoud A

2015-12-01

Monoamine oxidases (MAO) are important components of the homeostatic machinery that maintains the levels of monoamine neurotransmitters, including dopamine, in balance. Given the imbalance in dopamine levels observed in Huntington disease (HD), the aim of this study was to examine MAO activity in a mouse striatal cell model of HD and in human neural cells differentiated from control and HD patient-derived induced pluripotent stem cell (hiPSC) lines. We show that mouse striatal neural cells expressing mutant huntingtin (HTT) exhibit increased MAO expression and activity. We demonstrate using luciferase promoter assays that the increased MAO expression reflects enhanced epigenetic activation in striatal neural cells expressing mutant HTT. Using cellular stress paradigms, we further demonstrate that the increase in MAO activity in mutant striatal neural cells is accompanied by enhanced susceptibility to oxidative stress and impaired viability. Treatment of mutant striatal neural cells with MAO inhibitors ameliorated oxidative stress and improved cellular viability. Finally, we demonstrate that human HD neural cells exhibit increased MAO-A and MAO-B expression and activity. Altogether, this study demonstrates abnormal MAO expression and activity and suggests a potential use for MAO inhibitors in HD.

The Effects of Injury Magnitude on the Kinetics of the Acute Phase Response

PubMed Central

Bauzá, Graciela; Miller, Glenn; Kaseje, Neema; Wigner, Nathan A.; Wang, Zhongyan; Gerstenfeld, Louis C.; Burke, Peter A.

2013-01-01

Background The acute-phase response (APR) is critical to the body's ability to successfully respond to injury. A murine model of closed unilateral femur fractures and bilateral femur fracture were used to study the effect of injury magnitude on this response. Methods Standardized unilateral femur fracture and bilateral femur fracture in mice were performed. The femur fracture sites, livers, and serum were harvested over time after injury. Changes in mRNA expression of cytokines, hepatic acute-phase proteins, and serum cytokines overtime were measured. Results There was a rapid and short-lived hepatic APR to fracture injuries. The overall pattern in both models was similar. Both acute-phase proteins' mRNA (fibrinogen-γ and serum amyloid A-3) showed increased mRNA expression over baseline within the first 48 hours and their levels positively correlated with the extent of injury. However, increased severity of injury resulted in a delayed induction of the APR. A similar effect on the gene expression of cytokines (interleukin [IL]-1β, IL-6, and tumor necrosis factor-α) at the fracture site was seen. Serum IL-6 levels increased with increased injury and showed no delay between injury models. Conclusions Greater severity of injury resulted in a delayed induction of the liver's APR and a diminished expression of cytokines at the fracture site. Serum IL-6 levels were calibrated to the extent of the injury, and changes may represent mechanisms by which the local organ responses to injury are regulated by the injury magnitude. PMID:20693926

Effect of Sirtuin-1 on Synaptic Plasticity in Nucleus Accumbens in a Rat Model of Heroin Addiction.

PubMed

Xia, Baijuan; Li, Yixin; Li, Rongrong; Yin, Dan; Chen, Xingqiang; Li, Jie; Liang, Wenmei

2018-06-05

BACKGROUND Synaptic plasticity plays an important role in the process of addiction. This study investigated the relationship between synaptic plasticity and changes in addictive behavior and examined the expression of synaptic plasticity-associated proteins and genes in the nucleus accumbens (NAc) region in different rat models. MATERIAL AND METHODS Heroin addiction, SIRT1-overexpression, and SIRT1-silenced rat models were established. Polymerase chain reaction gene chip technology, immunohistochemistry, Western blotting, and transmission electron microscopy were used to detect changes in synaptic plasticity-related gene and protein expression, and changes in the ultrastructure of synapses, in the NAc. RESULTS Naloxone withdrawal symptoms appeared in the SIRT1-overexpression group. In the SIRT1-silenced group the symptoms were reduced. Immunohistochemistry and Western blotting results showed that FOXO1 expression decreased in the heroin addiction (HA) group but increased in the SIRT1-silenced group (p<0.05). The expression of Cdk5, Nf-κB, PSD95, and Syn was enhanced in the HA group (p<0.05) and further increased in the SIRT1-overexpression group but were reduced in the SIRT1-silenced group (p<0.05). The number of synapses increased in the HA group (p<0.05) along with mitochondrial swelling in the presynaptic membrane and obscuring of the synaptic cleft. CONCLUSIONS SIRT1 and other synaptic plasticity-related genes in NAc are involved in the regulation of heroin addiction. SIRT1 overexpression can increase behavioral sensitization in the NAc of rats, and SIRT1 silencing might ease withdrawal symptoms and reduce conditioned place preferences.

Modulation of Immune Signaling and Metabolism Highlights Host and Fungal Transcriptional Responses in Mouse Models of Invasive Pulmonary Aspergillosis.

PubMed

Kale, Shiv D; Ayubi, Tariq; Chung, Dawoon; Tubau-Juni, Nuria; Leber, Andrew; Dang, Ha X; Karyala, Saikumar; Hontecillas, Raquel; Lawrence, Christopher B; Cramer, Robert A; Bassaganya-Riera, Josep

2017-12-06

Incidences of invasive pulmonary aspergillosis, an infection caused predominantly by Aspergillus fumigatus, have increased due to the growing number of immunocompromised individuals. While A. fumigatus is reliant upon deficiencies in the host to facilitate invasive disease, the distinct mechanisms that govern the host-pathogen interaction remain enigmatic, particularly in the context of distinct immune modulating therapies. To gain insights into these mechanisms, RNA-Seq technology was utilized to sequence RNA derived from lungs of 2 clinically relevant, but immunologically distinct murine models of IPA on days 2 and 3 post inoculation when infection is established and active disease present. Our findings identify notable differences in host gene expression between the chemotherapeutic and steroid models at the interface of immunity and metabolism. RT-qPCR verified model specific and nonspecific expression of 23 immune-associated genes. Deep sequencing facilitated identification of highly expressed fungal genes. We utilized sequence similarity and gene expression to categorize the A. fumigatus putative in vivo secretome. RT-qPCR suggests model specific gene expression for nine putative fungal secreted proteins. Our analysis identifies contrasting responses by the host and fungus from day 2 to 3 between the two models. These differences may help tailor the identification, development, and deployment of host- and/or fungal-targeted therapeutics.

Exercise-Induced Hypertrophic and Oxidative Signaling Pathways and Myokine Expression in Fast Muscle of Adult Zebrafish

PubMed Central

Rovira, Mireia; Arrey, Gerard; Planas, Josep V.

2017-01-01

Skeletal muscle is a plastic tissue that undergoes cellular and metabolic adaptations under conditions of increased contractile activity such as exercise. Using adult zebrafish as an exercise model, we previously demonstrated that swimming training stimulates hypertrophy and vascularization of fast muscle fibers, consistent with the known muscle growth-promoting effects of exercise and with the resulting increased aerobic capacity of this tissue. Here we investigated the potential involvement of factors and signaling mechanisms that could be responsible for exercise-induced fast muscle remodeling in adult zebrafish. By subjecting zebrafish to swimming-induced exercise, we observed an increase in the activity of mammalian target of rapamycin (mTOR) and Mef2 protein levels in fast muscle. We also observed an increase in the protein levels of the mitotic marker phosphorylated histone H3 that correlated with an increase in the protein expression levels of Pax7, a satellite-like cell marker. Furthermore, the activity of AMP-activated protein kinase (AMPK) was also increased by exercise, in parallel with an increase in the mRNA expression levels of pgc1α and also of pparda, a β-oxidation marker. Changes in the mRNA expression levels of slow and fast myosin markers further supported the notion of an exercise-induced aerobic phenotype in zebrafish fast muscle. The mRNA expression levels of il6, il6r, apln, aplnra and aplnrb, sparc, decorin and igf1, myokines known in mammals to be produced in response to exercise and to signal through mTOR/AMPK pathways, among others, were increased in fast muscle of exercised zebrafish. These results support the notion that exercise increases skeletal muscle growth and myogenesis in adult zebrafish through the coordinated activation of the mTOR-MEF2 and AMPK-PGC1α signaling pathways. These results, coupled with altered expression of markers for oxidative metabolism and fast-to-slow fiber-type switch, also suggest improved aerobic capacity as a result of swimming-induced exercise. Finally, the induction of myokine expression by swimming-induced exercise support the hypothesis that these myokines may have been produced and secreted by the exercised zebrafish muscle and acted on fast muscle cells to promote metabolic remodeling. These results support the use of zebrafish as a suitable model for studies on muscle remodeling in vertebrates, including humans. PMID:29326600

MicroRNA-155 deficiency promotes nephrin acetylation and attenuates renal damage in hyperglycemia-induced nephropathy.

PubMed

Lin, Xu; You, Yanwu; Wang, Jie; Qin, Youling; Huang, Peng; Yang, Fafen

2015-04-01

MiR-155 has been reported to be involved in both innate and adaptive immune responses. But the role of miR-155 in hyperglycemia-induced nephropathy is still unknown. In our current study, 3-month-old male wild-type C57 mice and Mir-155(-/-) mice were used to establish hyperglycemia-induced nephropathy. In our hyperglycemia-induced nephropathy model, the expression of podocyte injury marker desmin was markedly increased in the diabetes group when compared with control. Diabetes also significantly decreased the levels of nephrin and acetylated nephrin, whereas the expression of miR-155 was markedly increased in diabetes group when compared with control. MiR-155(-/-) mice showed significantly increased expression of nephrin, acetylated nephrin, and Wilm's tumor-1 protein (WT-1) when compared with wild-type control. MiR-155 deficiency results in significantly decrease in IL-17A expression both in vivo and in vitro. And the increased expression of WT-1, nephrin, and ac-nephrin was reversed with additional treatment of rmIL-17. Furthermore, we found that the inhibited Th17 differentiation induced by miR-155 deficiency was dependent on increased expression of SOCS1. In conclusion, miR-155 deficiency promotes nephrin acetylation and attenuates renal damage in hyperglycemia-induced nephropathy. This was associated with inhibited IL-17 production through enhancement of SOCS1 expression.

Baicalin increases developmental competence of mouse embryos in vitro by inhibiting cellular apoptosis and modulating HSP70 and DNMT expression

PubMed Central

QI, Xiaonan; LI, Huatao; CONG, Xia; WANG, Xin; JIANG, Zhongling; CAO, Rongfeng; TIAN, Wenru

2016-01-01

Scutellaria baicalensis has been effectively used in Chinese traditional medicine to prevent miscarriages. However, little information is available on its mechanism of action. This study is designed specifically to reveal how baicalin, the main effective ingredient of S. baicalensis, improves developmental competence of embryos in vitro, using the mouse as a model. Mouse pronuclear embryos were cultured in KSOM medium supplemented with (0, 2, 4 and 8 μg/ml) baicalin. The results demonstrated that in vitro culture conditions significantly decreased the blastocyst developmental rate and blastocyst quality, possibly due to increased cellular stress and apoptosis. Baicalin (4 µg/ml) significantly increased 2- and 4-cell cleavage rates, morula developmental rate, and blastocyst developmental rate and cell number of in vitro-cultured mouse embryos. Moreover, baicalin increased the expression of Gja1, Cdh1, Bcl-2, and Dnmt3a genes, decreased the expression of Dnmt1 gene, and decreased cellular stress and apoptosis as it decreased the expression of HSP70, CASP3, and BAX and increased BCL-2 expression in blastocysts cultured in vitro. In conclusion, baicalin improves developmental competence of in vitro-cultured mouse embryos through inhibition of cellular apoptosis and HSP70 expression, and improvement of DNA methylation. PMID:27478062

Theoretical model of the effect of potassium on the uptake of radiocesium by rice.

PubMed

Fujimura, Shigeto; Ishikawa, Junko; Sakuma, Yuuki; Saito, Takashi; Sato, Mutsuto; Yoshioka, Kunio

2014-12-01

After the accident at the Fukushima Dai-ichi Nuclear Power Plant owned by Tokyo Electric Power Company on 11 March 2011, potassium was applied to fields in the Tohoku and Kanto areas of Japan to reduce radiocesium uptake by crops. Despite the intense studies relating to the effect of potassium application on availability of radiocesium in the soil, physiological changes of radiocesium uptake by crops in response to K(+) concentration around roots remains elusive. In the present study, we developed physiological models describing the effect of K(+) on the uptake of radiocesium by rice. Two Cs(+):K(+) competition models were evaluated using a wide range of data obtained from pot and field experiments: the model assuming a uniformity in the gene expression of K(+) transporter (Model I) and the model assuming the increase in the gene expression of K(+) transporter in response to K(+) concentration below threshold (Model II). The root-mean-square deviation between the measured and estimated values was larger in Model I than in Model II. Residuals were positively correlated with K(+) in Model I but showed no deflection in Model II. These results indicate that Model II explains the effect of K(+) on the uptake of radiocesium better than Model I. Model II may provide the appropriate countermeasures in inhibiting the transfer of radiocesium from soil to crop. The effect of changes in the variables in Model II on the relationship between available K(+) in soil and (137)Cs uptake by plant was simulated. An increase in available (137)Cs(+) in soil enhanced the response of (137)Cs uptake to K(+). The effects of Michaelis-Menten constant for Cs(+) were the inverse of the (137)Cs(+) effect. The effect of Michaelis-Menten constant for K(+) showed the same tendency as that of (137)Cs(+), but the effect was much less than that of (137)Cs(+). An increase in the threshold of K(+) below which the gene expression of K(+) transporter increases enhanced the response of (137)Cs uptake to K(+) in the high-K(+) range. Copyright © 2014 Elsevier Ltd. All rights reserved.

Computational Model of the Modulation of Gene Expression Following DNA Damage

NASA Technical Reports Server (NTRS)

Cucinotta, F. A.; Dicello, J. F.; Nikjoo, H.; Cherubini, R.

2002-01-01

High linear energy transfer (LET) radiation, such as heavy ions or neutrons, has an increased biological effectiveness compared to X rays for gene mutation, genomic instability, and carcinogenesis. In the traditional paradigm, mutations or chromosomal aberrations are causative of late effects. However, in recent years experimental evidence has demonstrated the important role of the description of the modification of gene expression by radiation in understanding the mechanisms of radiation action. In this report, approaches are discussed to the mathematical description of mRNA and protein expression kinetics following DNA damage. Several hypotheses for models of radiation modulation of protein expression are discussed including possible non-linear processes that evolve from the linear dose responses that follow the initial DNA damage produced by radiation.

Impact of chronic unpredicted mild stress-induced depression on repaglinide fate via glucocorticoid signaling pathway.

PubMed

Wei, Hongyan; Zhou, Ting; Tan, Boyu; Zhang, Lei; Li, Mingming; Xiao, Zhijun; Xu, Feng

2017-07-04

Chronic unpredicted mild stress (CUMS)-induced depression could alter the pharmacokinetics of many drugs in rats, however, the underlying mechanism is not clear. In this work we studied the pharmacokinetics of repaglinide, and explored the role of glucocorticoid and adrenergic signaling pathway in regulating drug metabolizing enzymes (DMEs) in GK rats and BRL 3A cells. The plasma cortisol and epinephrine levels were increased, meanwhile the pharmacokinetics of repaglinide were altered significantly in depression model rats. Forty-nine genes in liver of model rats displayed significant difference comparing to control rats. The differentially expressed genes enriched in the drug metabolism and steroid hormone biosynthesis pathway significantly, and Nr1i3 matched 335 connectivity genes. CAR and Ugt1a1 protein expression were enhanced significantly in liver of model rats. The mRNA expression of Ugt1a1 and Nr1i2 were increased 2 and 4 times respectively with dexamethasone (DEX) and 8-Br-cAMP co-treatment in BRL 3A cells. The protein expression of PXR was up-regulated, too. However, RU486 reversed the up-regulated effect. The adrenergic receptor agonists had little impact on the DMEs in BRL 3A. Our data suggested that CUMS-induced depression might up-regulate DMEs expression via glucocorticoid signaling pathway, and accelerate the fate of the repaglinide in spontaneous diabetes rats.

Impact of chronic unpredicted mild stress-induced depression on repaglinide fate via glucocorticoid signaling pathway

PubMed Central

Tan, Boyu; Zhang, Lei; Li, Mingming; Xiao, Zhijun; Xu, Feng

2017-01-01

Chronic unpredicted mild stress (CUMS)-induced depression could alter the pharmacokinetics of many drugs in rats, however, the underlying mechanism is not clear. In this work we studied the pharmacokinetics of repaglinide, and explored the role of glucocorticoid and adrenergic signaling pathway in regulating drug metabolizing enzymes (DMEs) in GK rats and BRL 3A cells. The plasma cortisol and epinephrine levels were increased, meanwhile the pharmacokinetics of repaglinide were altered significantly in depression model rats. Forty-nine genes in liver of model rats displayed significant difference comparing to control rats. The differentially expressed genes enriched in the drug metabolism and steroid hormone biosynthesis pathway significantly, and Nr1i3 matched 335 connectivity genes. CAR and Ugt1a1 protein expression were enhanced significantly in liver of model rats. The mRNA expression of Ugt1a1 and Nr1i2 were increased 2 and 4 times respectively with dexamethasone (DEX) and 8-Br-cAMP co-treatment in BRL 3A cells. The protein expression of PXR was up-regulated, too. However, RU486 reversed the up-regulated effect. The adrenergic receptor agonists had little impact on the DMEs in BRL 3A. Our data suggested that CUMS-induced depression might up-regulate DMEs expression via glucocorticoid signaling pathway, and accelerate the fate of the repaglinide in spontaneous diabetes rats. PMID:28574832

20180311 - Differential Gene Expression and Concentration-Response Modeling Workflow for High-Throughput Transcriptomic (HTTr) Data: Results From MCF7 Cells (SOT)

EPA Science Inventory

Increasing efficiency and declining cost of generating whole transcriptome profiles has made high-throughput transcriptomics a practical option for chemical bioactivity screening. The resulting data output provides information on the expression of thousands of genes and is amenab...

Music and the Expressive Arts with Children Experiencing Trauma

ERIC Educational Resources Information Center

Davis, Keith M.

2010-01-01

The creative and expressive use of music can be a powerful therapeutic intervention with children and adolescents who have experienced trauma. In this article, a model for increasing self-awareness and self-understanding including materials, facilitation, and processing of musical activities in group format is presented. Creative activities such…

Instrumentality, Expressivity, and Relational Qualities in the Same-Sex Friendships of College Women and Men

ERIC Educational Resources Information Center

Frey, Lisa L.; Beesley, Denise; Hurst, Rebecca; Saldana, Star; Licuanan, Brian

2016-01-01

Using the relational-cultural model (Jordan, Kaplan, Miller, Stiver, & Surrey, 1991), the authors hypothesized that instrumentality, expressivity, and the individual affective experience of same-sex friendships would predict increased relationship mutuality, with college women and men showing different predictive patterns. Overall, results…

Differential Gene Expression and Concentration-Response Modeling Workflow for High-Throughput Transcriptomic (HTTr) Data: Results From MCF7 Cells

EPA Science Inventory

Increasing efficiency and declining cost of generating whole transcriptome profiles has made high-throughput transcriptomics a practical option for chemical bioactivity screening. The resulting data output provides information on the expression of thousands of genes and is amenab...

Nitric oxide increases Wnt-induced secreted protein-1 (WISP-1/CCN4) expression and function in colitis.

PubMed

Wang, Hongying; Zhang, Rui; Wen, Shoubin; McCafferty, Donna-Marie; Beck, Paul L; MacNaughton, Wallace K

2009-04-01

Nitric oxide (NO) derived from the inducible NO synthase (iNOS) is an important and complex mediator of inflammation in the intestine. Wnt-inducible secreted protein (WISP)-1 (CCN4), a member of the connective tissue growth factor family, is involved in tissue repair. We sought to determine the relationship between iNOS and WISP-1 in colitis. By analyzing human colonic biopsy samples, we showed that the expression of mRNA for both iNOS and WISP-1 was significantly higher in ulcerative colitis samples compared with control tissue. The upregulation of WISP-1 was positively correlated with iNOS expression in two models of colitis, induced by intrarectal trinitrobenzenesulfonic acid (TNBS) or occurring spontaneously in IL-10 deficient mice. Loss of iNOS, studied using iNOS(-/-) mice in both TNBS-induced and IL-10(-/-) colitis models, significantly attenuated the colitis-related WISP-1 increase. In human colonic epithelial cell lines, the NO donor, DETA-NONOate, elevated WISP-1 mRNA and protein expression through a beta-catenin and CREB-dependent, but Wnt-1-independent, pathway. In addition, NO-induced WISP-1 directly induced secretion of soluble collagen in colonic fibroblast cells. NO increases WISP-1 expression both in vitro and in vivo, suggesting a new role for iNOS and NO in colitis.

Fibroblast spheroids as a model to study sustained fibroblast quiescence and their crosstalk with tumor cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Salmenperä, Pertteli, E-mail: pertteli.salmenpera@helsinki.fi; Karhemo, Piia-Riitta; Räsänen, Kati

Stromal fibroblasts have an important role in regulating tumor progression. Normal and quiescent fibroblasts have been shown to restrict and control cancer cell growth, while cancer-associated, i. e. activated fibroblasts have been shown to enhance proliferation and metastasis of cancer cells. In this study we describe generation of quiescent fibroblasts in multicellular spheroids and their effects on squamous cell carcinoma (SCC) growth in soft-agarose and xenograft models. Quiescent phenotype of fibroblasts was determined by global down-regulation of expression of genes related to cell cycle and increased expression of p27. Interestingly, microarray analysis showed that fibroblast quiescence was associated with similarmore » secretory phenotype as seen in senescence and they expressed senescence-associated-β-galactosidase. Quiescent fibroblasts spheroids also restricted the growth of RT3 SCC cells both in soft-agarose and xenograft models unlike proliferating fibroblasts. Restricted tumor growth was associated with marginally increased tumor cell senescence and cellular differentiation, showed with senescence-associated-β-galactosidase and cytokeratin 7 staining. Our results show that the fibroblasts spheroids can be used as a model to study cellular quiescence and their effects on cancer cell progression. - Highlights: • Fibroblasts acquire a sustained quiescence when grown as multicellular spheroids. • This quiescence is associated with drastic change in gene expression. • Fibroblasts spheroids secrete various inflammation-linked cytokines and chemokines. • Fibroblasts spheroids reduced growth of RT3 SCC cells in xenograft model.« less

Effect of Tripterygium Wilfordii Polyglycoside on Experimental Prostatitis Caused by Ureaplasma Urealyticum in Rats

PubMed Central

Shan, Pingnan; Lu, Zhiyong; Ye, Lihong; Fang, Yaqin; Tan, Suhong; Xuan, Guohong; Ru, Jincheng; Mao, Liming

2016-01-01

Background Prostatitis is a common and refractory urological disease with complicated etiology. Ureaplasma urealyticum (UU) has a close relationship with human urinary tract infection that can induce nonbacterial prostatitis. Tripterygium wilfordii polyglycoside (TWP) is a non-steroidal immune inhibitor that causes significant immune suppression and anti-inflammatory effects. Its role in prostatitis caused by UU has not yet been established. The aim of this study was to investigate the effect of TWP on UU-infected prostatitis in a rat model. Material/Methods UU-infected prostatitis SD model rats were randomly divided into 2 groups: the prostatitis group (model group) and the TWP treatment group (treatment group). At 7 days after treatment, prostate weight, leucocyte count, lecithin corpuscles, UU infection rate, and UU microbe count were compared between the 2 groups. Serum inflammatory cytokines TNF-α was determined by ELISA, and ICAM-1 and NF-κB expression were detected. Results UU infection rate was 80% after modeling. The rat prostate weight and leucocyte count in the model group increased significantly, while lecithin corpuscles decreased. Compared with controls, inflammatory factor TNF-α, ICAM-1, and NF-κB expression were obviously higher (P<0.05). TWP markedly reduced prostate weight and leucocyte count, increased lecithin corpuscles, and decreased UU microbe count and TNF-α, ICAM-1, and NF-κB expression (P<0.05). Conclusions TWP can inhibit expression of inflammatory factors and may be useful in treating UU-infected prostatitis through reducing UU infection rate. PMID:27743513

Effect of Tripterygium Wilfordii Polyglycoside on Experimental Prostatitis Caused by Ureaplasma Urealyticum in Rats.

PubMed

Shan, Pingnan; Lu, Zhiyong; Ye, Lihong; Fang, Yaqin; Tan, Suhong; Xuan, Guohong; Ru, Jincheng; Mao, Liming

2016-10-15

BACKGROUND Prostatitis is a common and refractory urological disease with complicated etiology. Ureaplasma urealyticum (UU) has a close relationship with human urinary tract infection that can induce nonbacterial prostatitis. Tripterygium wilfordii polyglycoside (TWP) is a non-steroidal immune inhibitor that causes significant immune suppression and anti-inflammatory effects. Its role in prostatitis caused by UU has not yet been established. The aim of this study was to investigate the effect of TWP on UU-infected prostatitis in a rat model. MATERIAL AND METHODS UU-infected prostatitis SD model rats were randomly divided into 2 groups: the prostatitis group (model group) and the TWP treatment group (treatment group). At 7 days after treatment, prostate weight, leucocyte count, lecithin corpuscles, UU infection rate, and UU microbe count were compared between the 2 groups. Serum inflammatory cytokines TNF-α was determined by ELISA, and ICAM-1 and NF-κB expression were detected. RESULTS UU infection rate was 80% after modeling. The rat prostate weight and leucocyte count in the model group increased significantly, while lecithin corpuscles decreased. Compared with controls, inflammatory factor TNF-α, ICAM-1, and NF-κB expression were obviously higher (P<0.05). TWP markedly reduced prostate weight and leucocyte count, increased lecithin corpuscles, and decreased UU microbe count and TNF-α, ICAM-1, and NF-κB expression (P<0.05). CONCLUSIONS TWP can inhibit expression of inflammatory factors and may be useful in treating UU-infected prostatitis through reducing UU infection rate.

Development of Urinary Bladder Pre-Neoplasia by Schistosoma haematobium Eggs and Chemical Carcinogen in Mice

PubMed Central

Chala, Bayissa; Choi, Min-Ho; Moon, Kyung Chul; Kim, Hyung Suk; Kwak, Cheol; Hong, Sung-Tae

2017-01-01

Schistosoma haematobium is a biocarcinogen of human urinary bladder (UB). The present study investigated developing UB cancer mouse model by injecting S. haematobium eggs into the bladder wall and introduction of chemical carcinogens. Histopathological findings showed mild hyperplasia to epithelial vacuolar change, and high grade dysplasia. Squamous metaplasia was observed in the S. haematobium eggs+NDMA group at week 12 but not in other groups. Immunohistochemistry revealed significantly high expression of Ki-67 in urothelial epithelial cells of the S. haematobium eggs+BBN group at week 20. The qRT-PCR showed high expression of p53 gene in S. haematobium eggs group at week 4 and S. haematobium eggs+BBN group at week 20. E-cadherin and vimentin showed contrasting expression in S. haematobium eggs+BBN group. Such inverse expression of E-cadherin and vimentin may indicate epithelial mesenchymal transition in the UB tissue. In conclusion, S. haematobium eggs and nitrosamines may transform UB cells into squamous metaplasia and dysplasia in correlation with increased expression of Ki-67. Marked decrease in E-cadherin and increase in p53 and vimentin expressions may support the transformation. The present study introduces a promising modified animal model for UB cancer study using S. haematobium eggs. PMID:28285503

Experimental type II diabetes and related models of impaired glucose metabolism differentially regulate glucose transporters at the proximal tubule brush border membrane.

PubMed

Chichger, Havovi; Cleasby, Mark E; Srai, Surjit K; Unwin, Robert J; Debnam, Edward S; Marks, Joanne

2016-06-01

What is the central question of this study? Although SGLT2 inhibitors represent a promising treatment for patients suffering from diabetic nephropathy, the influence of metabolic disruption on the expression and function of glucose transporters is largely unknown. What is the main finding and its importance? In vivo models of metabolic disruption (Goto-Kakizaki type II diabetic rat and junk-food diet) demonstrate increased expression of SGLT1, SGLT2 and GLUT2 in the proximal tubule brush border. In the type II diabetic model, this is accompanied by increased SGLT- and GLUT-mediated glucose uptake. A fasted model of metabolic disruption (high-fat diet) demonstrated increased GLUT2 expression only. The differential alterations of glucose transporters in response to varying metabolic stress offer insight into the therapeutic value of inhibitors. SGLT2 inhibitors are now in clinical use to reduce hyperglycaemia in type II diabetes. However, renal glucose reabsorption across the brush border membrane (BBM) is not completely understood in diabetes. Increased consumption of a Western diet is strongly linked to type II diabetes. This study aimed to investigate the adaptations that occur in renal glucose transporters in response to experimental models of diet-induced insulin resistance. The study used Goto-Kakizaki type II diabetic rats and normal rats rendered insulin resistant using junk-food or high-fat diets. Levels of protein kinase C-βI (PKC-βI), GLUT2, SGLT1 and SGLT2 were determined by Western blotting of purified renal BBM. GLUT- and SGLT-mediated d-[(3) H]glucose uptake by BBM vesicles was measured in the presence and absence of the SGLT inhibitor phlorizin. GLUT- and SGLT-mediated glucose transport was elevated in type II diabetic rats, accompanied by increased expression of GLUT2, its upstream regulator PKC-βI and SGLT1 protein. Junk-food and high-fat diet feeding also caused higher membrane expression of GLUT2 and its upstream regulator PKC-βI. However, the junk-food diet also increased SGLT1 and SGLT2 levels at the proximal tubule BBM. Glucose reabsorption across the proximal tubule BBM, via GLUT2, SGLT1 and SGLT2, is not solely dependent on glycaemic status, but is also influenced by diet-induced changes in glucose metabolism. We conclude that different metabolic disturbances result in complex adaptations in renal glucose transporter protein levels and function. © 2016 The Authors. Experimental Physiology © 2016 The Physiological Society.

Paeonia lactiflora Enhances the Adhesion of Trophoblast to the Endometrium via Induction of Leukemia Inhibitory Factor Expression

PubMed Central

Park, Mi-Ju; Lee, Kyu Sup; Yoon, Youngjin; Kim, Hyung Sik; Lee, Jun Hee; Kwon, Sang-Mo; Lee, Syng-Ook; Kim, Keuk-Jun; Baek, Jin-Ho; Ha, Ki-Tae

2016-01-01

In the present study, we investigated the role of Paeonia lactiflora Pall. extract on embryo implantation in vitro and in vivo. A polysaccharides depleted-water extract of P. lactiflora (PL-PP) increased LIF expression in human endometrial Ishikawa cells at non-cytotoxic doses. PL-PP significantly increased the adhesion of the human trophectoderm-derived JAr spheroids to endometrial Ishikawa cells. PL-PP-induced LIF expression was decreased in the presence of a p38 kinase inhibitor SB203580 and an MEK/ERK inhibitor U0126. Furthermore, endometrial LIF knockdown by shRNA reduced the expression of integrins β3 and β5 and adhesion of JAr spheroids to Ishikawa cells. In vivo administration of PL-PP restored the implantation of mouse blastocysts in a mifepristone-induced implantation failure mice model. Our results demonstrate that PL-PP increases LIF expression via the p38 and MEK/ERK pathways and favors trophoblast adhesion to endometrial cells. PMID:26839969

Iron Overload Accelerates the Progression of Diabetic Retinopathy in Association with Increased Retinal Renin Expression.

PubMed

Chaudhary, Kapil; Promsote, Wanwisa; Ananth, Sudha; Veeranan-Karmegam, Rajalakshmi; Tawfik, Amany; Arjunan, Pachiappan; Martin, Pamela; Smith, Sylvia B; Thangaraju, Muthusamy; Kisselev, Oleg; Ganapathy, Vadivel; Gnana-Prakasam, Jaya P

2018-02-14

Diabetic retinopathy (DR) is a leading cause of blindness among working-age adults. Increased iron accumulation is associated with several degenerative diseases. However, there are no reports on the status of retinal iron or its implications in the pathogenesis of DR. In the present study, we found that retinas of type-1 and type-2 mouse models of diabetes have increased iron accumulation compared to non-diabetic retinas. We found similar iron accumulation in postmortem retinal samples from human diabetic patients. Further, we induced diabetes in HFE knockout (KO) mice model of genetic iron overload to understand the role of iron in the pathogenesis of DR. We found increased neuronal cell death, vascular alterations and loss of retinal barrier integrity in diabetic HFE KO mice compared to diabetic wildtype mice. Diabetic HFE KO mouse retinas also exhibited increased expression of inflammation and oxidative stress markers. Severity in the pathogenesis of DR in HFE KO mice was accompanied by increase in retinal renin expression mediated by G-protein-coupled succinate receptor GPR91. In light of previous reports implicating retinal renin-angiotensin system in DR pathogenesis, our results reveal a novel relationship between diabetes, iron and renin-angiotensin system, thereby unraveling new therapeutic targets for the treatment of DR.

Methionine Deprivation Induces a Targetable Vulnerability in Triple-negative Breast Cancer Cells by Enhancing TRAIL Receptor-2 Expression

PubMed Central

Strekalova, Elena; Malin, Dmitry; Good, David M.; Cryns, Vincent L.

2015-01-01

Purpose Many neoplasms are vulnerable to methionine deficiency by mechanisms that are poorly understood. Because gene profiling studies have revealed that methionine depletion increases TNF-related apoptosis-inducing ligand receptor-2 (TRAIL-R2) mRNA, we postulated that methionine stress sensitizes breast cancer cells to proapoptotic TRAIL-R2 agonists. Experimental Design Human triple (ER/PR/HER2)-negative breast carcinoma cell lines were cultured in control or methionine-free media. The effects of methionine depletion on TRAIL receptor expression and sensitivity to chemotherapy or a humanized agonistic TRAIL-R2 monoclonal antibody (lexatumumab) were determined. The melanoma-associated antigen MAGED2 was silenced to delineate its functional role in sensitizing TNBC cells to methionine stress. An orthotopic TNBC model was utilized to evaluate the effects of dietary methionine deficiency, lexatumumab or the combination. Results Methionine depletion sensitized TNBC cells to lexatumumab-induced caspase activation and apoptosis by increasing TRAIL-R2 mRNA and cell surface expression. MCF-10A cells transformed by oncogenic H-Ras, but not untransformed cells, and matrix-detached TNBC cells were highly sensitive to the combination of lexatumumab and methionine depletion. Proteomics analyses revealed that MAGED2, which has been reported to reduce TRAIL-R2 expression, was suppressed by methionine stress. Silencing MAGED2 recapitulated features of methionine deprivation, including enhanced mRNA and cell surface expression of TRAIL receptors and increased sensitivity to TRAIL receptor agonists. Dietary methionine deprivation enhanced the antitumor effects of lexatumumab in an orthotopic metastatic TNBC model. Conclusion Methionine depletion exposes a targetable defect in TNBC cells by increasing TRAIL-R2 expression. Our findings provide the foundation for a clinical trial combining dietary methionine restriction and TRAIL-R2 agonists. PMID:25724522

PDGF-α stimulates intestinal epithelial cell turnover after massive small bowel resection in a rat.

PubMed

Sukhotnik, Igor; Mogilner, Jorge G; Pollak, Yulia; Blumenfeld, Shiri; Bejar, Jacob; Coran, Arnold G

2012-06-01

Numerous cytokines have been shown to affect epithelial cell differentiation and proliferation through epithelial-mesenchymal interaction. Growing evidence suggests that platelet-derived growth factor (PDGF) signaling is an important mediator of these interactions. The purpose of this study was to evaluate the effect of PDGF-α on enterocyte turnover in a rat model of short bowel syndrome (SBS). Male rats were divided into four groups: Sham rats underwent bowel transection, Sham-PDGF-α rats underwent bowel transection and were treated with PDGF-α, SBS rats underwent a 75% bowel resection, and SBS-PDGF-α rats underwent bowel resection and were treated with PDGF-α. Parameters of intestinal adaptation, enterocyte proliferation and apoptosis were determined at euthanasia. Illumina's Digital Gene Expression analysis was used to determine PDGF-related gene expression profiling. PDGF-α and PDGF-α receptor (PDGFR-α) expression was determined by real-time PCR. Western blotting was used to determine p-ERK, Akt1/2/3, bax, and bcl-2 protein levels. SBS rats demonstrated a significant increase in PDGF-α and PDGFR-α expression in jejunum and ileum compared with sham animals. SBS-PDGF-α rats demonstrated a significant increase in bowel and mucosal weight, villus height, and crypt depth in jejunum and ileum compared with SBS animals. PDGF-α receptor expression in crypts increased in SBS rats (vs. sham) and was accompanied by an increased cell proliferation following PDGF-α administration. A significant decrease in cell apoptosis in this group was correlated with lower bax protein levels. In conclusion, in a rat model of SBS, PDGF-α stimulates enterocyte turnover, which is correlated with upregulated PDGF-α receptor expression in the remaining small intestine.

Gene Expression Profiles of Human Dendritic Cells Interacting with Aspergillus fumigatus in a Bilayer Model of the Alveolar Epithelium/Endothelium Interface

PubMed Central

Morton, Charles Oliver; Fliesser, Mirjam; Dittrich, Marcus; Mueller, Tobias; Bauer, Ruth; Kneitz, Susanne; Hope, William; Rogers, Thomas Richard; Einsele, Hermann; Loeffler, Juergen

2014-01-01

The initial stages of the interaction between the host and Aspergillus fumigatus at the alveolar surface of the human lung are critical in the establishment of aspergillosis. Using an in vitro bilayer model of the alveolus, including both the epithelium (human lung adenocarcinoma epithelial cell line, A549) and endothelium (human pulmonary artery epithelial cells, HPAEC) on transwell membranes, it was possible to closely replicate the in vivo conditions. Two distinct sub-groups of dendritic cells (DC), monocyte-derived DC (moDC) and myeloid DC (mDC), were included in the model to examine immune responses to fungal infection at the alveolar surface. RNA in high quantity and quality was extracted from the cell layers on the transwell membrane to allow gene expression analysis using tailored custom-made microarrays, containing probes for 117 immune-relevant genes. This microarray data indicated minimal induction of immune gene expression in A549 alveolar epithelial cells in response to germ tubes of A. fumigatus. In contrast, the addition of DC to the system greatly increased the number of differentially expressed immune genes. moDC exhibited increased expression of genes including CLEC7A, CD209 and CCL18 in the absence of A. fumigatus compared to mDC. In the presence of A. fumigatus, both DC subgroups exhibited up-regulation of genes identified in previous studies as being associated with the exposure of DC to A. fumigatus and exhibiting chemotactic properties for neutrophils, including CXCL2, CXCL5, CCL20, and IL1B. This model closely approximated the human alveolus allowing for an analysis of the host pathogen interface that complements existing animal models of IA. PMID:24870357

Manganese Superoxide Dismutase Is a Promising Target for Enhancing Chemosensitivity of Basal-Like Breast Carcinoma

PubMed Central

Kumar, Alan Prem; Loo, Ser Yue; Shin, Sung Won; Tan, Tuan Zea; Eng, Chon Boon; Singh, Rajeev; Putti, Thomas Choudary; Ong, Chee Wee; Salto-Tellez, Manuel; Goh, Boon Cher; Park, Joo In; Thiery, Jean Paul; Pervaiz, Shazib

2014-01-01

Abstract Aims: Although earlier reports highlighted a tumor suppressor role for manganese superoxide dismutase (MnSOD), recent evidence indicates increased expression in a variety of human cancers including aggressive breast carcinoma. In the present article, we hypothesized that MnSOD expression is significantly amplified in the aggressive breast carcinoma basal subtype, and targeting MnSOD could be an attractive strategy for enhancing chemosensitivity of this highly aggressive breast cancer subtype. Results: Using MDA-MB-231 and BT549 as a model of basal breast cancer cell lines, we show that knockdown of MnSOD decreased the colony-forming ability and sensitized the cells to drug-induced cell death, while drug resistance was associated with increased MnSOD expression. In an attempt to develop a clinically relevant approach to down-regulate MnSOD expression in patients with basal breast carcinoma, we employed activation of the peroxisome proliferator-activated receptor gamma (PPARγ) to repress MnSOD expression; PPARγ activation significantly reduced MnSOD expression, increased chemosensitivity, and inhibited tumor growth. Moreover, as a proof of concept for the clinical use of PPARγ agonists to decrease MnSOD expression, biopsies derived from breast cancer patients who had received synthetic PPARγ ligands as anti-diabetic therapy had significantly reduced MnSOD expression. Finally, we provide evidence to implicate peroxynitrite as the mechanism involved in the increased sensitivity to chemotherapy induced by MnSOD repression. Innovation and Conclusion: These data provide evidence to link increased MnSOD expression with the aggressive basal breast cancer, and underscore the judicious use of PPARγ ligands for specifically down-regulating MnSOD to increase the chemosensitivity of this subtype of breast carcinoma. Antioxid. Redox Signal. 20, 2326–2346. PMID:23964924

Epidermal Cadm1 expression promotes autoimmune alopecia via enhanced T cell adhesion and cytotoxicity.

PubMed

Giangreco, Adam; Hoste, Esther; Takai, Yoshimi; Rosewell, Ian; Watt, Fiona M

2012-02-01

Autoimmune alopecia is characterized by an extensive epidermal T cell infiltrate that mediates hair follicle destruction. We have investigated the role of cell adhesion molecule 1 (Cadm1; Necl2) in this disease. Cadm1 is expressed by epidermal cells and mediates heterotypic adhesion to lymphocytes expressing class 1-restricted T cell-associated molecule (CRTAM). Using a murine autoimmune alopecia model, we observed an increase in early-activated cytotoxic (CD8-restricted, CRTAM-expressing) T cells, which preferentially associated with hair follicle keratinocytes expressing Cadm1. Coculture with Cadm1-transduced MHC-matched APCs stimulated alopecic lymph node cells to release IL-2 and IFN-γ. Overexpression of Cadm1 in cultured human keratinocytes did not promote cytokine secretion, but led to increased adhesion of alopecic cytotoxic T cells and enhanced T cell cytotoxicity in an MHC-independent manner. Epidermal overexpression of Cadm1 in transgenic mice led to increased autoimmune alopecia susceptibility relative to nontransgenic littermate controls. Our findings reveal that Cadm1 expression in the hair follicle plays a role in autoimmune alopecia.

Reduced expression of brain cannabinoid receptor 1 (Cnr1) is coupled with an increased complementary micro-RNA (miR-26b) in a mouse model of fetal alcohol spectrum disorders.

PubMed

Stringer, Randa L; Laufer, Benjamin I; Kleiber, Morgan L; Singh, Shiva M

2013-08-02

Prenatal alcohol exposure is known to result in fetal alcohol spectrum disorders, a continuum of physiological, behavioural, and cognitive phenotypes that include increased risk for anxiety and learning-associated disorders. Prenatal alcohol exposure results in life-long disorders that may manifest in part through the induction of long-term gene expression changes, potentially maintained through epigenetic mechanisms. Here we report a decrease in the expression of Canabinoid receptor 1 (Cnr1) and an increase in the expression of the regulatory microRNA miR-26b in the brains of adult mice exposed to ethanol during neurodevelopment. Furthermore, we show that miR-26b has significant complementarity to the 3'-UTR of the Cnr1 transcript, giving it the potential to bind and reduce the level of Cnr1 expression. These findings elucidate a mechanism through which some genes show long-term altered expression following prenatal alcohol exposure, leading to persistent alterations to cognitive function and behavioural phenotypes observed in fetal alcohol spectrum disorders.

A Protein Chimera Strategy Supports Production of a Model "Difficult-to-Express" Recombinant Target.

PubMed

Hussain, Hirra; Fisher, David I; Roth, Robert G; Abbott, W Mark; Carballo-Amador, Manuel Alejandro; Warwicker, Jim; Dickson, Alan J

2018-06-22

Due in part to the needs of the biopharmaceutical industry, there has been an increased drive to generate high quality recombinant proteins in large amounts. However, achieving high yields can be a challenge as the novelty and increased complexity of new targets often makes them 'difficult-to-express'. This study aimed to define the molecular features that restrict the production of a model 'difficult-to-express' recombinant protein, Tissue Inhibitor Metalloproteinase-3 (TIMP-3). Building from experimental data, computational approaches were used to rationalise the re-design of this recombinant target to generate a chimera with enhanced secretion. The results highlight the importance of early identification of unfavourable sequence attributes, enabling the generation of engineered protein forms that bypass 'secretory' bottlenecks and result in efficient recombinant protein production. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

Carboxymethylcellulose-based and docetaxel-loaded nanoparticles circumvent P-glycoprotein mediated multidrug resistance

PubMed Central

Roy, Aniruddha; Murakami, Mami; Ernsting, Mark J.; Hoang, Bryan; Undzys, Elijus; Li, Shyh-Dar

2014-01-01

Taxanes are a class of anticancer agents with a broad spectrum and have been widely used to treat a variety of cancer. However, its long term use has been hampered by accumulating toxicity and development of drug resistance. The most extensively reported mechanism of resistance is the overexpression of P-glycoprotein (Pgp). We have developed a PEGylated carboxymethylcellulose conjugate of docetaxel (Cellax), which condenses into ~120 nm nanoparticles. Here we demonstrated that Cellax therapy did not upregulate Pgp expression in MDA-MB-231 and EMT-6 breast tumor cells whereas a significant increase in Pgp expression was measured with native docetaxel (DTX) treatment. Treatment with DTX led to 4 to 7-fold higher Pgp mRNA expression and 2-fold higher Pgp protein expression compared to Cellax treatment in the in vitro and in vivo system respectively. Cellax also exhibited significantly increased efficacy compared to DTX in a taxane-resistant breast tumor model. Against the highly Pgp expressing EMT6/AR1 cells, Cellax exhibited a 6.5 times lower IC50 compared to native DTX, and in the in vivo model, Cellax exhibited 90% tumor growth inhibition, while native DTX had no significant antitumor activity. PMID:24564177

Hydrocortisone-induced parkin prevents dopaminergic cell death via CREB pathway in Parkinson's disease model.

PubMed

Ham, Sangwoo; Lee, Yun-Il; Jo, Minkyung; Kim, Hyojung; Kang, Hojin; Jo, Areum; Lee, Gum Hwa; Mo, Yun Jeong; Park, Sang Chul; Lee, Yun Song; Shin, Joo-Ho; Lee, Yunjong

2017-04-03

Dysfunctional parkin due to mutations or post-translational modifications contributes to dopaminergic neurodegeneration in Parkinson's disease (PD). Overexpression of parkin provides protection against cellular stresses and prevents dopamine cell loss in several PD animal models. Here we performed an unbiased high-throughput luciferase screening to identify chemicals that can increase parkin expression. Among promising parkin inducers, hydrocortisone possessed the most favorable profiles including parkin induction ability, cell protection ability, and physicochemical property of absorption, distribution, metabolism, and excretion (ADME) without inducing endoplasmic reticulum stress. We found that hydrocortisone-induced parkin expression was accountable for cell protection against oxidative stress. Hydrocortisone-activated parkin expression was mediated by CREB pathway since gRNA to CREB abolished hydrocortisone's ability to induce parkin. Finally, hydrocortisone treatment in mice increased brain parkin levels and prevented 6-hydroxy dopamine induced dopamine cell loss when assessed at 4 days after the toxin's injection. Our results showed that hydrocortisone could stimulate parkin expression via CREB pathway and the induced parkin expression was accountable for its neuroprotective effect. Since glucocorticoid is a physiological hormone, maintaining optimal levels of glucocorticoid might be a potential therapeutic or preventive strategy for Parkinson's disease.

Three distinct cell populations express extracellular matrix proteins and increase in number during skeletal muscle fibrosis.

PubMed

Chapman, Mark A; Mukund, Kavitha; Subramaniam, Shankar; Brenner, David; Lieber, Richard L

2017-02-01

Tissue extracellular matrix (ECM) provides structural support and creates unique environments for resident cells (Bateman JF, Boot-Handford RP, Lamandé SR. Nat Rev Genet 10: 173-183, 2009; Kjaer M. Physiol Rev 84: 649-98, 2004). However, the identities of cells responsible for creating specific ECM components have not been determined. In striated muscle, the identity of these cells becomes important in disease when ECM changes result in fibrosis and subsequent increased tissue stiffness and dysfunction. Here we describe a novel approach to isolate and identify cells that maintain the ECM in both healthy and fibrotic muscle. Using a collagen I reporter mouse, we show that there are three distinct cell populations that express collagen I in both healthy and fibrotic skeletal muscle. Interestingly, the number of collagen I-expressing cells in all three cell populations increases proportionally in fibrotic muscle, indicating that all cell types participate in the fibrosis process. Furthermore, while some profibrotic ECM and ECM-associated genes are significantly upregulated in fibrotic muscle, the fibrillar collagen gene expression profile is not qualitatively altered. This suggests that muscle fibrosis in this model results from an increased number of collagen I-expressing cells and not the initiation of a specific fibrotic collagen gene expression program. Finally, in fibrotic muscle, we show that these collagen I-expressing cell populations differentially express distinct ECM proteins-fibroblasts express the fibrillar components of ECM, fibro/adipogenic progenitors cells differentially express basal laminar proteins, and skeletal muscle progenitor cells differentially express genes important for the satellite cell. Copyright © 2017 the American Physiological Society.

Three distinct cell populations express extracellular matrix proteins and increase in number during skeletal muscle fibrosis

PubMed Central

Chapman, Mark A.; Mukund, Kavitha; Subramaniam, Shankar; Brenner, David

2017-01-01

Tissue extracellular matrix (ECM) provides structural support and creates unique environments for resident cells (Bateman JF, Boot-Handford RP, Lamandé SR. Nat Rev Genet 10: 173–183, 2009; Kjaer M. Physiol Rev 84: 649–98, 2004). However, the identities of cells responsible for creating specific ECM components have not been determined. In striated muscle, the identity of these cells becomes important in disease when ECM changes result in fibrosis and subsequent increased tissue stiffness and dysfunction. Here we describe a novel approach to isolate and identify cells that maintain the ECM in both healthy and fibrotic muscle. Using a collagen I reporter mouse, we show that there are three distinct cell populations that express collagen I in both healthy and fibrotic skeletal muscle. Interestingly, the number of collagen I-expressing cells in all three cell populations increases proportionally in fibrotic muscle, indicating that all cell types participate in the fibrosis process. Furthermore, while some profibrotic ECM and ECM-associated genes are significantly upregulated in fibrotic muscle, the fibrillar collagen gene expression profile is not qualitatively altered. This suggests that muscle fibrosis in this model results from an increased number of collagen I-expressing cells and not the initiation of a specific fibrotic collagen gene expression program. Finally, in fibrotic muscle, we show that these collagen I-expressing cell populations differentially express distinct ECM proteins—fibroblasts express the fibrillar components of ECM, fibro/adipogenic progenitors cells differentially express basal laminar proteins, and skeletal muscle progenitor cells differentially express genes important for the satellite cell. PMID:27881411

Angiotensin II alters the expression of duodenal iron transporters, hepatic hepcidin, and body iron distribution in mice.

PubMed

Tajima, Soichiro; Ikeda, Yasumasa; Enomoto, Hideaki; Imao, Mizuki; Horinouchi, Yuya; Izawa-Ishizawa, Yuki; Kihira, Yoshitaka; Miyamoto, Licht; Ishizawa, Keisuke; Tsuchiya, Koichiro; Tamaki, Toshiaki

2015-08-01

Angiotensin II (ANG II) has been shown to affect iron metabolism through alteration of iron transporters, leading to increased cellular and tissue iron contents. Serum ferritin, a marker of body iron storage, is elevated in various cardiovascular diseases, including hypertension. However, the associated changes in iron absorption and the mechanism underlying increased iron content in a hypertensive state remain unclear. The C57BL6/J mice were treated with ANG II to generate a model of hypertension. Mice were divided into three groups: (1) control, (2) ANG II-treated, and (3) ANG II-treated and ANG II receptor blocker (ARB)-administered (ANG II-ARB) groups. Mice treated with ANG II showed increased serum ferritin levels compared to vehicle-treated control mice. In ANG II-treated mice, duodenal divalent metal transporter-1 and ferroportin (FPN) expression levels were increased and hepatic hepcidin mRNA expression and serum hepcidin concentration were reduced. The mRNA expression of bone morphogenetic protein 6 and CCAAT/enhancer-binding protein alpha, which are regulators of hepcidin, was also down-regulated in the livers of ANG II-treated mice. In terms of tissue iron content, macrophage iron content and renal iron content were increased by ANG II treatment, and these increases were associated with reduced expression of transferrin receptor 1 and FPN and increased expression of ferritin. These changes induced by ANG II treatment were ameliorated by the administration of an ARB. Angiotensin II (ANG II) altered the expression of duodenal iron transporters and reduced hepcidin levels, contributing to the alteration of body iron distribution.

A loss of Pdxk model of Parkinson disease in Drosophila can be suppressed by Buffy.

PubMed

M'Angale, P Githure; Staveley, Brian E

2017-06-12

The identification of a DNA variant in pyridoxal kinase (Pdxk) associated with increased risk to Parkinson disease (PD) gene led us to study the inhibition of this gene in the Dopa decarboxylase (Ddc)-expressing neurons of the well-studied model organism Drosophila melanogaster. The multitude of biological functions attributable to the vitamers catalysed by this kinase reveal an overabundance of possible links to PD, that include dopamine synthesis, antioxidant activity and mitochondrial function. Drosophila possesses a single homologue of Pdxk and we used RNA interference to inhibit the activity of this kinase in the Ddc-Gal4-expressing neurons. We further investigated any association between this enhanced disease risk gene with the established PD model induced by expression of α-synuclein in the same neurons. We relied on the pro-survival functions of Buffy, an anti-apoptotic Bcl-2 homologue, to rescue the Pdxk-induced phenotypes. To drive the expression of Pdxk RNA interference in DA neurons of Drosophila, we used Ddc-Gal4 which drives expression in both dopaminergic and serotonergic neurons, to result in decreased longevity and compromised climbing ability, phenotypes that are strongly associated with Drosophila models of PD. The inhibition of Pdxk in the α-synuclein-induced Drosophila model of PD did not alter longevity and climbing ability of these flies. It has been previously shown that deficiency in vitamers lead to mitochondrial dysfunction and neuronal decay, therefore, co-expression of Pdxk-RNAi with the sole pro-survival Bcl-2 homologue Buffy in the Ddc-Gal4-expressing neurons, resulted in increased survival and a restored climbing ability. In a similar manner, when we inhibited Pdxk in the developing eye using GMR-Gal4, we found that there was a decrease in the number of ommatidia and the disruption of the ommatidial array was more pronounced. When Pdxk was inhibited with the α-synuclein-induced developmental eye defects, the eye phenotypes were unaltered. Interestingly co-expression with Buffy restored ommatidia number and decreased the severity of disruption of the ommatidial array. Though Pdxk is not a confirmed Parkinson disease gene, the inhibition of this kinase recapitulated the PD-like symptoms of decreased lifespan and loss of locomotor function, possibly producing a new model of PD.

The Stimulation of Macrophages with TLR Ligands Supports Increased IL-19 Expression in Inflammatory Bowel Disease Patients and in Colitis Models.

PubMed

Steinert, Anna; Linas, Ioannis; Kaya, Berna; Ibrahim, Mohamed; Schlitzer, Andreas; Hruz, Petr; Radulovic, Katarina; Terracciano, Luigi; Macpherson, Andrew J; Niess, Jan Hendrik

2017-10-01

IL-19, a member of the IL-10 cytokine family that signals through the IL-20 receptor type I (IL-20Rα:IL-20Rβ), is a cytokine whose function is not completely known. In this article, we show that the expression of IL19 in biopsies of patients with active ulcerative colitis was increased compared with patients with quiescent ulcerative colitis and that colitis was attenuated in IL-19-deficient mice. The disruption of the epithelial barrier with dextran sodium sulfate leads to increased IL-19 expression. Attenuated colitis in IL-19-deficient animals was associated with reduced numbers of IL-6-producing macrophages in the inflamed colonic lamina propria. Microbial-driven expression of IL-19 by intestinal macrophages may contribute to the pathogenesis of inflammatory bowel disease. Copyright © 2017 by The American Association of Immunologists, Inc.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Dar, Roy; Shaffer, Sydney M.; Singh, Abhyudai

Recent analysis demonstrates that the HIV-1 Long Terminal Repeat (HIV LTR) promoter exhibits a range of possible transcriptional burst sizes and frequencies for any mean-expression level. However, these results have also been interpreted as demonstrating that cell-tocell expression variability (noise) and mean are uncorrelated, a significant deviation from previous results. Here, we re-examine the available mRNA and protein abundance data for the HIV LTR and find that noise in mRNA and protein expression scales inversely with the mean along analytically predicted transcriptional burst-size manifolds. We then experimentally perturb transcriptional activity to test a prediction of the multiple burst-size model: thatmore » increasing burst frequency will cause mRNA noise to decrease along given burst-size lines as mRNA levels increase. In conclusion, the data show that mRNA and protein noise decrease as mean expression increases, supporting the canonical inverse correlation between noise and mean.« less

Proteome map of Aspergillus nidulans during osmoadaptation.

PubMed

Kim, Yonghyun; Nandakumar, M P; Marten, Mark R

2007-09-01

The model filamentous fungus Aspergillus nidulans, when grown in a moderate level of osmolyte (+0.6M KCl), was previously found to have a significantly reduced cell wall elasticity (Biotech Prog, 21:292, 2005). In this study, comparative proteomic analysis via two-dimensional gel electrophoresis (2de) and matrix-assisted laser desorption ionization/time-of-flight (MALDI-TOF) mass spectrometry was used to assess molecular level events associated with this phenomenon. Thirty of 90 differentially expressed proteins were identified. Sequence homology and conserved domains were used to assign probable function to twenty-one proteins currently annotated as "hypothetical." In osmoadapted cells, there was an increased expression of glyceraldehyde-3-phosphate dehydrogenase and aldehyde dehydrogenase, as well as a decreased expression of enolase, suggesting an increased glycerol biosynthesis and decreased use of the TCA cycle. There also was an increased expression of heat shock proteins and Shp1-like protein degradation protein, implicating increased protein turnover. Five novel osmoadaptation proteins of unknown functions were also identified.

[The effects of electro-acupuncture on the signaling pathway of TLR/MYD88 in ankle joint synovial tissue of acute gouty arthritis rats].

PubMed

Zhang, Chao-nan; Huang, Xue-kuan; Luo, Yan; Jiang, Juan; Wan, Lei; Wang, Ling

2014-11-01

To investigate the effects of electro-acupuncture ( EA) on the related protein expression of the signaling pathway of the toll-like receptor2 (TLR2)/myeloid differentiation factor (MYD) 88 in ankle joint synovial tissue of acute gouty arthritis (AGA) rats. Fifty male SD rats were randomly divided into 5 groups: normal group, SMD group, AGA model group, medication group and EA group, 10 rats in each group. SMD group established model by inducing SMD, other groups established AGA model by inducing monosodium urate, except the normal group. Two days before model was established, normal and SMD and AGA model groups were lavaged with normal saline (20 mL/kg), medication group was lavaged with colchicine solution (1 mg/kg), EA (1. 5-2 Hz, D.-D. wave, 9 V, 1-3 mA) was applied to"Sanyinjiao" (SP6),"Jiexi"(ST41) and "kunlun" (BL60) for 20 min, once daily, continuously for 9 days. Then the join sewlling index was observed periodically, the protein expression of TLR2 and MYD88 was determined by immunohistochemistry. Compared to the normal group, the join sewlling of the SMD group in test join increased significantly (P<0. 05) and the protein expression of TLR2 and MYD88 in synovial tissue has not statistically significant (P>0.05), the oin sewlling and protein expression of TLR2 and MYD88 in synovial tissue of model group increased significantly P<0. 05); The medication and EA group compared to the model group, the protein expression of TLR2 and MYD88 in synovial tissue decreased significantly (P <0. 05), the join sewlling in test join decreased significantly P<1. 05); There were not statistically significant between the EA group and the medication group (P>0.05). EA can alleviate the symptoms of AGA, which may be related to regulation of the protein expression Y TRI and MYD88 in the TLR/MYD88 signaling pathway.

Increased expression of stress inducible protein 1 in glioma-associated microglia/macrophages.

PubMed

Carvalho da Fonseca, Anna Carolina; Wang, Huaqing; Fan, Haitao; Chen, Xuebo; Zhang, Ian; Zhang, Leying; Lima, Flavia Regina Souza; Badie, Behnam

2014-09-15

Factors released by glioma-associated microglia/macrophages (GAMs) play an important role in the growth and infiltration of tumors. We have previously demonstrated that the co-chaperone stress-inducible protein 1 (STI1) secreted by microglia promotes proliferation and migration of human glioblastoma (GBM) cell lines in vitro. In the present study, in order to investigate the role of STI1 in a physiological context, we used a glioma model to evaluate STI1 expression in vivo. Here, we demonstrate that STI1 expression in both the tumor and in the infiltrating GAMs and lymphocytes significantly increased with tumor progression. Interestingly, high expression of STI1 was observed in macrophages and lymphocytes that infiltrated brain tumors, whereas STI1 expression in the circulating blood monocytes and lymphocytes remained unchanged. Our results correlate, for the first time, the expression of STI1 and glioma progression, and suggest that STI1 expression in GAMs and infiltrating lymphocytes is modulated by the brain tumor microenvironment. Copyright © 2014 Elsevier B.V. All rights reserved.

Increased Expression of Stress Inducible Protein 1 in Glioma-Associated Microglia/Macrophages

PubMed Central

da Fonseca, Anna Carolina Carvalho; Wang, Huaqing; Fan, Haitao; Chen, Xuebo; Zhang, Ian; Zhang, Leying; Lima, Flavia Regina Souza; Badie, Behnam

2014-01-01

Factors released by glioma-associated microglia/macrophages (GAMs) play an important role in the growth and infiltration of tumors. We have previously demonstrated that the co-chaperone stress-inducible protein 1 (STI1) secreted by microglia promotes proliferation and migration of human glioblastoma (GBM) cell lines in vitro. In the present study, in order to investigate the role of STI1 in a physiological context, we used a glioma model to evaluate STI1 expression in vivo. Here, we demonstrate that STI1 expression in both the tumor and in the infiltrating GAMs and lymphocytes significantly increased with tumor progression. Interestingly, high expression of STI1 was observed in macrophages and lymphocytes that infiltrated brain tumors, whereas STI1 expression in the circulating blood monocytes and lymphocytes remained unchanged. Our results correlate, for the first time, the expression of STI1 and glioma progression, and suggest that STI1 expression in GAMs and infiltrating lymphocytes is modulated by the brain tumor microenvironment. PMID:25042352

EZH2 Modulates Angiogenesis In Vitro and in a Mouse Model of Limb Ischemia

PubMed Central

Mitić, Tijana; Caporali, Andrea; Floris, Ilaria; Meloni, Marco; Marchetti, Micol; Urrutia, Raul; Angelini, Gianni D; Emanueli, Costanza

2015-01-01

Epigenetic mechanisms may regulate the expression of pro-angiogenic genes, thus affecting reparative angiogenesis in ischemic limbs. The enhancer of zest homolog-2 (EZH2) induces thtrimethylation of lysine 27 on histone H3 (H3K27me3), which represses gene transcription. We explored (i) if EZH2 expression is regulated by hypoxia and ischemia; (ii) the impact of EZH2 on the expression of two pro-angiogenic genes: eNOS and BDNF; (iii) the functional effect of EZH2 inhibition on cultured endothelial cells (ECs); (iv) the therapeutic potential of EZH2 inhibition in a mouse model of limb ischemia (LI). EZH2 expression was increased in cultured ECs exposed to hypoxia (control: normoxia) and in ECs extracted from mouse ischemic limb muscles (control: absence of ischemia). EZH2 increased the H3K27me3 abundance onto regulatory regions of eNOS and BDNF promoters. In vitro RNA silencing or pharmacological inhibition by 3-deazaneplanocin (DZNep) of EZH2 increased eNOS and BDNF mRNA and protein levels and enhanced functional capacities (migration, angiogenesis) of ECs under either normoxia or hypoxia. In mice with experimentally induced LI, DZNep increased angiogenesis in ischaemic muscles, the circulating levels of pro-angiogenic hematopoietic cells and blood flow recovery. Targeting EZH2 for inhibition may open new therapeutic avenues for patients with limb ischemia. PMID:25189741

Icariin protects rats against 5/6 nephrectomy-induced chronic kidney failure by increasing the number of renal stem cells.

PubMed

Huang, Zhongdi; He, Liqun; Huang, Di; Lei, Shi; Gao, Jiandong

2015-10-21

Chronic kidney disease poses a serious health problem worldwide with increasing prevalence and lack of effective treatment. This study aimed to investigate the mechanism of icariin in alleviating chronic renal failure induced by 5/6 nephrectomy in rats. The chronic renal failure model was established by a two-phased 5/6 nephrectomy procedure. The model rats were given daily doses of water or icariin for 8 weeks. The kidney morphology was checked by HE staining. The levels of blood urea nitrogen, serum creatinine, and serum uric acid were measured by colometric methods. The expression of specified genes was analyzed by quantitative real-time PCR and immunohistochemical staining. The number of renal stem/progenitor cells was analyzed by CD133 and CD24 immunohistochemical staining. Icariin protected against CDK-caused damages to kidney histology and improved renal function, significantly reduced levels of BUN, creatinine, and uric acid. Icariin inhibited the expression level of TGF-β1 whereas upregulated HGF, BMP-7, WT-1, and Pax2 expression. Moreover, ccariin significantly increased the expression of CD24, CD133, Osr1, and Nanog in remnant kidney and the numbers of CD133(+)/CD24(+) renal stem/progenitor cells. These data demonstrated that icariin effectively alleviated 5/6 nephrectomy induced chronic renal failure through increasing renal stem/progenitor cells.

Substance P as an immunomodulatory neuropeptide in a mouse model for autoimmune hair loss (alopecia areata).

PubMed

Siebenhaar, Frank; Sharov, Andrey A; Peters, Eva M J; Sharova, Tatyana Y; Syska, Wolfgang; Mardaryev, Andrei N; Freyschmidt-Paul, Pia; Sundberg, John P; Maurer, Marcus; Botchkarev, Vladimir A

2007-06-01

Alopecia areata (AA) is an autoimmune disorder of the hair follicle characterized by inflammatory cell infiltrates around actively growing (anagen) hair follicles. Substance P (SP) plays a critical role in the cutaneous neuroimmune network and influences immune cell functions through the neurokinin-1 receptor (NK-1R). To better understand the role of SP as an immunomodulatory neuropeptide in AA, we studied its expression and effects on immune cells in a C3H/HeJ mouse model for AA. During early stages of AA development, the number of SP-immunoreactive nerve fibers in skin is increased, compared to non-affected mice. However, during advanced stages of AA, the number of SP-immunoreactive nerves and SP protein levels in skin are decreased, whereas the expression of the SP-degrading enzyme neutral endopeptidase (NEP) is increased, compared to control skin. In AA, NK-1R is expressed on CD8+ lymphocytes and macrophages accumulating around affected hair follicles. Additional SP supply to the skin of AA-affected mice leads to a significant increase of mast cell degranulation and to accelerated hair follicle regression (catagen), accompanied by an increase of CD8+ cells-expressing granzyme B. These data suggest that SP, NEP, and NK-1R serve as important regulators in the molecular signaling network modulating inflammatory response in autoimmune hair loss.

Proximal Tubules Have the Capacity to Regulate Uptake of Albumin.

PubMed

Wagner, Mark C; Campos-Bilderback, Silvia B; Chowdhury, Mahboob; Flores, Brittany; Lai, Xianyin; Myslinski, Jered; Pandit, Sweekar; Sandoval, Ruben M; Wean, Sarah E; Wei, Yuan; Satlin, Lisa M; Wiggins, Roger C; Witzmann, Frank A; Molitoris, Bruce A

2016-02-01

Evidence from multiple studies supports the concept that both glomerular filtration and proximal tubule (PT) reclamation affect urinary albumin excretion rate. To better understand these roles of glomerular filtration and PT uptake, we investigated these processes in two distinct animal models. In a rat model of acute exogenous albumin overload, we quantified glomerular sieving coefficients (GSC) and PT uptake of Texas Red-labeled rat serum albumin using two-photon intravital microscopy. No change in GSC was observed, but a significant decrease in PT albumin uptake was quantified. In a second model, loss of endogenous albumin was induced in rats by podocyte-specific transgenic expression of diphtheria toxin receptor. In these albumin-deficient rats, exposure to diphtheria toxin induced an increase in albumin GSC and albumin filtration, resulting in increased exposure of the PTs to endogenous albumin. In this case, PT albumin reabsorption was markedly increased. Analysis of known albumin receptors and assessment of cortical protein expression in the albumin overload model, conducted to identify potential proteins and pathways affected by acute protein overload, revealed changes in the expression levels of calreticulin, disabled homolog 2, NRF2, angiopoietin-2, and proteins involved in ATP synthesis. Taken together, these results suggest that a regulated PT cell albumin uptake system can respond rapidly to different physiologic conditions to minimize alterations in serum albumin level. Copyright © 2016 by the American Society of Nephrology.

Proximal Tubules Have the Capacity to Regulate Uptake of Albumin

PubMed Central

Wagner, Mark C.; Campos-Bilderback, Silvia B.; Chowdhury, Mahboob; Flores, Brittany; Lai, Xianyin; Myslinski, Jered; Pandit, Sweekar; Sandoval, Ruben M.; Wean, Sarah E.; Wei, Yuan; Satlin, Lisa M.; Wiggins, Roger C.; Witzmann, Frank A.

2016-01-01

Evidence from multiple studies supports the concept that both glomerular filtration and proximal tubule (PT) reclamation affect urinary albumin excretion rate. To better understand these roles of glomerular filtration and PT uptake, we investigated these processes in two distinct animal models. In a rat model of acute exogenous albumin overload, we quantified glomerular sieving coefficients (GSC) and PT uptake of Texas Red-labeled rat serum albumin using two-photon intravital microscopy. No change in GSC was observed, but a significant decrease in PT albumin uptake was quantified. In a second model, loss of endogenous albumin was induced in rats by podocyte-specific transgenic expression of diphtheria toxin receptor. In these albumin-deficient rats, exposure to diphtheria toxin induced an increase in albumin GSC and albumin filtration, resulting in increased exposure of the PTs to endogenous albumin. In this case, PT albumin reabsorption was markedly increased. Analysis of known albumin receptors and assessment of cortical protein expression in the albumin overload model, conducted to identify potential proteins and pathways affected by acute protein overload, revealed changes in the expression levels of calreticulin, disabled homolog 2, NRF2, angiopoietin-2, and proteins involved in ATP synthesis. Taken together, these results suggest that a regulated PT cell albumin uptake system can respond rapidly to different physiologic conditions to minimize alterations in serum albumin level. PMID:26054544

Elicited soybean (Glycine max) extract effect on improving levels of Ter-119+Cd59+ in a mouse model fed a high fat-fructose diet

NASA Astrophysics Data System (ADS)

Safitri, Yunita Diyah; Widyarti, Sri; Rifa'i, Muhaimin

2017-05-01

People who have unbalanced lifestyles and habits such as consuming high fat and sugar foods, as well as the lack of physical activity, have an increased risk of obesity and related metabolic diseases. The condition of obesity occurs due to an excess of nutrients which leads to low-grade inflammation. Inflammation induced by obesity causes unstable bone marrow homeostasis which is associated with proliferation and differentiation of Hematopoietic Stem Cells (HSCs). This study aimed to observe the erythroid progenitor (TER-119) and complement regulator (CD59) on bone marrow cells in mouse models fed a high fat-fructose diet (HFFD). This research was conducted by modeling obese mice using high fat and fructose food for 20 weeks, and then treating them with elicited soybean extract (ESE) for four weeks with several doses: low dose (78 mg/kgBB), moderate dose (104 mg/kgBB) and high dose (130 mg/kgBB). Cell TER119+CD59+ expression decreased in the HFFD group compared to the normal group. In the low, moderate and high dose group, TER119+CD59+ expression significantly increased compared to the HFFD group. These results demonstrate that soybean elicited extract can improve the hematopoietic system by increasing TER119+CD59+ expression in a high fat and fructose diet mouse model.

The Circular RNA Interacts with STAT3, Increasing Its Nuclear Translocation and Wound Repair by Modulating Dnmt3a and miR-17 Function.

PubMed

Yang, Zhen-Guo; Awan, Faryal Mehwish; Du, William W; Zeng, Yan; Lyu, Juanjuan; Wu, De; Gupta, Shaan; Yang, Weining; Yang, Burton B

2017-09-06

Delayed or impaired wound healing is a major health issue worldwide, especially in patients with diabetes and atherosclerosis. Here we show that expression of the circular RNA circ-Amotl1 accelerated healing process in a mouse excisional wound model. Further studies showed that ectopic circ-Amotl1 increased protein levels of Stat3 and Dnmt3a. The increased Dnmt3a then methylated the promoter of microRNA miR-17, decreasing miR-17-5p levels but increasing fibronectin expression. We found that Stat3, similar to Dnmt3a and fibronectin, was a target of miR-17-5p. Decreased miR-17-5p levels would increase expression of fibronectin, Dnmt3a, and Stat3. All of these led to increased cell adhesion, migration, proliferation, survival, and wound repair. Furthermore, we found that circ-Amotl1 not only increased Stat3 expression but also facilitated Stat3 nuclear translocation. Thus, the ectopic expressed circ-Amotl1 and Stat3 were mainly translocated to nucleus. In the presence of circ-Amotl1, Stat3 interacted with Dnmt3a promoter with increased affinity, facilitating Dnmt3a transcription. Ectopic application of circ-Amotl1 accelerating wound repair may shed light on skin wound healing clinically. Copyright © 2017. Published by Elsevier Inc.

Immunomodulatory Effects of Deokgu Thermomineral Water Balneotherapy on Oxazolone-Induced Atopic Dermatitis Murine Model

PubMed Central

Lee, Young Bok; Kim, Su Jin; Park, Sae Mi; Lee, Kyung Ho; Han, Hyung Jin; Yu, Dong Soo; Woo, So Youn; Yun, Seong Taek; Hamm, Se-Yeong; Kim, Hong Jig

2016-01-01

Background Although the therapeutic mechanism of balneotherapy for atopic dermatitis has not been clarified, many atopic patients who visit thermomineral springs have shown clinical improvements. Objective This study was aimed to evaluate the immunomodulatory effect of thermomineral water balneotherapy on the atopic dermatitis murine model. Methods The oxazolone-induced atopic dermatitis murine model was used to evaluate the therapeutic effect of balneotherapy with Deokgu thermomineral water compared with distilled water. Histologic evaluation and confocal microscopic imaging were performed to analyze the lesional expression of cluster-of-differentiation (CD)4 and forkhead box p3 (Foxp3). Lesional mRNA expression of interleukin (IL) 33, thymic stromal lymphopoietin (TSLP), and Foxp3 was evaluated by real-time reverse transcription polymerase chain reaction. Results Compared with the distilled water bath group, confocal microscopic evaluation of CD4 and Foxp3 merged images showed increased expression of regulatory T cells in the thermomineral balneotherapy group. The lesional mRNA level of IL-33 showed a reduced trend in the thermomineral balneotherapy group, whereas the level of mRNA of Foxp3 was increased. TSLP showed a decreased trend in both distilled water and thermomineral water bath groups. There was a trend of reduced expression in lesional IL-33 mRNA but increased cell count of CD4+ Foxp3+ regulatory T cells in thermomineral balneotherapy compared with distilled water bath. Conclusion Therefore, thermomineral balneotherapy can be an effective and safe adjuvant therapeutic option for atopic dermatitis. PMID:27081266

Immunomodulatory Effects of Deokgu Thermomineral Water Balneotherapy on Oxazolone-Induced Atopic Dermatitis Murine Model.

PubMed

Lee, Young Bok; Kim, Su Jin; Park, Sae Mi; Lee, Kyung Ho; Han, Hyung Jin; Yu, Dong Soo; Woo, So Youn; Yun, Seong Taek; Hamm, Se-Yeong; Kim, Hong Jig; Kim, Jin-Wou

2016-04-01

Although the therapeutic mechanism of balneotherapy for atopic dermatitis has not been clarified, many atopic patients who visit thermomineral springs have shown clinical improvements. This study was aimed to evaluate the immunomodulatory effect of thermomineral water balneotherapy on the atopic dermatitis murine model. The oxazolone-induced atopic dermatitis murine model was used to evaluate the therapeutic effect of balneotherapy with Deokgu thermomineral water compared with distilled water. Histologic evaluation and confocal microscopic imaging were performed to analyze the lesional expression of cluster-of-differentiation (CD)4 and forkhead box p3 (Foxp3). Lesional mRNA expression of interleukin (IL) 33, thymic stromal lymphopoietin (TSLP), and Foxp3 was evaluated by real-time reverse transcription polymerase chain reaction. Compared with the distilled water bath group, confocal microscopic evaluation of CD4 and Foxp3 merged images showed increased expression of regulatory T cells in the thermomineral balneotherapy group. The lesional mRNA level of IL-33 showed a reduced trend in the thermomineral balneotherapy group, whereas the level of mRNA of Foxp3 was increased. TSLP showed a decreased trend in both distilled water and thermomineral water bath groups. There was a trend of reduced expression in lesional IL-33 mRNA but increased cell count of CD4(+) Foxp3(+) regulatory T cells in thermomineral balneotherapy compared with distilled water bath. Therefore, thermomineral balneotherapy can be an effective and safe adjuvant therapeutic option for atopic dermatitis.

Abnormal platelet-derived growth factor signaling accounting for lung hypoplasia in experimental congenital diaphragmatic hernia.

PubMed

Dingemann, Jens; Doi, Takashi; Ruttenstock, Elke; Puri, Prem

2010-10-01

The pathogenesis of pulmonary hypoplasia in congenital diaphragmatic hernia (CDH) is not fully understood. Platelet-derived growth factor A (PDGFA) and platelet-derived growth factor receptor α (PDGFRα) play a crucial role in lung development. It has been reported that PDGF induces H(2)O(2)-production and that oxidative stress may be an important mechanism for the impaired lung development in the nitrofen rat model. We hypothesized that pulmonary expression of PDGFA and PDGFRα is altered in the nitrofen induced CDH model. Pregnant rats received 100 mg nitrofen or vehicle on gestational day 9 (D9) and were sacrificed on D15, D18 or D21. RNA was extracted from fetal left lungs and mRNA levels of PDGFA and PDGFRα were determined using real-time polymerase chain reaction. Immunohistochemistry for protein expression of PDGFA and PDGFRα was performed. Pulmonary H(2)O(2) was measured colorimetrically. mRNA levels of PDGFRα at D15 (4.50 ± 0.87) and PDGFA at D18 (2.90 ± 1.38) were increased in the nitrofen group (P < .05). Immunohistochemistry revealed increased pulmonary expression of PDGFRα and PDGFA. H(2)O(2) content was significantly higher in the nitrofen group. Increased expression of PDGFA and PDGFRα suggests that pulmonary hypoplasia in the nitrofen CDH model may be owing to PDGF-induced oxidative stress during lung development. Copyright © 2010 Elsevier Inc. All rights reserved.

Gas6 Deficiency Increases Oligodendrocyte Loss and Microglial Activation in Response to Cuprizone-Induced Demyelination

PubMed Central

Binder, Michele D.; Cate, Holly S.; Prieto, Anne L.; Kemper, Dennis; Butzkueven, Helmut; Gresle, Melissa M.; Cipriani, Tania; Jokubaitis, Vilija G.; Carmeliet, Peter

2008-01-01

The TAM family of receptor protein tyrosine kinases comprises three known members, namely Tyro3, Axl, and Mer. These receptors are widely expressed in the nervous system, including by oligodendrocytes, the cell type responsible for myelinating the CNS. We examined the potential role of the TAM family and of their principle cognate ligand, Gas6 (growth arrest gene 6), in modulating the phenotype of the cuprizone model of demyelination. We found that the expression profiles of Axl, Mer, and Gas6 mRNA were increased in the corpus callosum in a temporal profile correlating with the increased migration and proliferation of microglia/macrophages in this model. In contrast, expression of Tyro3 decreased, correlating with the loss of oligodendrocytes. Gas6 both promoted in vitro survival of oligodendrocytes (39.3 ± 3.1 vs 11.8 ± 2.4%) and modulated markers of activation in purified cultures of microglia (tumor necrosis factor α mRNA expression was reduced ∼48%). In Gas6−/− mice subjected to cuprizone-challenge, demyelination was greater than in control mice, within the rostral region of the corpus callosum, as assessed by luxol fast blue staining (myelination reduced by 36%) and by ultrastructural analysis. An increased loss of Gst-π (glutathione S-transferase-π)-positive oligodendrocytes was also identified throughout the corpus callosum of Gas6−/− mice. Microglial marker expression (ionized calcium-binding adapter molecule 1) was increased in Gas6−/− mice but was restricted to the rostral corpus callosum. Therefore, TAM receptor activation and regulation can independently influence both oligodendrocyte survival and the microglial response after CNS damage. PMID:18480276

Immunohistochemical Characterization of Connexin43 Expression in a Mouse Model of Diabetic Retinopathy and in Human Donor Retinas

PubMed Central

Mugisho, Odunayo O.; Green, Colin R.; Zhang, Jie; Binz, Nicolette; Acosta, Monica L.; Rakoczy, Elizabeth

2017-01-01

Diabetic retinopathy (DR) develops due to hyperglycemia and inflammation-induced vascular disruptions in the retina with connexin43 expression patterns in the disease still debated. Here, the effects of hyperglycemia and inflammation on connexin43 expression in vitro in a mouse model of DR and in human donor tissues were evaluated. Primary human retinal microvascular endothelial cells (hRMECs) were exposed to high glucose (HG; 25 mM) or pro-inflammatory cytokines IL-1β and TNF-α (10 ng/mL each) or both before assessing connexin43 expression. Additionally, connexin43, glial fibrillary acidic protein (GFAP), and plasmalemma vesicular associated protein (PLVAP) were labeled in wild-type (C57BL/6), Akita (diabetic), and Akimba (DR) mouse retinas. Finally, connexin43 and GFAP expression in donor retinas with confirmed DR was compared to age-matched controls. Co-application of HG and cytokines increased connexin43 expression in hRMECs in line with results seen in mice, with no significant difference in connexin43 or GFAP expression in Akita but higher expression in Akimba compared to wild-type mice. On PLVAP-positive vessels, connexin43 was higher in Akimba but unchanged in Akita compared to wild-type mice. Connexin43 expression appeared higher in donor retinas with confirmed DR compared to age-matched controls, similar to the distribution seen in Akimba mice and correlating with the in vitro results. Although connexin43 expression seems reduced in diabetes, hyperglycemia and inflammation present in the pathology of DR seem to increase connexin43 expression, suggesting a causal role of connexin43 channels in the disease progression. PMID:29186067

Osteoarticular Expression of Musashi-1 in an Experimental Model of Arthritis

PubMed Central

O'Valle, Francisco; Peregrina, Magdalena; Crespo-Lora, Vicente; Galindo-Moreno, Pablo; Roman, Maria; Aneiros-Fernandez, Jose; Aguilar, David; Gonzalez-Rey, Elena; Delgado, Mario; Hernandez-Cortes, Pedro

2015-01-01

Background. Collagen-induced arthritis (CIA), a murine experimental disease model induced by immunization with type II collagen (CII), is used to evaluate novel therapeutic strategies for rheumatoid arthritis. Adult stem cell marker Musashi-1 (Msi1) plays an important role in regulating the maintenance and differentiation of stem/precursor cells. The objectives of this investigation were to perform a morphological study of the experimental CIA model, evaluate the effect of TNFα-blocker (etanercept) treatment, and determine the immunohistochemical expression of Msi1 protein. Methods. CIA was induced in 50 male DBA1/J mice for analyses of tissue and serum cytokine; clinical and morphological lesions in limbs; and immunohistochemical expression of Msi1. Results. Clinically, TNFα-blocker treatment attenuated CIA on day 32 after immunization (P < 0.001). Msi1 protein expression was significantly higher in joints damaged by CIA than in those with no lesions (P < 0.0001) and was related to the severity of the lesions (Spearman's rho = 0.775, P = 0.0001). Conclusions. Treatment with etanercept attenuates osteoarticular lesions in the murine CIA model. Osteoarticular expression of Msi1 protein is increased in joints with CIA-induced lesion and absent in nonlesioned joints, suggesting that this protein is expressed when the lesion is produced in order to favor tissue repair. PMID:26064941

Endosperm-specific co-expression of recombinant soybean ferritin and Aspergillus phytase in maize results in significant increases in the levels of bioavailable iron.

PubMed

Drakakaki, Georgia; Marcel, Sylvain; Glahn, Raymond P; Lund, Elizabeth K; Pariagh, Sandra; Fischer, Rainer; Christou, Paul; Stoger, Eva

2005-12-01

We have generated transgenic maize plants expressing Aspergillus phytase either alone or in combination with the iron-binding protein ferritin. Our aim was to produce grains with increased amounts of bioavailable iron in the endosperm. Maize seeds expressing recombinant phytase showed enzymatic activities of up to 3 IU per gram of seed. In flour paste prepared from these seeds, up to 95% of the endogenous phytic acid was degraded, with a concomitant increase in the amount of available phosphate. In seeds expressing ferritin in addition to phytase, the total iron content was significantly increased. To evaluate the impact of the recombinant proteins on iron absorption in the human gut, we used an in vitro digestion/Caco-2 cell model. We found that phytase in the maize seeds was associated with increased cellular iron uptake, and that the rate of iron uptake correlated with the level of phytase expression regardless of the total iron content of the seeds. We also investigated iron bioavailability under more complex meal conditions by adding ascorbic acid, which promotes iron uptake, to all samples. This resulted in a further increase in iron absorption, but the effects of phytase and ascorbic acid were not additive. We conclude that the expression of recombinant ferritin and phytase could help to increase iron availability and enhance the absorption of iron, particularly in cereal-based diets that lack other nutritional components.

Deletion of the Inflammasome Sensor Aim2 Mitigates Aβ Deposition and Microglial Activation but Increases Inflammatory Cytokine Expression in an Alzheimer Disease Mouse Model.

PubMed

Wu, Pei-Jung; Hung, Yun-Fen; Liu, Hsin-Yu; Hsueh, Yi-Ping

2017-01-01

Inflammation is clearly associated with Alzheimer disease (AD). Knockout of Nlrp3, a gene encoding an inflammasome sensor, has been shown to ameliorate AD pathology in a mouse model. Because AIM2 is the most dominant inflammasome sensor expressed in mouse brains, here we investigate whether Aim2 deletion also influences the phenotype of a 5XFAD AD mouse model. Quantitative RT-PCR, immunostaining, immunoblotting, and behavioral analyses were applied to compare wild-type, Aim2-/-, 5XFAD, and Aim2-/-;5XFAD mice. We found that Aim2 knockout mitigates Aβ deposition in the cerebral cortex and hippocampus of 5XFAD mice. The activation of microglial cells is also reduced in Aim2-/-;5XFAD brains compared with 5XFAD brains. However, Aim2 knockout does not improve memory and anxiety phenotypes of 5XFAD mice in an open field, cued Y-maze, or Barnes maze. Compared with 5XFAD mice, Il-1 expression levels are not reduced in Aim2-/-;5XFAD mice. Unexpectedly, Il-6 and Il-18 expression levels in 5XFAD brains were further increased when Aim2 was deleted. Thus, inflammatory cytokine expression in 5XFAD brains is upregulated by Aim2 deletion through an unknown mechanism. Although Aim2 knockout mitigates Aβ deposition and microglial activation, Aim2 deletion does not have a beneficial effect on the spatial memory or cytokine expression of 5XFAD mice. Our findings suggest that Aβ aggregation and microglial activation may not always be correlated with the expression of inflammatory cytokines or cognitive function of 5XFAD mice. Our study also implies that different inflammasomes likely perform distinct roles in different physiological and/or pathological events. © 2017 S. Karger AG, Basel.

SENP1 attenuates the liver fibrosis through down-regulating the expression of SMAD2.

PubMed

Wu, Linshi; Qiu, Weiqing; Sun, Jianhua; Wang, Jian

2018-01-01

To investigate whether SENP1 could play a regulating role in the liver fibrosis process, the Sprague-Dawley (SD) rats were used to establish the liver fibrosis rat models by intraperitoneally injecting with 1 ml/kg of 10% CCl 4 , while the control normal rats were injected with olive oil. Then confirmation experiments to verify the successful establishment of these models were conducted by detecting the cellular and lobular architecture, and liver function indexes using hematoxylin-eosin staining, Masson's trichrome staining and microplate method, respectively. In addition, the expression levels of fibrosis markers including collagen I, collagen III, α-SMA and TGF-β1 were inspected using quantitative real-time PCR (qRT-PCR), as well as SMAD2. Subsequently, the relative mRNA and protein level of SENP1 was also determined via qRT-PCR and western blot analysis. Next, the HSC-T6 cells of SENP1 knock-down were constructed and used to test the relative protein expression levels of α-SMA and SMAD2 in these cells. The results of hematoxylin-eosin staining, Masson's trichrome staining and microplate method turned out that the rat liver fibrosis models were constructed successfully, which was further confirmed by the increased expression of collagen I, collagen III, α-SMA and TGF-β1 in mRNA and protein level, as well as SMAD2. Then the expression of SENP1 was overexpressed in the rat liver fibrosis models induced by CCl 4 and the TGF-β1 treatment could increase the protein expression level of collagen I, collagen III and α-SMA. Lastly, the SENP1 knockdown HSC-T6 cells were successfully constructed, while the silence of SENP1 down-regulated the protein expression of α-SMA and SMAD2. In conclusion, this study provided a new regulation mechanism about the liver fibrosis process. Copyright © 2017 Elsevier Inc. All rights reserved.

Preventing mutant huntingtin proteolysis and intermittent fasting promote autophagy in models of Huntington disease.

PubMed

Ehrnhoefer, Dagmar E; Martin, Dale D O; Schmidt, Mandi E; Qiu, Xiaofan; Ladha, Safia; Caron, Nicholas S; Skotte, Niels H; Nguyen, Yen T N; Vaid, Kuljeet; Southwell, Amber L; Engemann, Sabine; Franciosi, Sonia; Hayden, Michael R

2018-03-06

Huntington disease (HD) is caused by the expression of mutant huntingtin (mHTT) bearing a polyglutamine expansion. In HD, mHTT accumulation is accompanied by a dysfunction in basal autophagy, which manifests as specific defects in cargo loading during selective autophagy. Here we show that the expression of mHTT resistant to proteolysis at the caspase cleavage site D586 (C6R mHTT) increases autophagy, which may be due to its increased binding to the autophagy adapter p62. This is accompanied by faster degradation of C6R mHTT in vitro and a lack of mHTT accumulation the C6R mouse model with age. These findings may explain the previously observed neuroprotective properties of C6R mHTT. As the C6R mutation cannot be easily translated into a therapeutic approach, we show that a scheduled feeding paradigm is sufficient to lower mHTT levels in YAC128 mice expressing cleavable mHTT. This is consistent with a previous model, where the presence of cleavable mHTT impairs basal autophagy, while fasting-induced autophagy remains functional. In HD, mHTT clearance and autophagy may become increasingly impaired as a function of age and disease stage, because of gradually increased activity of mHTT-processing enzymes. Our findings imply that mHTT clearance could be enhanced by a regulated dietary schedule that promotes autophagy.

Effects of BDNF-Transfected BMSCs on Neural Functional Recovery and Synaptophysin Expression in Rats with Cerebral Infarction.

PubMed

Zhang, Yongming; Qiu, Binghui; Wang, Jinbiao; Yao, Yi; Wang, Chunlin; Liu, Jiachuan

2017-07-01

The purpose of this study was to investigate the effects of brain-derived neurotrophic factor (BDNF)-transfected bone marrow mesenchymal stem cells (BMSCs) on neural functional recovery and synaptophysin expression in rats with cerebral infarction (CI). A total of 120 healthy Sprague Dawley rats were randomly divided into sham group, control group, and model group. Craniotomy was conducted and neurological function defect scoring was used to verify the model. BDNF containing recombinant plasmid was transfected into rat BMSCs, which was verified by flow cytometry and Western Blot. After injection of the transfected BMSCs, neural functional recovery of the CI rats and synaptophysin expression were measured. After the CI rat model was established, magnetic resonance (MR) imaging, 2, 3, 5- triphenyl tetrazolium chloride (TTC) staining, and the neurological function defect scoring determined the success of the model. CD34 (-), CD45 (-), CD29 (+), and CD90 (+) cells detected showed that the obtained BMSCs have high purity. BDNF protein was highly expressed in the BMSCs successfully transfected with the recombinant plasmid. Balance beam walking score, rotating bar walking score, and screen test score were significantly lower, while synaptophysin expression was higher in the BDNF model group than those in the non-BDNF model group and sham group with time extension. BDNF can increase synaptic plasticity and neurogenesis and have a promotional role in neural functional recovery and synaptophysin expression in rats with CI. BDNF-transfected BMSCs may therefore have better treatment efficacy for CI clinically.

Effects of cevimeline on the immunolocalization of aquaporin-5 and the ultrastructure of salivary glands in Sjögren's syndrome model mice.

PubMed

Nishimura, Hidehiko; Yakeishi, Akira; Saga, Tsuyoshi; Yamaki, Koh-Ichi

2009-01-01

Sjögren's syndrome (SS) is an autoimmune disorder whose main symptoms include xerostomia and dry eyes. It has been demonstrated that abnormal expression of aquaporin (AQP)-5 in the parotid and submandibular glands in SS model mice was corrected by cevimeline. In the present study, we orally administered cevimeline hydrochloride (cevimeline) to female MRL/l mice, which are widely used as a model for SS, to immunohistochemically investigate the localization of AQP-5 in the salivary glands. We also assessed the ultrastructure of acinar cells in the submandibular glands. AQP-5 was expressed in the apical and lateral cell membranes of acinar cells in the parotid and submandibular glands of normal mice, but not in the sublingual glands. In contrast, AQP-5 was expressed not only in the cell membranes in the apical domains but also in the cytoplasm in the SS model mice, indicating that the localization of AQP-5 was disordered in the SS model mice. After administration of cevimeline, AQP-5 was predominantly localized in the cellular apical domains of the acinar cells. Electron microscopy revealed that administration of cevimeline to the SS model mice and normal mice markedly reduced the number of secretory granules, increased the area of the rough endoplasmic reticulum, and expanded the intercellular gaps in the cells of the submandibular acini. Condensed vacuoles were also observed in the Golgi apparatuses, indicating that secondary enhancement of secretion and production of saliva had occurred. Consequently, the results of the present study demonstrate that the administration of cevimeline to the SS model mice increased salivary secretion in the submandibular glands. Furthermore, cevimeline transiently normalized the localization of AQP-5 expression in the parotid and submandibular glands.

Assessment of brain metabolite correlates of adeno-associated virus-mediated over-expression of human alpha-synuclein in cortical neurons by in vivo (1) H-MR spectroscopy at 9.4 T.

PubMed

Cuellar-Baena, Sandra; Landeck, Natalie; Sonnay, Sarah; Buck, Kerstin; Mlynarik, Vladimir; In 't Zandt, René; Kirik, Deniz

2016-06-01

In this study, we used proton-localized spectroscopy ((1) H-MRS) for the acquisition of the neurochemical profile longitudinally in a novel rat model of human wild-type alpha-synuclein (α-syn) over-expression. Our goal was to find out if the increased α-syn load in this model could be linked to changes in metabolites in the frontal cortex. Animals injected with AAV vectors encoding for human α-syn formed the experimental group, whereas green fluorescent protein expressing animals were used as the vector-treated control group and a third group of uninjected animals were used as naïve controls. Data were acquired at 2, 4, and 8 month time points. Nineteen metabolites were quantified in the MR spectra using LCModel software. On the basis of 92 spectra, we evaluated any potential gender effect and found that lactate (Lac) levels were lower in males compared to females, while the opposite was observed for ascorbate (Asc). Next, we assessed the effect of age and found increased levels of GABA, Tau, and GPC+PCho. Finally, we analyzed the effect of treatment and found that Lac levels (p = 0.005) were specifically lower in the α-syn group compared to the green fluorescent protein and control groups. In addition, Asc levels (p = 0.05) were increased in the vector-injected groups, whereas glucose levels remained unchanged. This study indicates that the metabolic switch between glucose-lactate could be detectable in vivo and might be modulated by Asc. No concomitant changes were found in markers of neuronal integrity (e.g., N-acetylaspartate) consistent with the fact that α-syn over-expression in cortical neurons did not result in neurodegeneration in this model. We acquired the neurochemical profile longitudinally in a rat model of human wild-type alpha-synuclein (α-syn) over-expression in cortical neurons. We found that Lactate levels were reduced in the α-syn group compared to the control groups and Ascorbate levels were increased in the vector-injected groups. No changes were found in markers of neuronal integrity consistent with the fact that α-syn over-expression did not result in frank neurodegeneration. © 2016 International Society for Neurochemistry.

Effects of high-pressure oxygen therapy on brain tissue water content and AQP4 expression in rabbits with cerebral hemorrhage.

PubMed

Wu, Jing; Chen, Jiong; Guo, Hua; Peng, Fang

2014-12-01

To investigate the effects of different atmosphere absolutes (ATA) of high-pressure oxygen (HPO) on brain tissue water content and Aquaporin-4 (AQP4) expression in rabbits with cerebral hemorrhage. 180 New Zealand white rabbits were selected and randomly divided into normal group (n = 30), control group (n = 30) and cerebral hemorrhage group (n = 120), and cerebral hemorrhage group was divided into group A, B, C and D with 30 rabbits in each group. The groups received 1.0, 1.8, 2.0 and 2.2 ATA of HPO treatments, respectively. Ten rabbits in each group were killed at first, third and fifth day to detect the brain tissue water content and change of AQP4 expression. In cerebral hemorrhage group, brain tissue water content and AQP4 expression after model establishment were first increased, then decreased and reached the maximum on third day (p < 0.05). Brain tissue water content and AQP4 expression in control group and cerebral hemorrhage group were significantly higher than normal group at different time points (p < 0.05). In contrast, brain tissue water content and AQP4 expression in group C were significantly lower than in group A, group B, group D and control group (p < 0.05). In control group, AQP4-positive cells significantly increased after model establishment, which reached maximum on third day, and positive cells in group C were significantly less than in group A, group B and group D. We also found that AQP4 expression were positively correlated with brain tissue water content (r = 0.719, p < 0.05) demonstrated by significantly increased AQP4 expression along with increased brain tissue water content. In conclusion, HPO can decrease AQP4 expression in brain tissue of rabbits with cerebral hemorrhage to suppress the progression of brain edema and promote repairing of injured tissue. 2.0 ATA HPO exerts best effects, which provides an experimental basis for ATA selection of HPO in treating cerebral hemorrhage.

Changes in beta-actin mRNA expression in remodeling canine myocardium.

PubMed

Carlyle, W C; Toher, C A; Vandervelde, J R; McDonald, K M; Homans, D C; Cohn, J N

1996-01-01

Beta-actin, a cytoskeletal protein important in the maintenance of cytoarchitecture, has long been thought to be expressed constitutively in myocardial tissue. As such, beta-actin mRNA has been used as a control gene in a wide range of experiments. However, we have uncovered consistent changes in beta-actin mRNA expression in canine myocardium remodeling as a result of insult to the left ventricle. The experimental canine models used were either DC shock damage to the left ventricle or volume overload resulting from severe mitral regurgitation. The remodeling process in both canine models is characterized by an increase in left ventricular mass. PCR amplification using primers designed to selectively amplify the 3' end and a portion of the 3' untranslated region of beta-actin mRNA resulted in the generation of a 297 base pair product predominant only in normal canine myocardium and a 472 base pair product that became increasingly prominent from 1 to 30 days after DC shock damage to the left ventricle and from 10 to 90 days after creation of mitral regurgitation. Northern analysis showed a three-fold increase in beta-actin mRNA after either DC shock or creation of mitral regurgitation. Western analysis revealed an early increase in beta-actin protein followed by an apparent decrease to below baseline levels. These observations suggest that changes in beta-actin mRNA expression accompany the structural alterations that occur in response to myocardial damage. Whether or not the changes in beta-actin mRNA expression play a role in mediating these structural alterations remains to be determined.

MLCK-mediated intestinal permeability promotes immune activation and visceral hypersensitivity in PI-IBS mice.

PubMed

Long, Y; Du, L; Kim, J J; Chen, B; Zhu, Y; Zhang, Y; Yao, S; He, H; Zheng, X; Huang, Z; Dai, N

2018-04-11

Alterations in intestinal permeability regulated by tight junctions (TJs) are associated with immune activation and visceral hypersensitivity in irritable bowel syndrome (IBS). Myosin light chain kinase (MLCK) is an important mediator of epithelial TJ. The aim of this study is to investigate the role of MLCK in the pathogenesis of IBS using a post infectious IBS (PI-IBS) mouse model. Trichinella spiralis-infected PI-IBS mouse model was used. Urine lactulose/mannitol ratio was measured to assess intestinal epithelial permeability. Western blotting was used to evaluate intestinal TJ protein (zonula occludens-1) and MLCK-associated protein expressions. Immune profile was assessed by measuring Th (T helper) 1/Th2 cytokine expression. Visceral sensitivity was determined by abdominal withdrawal reflex in response to colorectal distension. Eight weeks after inoculation with T. spiralis, PI-IBS mice developed decreased pain and volume thresholds during colorectal distention, increased urine lactulose/mannitol ratio, elevated colonic Th1/Th2 cytokine ratio, and decreased zonula occludens-1 expression compared to the control mice. MLCK expression was dramatically elevated in the colonic mucosa of PI-IBS mice compared to the control mice, alongside increased pMLC/MLC and decreased MLCP expression. Administration of MLCK inhibitor and TJ blocker both reversed the increased intestinal permeability, visceral hypersensitivity, and Th1-dominant immune profile in PI-IBS mice. MLCK is a pivotal step in inducing increased intestinal permeability promoting low-grade intestinal immune activation and visceral hypersensitivity in PI-IBS mice. MLCK inhibitor may provide a potential therapeutic option in the treatment of IBS. © 2018 John Wiley & Sons Ltd.

Role of TGF-β in a Mouse Model of High Turnover Renal Osteodystrophy†

PubMed Central

Liu, Shiguang; Song, Wenping; Boulanger, Joseph H; Tang, Wen; Sabbagh, Yves; Kelley, Brian; Gotschall, Russell; Ryan, Susan; Phillips, Lucy; Malley, Katie; Cao, Xiaohong; Xia, Tai-He; Zhen, Gehua; Cao, Xu; Ling, Hong; Dechow, Paul C; Bellido, Teresita M; Ledbetter, Steven R; Schiavi, Susan C

2014-01-01

Altered bone turnover is a key pathologic feature of chronic kidney disease-mineral and bone disorder (CKD-MBD). Expression of TGF-β1, a known regulator of bone turnover, is increased in bone biopsies from individuals with CKD. Similarly, TGF-β1 mRNA and downstream signaling is increased in bones from jck mice, a model of high-turnover renal osteodystropy. A neutralizing anti-TGF-β antibody (1D11) was used to explore TGF-βs role in renal osteodystrophy. 1D11 administration to jck significantly attenuated elevated serum osteocalcin and type I collagen C-telopeptides. Histomorphometric analysis indicated that 1D11 administration increased bone volume and suppressed the elevated bone turnover in a dose-dependent manner. These effects were associated with reductions in osteoblast and osteoclast surface areas. μCT confirmed the observed increase in trabecular bone volume and demonstrated improvements in trabecular architecture and increased cortical thickness. 1D11 administration was associated with significant reductions in expression of osteoblast marker genes (Runx2, alkaline phosphatase, osteocalcin) and the osteoclast marker gene, Trap5. Importantly, in this model, 1D11 did not improve kidney function or reduce serum PTH levels indicating that 1D11 effects on bone are independent of changes in renal or parathyroid function. 1D11 also significantly attenuated high turnover bone disease in the adenine-induced uremic rat model. Antibody administration was associated with a reduction in pSMAD2/SMAD2 in bone but not bone marrow as assessed by quantitative immunoblot analysis. Immunostaining revealed pSMAD staining in osteoblasts and osteocytes but not osteoclasts, suggesting 1D11 effects on osteoclasts may be indirect. Immunoblot and whole genome mRNA expression analysis confirmed our previous observation that repression of Wnt/β catenin expression in bone is correlated with increased osteoclast activity in jck mice and bone biopsies from CKD patients. Furthermore, our data suggests that elevated TGF-β may contribute to the pathogenesis of high turnover disease partially through inhibition of β-catenin signaling. PMID:24166835

Role of TGF-β in a mouse model of high turnover renal osteodystrophy.

PubMed

Liu, Shiguang; Song, Wenping; Boulanger, Joseph H; Tang, Wen; Sabbagh, Yves; Kelley, Brian; Gotschall, Russell; Ryan, Susan; Phillips, Lucy; Malley, Katie; Cao, Xiaohong; Xia, Tai-He; Zhen, Gehua; Cao, Xu; Ling, Hong; Dechow, Paul C; Bellido, Teresita M; Ledbetter, Steven R; Schiavi, Susan C

2014-01-01

Altered bone turnover is a key pathologic feature of chronic kidney disease-mineral and bone disorder (CKD-MBD). Expression of TGF-β1, a known regulator of bone turnover, is increased in bone biopsies from individuals with CKD. Similarly, TGF-β1 mRNA and downstream signaling is increased in bones from jck mice, a model of high-turnover renal osteodystrophy. A neutralizing anti-TGF-β antibody (1D11) was used to explore TGF-β's role in renal osteodystrophy. 1D11 administration to jck significantly attenuated elevated serum osteocalcin and type I collagen C-telopeptides. Histomorphometric analysis indicated that 1D11 administration increased bone volume and suppressed the elevated bone turnover in a dose-dependent manner. These effects were associated with reductions in osteoblast and osteoclast surface areas. Micro-computed tomography (µCT) confirmed the observed increase in trabecular bone volume and demonstrated improvements in trabecular architecture and increased cortical thickness. 1D11 administration was associated with significant reductions in expression of osteoblast marker genes (Runx2, alkaline phosphatase, osteocalcin) and the osteoclast marker gene, Trap5. Importantly, in this model, 1D11 did not improve kidney function or reduce serum parathyroid hormone (PTH) levels, indicating that 1D11 effects on bone are independent of changes in renal or parathyroid function. 1D11 also significantly attenuated high-turnover bone disease in the adenine-induced uremic rat model. Antibody administration was associated with a reduction in pSMAD2/SMAD2 in bone but not bone marrow as assessed by quantitative immunoblot analysis. Immunostaining revealed pSMAD staining in osteoblasts and osteocytes but not osteoclasts, suggesting 1D11 effects on osteoclasts may be indirect. Immunoblot and whole genome mRNA expression analysis confirmed our previous observation that repression of Wnt/β-catenin expression in bone is correlated with increased osteoclast activity in jck mice and bone biopsies from CKD patients. Furthermore, our data suggest that elevated TGF-β may contribute to the pathogenesis of high-turnover disease partially through inhibition of β-catenin signaling. © 2014 American Society for Bone and Mineral Research.

Protective effect of active perfusion in porcine models of acute myocardial ischemia

PubMed Central

Feng, Zanxiang; Mao, Zhifu; Dong, Shengjun; Liu, Baohui

2016-01-01

Mortality rates associated with off-pump coronary artery bypass (CAB) are relatively high, as the majority of patients requiring CAB are at a high risk for cardiac events. The present study aimed to establish porcine models of acute myocardial ischemia, and evaluate the protective role of shunt and active perfusion. A total of 30 pigs were randomly assigned to five groups, as follows: i) Sham (control); ii) A1 (shunt; stenosis rate, 55%); iii) A2 (shunt; stenosis rate, 75%); iv) B1 (active perfusion; stenosis rate, 55%); and v) B2 (active perfusion; stenosis rate, 75%) groups. Aortic pressure (P0), left anterior descending coronary pressure (P1), and coronary effective perfusion pressure (P1/P0) were measured. The expression levels of tumor necrosis factor-α (TNF-α), cardiac troponin (cTnI), creatine kinase-myocardial band (CK-MB), interleukin (IL)-6, IL-10, B-cell lymphoma 2 (Bcl-2), and caspase-3 were detected using enzyme-linked immunosorbent assay or western blotting. The myocardial apoptosis rate was determined using the terminal deoxynucleotidyl transferase dUTP nick end labeling assay. Ischemia models with stenosis rates of 55 and 75% were successfully constructed following suturing of the descending artery. Compared with the control, the 55 and 75% stenosis groups demonstrated significantly decreased P1/P0, increased expression levels of TNF-α, cTnI, CK-MB, IL-6, IL-10 and caspase-3, an increased rate of myocardial apoptosis, and a decreased expression level of anti-apoptotic protein, Bcl-2. At 30 min following successful establishment of the model (ST segment elevation to 1 mm), group B demonstrated significantly increased P1/P0, decreased expression levels of TNF-α, cTnI, CK-MB, IL-6, IL-10 and caspase-3, a decreased rate of myocardial apoptosis, and an increased expression level of anti-apoptotic protein, Bcl-2. Furthermore, the current study indicated that active perfusion was more efficacious in maintaining myocardial perfusion and alleviating ischemic injury when compared with traditional shunt perfusion. PMID:27573177

Totarol prevents neuronal injury in vitro and ameliorates brain ischemic stroke: Potential roles of Akt activation and HO-1 induction

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gao, Yuanxue; Xu, Xiaojun; Chang, Sai

The natural product totarol, a phenolic diterpenoid and a major constituent isolated from the sap of Podocarpus totara, has been reported to have a potent antimicrobial activity. In this study, we determined whether totarol possessed an additional neuroprotective activity in vitro and in vivo. We found that totarol prevented glutamate- and oxygen and glucose deprivation-induced neuronal death in primary rat cerebellar granule neuronal cells and cerebral cortical neurons. Totarol increased Akt and GSK-3β phosphorylation, Nrf2 and heme oxygenase-1 (HO-1) protein expressions and suppressed oxidative stress by increasing GSH and SOD activities. The PI3K/Akt inhibitor LY294002 prevented totarol neuroprotective effect bymore » suppressing the totarol-induced changes in HO-1 expression and the activities of GSH and SOD. The HO-1 inhibitor ZnPPIX also prevented totarol-increased GSH and SOD activities. In a model of acute cerebral ischemic injury in Sprague–Dawley rats, produced by occlusion of the middle cerebral artery for 2 h followed by 22 h or 46 h of reperfusion, totarol significantly reduced infarct volume and improved the neurological deficit. In this model, totarol increased HO-1 expression and the activities of GSH and SOD. These observations suggest that totarol may be a novel activator of the Akt/HO-1 pathway protecting against ischemic stroke through reduction of oxidative stress. - Graphical abstract: It is unknown whether the natural product totarol has neuroprotective effects in vitro and in vivo. This study underscores that totarol prevents neuronal injury in vitro, not only by activating PI3K/Akt pathway, but also via induction of Nrf2, HO-1, GSH and SOD expressions. Totarol also ameliorated acute cerebral ischemic injury in a rat ischemic stroke model. The findings highlight that totarol may be exploited for protecting against ischemic stroke through Akt/HO-1 pathway. - Highlights: • Totarol protects glutamate- and OGD-induced neuronal injury in vitro. • Totarol activates PI3K/Akt pathway in neurons. • Totarol induces HO-1, GSH and SOD expression in vitro and in vivo. • Totarol exhibits neuroprotective effects in rat brain ischemic stroke model.« less

[Effect of tongluo xingnao effervescent tablet on learning and memory of AD rats and expression of insulin-degrading enzyme in hippocampus].

PubMed

Zhang, Yin-Jie; Dai, Yuan; Hu, Yong; Ma, Yun-Tong; Xu, Shi-Jun; Wang, Yong-Yan

2013-09-01

To study the effect of Tongluo Xingnao effervescent tablet on learning and memory of dementia rats induced by injection of Abeta25-35 in hippocampus and expression of insulin-degrading enzyme in hippocampus, in order to provide basis for preventing and treating senile dementia. The dementia rat model was established by injecting Abeta25-35 in hippocampus. The rats were divided into the model control group, the Aricept (1.4 mg x kg(-1)) group, and Tongluo Xingnao effervescent tablet high dose (7.56 g x kg(-1)), middle dose (3.78 g x kg(-1)) and low dose (1.59 g x kg(-1)) groups. A sham operation group was established by injecting normal saline in hippocampus. The rats were orally given drugs for 90 days, once a day. Their learning and memory were tested by using Morris water maze. Immunohistochemistry and image analysis were utilized for a quantitative analysis on the expression of insulin-degrading enzyme in hippocampus. Tongluo Xingnao effervescent tablet could significantly shorten the escape latency of rats in the directional navigation test, prolong the retention time in the first quadrant dwell, decrease the retention time in the third quadrant dwell, increase the frequency of crossing the platform, show a more notable statistical significance than the model control group (P < 0.05). Additionally, it could also remarkably increase the average optical density of insulin-degrading enzyme in hippocampus, promote the expression of insulin-degrading enzyme in hippocampus, and show a more notable statistical significance than the model control group (P < 0.05). Tongluo Xingnao effervescent tablet has the effects of improving learning and memory capacity of AD rats and promoting the expression of insulin-degrading enzyme in hippocampus. Its effect in promoting intelligence will be related to increased insulin-degrading enzyme in hippocampus.

Changes in the renin angiotensin system during the development of colorectal cancer liver metastases.

PubMed

Neo, Jaclyn H; Ager, Eleanor I; Angus, Peter W; Zhu, Jin; Herath, Chandana B; Christophi, Christopher

2010-04-10

Blockade of the renin angiotensin system (RAS) via angiotensin I converting enzyme (ACE) inhibition reduces growth of colorectal cancer (CRC) liver metastases in a mouse model. In this work we defined the expression of the various components of the RAS in both tumor and liver during the progression of this disease. Immunohistochemistry and quantitative RT-PCR was used to examine RAS expression in a mouse CRC liver metastases model. CRC metastases and liver tissue was assessed separately at key stages of CRC liver metastases development in untreated (control) mice and in mice treated with the ACE inhibitor captopril (750 mg/kg/day). Non-tumor induced (sham) mice indicated the effect of tumors on normal liver RAS. The statistical significance of multiple comparisons was determined using one-way analysis of variance followed by Bonferroni adjustment with SAS/STAT software. Reduced volume of CRC liver metastases with captopril treatment was evident. Local RAS of CRC metastases differed from the surrounding liver, with lower angiotensin II type 1 receptor (AT1R) expression but increased ANG-(1-7) receptor (MasR) compared to the liver. The AT1R localised to cancer and stromal infiltrating cells, while other RAS receptors were detected in cancer cells only. Tumor induction led to an initial increase in AT1R and ACE expression while captopril treatment significantly increased ACE expression in the final stages of tumor growth. Conversely, captopril treatment decreased expression of AT1R and angiotensinogen. These results demonstrate significant changes in RAS expression in the tumor-bearing captopril treated liver and in CRC metastases. The data suggests the existence of a tumor-specific RAS that can be independently targeted by RAS blockade.

Changes in the renin angiotensin system during the development of colorectal cancer liver metastases

PubMed Central

2010-01-01

Background Blockade of the renin angiotensin system (RAS) via angiotensin I converting enzyme (ACE) inhibition reduces growth of colorectal cancer (CRC) liver metastases in a mouse model. In this work we defined the expression of the various components of the RAS in both tumor and liver during the progression of this disease. Methods Immunohistochemistry and quantitative RT-PCR was used to examine RAS expression in a mouse CRC liver metastases model. CRC metastases and liver tissue was assessed separately at key stages of CRC liver metastases development in untreated (control) mice and in mice treated with the ACE inhibitor captopril (750 mg/kg/day). Non-tumor induced (sham) mice indicated the effect of tumors on normal liver RAS. The statistical significance of multiple comparisons was determined using one-way analysis of variance followed by Bonferroni adjustment with SAS/STAT software. Results Reduced volume of CRC liver metastases with captopril treatment was evident. Local RAS of CRC metastases differed from the surrounding liver, with lower angiotensin II type 1 receptor (AT1R) expression but increased ANG-(1-7) receptor (MasR) compared to the liver. The AT1R localised to cancer and stromal infiltrating cells, while other RAS receptors were detected in cancer cells only. Tumor induction led to an initial increase in AT1R and ACE expression while captopril treatment significantly increased ACE expression in the final stages of tumor growth. Conversely, captopril treatment decreased expression of AT1R and angiotensinogen. Conclusions These results demonstrate significant changes in RAS expression in the tumor-bearing captopril treated liver and in CRC metastases. The data suggests the existence of a tumor-specific RAS that can be independently targeted by RAS blockade. PMID:20380732

Effects of electroacupuncture on luteal regression and steroidogenesis in ovarian hyperstimulation syndrome model rat.

PubMed

Huang, Xuan; Chen, Li; Xia, You-Bing; Xie, Min; Sun, Qin; Yao, Bing

2018-03-15

Electroacupuncture (EA) is an effective and safe therapeutic method widely used for treating clinical diseases. Previously, we found that EA could decrease serum hormones and reduce ovarian size in ovarian hyperstimulation syndrome (OHSS) rat model. Nevertheless, the mechanisms that contribute to these improvements remain unclear. HE staining was used to count the number of corpora lutea (CL) and follicles. Immunohistochemical and ELISA were applied to examine luteal functional and structural regression. Immunoprecipitation was used for analyzing the interaction between NPY (neuropeptide Y) and COX-2; western blotting and qRT-PCR were used to evaluate the expressions of steroidogenic enzymes and PKA/CREB pathway. EA treatment significantly reduced the ovarian weight and the number of CL, also decreased ovarian and serum levels of PGE2 and COX-2 expression; increased ovarian PGF2α levels and PGF2α/PGE2 ratio; decreased PCNA expression and distribution; and increased cyclin regulatory inhibitor p27 expression to have further effect on the luteal formation, and promote luteal functional and structural regression. Moreover, expression of COX-2 in ovaries was possessed interactivity increased expression of NPY. Furthermore, EA treatment lowered the serum hormone levels, inhibited PKA/CREB pathway and decreased the expressions of steroidogenic enzymes. Hence, interaction with COX-2, NPY may affect the levels of PGF2α and PGE2 as well as impact the proliferation of granulosa cells in ovaries, thus further reducing the luteal formation, and promoting luteal structural and functional regression, as well as the ovarian steroidogenesis following EA treatment. EA treatment could be an option for preventing OHSS in ART. Copyright © 2018 Elsevier Inc. All rights reserved.

Increased IL-27/IL-27R expression in association with the immunopathology of murine ocular toxoplasmosis.

PubMed

Tong, Xinxin; Chen, Shengjie; Zheng, Huanqin; Huang, Shiguang; Lu, Fangli

2018-05-19

Interleukin 27 (IL-27) is a member of the IL-6/IL-12 family, and IL-27 receptor (IL-27R) consists of WSX-1 (the IL-27Rα subunit) and the signal-transducing subunit gp130. Human and mouse mast cells (MCs) express the IL-27R. To explore the expressions of IL-27/IL-27R subunits (WSX-1 and gp130) during acute ocular toxoplasmosis (OT), we established mouse model by intraocular injection of 500 Toxoplasma gondii RH strain tachyzoites. Histopathological changes were analyzed, MCs were counted by toluidine blue staining, and tryptase + /IL-27 + MCs were examined by immunofluorescence double-staining in the eyes and cervical lymph nodes (CLNs) of T. gondii-infected mice. The mRNA expressions of IL-27p28, WSX-1, gp130, and tachyzoite specific surface antigen 1 (SAG1) in the eyes and CLNs of T. gondii-infected mice, and the expressions of WSX-1 and gp130 in the murine mastocytoma cell line P815 infected with T. gondii tachyzoites in vitro were examined by using quantitative real-time reverse transcription-polymerase chain reaction. Our results showed that, after T. gondii infection, severe histopathological changes, increased numbers of total MCs and degranulated MCs, elevated expressions of IL-27p28, WSX-1, and gp130 were found in the eyes and CLNs, and significant correlations between the levels of IL-27 and SAG1 existed in the eyes and CLNs of T. gondii-infected mice. In addition, increased levels of WSX-1 and gp130 were examined in T. gondii-infected P815 cells. Our data suggested that IL-27/IL-27R expression induced by T. gondii infection may regulate MC-mediated immune response during acute OT in mouse model.

Expression and activity of Rac1 is negatively affected in the dehydroepiandrosterone induced polycystic ovary of mouse

PubMed Central

2014-01-01

Background Polycystic ovarian syndrome (PCOS) is characterized by the presence of multiple follicular cysts, giving rise to infertility due to anovulation. This syndrome affects about 10% of women, worldwide. The exact molecular mechanism leading to PCOS remains obscure. RhoGTPase has been associated with oogenesis, but its role in PCOS remains unexplored. Therefore, we attempted to elucidate the Vav-Rac1 signaling in PCOS mice model. Methods We generated a PCOS mice model by injecting dehydroepiandrosterone (DHEA) for a period of 20 days. The expression levels of Rac1, pRac1, Vav, pVav and Caveolin1 were analyzed by employing immuno-blotting and densitometry. The association between Vav and Rac1 proteins were studied by immuno-precipitation. Furthermore, we analyzed the activity of Rac1 and levels of inhibin B and 17β-estradiol in ovary using biochemical assays. Results The presence of multiple follicular cysts in ovary were confirmed by histology. The activity of Rac1 (GTP bound state) was significantly reduced in the PCOS ovary. Similarly, the expression levels of Rac1 and its phosphorylated form (pRac1) were decreased in PCOS in comparison to the sham ovary. The expression level and activity (phosphorylated form) of guanine nucleotide exchanger of Rac1, Vav, was moderately down-regulated. We observed comparatively increased expressions of Caveolin1, 17β-estradiol, and inhibin B in the polycystic ovary. Conclusion We conclude that hyperandrogenization (PCOS) by DHEA diminishes ovarian Rac1 and Vav expression and activity along with an increase in expression of Caveolin1. This is accompanied by an increase in the intra-ovarian level of '17 β-estradiol and inhibin B. PMID:24628852

Prostaglandin H2 synthase-1 and -2 expression in guinea pig gestational tissues during late pregnancy and parturition

PubMed Central

Welsh, Toni; Mitchell, Carolyn M; Walters, William A; Mesiano, Sam; Zakar, Tamas

2005-01-01

Increased intrauterine prostaglandin (PG) production is crucial for the initiation of parturition. To investigate the mechanisms controlling intrauterine PG synthesis, we examined the expression of the key PG biosynthetic isoenzymes, PG-H2 synthase (PTGS)-1 and -2, in the amnion, visceral yolk sac (VYS), placenta and myo-endometrium of pregnant guinea pigs. This animal model was chosen because the hormonal milieu of pregnancy and the role of PGs in the hormonal control of parturition are similar to those in the human. PTGS1 mRNA abundance, measured by real-time RT-PCR, increased in the amnion and the placenta during the last third of gestation. During labour, PTGS1 mRNA levels decreased precipitously in all four tissues. PTGS1 protein abundance, assessed by immunoblotting, increased to high levels in the amnion and the placenta by the end of pregnancy and remained high during labour. PTGS2 mRNA expression was higher in the placenta than in the other tissues, but did not change before and during labour. PTGS2 protein expression decreased in the placenta and remained low in the other tissues during labour. Immunohistochemistry showed pervasive PTGS1 protein expression in the amnion and strong expression in the parietal yolk sac membrane (PYS) covering the placenta. PTGS2 was expressed in the PYS and the endometrium. The PTGS inhibitor piroxicam, administered in doses that inhibited PTGS1 but not PTGS2, significantly prolonged gestation. These data suggest that PGs generated by intrauterine PTGS1 are involved in the timing of birth in guinea pigs. The induction of PTGS1 in the amnion and the PYS is a critical event leading to labour in guinea pigs and models analogous changes in the human gestational tissues before labour. PMID:16239272

Relationship between quantitative GRB7 RNA expression and recurrence after adjuvant anthracycline chemotherapy in triple-negative breast cancer.

PubMed

Sparano, Joseph A; Goldstein, Lori J; Childs, Barrett H; Shak, Steven; Brassard, Diana; Badve, Sunil; Baehner, Frederick L; Bugarini, Roberto; Rowley, Steve; Perez, Edith A; Shulman, Lawrence N; Martino, Silvana; Davidson, Nancy E; Kenny, Paraic A; Sledge, George W; Gray, Robert

2011-11-15

To conduct an exploratory analysis of the relationship between gene expression and recurrence in patients with operable triple-negative breast cancer (TNBC) treated with adjuvant doxorubicin-containing chemotherapy. RNA was extracted from archived tumor samples derived from 246 patients with stage I-III TNBC treated with adjuvant doxorubicin-containing chemotherapy, and was analyzed by quantitative reverse transcriptase PCR for a panel of 374 genes. The relationship between gene expression and recurrence was evaluated using weighted Cox proportional hazards model score tests. Growth factor receptor bound protein 7 (GRB7) was the only gene for which higher expression was significantly associated with increased recurrence in TNBC (Korn's adjusted P value = 0.04). In a Cox proportional hazards model adjusted for clinicopathologic features, higher GRB7 expression was associated with an increased recurrence risk (HR = 2.31; P = 0.04 using the median as the split). The 5-year recurrence rates were 10.5% [95% confidence intervals (CI), 7.8-14.1] in the low and 20.4% (95% CI, 16.5-25.0) in the high GRB7 groups. External validation in other datasets indicated that GRB7 expression was not prognostic in two adjuvant trials including variable systemic therapy, but in two other trials showed that high GBR7 expression was associated with resistance to neoadjuvant doxorubicin and taxane therapy. GRB7 was associated with an increased risk of recurrence in TNBC, suggesting that GRB7 or GRB7-dependent pathways may serve as potential biomarkers for therapeutic targets. Therapeutic targeting of one or more factors identified which function as interaction nodes or effectors should also be considered.

Relationship Between Quantitative GRB7 RNA Expression and Recurrence after Adjuvant Anthracycline Chemotherapy in Triple Negative Breast Cancer

PubMed Central

Sparano, Joseph A.; Goldstein, Lori J.; Childs, Barrett H.; Shak, Steven; Brassard, Diana; Badve, Sunil; Baehner, Frederick L.; Bugarini, Roberto; Rowley, Steve; Perez, Edith; Shulman, Lawrence N.; Martino, Silvana; Davidson, Nancy E.; Kenny, Paraic A.; Sledge, George W.; Gray, Robert

2012-01-01

Purpose To perform an exploratory analysis of the relationship between gene expression and recurrence in patients with operable triple negative breast cancer (TNBC) treated with adjuvant doxorubicin-containing chemotherapy. Experimental design RNA was extracted from archived tumor samples derived from 246 patients with stage I-III TNBC treated with adjuvant doxorubicin-containing chemotherapy, and was analyzed by quantitative RT-PCR for a panel of 374 genes. The relationship between gene expression and recurrence was evaluated using weighted Cox proportional hazards model score tests. Results GRB7 was the only gene for which higher expression was significantly associated with increased recurrence in TNBC (Korn’s adjusted p value=0.04). In a Cox proportional hazards model adjusted for clinicopathologic features, higher GRB7 expression was associated with an increased recurrence risk (HR 2.31, p=0.04 using the median as the split). The 5-year recurrence rates were 10.5% (95% confidence intervals [CI] 7.8%, 14.1%) in the low and 20.4% (95% CI 16.5%, 25.0%) in the high GRB7 groups. External validation in other datasets indicated that GRB7 expression was not prognostic in two adjuvant trials including variable systemic therapy, but in two other trials showed that high GBR7 expression was associated with resistance to neoadjuvant doxorubicin and taxane therapy. Conclusions GRB7 was associated with an increased risk of recurrence in TNBC, suggesting that GRB7 or GRB7-dependent pathways may serve as potential biomarkers for therapeutic targets. Therapeutic targeting of one or more factors identified which function as interaction nodes or effectors should also be considered. PMID:21933890

Bexarotene, a Selective RXRα Agonist, Reverses Hypotension Associated with Inflammation and Tissue Injury in a Rat Model of Septic Shock.

PubMed

Tunctan, Bahar; Kucukkavruk, Sefika P; Temiz-Resitoglu, Meryem; Guden, Demet S; Sari, Ayse N; Sahan-Firat, Seyhan

2018-02-01

Peroxisome proliferator-activated receptors (PPARs) are nuclear receptors that can activate or inhibit the expression of many target genes by forming a heterodimer complex with the retinoid X receptor (RXR). The aim of this study was to investigate effects of bexarotene, a selective RXRα agonist, on the changes in renal, cardiac, hepatic, and pulmonary expression/activity of inducible nitric oxide synthase (iNOS) and cytochrome P450 (CYP) 4F6 in relation to PPARα/β/γ-RXRα heterodimer formation in a rat model of septic shock. Rats were injected with dimethyl sulfoxide or bexarotene 1 h after administration of saline or lipopolysaccharide (LPS). Mean arterial pressure (MAP) and heart rate (HR) were recorded from rats, which had received either saline or LPS before and after 1, 2, 3, and 4 h. Serum iNOS, LTB 4 , myeloperoxidase (MPO), and lactate dehydrogenase (LDH) levels as well as tissue iNOS and CYP4F6 mRNA expression in addition to PPARα/β/γ and RXRα proteins were measured. LPS-induced decrease in MAP and increase in HR were associated with a decrease in PPARα/β/γ-RXRα heterodimer formation and CYP4F6 mRNA expression. LPS also caused an increase in systemic iNOS, LTB 4 , MPO, and LDH levels as well as iNOS mRNA expression. Bexarotene at 0.1 mg/kg (i.p.) prevented the LPS-induced changes, except tachycardia. The results suggest that increased formation of PPARα/β/γ-RXRα heterodimers and CYP4F6 expression/activity in addition to decreased iNOS expression contributes to the beneficial effect of bexarotene to prevent the hypotension associated with inflammation and tissue injury during rat endotoxemia.

The expression and proangiogenic effect of nucleolin during the recovery of heat-denatured HUVECs.

PubMed

Liang, Pengfei; Jiang, Bimei; Lv, Chunliu; Huang, Xu; Sun, Li; Zhang, Pihong; Huang, Xiaoyuan

2013-10-01

The present study aims to examine the expression patterns and roles of nucleolin during the recovery of heat-denatured human umbilical vein endothelial cells (HUVECs). Deep partial thickness burn model in Sprague-Dawley rats and the heat denatured cell model (52°C, 35s) were used. The expression of nucleolin was measured using Western blot analysis and real-time PCR. Angiogenesis was assessed using in vitro parameters including endothelial cell proliferation, transwell migration assay, and scratched wound healing. Gene transfection and RNA interference approaches were employed to investigate the roles of nucleolin. Nucleolin mRNA and protein expression showed a time-dependent increase during the recovery of heat-denatured dermis and HUVECs. Heat-denaturation time-dependently promoted cell growth, adhesion, migration, scratched wound healing and formation of tube-like structures in HUVECs. These effects of heat denaturation on endothelial wound healing and formation of tube-like structures were prevented by knockdown of nucleolin, whereas over-expression of nucleolin increased cell growth, migration, and formation of tube-like structures in cultured HUVEC endothelial cells. In addition, we found that the expression of vascular endothelial growth factor (VEGF) increased during the recovery of heat-denatured dermis and HUVECs, and nucleolin up-regulated VEGF in HUVECs. The present study reveals that the expression of nucleolin is up-regulated, and plays a pro-angiogenic role during the recovery of heat-denatured dermis and its mechanism is probably dependent on production of VEGF. We find a novel and important pro-angiogenic role of nucleolin during the recovery of heat-denatured dermis. Copyright © 2013 Elsevier B.V. All rights reserved.

Simple, Flexible, Trigonometric Taper Equations

Treesearch

Charles E. Thomas; Bernard R. Parresol

1991-01-01

There have been numerous approaches to modeling stem form in recent decades. The majority have concentrated on the simpler coniferous bole form and have become increasingly complex mathematical expressions. Use of trigonometric equations provides a simple expression of taper that is flexible enough to fit both coniferous and hard-wood bole forms. As an illustration, we...

Upregulation of endothelin receptors A and B in the nitrofen induced hypoplastic lung occurs early in gestation.

PubMed

Dingemann, Jens; Doi, Takashi; Ruttenstock, Elke; Puri, Prem

2010-01-01

Pulmonary hypoplasia and persistent pulmonary hypertension (PPH) aggravate clinical courses in congenital diaphragmatic hernia (CDH). Endothelin 1 enhances PPH by vasoconstriction and proliferation of vessel walls. Up-regulation of pulmonary Endothelin Receptors A and B (EDNRA, EDNRB) has been reported in human CDH and animal models, but the onset of those alterations during lung development remains unclear. We hypothesized that pulmonary expression of EDNRA and EDNRB is up-regulated at early gestational stages in the nitrofen model. Pregnant rats were exposed to nitrofen or vehicle on gestational day 9 (D9). Embryos were sacrificed on D15, D18 and D21 and divided into nitrofen- and control group. Pulmonary RNA was extracted and mRNA levels of EDNRA and EDNRB were determined by real-time PCR. Immunohistochemistry for protein expression of both receptors was performed. mRNA levels of EDNRA and EDNRB were significantly increased in the nitrofen group on D15, D18 and D21. Immunohistochemistry revealed increased pulmonary vascular expression of EDNRA and EDNRB compared to controls. Altered expression of EDNRA and EDNRB is an early event in lung morphogenesis in the nitrofen model. We speculate that pulmonary arteries in CDH become excessively muscularised in early fetal life, becoming unable to adapt normally at birth.

MiR-19a targets suppressor of cytokine signaling 1 to modulate the progression of neuropathic pain

PubMed Central

Wang, Conghui; Jiang, Qi; Wang, Min; Li, Dong

2015-01-01

Purpose: we aimed to investigate whether miR-19a is associated with neuropathic pain and elucidate the underlying regulatory mechanism. Methods: We established a neuropathic pain model of bilateral chronic constriction injury (bCCI). Then bCCI rats were injected with mo-miR-19a, siR-SOCS1 or blank expression vector through a microinjection syringe via an intrathecal catheter on 3 day before surgery and after surgery. Behavioral tests, such as mechanical allodynia, thermal hyperalgesia and acetone induced cold allodynia, were performed to evaluate the pain threshold. Besides, quantitative real-time polymerase chain reaction (qRT-PCR) was performed to determine the expression of miR-19a and western blotting was carried out to measure the expression of SOCS1. Results: miR-19a expression levels were markedly increased in neuropathic pain models. Moreover, miR-19a significantly attenuated mechanical allodynia and thermal hyperalgesia, and similar results were obtained after knockdown of SOCS1 expression. However, miR-19a markedly increased the times that the rats appeared a sign of cold allodynia, and knockdown of SOCS1 expression had similar effects. Besides, the results of bioinformatics analysis and western blotting analysis were all confirmed that SOCS1 was a direct target of miR-19a in neuropathic pain models. Conclusions: Our finding indicate that SOCS1 is a direct target of miR-19a in neuropathic pain rats and miR-19a may play a critical role in regulating of neuropathic pain via targeting SOCS1. PMID:26617805

MiR-19a targets suppressor of cytokine signaling 1 to modulate the progression of neuropathic pain.

PubMed

Wang, Conghui; Jiang, Qi; Wang, Min; Li, Dong

2015-01-01

We aimed to investigate whether miR-19a is associated with neuropathic pain and elucidate the underlying regulatory mechanism. We established a neuropathic pain model of bilateral chronic constriction injury (bCCI). Then bCCI rats were injected with mo-miR-19a, siR-SOCS1 or blank expression vector through a microinjection syringe via an intrathecal catheter on 3 day before surgery and after surgery. Behavioral tests, such as mechanical allodynia, thermal hyperalgesia and acetone induced cold allodynia, were performed to evaluate the pain threshold. Besides, quantitative real-time polymerase chain reaction (qRT-PCR) was performed to determine the expression of miR-19a and western blotting was carried out to measure the expression of SOCS1. miR-19a expression levels were markedly increased in neuropathic pain models. Moreover, miR-19a significantly attenuated mechanical allodynia and thermal hyperalgesia, and similar results were obtained after knockdown of SOCS1 expression. However, miR-19a markedly increased the times that the rats appeared a sign of cold allodynia, and knockdown of SOCS1 expression had similar effects. Besides, the results of bioinformatics analysis and western blotting analysis were all confirmed that SOCS1 was a direct target of miR-19a in neuropathic pain models. Our finding indicate that SOCS1 is a direct target of miR-19a in neuropathic pain rats and miR-19a may play a critical role in regulating of neuropathic pain via targeting SOCS1.

The use of expressive methods for developing empathic skills.

PubMed

Ozcan, Neslihan Keser; Bilgin, Hülya; Eracar, Nevin

2011-01-01

Empathy is one of the fundamental concepts in nursing, and it is an ability that can be learned. Various education models have been tested for improving empathic skills. Research has focused on using oral presentations, videos, modeling, practiced negotiation based on experiences, and psychodrama methods, such as role playing, as ways to improve empathy in participants. This study looked at the use of expressive arts to improve empathic skills of nursing students. The study was conducted with 48 students who were separated into five different groups. All groups lasted for two hours, and met for 12 weeks. Expressive art and psychodrama methods were used in the group studies. The Scale of Empathic Skill was administered to participants before and after the group studies. Before the group study took place, the average score for empathic skill was 127.97 (SD = 21.26). After the group study, it increased to 138.87 (SD = 20.40). The average score for empathic skill increased after the group (t = 3.996, p = .000). Results show that expressive methods are easier, more effective, and enjoyable processes in nursing training.

Study of effects of radio-wave frequency radiation emitted from cellular telephones on embryonic development of danio rerio

NASA Astrophysics Data System (ADS)

Vagula, Mary; Harkless, Ryan

2013-05-01

Radio wave frequency (RF) radiation emitted from cellular telephones has become increasingly ubiquitous as a result of the popularity of these phones. With the increasing and unavoidable exposure to RF radiation a reality, it is imperative that the effects of such radiation on living tissue be well understood. In particular, it is critical to understand any effects that RF radiation may have as a carcinogen and on embryonic development, as pregnant women are not exempt from such exposure. As a model organism, zebrafish (Danio rerio) have been studied extensively, and their value in studies of gene expression cannot be overstated. This study observed the effects of RF radiation on the embryonic development of zebrafish. The expression of two genes, shha and hoxb9a, that are key to the early development of the fish was examined. Both genes have homologs in humans as well as in other model organisms. Preliminary results suggest that exposure to cell phone radiation might have an effect on the expression of shha in zebrafish embryos, causing under expression. More trials are necessary to validate these results.

Associations between expression levels of nucleotide excision repair proteins in lymphoblastoid cells and risk of squamous cell carcinoma of the head and neck.

PubMed

Han, Peng; Liu, Hongliang; Shi, Qiong; Liu, Zhensheng; Troy, Jesse D; Lee, Walter T; Zevallos, Jose P; Li, Guojun; Sturgis, Erich M; Wei, Qingyi

2018-06-01

Squamous cell carcinoma of head and neck (SCCHN) is one of the most common malignancies worldwide, and nucleotide excision repair (NER) is involved in SCCHN susceptibility. In this analysis of 349 newly diagnosed SCCHN patients and 295 cancer-free controls, we investigated whether expression levels of eight core NER proteins were associated with risk of SCCHN. We quantified NER protein expression levels in cultured peripheral lymphocytes using a reverse-phase protein microarray. Compared with the controls, SCCHN patients had statistically significantly lower expression levels of ERCC3 and XPA (P = 0.001 and 0.001, respectively). After dividing the subjects by controls' median values of expression levels, we found a dose-dependent association between an increased risk of SCCHN and low expression levels of ERCC3 (adjusted OR, 1.75, and 95% CI: 1.26-2.42; P trend  = 0.008) and XPA (adjusted OR, 1.88; 95% CI, 1.35-2.60; P trend  = 0.001). We also identified a significant multiplicative interaction between smoking status and ERCC3 expression levels (P = 0.014). Finally, after integrating demographic and clinical variables, we found that the addition of ERCC3 and XPA expression levels to the model significantly improved the sensitivity of the expanded model on SCCHN risk. In conclusion, reduced protein expression levels of ERCC3 and XPA were associated with an increased risk of SCCHN. However, these results need to be confirmed in additional large studies. © 2018 Wiley Periodicals, Inc.

Changes in gene expression in chronic allergy mouse model exposed to natural environmental PM2.5-rich ambient air pollution.

PubMed

Ouyang, Yuhui; Xu, Zhaojun; Fan, Erzhong; Li, Ying; Miyake, Kunio; Xu, Xianyan; Zhang, Luo

2018-04-20

Particulate matter (PM) air pollution has been associated with an increase in the incidence of chronic allergic diseases; however, the mechanisms underlying the effect of exposure to natural ambient air pollution in chronic allergic diseases have not been fully elucidated. In the present study, we aimed to investigate the cellular responses induced by exposure to natural ambient air pollution, employing a mouse model of chronic allergy. The results indicated that exposure to ambient air pollution significantly increased the number of eosinophils in the nasal mucosa. The modulation of gene expression profile identified a set of regulated genes, and the Triggering Receptor Expressed on Myeloid cells1(TREM1) signaling canonical pathway was increased after exposure to ambient air pollution. In vitro, PM2.5 increased Nucleotide-binding oligomerization domain-containing protein 1 (Nod1) and nuclear factor (NF)-κB signaling pathway activation in A549 and HEK293 cell cultures. These results suggest a novel mechanism by which, PM2.5 in ambient air pollution may stimulate the innate immune system through the PM2.5-Nod1-NF-κB axis in chronic allergic disease.

STAMP2 increases oxidative stress and is critical for prostate cancer

PubMed Central

Jin, Yang; Wang, Ling; Qu, Su; Sheng, Xia; Kristian, Alexandr; Mælandsmo, Gunhild M; Pällmann, Nora; Yuca, Erkan; Tekedereli, Ibrahim; Gorgulu, Kivanc; Alpay, Neslihan; Sood, Anil; Lopez-Berestein, Gabriel; Fazli, Ladan; Rennie, Paul; Risberg, Bjørn; Wæhre, Håkon; Danielsen, Håvard E; Ozpolat, Bulent; Saatcioglu, Fahri

2015-01-01

The six transmembrane protein of prostate 2 (STAMP2) is an androgen-regulated gene whose mRNA expression is increased in prostate cancer (PCa). Here, we show that STAMP2 protein expression is increased in human PCa compared with benign prostate that is also correlated with tumor grade and treatment response. We also show that STAMP2 significantly increased reactive oxygen species (ROS) in PCa cells through its iron reductase activity which also depleted NADPH levels. Knockdown of STAMP2 expression in PCa cells inhibited proliferation, colony formation, and anchorage-independent growth, and significantly increased apoptosis. Furthermore, STAMP2 effects were, at least in part, mediated by activating transcription factor 4 (ATF4), whose expression is regulated by ROS. Consistent with in vitro findings, silencing STAMP2 significantly inhibited PCa xenograft growth in mice. Finally, therapeutic silencing of STAMP2 by systemically administered nanoliposomal siRNA profoundly inhibited tumor growth in two established preclinical PCa models in mice. These data suggest that STAMP2 is required for PCa progression and thus may serve as a novel therapeutic target. PMID:25680860

Oncogenic RAS pathway activation promotes resistance to anti-VEGF therapy through G-CSF–induced neutrophil recruitment

PubMed Central

Phan, Vernon T.; Wu, Xiumin; Cheng, Jason H.; Sheng, Rebecca X.; Chung, Alicia S.; Zhuang, Guanglei; Tran, Christopher; Song, Qinghua; Kowanetz, Marcin; Sambrone, Amy; Tan, Martha; Meng, Y. Gloria; Jackson, Erica L.; Peale, Franklin V.; Junttila, Melissa R.; Ferrara, Napoleone

2013-01-01

Granulocyte-colony stimulating factor (G-CSF) promotes mobilization of CD11b+Gr1+ myeloid cells and has been implicated in resistance to anti-VEGF therapy in mouse models. High G-CSF production has been associated with a poor prognosis in cancer patients. Here we show that activation of the RAS/MEK/ERK pathway regulates G-CSF expression through the Ets transcription factor. Several growth factors induced G-CSF expression by a MEK-dependent mechanism. Inhibition of G-CSF release with a MEK inhibitor markedly reduced G-CSF production in vitro and synergized with anti-VEGF antibodies to reduce CD11b+Ly6G+ neutrophil mobilization and tumor growth and led to increased survival in animal models of cancer, including a genetically engineered mouse model of pancreatic adenocarcinoma. Analysis of biopsies from pancreatic cancer patients revealed increased phospho-MEK, G-CSF, and Ets expression and enhanced neutrophil recruitment compared with normal pancreata. These results provide insights into G-CSF regulation and on the mechanism of action of MEK inhibitors and point to unique anticancer strategies. PMID:23530240

A transcriptome-based model of central memory CD4 T cell death in HIV infection.

PubMed

Olvera-García, Gustavo; Aguilar-García, Tania; Gutiérrez-Jasso, Fany; Imaz-Rosshandler, Iván; Rangel-Escareño, Claudia; Orozco, Lorena; Aguilar-Delfín, Irma; Vázquez-Pérez, Joel A; Zúñiga, Joaquín; Pérez-Patrigeon, Santiago; Espinosa, Enrique

2016-11-22

Human central memory CD4 T cells are characterized by their capacity of proliferation and differentiation into effector memory CD4 T cells. Homeostasis of central memory CD4 T cells is considered a key factor sustaining the asymptomatic stage of Human Immunodeficiency Virus type 1 (HIV-1) infection, while progression to acquired immunodeficiency syndrome is imputed to central memory CD4 T cells homeostatic failure. We investigated if central memory CD4 T cells from patients with HIV-1 infection have a gene expression profile impeding proliferation and survival, despite their activated state. Using gene expression microarrays, we analyzed mRNA expression patterns in naive, central memory, and effector memory CD4 T cells from healthy controls, and naive and central memory CD4 T cells from patients with HIV-1 infection. Differentially expressed genes, defined by Log 2 Fold Change (FC) ≥ |0.5| and Log (odds) > 0, were used in pathway enrichment analyses. Central memory CD4 T cells from patients and controls showed comparable expression of differentiation-related genes, ruling out an effector-like differentiation of central memory CD4 T cells in HIV infection. However, 210 genes were differentially expressed in central memory CD4 T cells from patients compared with those from controls. Expression of 75 of these genes was validated by semi quantitative RT-PCR, and independently reproduced enrichment results from this gene expression signature. The results of functional enrichment analysis indicated movement to cell cycle phases G1 and S (increased CCNE1, MKI67, IL12RB2, ADAM9, decreased FGF9, etc.), but also arrest in G2/M (increased CHK1, RBBP8, KIF11, etc.). Unexpectedly, the results also suggested decreased apoptosis (increased CSTA, NFKBIA, decreased RNASEL, etc.). Results also suggested increased IL-1β, IFN-γ, TNF, and RANTES (CCR5) activity upstream of the central memory CD4 T cells signature, consistent with the demonstrated milieu in HIV infection. Our findings support a model where progressive loss of central memory CD4 T cells in chronic HIV-1 infection is driven by increased cell cycle entry followed by mitotic arrest, leading to a non-apoptotic death pathway without actual proliferation, possibly contributing to increased turnover.

KLF4-dependent epigenetic remodeling modulates podocyte phenotypes and attenuates proteinuria

PubMed Central

Hayashi, Kaori; Sasamura, Hiroyuki; Nakamura, Mari; Azegami, Tatsuhiko; Oguchi, Hideyo; Sakamaki, Yusuke; Itoh, Hiroshi

2014-01-01

The transcription factor Kruppel-like factor 4 (KLF4) has the ability, along with other factors, to reprogram somatic cells into induced pluripotent stem (iPS) cells. Here, we determined that KLF4 is expressed in kidney glomerular podocytes and is decreased in both animal models and humans exhibiting a proteinuric. Transient restoration of KLF4 expression in podocytes of diseased glomeruli in vivo, either by gene transfer or transgenic expression, resulted in a sustained increase in nephrin expression and a decrease in albuminuria. In mice harboring podocyte-specific deletion of Klf4, adriamycin-induced proteinuria was substantially exacerbated, although these animals displayed minimal phenotypical changes prior to adriamycin administration. KLF4 overexpression in cultured human podocytes increased expression of nephrin and other epithelial markers and reduced mesenchymal gene expression. DNA methylation profiling and bisulfite genomic sequencing revealed that KLF4 expression reduced methylation at the nephrin promoter and the promoters of other epithelial markers; however, methylation was increased at the promoters of genes encoding mesenchymal markers, suggesting selective epigenetic regulation of podocyte gene expression. Together, these results suggest that KLF4 epigenetically modulates podocyte phenotype and function and that the podocyte epigenome can be targeted for direct intervention and reduction of proteinuria. PMID:24812666

Increased expression of (immuno)proteasome subunits during epileptogenesis is attenuated by inhibition of the mammalian target of rapamycin pathway.

PubMed

Broekaart, Diede W M; van Scheppingen, Jackelien; Geijtenbeek, Karlijne W; Zuidberg, Mark R J; Anink, Jasper J; Baayen, Johannes C; Mühlebner, Angelika; Aronica, Eleonora; Gorter, Jan A; van Vliet, Erwin A

2017-08-01

Inhibition of the mammalian target of rapamycin (mTOR) pathway reduces epileptogenesis in various epilepsy models, possibly by inhibition of inflammatory processes, which may include the proteasome system. To study the role of mTOR inhibition in the regulation of the proteasome system, we investigated (immuno)proteasome expression during epileptogenesis, as well as the effects of the mTOR inhibitor rapamycin. The expression of constitutive (β1, β5) and immunoproteasome (β1i, β5i) subunits was investigated during epileptogenesis using immunohistochemistry in the electrical post-status epilepticus (SE) rat model for temporal lobe epilepsy (TLE). The effect of rapamycin was studied on (immuno)proteasome subunit expression in post-SE rats that were treated for 6 weeks. (Immuno)proteasome expression was validated in the brain tissue of patients who had SE or drug-resistant TLE and the effect of rapamycin was studied in primary human astrocyte cultures. In post-SE rats, increased (immuno)proteasome expression was detected throughout epileptogenesis in neurons and astrocytes within the hippocampus and piriform cortex and was most evident in rats that developed a progressive form of epilepsy. Rapamycin-treated post-SE rats had reduced (immuno)proteasome protein expression and a lower number of spontaneous seizures compared to vehicle-treated rats. (Immuno)proteasome expression was also increased in neurons and astrocytes within the human hippocampus after SE and in patients with drug-resistant TLE. In vitro studies using cultured human astrocytes showed that interleukin (IL)-1β-induced (immuno)proteasome gene expression could be attenuated by rapamycin. Because dysregulation of the (immuno)proteasome system is observed before the occurrence of spontaneous seizures in rats, is associated with progression of epilepsy, and can be modulated via the mTOR pathway, it may represent an interesting novel target for drug treatment in epilepsy. Wiley Periodicals, Inc. © 2017 International League Against Epilepsy.

Expression of hepatic lipid droplets is decreased in the nitrofen model of congenital diaphragmatic hernia.

PubMed

Takahashi, Hiromizu; Kutasy, Balazs; Friedmacher, Florian; Takahashi, Toshiaki; Puri, Prem

2016-02-01

Prenatal mortality in newborn infants with congenital diaphragmatic hernia (CDH) has been attributed to increased amounts of liver hernia ion through the diaphragmatic defect. Antenatal studies in human and rodent fetus with CDH further demonstrated a contribution of the developing liver in the pathogenesis of CDH. The abnormal hepatic growth in experimental animal models, therefore, indicates a disruption of normal liver development in CDH. However, the underlying structural, histological and functional changes in the liver of animals with CDH remain unclear. We design this study to test the hypothesis that the morphological and cellular liver development is altered in the nitrogen-induced CDH model. Pregnant rats were exposed to either olive oil or nitrofen on day 9 of gestation (D9). Livers and chest were harvested on D21 and divided into two groups: control (n = 8), nitrofen with CDH (CDH, n = 8). Haematoxylin-eosin (Straub et al. Histopathology 68:617-631, 2013) staining was performed to evaluate underlying morphological changes. Apoptosis was checked by using TUNEL staining and apoptotic cell number was counted on 16-16 slides in 25 fields by two independent viewers. Hepatic lipid droplet expressions were evaluated by hepatic adipose differentiation-related protein (ARDP) expression. Compared to controls markedly increased hypertrophy was seen in CDH group. Significantly increased apoptotic cell numbers were detected in CDH group compared to controls (5.1 ± 1.5 vs 2.1 ± 0.6) (p < 0.05). The relative mRNA expression levels of ARDP were significantly reduced in CDH group compared to controls. Immunohistochemistry showed markedly decreased hepatic ADRP immunoreactivity in CDH fetuses compared to controls. Our findings provide strong evidence of hepatic hypertrophy and increased cell apoptosis in the liver of nitrofen-induced CDH. These morphological changes may affect liver lipid droplet expression function.

MK-801 Impairs Cognitive Coordination on a Rotating Arena (Carousel) and Contextual Specificity of Hippocampal Immediate-Early Gene Expression in a Rat Model of Psychosis

PubMed Central

Kubík, Štěpán; Buchtová, Helena; Valeš, Karel; Stuchlík, Aleš

2014-01-01

Flexible behavior in dynamic, real-world environments requires more than static spatial learning and memory. Discordant and unstable cues must be organized in coherent subsets to give rise to meaningful spatial representations. We model this form of cognitive coordination on a rotating arena – Carousel where arena- and room-bound spatial cues are dissociated. Hippocampal neuronal ensemble activity can repeatedly switch between multiple representations of such an environment. Injection of tetrodotoxin into one hippocampus prevents cognitive coordination during avoidance of a stationary room-defined place on the Carousel and increases coactivity of previously unrelated neurons in the uninjected hippocampus. Place avoidance on the Carousel is impaired after systemic administration of non-competitive NMDAr blockers (MK-801) used to model schizophrenia in animals and people. We tested if this effect is due to cognitive disorganization or other effect of NMDAr antagonism such as hyperlocomotion, spatial memory impairment, or general learning deficit. We also examined if the same dose of MK-801 alters patterns of immediate-early gene (IEG) expression in the hippocampus. IEG expression is triggered in neuronal nuclei in a context-specific manner after behavioral exploration and it is used to map activity in neuronal populations. IEG expression is critical for maintenance of synaptic plasticity and memory consolidation. We show that the same dose of MK-801 that impairs spatial coordination of rats on the Carousel also eliminates contextual specificity of IEG expression in hippocampal CA1 ensembles. This effect is due to increased similarity between ensembles activated in different environments, consistent with the idea that it is caused by increased coactivity between neurons, which did not previously fire together. Our data support the proposition of the Hypersynchrony theory that cognitive disorganization in psychosis is due to increased coactivity between unrelated neurons. PMID:24659959

Expression of genes responsible for cell morphogenesis involved in differentiation in porcine buccal pouch mucosal cells during long-term primary culture and real-time proliferation in vitro.

PubMed

Dyszkiewicz-Konwińska, M; Bryja, A; Jopek, K; Budna, J; Khozmi, R; Jeseta, M; Bukowska, D; Antosik, P; Bruska, M; Nowicki, M; Zabel, M; Kempisty, B

2017-01-01

Recently, using experimental animal model, we demonstrated that porcine buccal pouch mucosal cells reflect increased proliferation capability during primary cultivation in vitro. Although the histological structure and morphogenesis in oral cavity is well recognized, the molecular mechanisms which regulate this process still need further investigation. This study was aimed to analyze the molecular marker expression profile involved in morphogenesis and differentiation capacity of porcine buccal pouch mucosal cells during their long-term primary cultivation in vitro. The experiment was performed on buccal pouch mucosal cells isolated from 80 pubertal crossbred Landrace gilts. After collection, the cells were treated enzymatically and transferred into a primary in vitro culture (IVC) system and cultured for 30 days. The cells were collected for RNA isolation after 7, 15 and 30 days of IVC and were checked for their real-time proliferative status using the RTCA system. We found an increased expression of FN1 and SOX9 genes when calculated against ACTB after 7, and 30 days of IVC, (P less than 0.01, P less than 0.001, respectively). The CXCL12 mRNA was down-regulated after 7, 15 and 30 days of IVC, but not statistically significant. Similar expression profile was observed when calculated against HPRT, however, DAB2 was found to be higher expressed at day 15 of IVC, (P less than 0.05). The cell index measured during real-time cell proliferation was substantially increased between 96 h and 147h of IVC and reached the log phase. Since FN1 and SOX9 revealed significant increase of expression after long-term culture in vitro, it is suggested that expression of these differentiation and stemness genes is accompanied by cell proliferation. Moreover, FN1 and SOX9 might be recognized as new markers of buccal pouch mucosal cell proliferation and differentiation in pigs in in vitro primary culture model.

The effect of feeding a low iron diet prior to and during gestation on fetal and maternal iron homeostasis in two strains of rat

PubMed Central

2013-01-01

Background Iron deficiency anaemia during pregnancy is a global problem, with short and long term consequences for maternal and child health. Animal models have demonstrated that the developing fetus is vulnerable to maternal iron restriction, impacting on postnatal metabolic and blood pressure regulation. Whilst long-term outcomes are similar across different models, the commonality in mechanistic events across models is unknown. This study examined the impact of iron deficiency on maternal and fetal iron homeostasis in two strains of rat. Methods Wistar (n=20) and Rowett Hooded Lister (RHL, n=19) rats were fed a control or low iron diet for 4 weeks prior to and during pregnancy. Tissues were collected at day 21 of gestation for analysis of iron content and mRNA/protein expression of regulatory proteins and transporters. Results A reduction in maternal liver iron content in response to the low iron diet was associated with upregulation of transferrin receptor expression and a reduction in hepcidin expression in the liver of both strains, which would be expected to promote increased iron absorption across the gut and increased turnover of iron in the liver. Placental expression of transferrin and DMT1+IRE were also upregulated, indicating adaptive responses to ensure availability of iron to the fetus. There were considerable differences in hepatic maternal and fetal iron content between strains. The higher quantity of iron present in livers from Wistar rats was not explained by differences in expression of intestinal iron transporters, and may instead reflect greater materno-fetal transfer in RHL rats as indicated by increased expression of placental iron transporters in this strain. Conclusions Our findings demonstrate substantial differences in iron homeostasis between two strains of rat during pregnancy, with variable impact of iron deficiency on the fetus. Whilst common developmental processes and pathways have been observed across different models of nutrient restriction during pregnancy, this study demonstrates differences in maternal adaptation which may impact on the trajectory of the programmed response. PMID:23635304

[Effect of electroacupuncture and moxibustion preconditioning on blood endothelin and creatine kinase contents and myocardial HSP 70 expression in rabbits with myocardial ischemia-reperfusion injury].

PubMed

Wang, Chao; Xie, Wen-juan; Liu, Mi; Yan, Jie; Zhang, Jia-li; Liu, Zhao; Guo, Li-na

2014-10-01

To observe the effect of electroacupuncture (EA) and moxibustion (Moxi) preconditioning of bi- lateral "Neiguan" (PC 6) on plasma endothelin (ET) and serum creatine kinase (CK) contents and myocardial hot shock protein 70 (HSP 70) expression in myocardial ischemia-reperfusion injury (MIRI) rabbits, so as to revel their mechanisms underlying prevention of myocardial ischemia. A total of 72 New Zealand rabbits were randomly divided into sham operation, MIRI model, EA preconditioning and Moxi preconditioning groups (n = 18/group). Each group was further divided into 0 h, 24 h and 48 h (time-point) subgroups (n=6 in each subgroup). The MIRI model was established by occlusion of the anterior descending branch of the left coronary artery for 40 min and reperfusion for 60 min. The contents of plasma ET and serum CK were detected by ELISA, and myocardial HSP 70 expression was detected by immunohistochemistry. EA and Moxi preconditioning were respectively applied to bilateral PC 6 for 20 min, once daily for 5 days. Following MIRI, contents of plasma ET and serum CK contents were significantly increased at 0 h, 24 h and 48 h in comparison with the sham group (P<0.01, P<0.05), while myo- cardial HSP 70 expression at the 3 time-points was moderately increased (P>0.05). Compared with the model groups, plasma ET contents at both 24 h and 48 h in the EA preconditioning group and at 48 h in the Moxi preconditioning group, CK contents at both 24 h and 48 h only in the EA preconditioning group were significantly down-regulated (P<0.01, P<0.05). Myocardial HSP 70 expression levels in the EA and Moxi preconditioning groups were considerably up-regulated at the three time-points in comparison with the model group(P<0.05, P<0.01). Acupuncture and moxibustion pretreatment may suppress MIRI-induced increase of plasma ET and serum CK and up-regulate myocardial HSP 70 protein expression in MIRI rabbits, suggesting a preventive protection action on ischemic myocardium.

Arctigenin Confers Neuroprotection Against Mechanical Trauma Injury in Human Neuroblastoma SH-SY5Y Cells by Regulating miRNA-16 and miRNA-199a Expression to Alleviate Inflammation.

PubMed

Song, Jie; Li, Na; Xia, Yang; Gao, Zhong; Zou, Sa-Feng; Yan, Yu-Hui; Li, Shao-Heng; Wang, Yue; Meng, Ya-Kun; Yang, Jing-Xian; Kang, Ting-Guo

2016-09-01

Mechanical trauma injury is a severe insult to neural cells. Subsequent secondary injury involves the release of inflammatory factors that have dramatic consequences for undamaged cells, leading to normal cell death after the initial injury. The present study investigated the capacity for arctigenin (ARC) to prevent secondary effects and evaluated the mechanism underlying the action of microRNA (miRNA)-199a and miRNA-16 in a mechanical trauma injury (MTI) model using SH-SY5Y cells in vitro. SH-SY5Y cells are often applied to in vitro models of neuronal function and differentiation. Recently, miRNAs have been demonstrated to play a crucial role in NF-κB and cholinergic signaling, which can regulate inflammation. The cell model was established by scratch-induced injury of human SH-SY5Y cells, which mimics the characteristics of MTI. A cell counting kit-8 (CCK-8), terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL), and immunocytochemistry were used to measure cell viability. Enzyme-linked immunosorbent assay (ELISA) was used to evaluate the inflammatory cytokine and cholinesterase (CHE) content. The lactate dehydrogenase (LDH) content was measured to assess the degree of cell injury. The mRNA levels were measured by RT-PCR to analyze ARC's mechanism of action. miRNA inhibitors and mimics were used to inhibit and strengthen the expression of miRNAs. Protein expression was detected by western blotting analysis. ARC treatment reduced the TNF-α and IL-6 levels as well as the number of TUNEL+ apoptotic SH-SY5Y cells surrounding the scratch and increased the IL-10 level compared to the controls. ARC attenuated the increase of the cell damage degree and LDH content induced by scratching, indicating increased cell survival. Mechanistic studies showed that ARC upregulated the miRNA-16 and miRNA-199a levels to reduce upstream protein (IKKα and IKKβ) expression and inhibit NF-κB signaling pathway activity; moreover, the increased miRNA-199a suppresses cholinesterases to increase cholinergic signaling, resulting in decreased expression of proinflammatory cytokines. ARC treatment confers protection for SH-SY5Y cells through positive regulation of miRNA expression, thereby reducing the inflammatory response. In turn, these effects accelerate injury repair in the scratch-induced injury model. These results might provide insights into the pharmacological role of ARC in anti-inflammation and neuroprotection in neural cells.

Peroxisome proliferator-activated receptor gamma modulation and lipogenic response in adipocytes of small-for-gestational age offspring

PubMed Central

2012-01-01

Background Small-for-gestational age (SGA) at birth increases risk of development of adult obesity and insulin resistance. A model of SGA rat offspring has been shown to exhibit increased adipose tissue expression of a key adipogenic transcription factor, peroxisome proliferator-activated receptor gamma (PPARγ), and increased fatty acid de novo synthesis during the nursing period, prior to onset of obesity. PPARγ agonists have been studied for potential use in the prevention of insulin resistance. Moreover, SGA adipocytes exhibit age-dependent differences in lipogenesis as mediated by PPARγ. The effects of PPARγ modulators on lipogenic gene expression and de novo lipogenesis on the age-dependent changes in SGA adipocytes are not known. The objectives of this study were: 1) to determine the adipogenic and lipogenic potential in SGA adipocytes at postnatal day 1 (p1) and day 21 (p21), 2) to determine how the PPARγ activator- and repressor-ligands affect the lipogenic potential, and 3) to determine the fatty acid metabolic response to PPARγ activator-ligand treatment. Methods Primary adipocyte cultures from p1 and p21 SGA and Control male offspring were established from a known maternal food-restriction model of SGA. Cell proliferation and Oil Red O (ORO) staining were quantified. Adipocytes were treated with increasing doses of rosiglitazone or bisphenol-A diglycidyl ether (BADGE). PPARγ and SREBP1 protein expression were determined. De novo lipogenesis with rosiglitazone treatment at p21 was studied using 50% U13C-glucose and gas chromatography/mass spectrometry. Results At p1 and p21, SGA demonstrated increased cell proliferation and increased ORO staining. At p21, SGA demonstrated increased lipogenic gene expression and increased glucose-mediated fatty acid de novo synthesis compared with Controls. In response to rosiglitazone, SGA adipocytes further increased glucose utilization for fatty acid synthesis. SGA lipogenic gene expression demonstrated resistance to BADGE treatment. Conclusions SGA adipocytes exhibit an enhanced adipogenic and lipogenic potential in early postnatal life. By p21, SGA demonstrated resistance to PPARγ repressor-ligand treatment, and selective response to high dose PPARγ activator-ligand treatment in adipogenic and lipogenic gene expression. p21 SGA adipocytes revealed increased fatty acid de novo synthesis through a complex relationship with glucose metabolism. PMID:22726273

Metallothionein-1 and nitric oxide expression are inversely correlated in a murine model of Chagas disease

PubMed Central

Gonzalez-Mejia, Martha Elba; Torres-Rasgado, Enrique; Porchia, Leonardo M; Salgado, Hilda Rosas; Totolhua, José-Luis; Ortega, Arturo; Hernández-Kelly, Luisa Clara Regina; Ruiz-Vivanco, Guadalupe; Báez-Duarte, Blanca G; Pérez-Fuentes, Ricardo

2014-01-01

Chagas disease, caused by Trypanosoma cruzi, represents an endemic among Latin America countries. The participation of free radicals, especially nitric oxide (NO), has been demonstrated in the pathophysiology of seropositive individuals with T. cruzi. In Chagas disease, increased NO contributes to the development of cardiomyopathy and megacolon. Metallothioneins (MTs) are efficient free radicals scavengers of NO in vitro and in vivo. Here, we developed a murine model of the chronic phase of Chagas disease using endemic T. cruzi RyCH1 in BALB/c mice, which were divided into four groups: infected non-treated (Inf), infected N-monomethyl-L-arginine treated (Inf L-NAME), non-infected L-NAME treated and non-infected vehicle-treated. We determined blood parasitaemia and NO levels, the extent of parasite nests in tissues and liver MT-I expression levels. It was observed that NO levels were increasing in Inf mice in a time-dependent manner. Inf L-NAME mice had fewer T. cruzi nests in cardiac and skeletal muscle with decreased blood NO levels at day 135 post infection. This affect was negatively correlated with an increase of MT-I expression (r = -0.8462, p < 0.0001). In conclusion, we determined that in Chagas disease, an unknown inhibitory mechanism reduces MT-I expression, allowing augmented NO levels. PMID:24676665

PD-1 expression by tumor-associated macrophages inhibits phagocytosis and tumor immunity

PubMed Central

Gordon, Sydney R.; Maute, Roy L.; Dulken, Ben W.; Hutter, Gregor; George, Benson M.; McCracken, Melissa N.; Gupta, Rohit; Tsai, Jonathan M.; Sinha, Rahul; Corey, Daniel; Ring, Aaron M.; Connolly, Andrew J.; Weissman, Irving L.

2017-01-01

Programmed cell death protein 1 (PD-1) is an immune checkpoint receptor that is upregulated on activated T cells to induce immune tolerance.1,2 Tumor cells frequently overexpress the ligand for PD-1, programmed cell death ligand 1 (PD-L1), facilitating escape from the immune system.3,4 Monoclonal antibodies blocking PD-1/PD-L1 have shown remarkable clinical efficacy in patients with a variety of cancers, including melanoma, colorectal cancer, non-small cell lung cancer, and Hodgkin’s lymphoma.5–9 Although it is well-established that PD-1/PD-L1 blockade activates T cells, little is known about the role that this pathway may have on tumor-associated macrophages (TAMs). Here we show that both mouse and human TAMs express PD-1. TAM PD-1 expression increases over time in mouse models, and with increasing disease stage in primary human cancers. TAM PD-1 expression negatively correlates with phagocytic potency against tumor cells, and blockade of PD-1/PD-L1 in vivo increases macrophage phagocytosis, reduces tumor growth, and lengthens survival in mouse models of cancer in a macrophage-dependent fashion. Our results suggest that PD-1/PD-L1 therapies may also function through a direct effect on macrophages, with significant implications for treatment with these agents. PMID:28514441

Leptin signaling enhances cell invasion and promotes the metastasis of human pancreatic cancer via increasing MMP-13 production.

PubMed

Fan, Yingchao; Gan, Yu; Shen, Yuling; Cai, Xiaojin; Song, Yanfang; Zhao, Fangyu; Yao, Ming; Gu, Jianren; Tu, Hong

2015-06-30

Emerging evidence has suggested that leptin, an adipokine related to energy homeostasis, plays a role in cancer growth and metastasis. However, its impact on pancreatic cancer is rarely studied. In this study, we found that leptin's functional receptor Ob-Rb was expressed in pancreatic cancer cell lines. Treatment with leptin enhanced the migration and invasion of pancreatic cancer cells but did not affect the proliferation of human pancreatic cancer cells. Leptin up-regulated the expression of matrix metalloproteinase-13 (MMP-13) via the JAK2/STAT3 signaling pathway. The overexpression of leptin was shown to significantly promote tumor growth and lymph node metastasis in a subcutaneous model and an orthotopic model of human pancreatic cancer, respectively. Furthermore, in human pancreatic cancer tissues, the expression of Ob-Rb was positively correlated with the MMP-13 level. The increased expression of either Ob-Rb or MMP-13 was significantly associated with lymph node metastasis and tended to be associated with the TNM stage in patients with pancreatic cancer. Our findings suggest that leptin enhances the invasion of pancreatic cancer through the increase in MMP-13 production, and targeting the leptin/MMP-13 axis could be an attractive therapeutic strategy for pancreatic cancer.

Leptin signaling enhances cell invasion and promotes the metastasis of human pancreatic cancer via increasing MMP-13 production

PubMed Central

Shen, Yuling; Cai, Xiaojin; Song, Yanfang; Zhao, Fangyu; Yao, Ming; Gu, Jianren; Tu, Hong

2015-01-01

Emerging evidence has suggested that leptin, an adipokine related to energy homeostasis, plays a role in cancer growth and metastasis. However, its impact on pancreatic cancer is rarely studied. In this study, we found that leptin's functional receptor Ob-Rb was expressed in pancreatic cancer cell lines. Treatment with leptin enhanced the migration and invasion of pancreatic cancer cells but did not affect the proliferation of human pancreatic cancer cells. Leptin up-regulated the expression of matrix metalloproteinase-13 (MMP-13) via the JAK2/STAT3 signaling pathway. The overexpression of leptin was shown to significantly promote tumor growth and lymph node metastasis in a subcutaneous model and an orthotopic model of human pancreatic cancer, respectively. Furthermore, in human pancreatic cancer tissues, the expression of Ob-Rb was positively correlated with the MMP-13 level. The increased expression of either Ob-Rb or MMP-13 was significantly associated with lymph node metastasis and tended to be associated with the TNM stage in patients with pancreatic cancer. Our findings suggest that leptin enhances the invasion of pancreatic cancer through the increase in MMP-13 production, and targeting the leptin/MMP-13 axis could be an attractive therapeutic strategy for pancreatic cancer. PMID:25948792

Altered ion transport by thyroid epithelia from CFTR−/− pigs suggests mechanisms for hypothyroidism in Cystic Fibrosis

PubMed Central

Li, Hui; Ganta, Suhasini; Fong, Peying

2010-01-01

Subclinical hypothyroidism has been linked to Cystic Fibrosis (CF), and the cystic fibrosis transmembrane conductance regulator (CFTR) shown to be expressed in the thyroid. The thyroid epithelium secretes Cl− and absorbs Na+ in response to cAMP. Chloride secretion may provide a counter-ion for the SLC26A4 (Pendrin)-mediated I− secretion which is required for the first step of thyroid hormonogenesis, thyroglobulin iodination. In contrast, few models exist to explain a role for Na+ absorption. Whether CFTR mediates the secretory Cl− current in thyroid epithelium has not been directly addressed. We used thyroids from a novel pig CFTR−/− model, generated primary pig thyroid epithelial cell cultures (pThECs), analyzed these cultures for preservation of thyroid-specific transcripts and proteins, and monitored 1) the Cl− secretory response to the cAMP agonist, isoproterenol and 2) the amiloride-sensitive Na+ current. Baseline short-circuit current (Isc) did not differ between CFTR+/+ and CFTR−/− cultures. Serosal isoproterenol increased Isc in CFTR+/+, but not CFTR−/−, monolayers. Compared to CFTR+/+ thyroid cultures, amiloride-sensitive Na+ absorption measured in CFTR−/− pThECs represented a greater fraction of the resting Isc. However, levels of transcripts encoding ENaC subunits did not differ between CFTR+/+ and CFTR−/− pThECs. Immunoblot analysis verified ENaC subunit protein expression, but quantification indicated no difference in expression levels. Our studies definitively demonstrate that CFTR mediates cAMP-stimulated Cl− secretion in a well-differentiated thyroid culture model, and that knockout of CFTR promotes increased Na+ absorption by a mechanism other than increased ENaC expression. These findings suggest several models for the mechanism of CF-associated hypothyroidism. PMID:20729267

Social coping by masking? Parental support and peer victimization as mediators of the relationship between depressive symptoms and expressive suppression in adolescents.

PubMed

Larsen, Junilla K; Vermulst, Ad A; Eisinga, Rob; English, Tammy; Gross, James J; Hofman, Elin; Scholte, Ron H J; Engels, Rutger C M E

2012-12-01

Expressive suppression is regarded as a generally ineffective emotion regulation strategy and appears to be associated with the development of depressive symptoms among adolescents. However, the mechanisms linking suppression to depressive symptoms are not well understood. The main aim of this study was to examine two potential mediators of the prospective relationship from depressive symptoms to expressive suppression among adolescents: parental support and peer victimization. Structural equation modelling was used to construct a three-wave cross-lagged model (n = 2,051 adolescents, 48.5 % female, at baseline; 1,465 with data at all three time points) with all possible longitudinal linkages. Depressive symptoms preceded decreases in perceived parental support 1 year later. Decreases in parental support mediated the relationship between depressive symptoms and increases in expressive suppression over a 2-year period. Multi-group analyses show that the mediation model tested was significant for girls, but not for boys. No evidence for other mediating models was found. Although initial suppression preceded increases in depressive symptoms 1 year later, we did not find any evidence for the reversed link from suppression to depressive symptoms. Clear evidence for a reciprocal relationship between depressive symptoms and parental support was found. However, only limited and inconsistent support was found for a reciprocal relationship between depressive symptoms and peer victimization. Finally, although some evidence for a unidirectional relationship from parental support to increases in suppression was found, no significant prospective relationship was found between peer victimization and suppression. The implications of our clear results for parental support, and mostly lacking results for peer victimization, are discussed.

TRPM5 mediates acidic extracellular pH signaling and TRPM5 inhibition reduces spontaneous metastasis in mouse B16-BL6 melanoma cells.

PubMed

Maeda, Toyonobu; Suzuki, Atsuko; Koga, Kaori; Miyamoto, Chihiro; Maehata, Yojiro; Ozawa, Shigeyuki; Hata, Ryu-Ichiro; Nagashima, Yoji; Nabeshima, Kazuki; Miyazaki, Kaoru; Kato, Yasumasa

2017-10-03

Extracellular acidity is a hallmark of solid tumors and is associated with metastasis in the tumor microenvironment. Acidic extracellular pH (pH e ) has been found to increase intracellular Ca 2+ and matrix metalloproteinase-9 (MMP-9) expression by activating NF-κB in the mouse B16 melanoma model. The present study assessed whether TRPM5, an intracellular Ca 2+ -dependent monovalent cation channel, is associated with acidic pH e signaling and induction of MMP-9 expression in this mouse melanoma model. Treatment of B16 cells with Trpm5 siRNA reduced acidic pH e -induced MMP-9 expression. Enforced expression of Trpm5 increased the rate of acidic pH e -induced MMP-9 expression, as well as increasing experimental lung metastasis. This genetic manipulation did not alter the pH e critical for MMP-9 induction but simply amplified the percentage of inducible MMP-9 at each pH e . Treatment of tumor bearing mice with triphenylphosphine oxide (TPPO), an inhibitor of TRPM5, significantly reduced spontaneous lung metastasis. In silico analysis of clinical samples showed that high TRPM5 mRNA expression correlated with poor overall survival rate in patients with melanoma and gastric cancer but not in patients with cancers of the ovary, lung, breast, and rectum. These results showed that TRPM5 amplifies acidic pH e signaling and may be a promising target for preventing metastasis of some types of tumor.

Dihydrotestosterone deteriorates cardiac insulin signaling and glucose transport in the rat model of polycystic ovary syndrome.

PubMed

Tepavčević, Snežana; Vojnović Milutinović, Danijela; Macut, Djuro; Žakula, Zorica; Nikolić, Marina; Božić-Antić, Ivana; Romić, Snježana; Bjekić-Macut, Jelica; Matić, Gordana; Korićanac, Goran

2014-05-01

It is supposed that women with polycystic ovary syndrome (PCOS) are prone to develop cardiovascular disease as a consequence of multiple risk factors that are mostly related to the state of insulin resistance and consequent hyperinsulinemia. In the present study, we evaluated insulin signaling and glucose transporters (GLUT) in cardiac cells of dihydrotestosterone (DHT) treated female rats as an animal model of PCOS. Expression of proteins involved in cardiac insulin signaling pathways and glucose transporters, as well as their phosphorylation or intracellular localization were studied by Western blot analysis in DHT-treated and control rats. Treatment with DHT resulted in increased body mass, absolute mass of the heart, elevated plasma insulin concentration, dyslipidemia and insulin resistance. At the molecular level, DHT treatment did not change protein expression of cardiac insulin receptor and insulin receptor substrate 1, while phosphorylation of the substrate at serine 307 was increased. Unexpectedly, although expression of downstream Akt kinase and its phosphorylation at threonine 308 were not altered, phosphorylation of Akt at serine 473 was increased in the heart of DHT-treated rats. In contrast, expression and phosphorylation of extracellular signal regulated kinases 1/2 were decreased. Plasma membrane contents of GLUT1 and GLUT4 were decreased, as well as the expression of GLUT4 in cardiac cells at the end of androgen treatment. The obtained results provide evidence for alterations in expression and especially in functional characteristics of insulin signaling molecules and glucose transporters in the heart of DHT-treated rats with PCOS, indicating impaired cardiac insulin action. Copyright © 2014 Elsevier Ltd. All rights reserved.

Neurodevelopmental Expression Profile of Dimeric and Monomeric Group 1 mGluRs: Relevance to Schizophrenia Pathogenesis and Treatment.

PubMed

Lum, Jeremy S; Fernandez, Francesca; Matosin, Natalie; Andrews, Jessica L; Huang, Xu-Feng; Ooi, Lezanne; Newell, Kelly A

2016-10-10

Group 1 metabotropic glutamate receptors (mGluR1/mGluR5) play an integral role in neurodevelopment and are implicated in psychiatric disorders, such as schizophrenia. mGluR1 and mGluR5 are expressed as homodimers, which is important for their functionality and pharmacology. We examined the protein expression of dimeric and monomeric mGluR1α and mGluR5 in the prefrontal cortex (PFC) and hippocampus throughout development (juvenile/adolescence/adulthood) and in the perinatal phencyclidine (PCP) model of schizophrenia. Under control conditions, mGluR1α dimer expression increased between juvenile and adolescence (209-328%), while monomeric levels remained consistent. Dimeric mGluR5 was steadily expressed across all time points; monomeric mGluR5 was present in juveniles, dramatically declining at adolescence and adulthood (-97-99%). The mGluR regulators, Homer 1b/c and Norbin, significantly increased with age in the PFC and hippocampus. Perinatal PCP treatment significantly increased juvenile dimeric mGluR5 levels in the PFC and hippocampus (37-50%) but decreased hippocampal mGluR1α (-50-56%). Perinatal PCP treatment also reduced mGluR1α dimer levels in the PFC at adulthood (-31%). These results suggest that Group 1 mGluRs have distinct dimeric and monomeric neurodevelopmental patterns, which may impact their pharmacological profiles at specific ages. Perinatal PCP treatment disrupted the early expression of Group 1 mGluRs which may underlie neurodevelopmental alterations observed in this model.

[Effect of electroacupuncture intervention on expression of extracellular matrix collagen and metabolic enzymes].

PubMed

Liao, Jun; Zhang, Le; Ke, Mei-gui; Xu, Teng

2013-12-01

To observe the effect of electroacupuncture (EA) at "Dazhui" (GV 14) on the contents of extracellular matrix (ECM), collagen type II (COL-II), collagen type V (COL-V), matrix metalloproteinase (MMP)-13, tissue inhibitor of metalloproteinase (TIMP)-1 in rats with cervicovertebral disc degeneration so as to explore its mechanism underlying relief of intervertebral disc degeneration. A total of 28 SD rats were randomly divided into sham group (n = 7), model group (n = 7), EA group (n = 7) and medication group (n = 7). The model of cervical intervertebral disc degeneration was established by trans-section of the deep neck splenius, the longest muscles of head, neck costocervicalis, head semi-spinatus muscle, supraspinous ligament and interspinal ligaments of cervical 2-7 segments, etc. to produce imbalance between the dynamic and static force. EA was applied to "Dazhui" (GV 14) for 30 min, once daily for 28 days, with a 2 days' interval between two courses. Animals of the medication group were treated by oral administration of meloxicam tablets (0.75 mg/kg) once daily for 28 days, with a 2 days' interval between two courses. Immunohistochemistry was used to measure the expression of ECM, COL- II, COL-V, MMP-13 and TIMP-1 in the cervicovertebral disc tissue. Compared with the sham group, the expression levels of ECM and COL-II proteins in the cervicovertebral disc tissue were significantly decreased in the model group (P < 0.01), while COL-V and MMP-13 expression levels in the model group were significantly increased (P < 0.01, P < 0.05). Compared with the model group, both ECM and COL-Il expression levels were considerably increased in the EA group and medication group (P < 0.01), while COL-V and MMP-13 expression levels were considerably down-regulated (P < 0.01, P < 0.05). No significant differences were found among the four groups in TIMP-1 expression levels (P > 0.05). EA of "Dazhui" (GV 14) can effectively regulate extracellular matrix system in rats with cervical intervertebral disc degeneration, which is possibly related to its effect in relieving cervical spondylosis.

Qishenyiqi Dropping Pill attenuates myocardial fibrosis in rats by inhibiting RAAS-mediated arachidonic acid inflammation.

PubMed

Wang, Jing; Lu, Linghui; Wang, Yong; Wu, Yan; Han, Jing; Wang, Wei; Li, Chun; Tu, Pengfei

2015-12-24

In China, Qishenyiqi Dropping Pill (QSDP), a Chinese medicine formula containing Astragalus membranaceus (Fisch.) Bunge, Salvia miltiorrhiza Bunge, Panax notoginseng (Burkill) F.H.Chen and Dalbergia odorifera T.C.Chen, has been used frequently in traditional folk medicine for treatment of coronary heart diseases (CHD) and heart failure (HF). Previous study has shown that QSDP has definite therapeutic effects on promoting the heart function on CHD patients. The present study was designed to study the anti-fibrosis effects of QSDP on HF rats and to explore the underlying molecular mechanisms. HF rat model was induced by left anterior descending (LAD) coronary artery ligation. Two-dimensional (2D) echocardiography was adopted to evaluate heart functions. Immunohistochemical (IHC) method and Western-blot were used to detect expression of critical proteins in renin-angiotensin-aldosterone system (RAAS) or arachidonic acid (AA) metabolic pathway. Heart functions were seriously injured in the model group. Expressions of fibrotic markers, such as collagen Ⅰ, collagen Ⅲ, matrix metallopeptidase 2 (MMP2) and MMP9 were elevated in the model group. RAAS pathway was activated. Interestingly, AA pathway was also up-regulated in the model group and it was down-regulated by angiotensin converting enzyme inhibitors (ACEIs) drug Captopril. Expressions of the important signal-transuding proteins, including NF-κB, JAK1/STAT3 and Akt, all increased remarkably in the model group. Treatment with QSDP could attenuate myocardial fibrosis by inhibiting RAAS-activated pathway, as indicated by decreased angiotensin type 1 receptor (AT1) and increased AT2 expression. Expressions of phospholipase A2 (PLA2), cyclooxygenase 1 (COX1) and COX2 were also down-regulated in the QSDP-treated group. In addition, "therapeutic" QSDP administration seemed to down-regulate expressions of NF-κB, JAK1/ STAT3 and Akt which may play important roles in myocardial fibrosis. QSDP can exert anti-fibrosis effect by down-regulating RAAS pathway, and subsequently inhibiting expressions of proteins in AA pathway. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Hypomethylation associated enhanced transcription of trefoil factor-3 mediates tamoxifen-stimulated oncogenicity of ER+ endometrial carcinoma cells.

PubMed

Pandey, Vijay; Zhang, Min; Chong, Qing-Yun; You, Mingliang; Raquib, Ainiah Rushdiana; Pandey, Amit K; Liu, Dong-Xu; Liu, Liang; Ma, Lan; Jha, Sudhakar; Wu, Zheng-Sheng; Zhu, Tao; Lobie, Peter E

2017-09-29

Tamoxifen (TAM) is widely used as an adjuvant therapy for women with breast cancer (BC). However, TAM possesses partial oestrogenic activity in the uterus and its use has been associated with an increased incidence of endometrial carcinoma (EC). The molecular mechanism for these observations is not well understood. Herein, we demonstrated that forced expression of Trefoil factor 3 ( TFF3) , in oestrogen receptor-positive (ER+) EC cells significantly increased cell cycle progression, cell survival, anchorage-independent growth, invasiveness and tumour growth in xenograft models. Clinically, elevated TFF3 protein expression was observed in EC compared with normal endometrial tissue, and its increased expression in EC was significantly associated with myometrial invasion. TAM exposure increased expression of TFF3 in ER+ EC cells and its elevated expression resulted in increased oncogenicity and invasiveness. TAM-stimulated expression of TFF3 in EC cells was associated with hypomethylation of the TFF3 promoter sequence and c-JUN/SP1-dependent transcriptional activation. In addition, small interfering ( si) RNA -mediated depletion or polyclonal antibody inhibition of TFF3 significantly abrogated oncogenicity and invasiveness in EC cells consequent to TAM induction or forced expression of TFF3. Hence, TAM-stimulated upregulation of TFF3 in EC cells was critical in promoting EC progression associated with TAM treatment. Importantly, inhibition of TFF3 function might be an attractive molecular modality to abrogate the stimulatory effects of TAM on endometrial tissue and to limit the progression of EC.

HO-1 and CO decrease platelet-derived growth factor-induced vascular smooth muscle cell migration via inhibition of Nox1

PubMed Central

Rodriguez, Andres I.; Gangopadhyay, Archana; Kelley, Eric E.; Pagano, Patrick J.; Zuckerbraun, Brian S.; Bauer, Philip M.

2009-01-01

Objective Heme oxygenase-1 (HO-1), via its enzymatic degradation products, exhibits cell and tissue protective effects in models of vascular injury and disease. The migration of vascular smooth muscle cells (VSMC) from the medial to the intimal layer of blood vessels plays an integral role in the development of a neointima in these models. Despite this, there are no studies addressing the effect of increased HO-1 expression on VSMC migration. Results and Methods The effects of increased HO-1 expression as well as biliverdin, bilirubin, and carbon monoxide (CO), were studied in in vitro models of VSMC migration. Induction of HO-1 or CO, but not biliverdin or bilirubin, inhibited VSMC migration. This effect was mediated by the inhibition of Nox1 as determined by a range of approaches including detection of intracellular superoxide, NADPH oxidase activity measurements, and siRNA experiments. Furthermore, CO decreased PDGF-stimulated, redox-sensitive signaling pathways. Conclusion Herein we demonstrate that increased HO-1 expression and CO decreases PDGF-stimulated VSMC migration via inhibition of Nox1 enzymatic activity. These studies reveal a novel mechanism by which HO-1 and CO may mediate their beneficial effects in arterial inflammation and injury. PMID:19875720

Lactobacillus rhamnosus GG treatment improves intestinal permeability and modulates inflammatory response and homeostasis of spleen and colon in experimental model of Pseudomonas aeruginosa pneumonia.

PubMed

Khailova, Ludmila; Baird, Christine H; Rush, Aubri A; Barnes, Christopher; Wischmeyer, Paul E

2017-12-01

Recent clinical trials and in vivo models demonstrate probiotic administration can reduce occurrence and improve outcome of pneumonia and sepsis, both major clinical challenges worldwide. Potential probiotic benefits include maintenance of gut epithelial barrier homeostasis and prevention of downstream organ dysfunction due to systemic inflammation. However, mechanism(s) of probiotic-mediated protection against pneumonia remain poorly understood. This study evaluated potential mechanistic targets in the maintenance of gut barrier homeostasis following Lactobacillus rhamnosus GG (LGG) treatment in a mouse model of pneumonia. Studies were performed in 6-8 week old FVB/N mice treated (o.g.) with or without LGG (10 9  CFU/ml) and intratracheally injected with Pseudomonas aeruginosa or saline. At 4, 12, and 24 h post-bacterial treatment spleen and colonic tissue were collected for analysis. Pneumonia significantly increased intestinal permeability and gut claudin-2. LGG significantly attenuated increased gut permeability and claudin-2 following pneumonia back to sham control levels. As mucin expression is key to gut barrier homeostasis we demonstrate that LGG can enhance goblet cell expression and mucin barrier formation versus control pneumonia animals. Further as Muc2 is a key gut mucin, we show LGG corrected deficient Muc2 expression post-pneumonia. Apoptosis increased in both colon and spleen post-pneumonia, and this increase was significantly attenuated by LGG. Concomitantly, LGG corrected pneumonia-mediated loss of cell proliferation in colon and significantly enhanced cell proliferation in spleen. Finally, LGG significantly reduced pro-inflammatory cytokine gene expression in colon and spleen post-pneumonia. These data demonstrate LGG can maintain intestinal barrier homeostasis by enhancing gut mucin expression/barrier formation, reducing apoptosis, and improving cell proliferation. This was accompanied by reduced pro-inflammatory cytokine expression in the gut and in a downstream organ (spleen). These may serve as potential mechanistic targets to explain LGG's protection against pneumonia in the clinical and in vivo setting. Copyright © 2016 Elsevier Ltd and European Society for Clinical Nutrition and Metabolism. All rights reserved.

Measuring trends of outpatient antibiotic use in Europe: jointly modelling longitudinal data in defined daily doses and packages.

PubMed

Bruyndonckx, Robin; Hens, Niel; Aerts, Marc; Goossens, Herman; Molenberghs, Geert; Coenen, Samuel

2014-07-01

To complement analyses of the linear trend and seasonal fluctuation of European outpatient antibiotic use expressed in defined daily doses (DDD) by analyses of data in packages, to assess the agreement between both measures and to study changes in the number of DDD per package over time. Data on outpatient antibiotic use, aggregated at the level of the active substance (WHO version 2011) were collected from 2000 to 2007 for 31 countries and expressed in DDD and packages per 1000 inhabitants per day (DID and PID, respectively). Data expressed in DID and PID were analysed separately using non-linear mixed models while the agreement between these measurements was analysed through a joint non-linear mixed model. The change in DDD per package over time was studied with a linear mixed model. Total outpatient antibiotic and penicillin use in Europe and their seasonal fluctuation significantly increased in DID, but not in PID. The use of combinations of penicillins significantly increased in DID and in PID. Broad-spectrum penicillin use did not increase significantly in DID and decreased significantly in PID. For all but one subgroup, country-specific deviations moved in the same direction whether measured in DID or PID. The correlations are not perfect. The DDD per package increased significantly over time for all but one subgroup. Outpatient antibiotic use in Europe shows contrasting trends, depending on whether DID or PID is used as the measure. The increase of the DDD per package corroborates the recommendation to adopt PID to monitor outpatient antibiotic use in Europe. © The Author 2014. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Expression-dependent susceptibility to face distortions in processing of facial expressions of emotion.

PubMed

Guo, Kun; Soornack, Yoshi; Settle, Rebecca

2018-03-05

Our capability of recognizing facial expressions of emotion under different viewing conditions implies the existence of an invariant expression representation. As natural visual signals are often distorted and our perceptual strategy changes with external noise level, it is essential to understand how expression perception is susceptible to face distortion and whether the same facial cues are used to process high- and low-quality face images. We systematically manipulated face image resolution (experiment 1) and blur (experiment 2), and measured participants' expression categorization accuracy, perceived expression intensity and associated gaze patterns. Our analysis revealed a reasonable tolerance to face distortion in expression perception. Reducing image resolution up to 48 × 64 pixels or increasing image blur up to 15 cycles/image had little impact on expression assessment and associated gaze behaviour. Further distortion led to decreased expression categorization accuracy and intensity rating, increased reaction time and fixation duration, and stronger central fixation bias which was not driven by distortion-induced changes in local image saliency. Interestingly, the observed distortion effects were expression-dependent with less deterioration impact on happy and surprise expressions, suggesting this distortion-invariant facial expression perception might be achieved through the categorical model involving a non-linear configural combination of local facial features. Copyright © 2018 Elsevier Ltd. All rights reserved.

Prenatal retinoic acid increases lipofibroblast expression in hypoplastic rat lungs with experimental congenital diaphragmatic hernia.

PubMed

Friedmacher, Florian; Fujiwara, Naho; Hofmann, Alejandro D; Takahashi, Hiromizu; Alvarez, Luis A J; Gosemann, Jan-Hendrik; Puri, Prem

2014-06-01

Prenatal administration of all-trans retinoic acid (ATRA) has been shown to stimulate alveolarization in nitrofen-induced pulmonary hypoplasia (PH) associated with congenital diaphragmatic hernia (CDH). Lipid-containing interstitial lipofibroblasts (LIFs), characterized by adipocyte differentiation-related protein (ADRP), play a critical role in alveolar development by coordinating lipid homeostasis. Previous studies have demonstrated that ATRA positively affects LIF expression in developing lungs. We hypothesized that pulmonary LIF expression is increased after prenatal ATRA treatment in the nitrofen model of CDH-associated PH. Timed-pregnant rats were treated with nitrofen or vehicle on E9.5, followed by injection of ATRA or placebo on E18.5, E19.5, and E20.5. Fetal lungs were dissected on E21.5 and divided into Control+Placebo, Control+ATRA, Nitrofen+Placebo, and Nitrofen+ATRA. Pulmonary gene expression levels of ADRP were analyzed by quantitative real-time polymerase chain reaction, and LIF expression was investigated by ADRP immunohistochemistry, oil-red-O-, and immunofluorescence-double-staining. Relative mRNA expression of pulmonary ADRP was significantly increased in Nitrofen+ATRA compared to Nitrofen+Placebo (0.31±0.02 vs. 0.08±0.01; P<0.0001). ADRP immunoreactivity and oil-red-O-staining were markedly increased in alveolar interstitium of Nitrofen+ATRA compared to Nitrofen+Placebo. Immunofluorescence-double-staining confirmed markedly increased LIF expression in alveolar walls of Nitrofen+ATRA compared to Nitrofen+Placebo. Increased LIF expression after prenatal treatment with ATRA in nitrofen-induced PH suggests that ATRA may have a therapeutic potential in attenuating CDH-associated PH by stimulating alveolar development. Copyright © 2014 Elsevier Inc. All rights reserved.

Constitutive androstane receptor activation evokes the expression of glycolytic genes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yarushkin, Andrei A.; Kazantseva, Yuliya A.; Prokopyeva, Elena A.

It is well-known that constitutive androstane receptor (CAR) activation by 1,4-bis[2-(3,5-dichloropyridyloxy)]benzene (TCPOBOP) increases the liver-to-body weight ratio. CAR-mediated liver growth is correlated with increased expression of the pleiotropic transcription factor cMyc, which stimulates cell cycle regulatory genes and drives proliferating cells into S phase. Because glycolysis supports cell proliferation and cMyc is essential for the activation of glycolytic genes, we hypothesized that CAR-mediated up-regulation of cMyc in mouse livers might play a role in inducing the expression of glycolytic genes. The aim of the present study was to examine the effect of long-term CAR activation on glycolytic genes in amore » mouse model not subjected to metabolic stress. We demonstrated that long-term CAR activation by TCPOBOP increases expression of cMyc, which was correlated with reduced expression of gluconeogenic genes and up-regulation of glucose transporter, glycolytic and mitochondrial pyruvate metabolising genes. These changes in gene expression after TCPOBOP treatment were strongly correlated with changes in levels of glycolytic intermediates in mouse livers. Moreover, we demonstrated a significant positive regulatory effect of TCPOBOP-activated CAR on both mRNA and protein levels of Pkm2, a master regulator of glucose metabolism and cell proliferation. Thus, our findings provide evidence to support the conclusion that CAR activation initiates a transcriptional program that facilitates the coordinated metabolic activities required for cell proliferation. - Highlights: • CAR-mediated liver growth is correlated with increased expression of cMyc. • CAR activation increased the expression of glycolytic genes in mouse livers. • CAR activation increased the level of Pkm2 in mouse livers.« less

UDP-Glucuronosyltransferase Expression in Mouse Liver Is Increased in Obesity- and Fasting-Induced Steatosis

PubMed Central

Xu, Jialin; Kulkarni, Supriya R.; Li, Liya

2012-01-01

UDP-glucuronosyltransferases (Ugt) catalyze phase II conjugation reactions with glucuronic acid, which enhances chemical polarity and the elimination from the body. Few studies have addressed whether Ugt expression and activity are affected by liver disease, such as steatosis. The purpose of this study was to determine whether steatosis induced by obesity or fasting could affect liver Ugt mRNA expression and activity. Male C57BL/6J and Lepob/ob (ob/ob) mice were fed ad libitum or food was withheld for 24 h. In steatotic livers of ob/ob mice, Ugt1a1, -1a6, -1a9, -2a3, -3a1, and -3a2 mRNA expression increased. Fasting, which also induced steatosis, increased hepatic Ugt1a1, -1a6, -1a7, -1a9, -2b1, -2b5, -2a3, -3a1, and -3a2 mRNA expression in mouse liver. Likewise, acetaminophen glucuronidation increased by 47% in hepatic microsomes from ob/ob mice compared with that in C57BL/6J mice, but not after fasting. In both steatosis models, Ugt induction was accompanied by increased aryl hydrocarbon receptor, constitutive androstane receptor (CAR), peroxisome proliferator-activated receptor (PPAR)-α, pregnane X receptor, nuclear factor (erythroid-derived 2)-like 2 (Nrf2), and peroxisome proliferator-activated receptor-γ coactivator-1α mRNA expression. In addition, fasting increased CAR, PPAR, and Nrf2 binding activity. The work points to hepatic triglyceride concentrations corresponding with nuclear receptor and Ugt expression. The findings indicate that steatosis significantly alters hepatic Ugt expression and activity, which could have a significant impact on determining circulating hormone levels, drug efficacy, and environmental chemical clearance. PMID:22031624

UDP-glucuronosyltransferase expression in mouse liver is increased in obesity- and fasting-induced steatosis.

PubMed

Xu, Jialin; Kulkarni, Supriya R; Li, Liya; Slitt, Angela L

2012-02-01

UDP-glucuronosyltransferases (Ugt) catalyze phase II conjugation reactions with glucuronic acid, which enhances chemical polarity and the elimination from the body. Few studies have addressed whether Ugt expression and activity are affected by liver disease, such as steatosis. The purpose of this study was to determine whether steatosis induced by obesity or fasting could affect liver Ugt mRNA expression and activity. Male C57BL/6J and Lep(ob/ob) (ob/ob) mice were fed ad libitum or food was withheld for 24 h. In steatotic livers of ob/ob mice, Ugt1a1, -1a6, -1a9, -2a3, -3a1, and -3a2 mRNA expression increased. Fasting, which also induced steatosis, increased hepatic Ugt1a1, -1a6, -1a7, -1a9, -2b1, -2b5, -2a3, -3a1, and -3a2 mRNA expression in mouse liver. Likewise, acetaminophen glucuronidation increased by 47% in hepatic microsomes from ob/ob mice compared with that in C57BL/6J mice, but not after fasting. In both steatosis models, Ugt induction was accompanied by increased aryl hydrocarbon receptor, constitutive androstane receptor (CAR), peroxisome proliferator-activated receptor (PPAR)-α, pregnane X receptor, nuclear factor (erythroid-derived 2)-like 2 (Nrf2), and peroxisome proliferator-activated receptor-γ coactivator-1α mRNA expression. In addition, fasting increased CAR, PPAR, and Nrf2 binding activity. The work points to hepatic triglyceride concentrations corresponding with nuclear receptor and Ugt expression. The findings indicate that steatosis significantly alters hepatic Ugt expression and activity, which could have a significant impact on determining circulating hormone levels, drug efficacy, and environmental chemical clearance.

Increased expression of placental growth factor in high-grade endometrial carcinoma

PubMed Central

COENEGRACHTS, LIEVE; SCHRAUWEN, STEFANIE; VAN BREE, RITA; DESPIERRE, EVELYN; LUYTEN, CATHERINE; JONCKX, BART; STASSEN, JEAN MARIE; VERGOTE, IGNACE; AMANT, FRÉDÉRIC

2013-01-01

Placental growth factor (PlGF), a homolog of vascular endothelial growth factor (VEGF), exerts pleiotropic functions in cancer by affecting tumor cells as well as endothelial and inflammatory cells. Moreover, PlGF expression correlates with tumor stage, recurrence, metastasis and patient outcome in different types of cancer. Recently, administration of anti-PlGF therapy reduced tumor growth and metastasis in preclinical tumor models. In the present study, we evaluated the diagnostic and prognostic value of systemic and local expression of PlGF in primary endometrial carcinomas. PlGF levels in tumor lysates (n=128) and serum (n=88) of patients with primary endometrial cancer were determined using ELISA. PlGF mRNA expression in endometrial carcinoma tissues was quantified by quantitative qRT-PCR. Results were compared to endometrial cancer stage and grade. Systemic PlGF levels were not altered in patients with endometrial cancer (FIGO stage I-II-III) as compared to healthy controls. Only in FIGO stage IV patients, serum PlGF levels were slightly increased. Local PlGF mRNA and protein expression in endometrial tumors progressively increased with tumor grade. In endometrioid carcinomas, PlGF mRNA expression was significantly increased in endometrioid grade 3 tumors as compared to normal endometrial tissue. PlGF protein expression was significantly increased in endometrioid grade 2 and 3 carcinomas and in serous carcinomas as compared to normal endometrial tissue. Our study showed that systemic/serum PlGF levels cannot be used as a diagnostic or prognostic marker in endometrial cancer. However, the increased local expression of PlGF, primarily in high-grade carcinomas, underscores the possibility for preclinical assessment of anti-PlGF therapy in endometrial cancer. PMID:23232836

Increased expression of placental growth factor in high-grade endometrial carcinoma.

PubMed

Coenegrachts, Lieve; Schrauwen, Stefanie; Van Bree, Rita; Despierre, Evelyn; Luyten, Catherine; Jonckx, Bart; Stassen, Jean Marie; Vergote, Ignace; Amant, Frédéric

2013-02-01

Placental growth factor (PlGF), a homolog of vascular endothelial growth factor (VEGF), exerts pleiotropic functions in cancer by affecting tumor cells as well as endothelial and inflammatory cells. Moreover, PlGF expression correlates with tumor stage, recurrence, metastasis and patient outcome in different types of cancer. Recently, administration of anti-PlGF therapy reduced tumor growth and metastasis in preclinical tumor models. In the present study, we evaluated the diagnostic and prognostic value of systemic and local expression of PlGF in primary endometrial carcinomas. PlGF levels in tumor lysates (n=128) and serum (n=88) of patients with primary endometrial cancer were determined using ELISA. PlGF mRNA expression in endometrial carcinoma tissues was quantified by quantitative qRT-PCR. Results were compared to endometrial cancer stage and grade. Systemic PlGF levels were not altered in patients with endometrial cancer (FIGO stage I-II-III) as compared to healthy controls. Only in FIGO stage IV patients, serum PlGF levels were slightly increased. Local PlGF mRNA and protein expression in endometrial tumors progressively increased with tumor grade. In endometrioid carcinomas, PlGF mRNA expression was significantly increased in endometrioid grade 3 tumors as compared to normal endometrial tissue. PlGF protein expression was significantly increased in endometrioid grade 2 and 3 carcinomas and in serous carcinomas as compared to normal endometrial tissue. Our study showed that systemic/serum PlGF levels cannot be used as a diagnostic or prognostic marker in endometrial cancer. However, the increased local expression of PlGF, primarily in high-grade carcinomas, underscores the possibility for preclinical assessment of anti-PlGF therapy in endometrial cancer.

The 18 kDa Translocator Protein (Peripheral Benzodiazepine Receptor) Expression in the Bone of Normal, Osteoprotegerin or Low Calcium Diet Treated Mice

PubMed Central

Kam, Winnie Wai-Ying; Meikle, Steven R.; Dunstan, Colin R.; Banati, Richard B.

2012-01-01

The presence of the translocator protein (TSPO), previously named as the mitochondrial or peripheral benzodiazepine receptor, in bone cells was studied in vitro and in situ using RT-qPCR, and receptor autoradiography using the selective TSPO ligand PK11195. In vitro, the TSPO is highly expressed in osteoblastic and osteoclastic cells. In situ, constitutive expression of TSPO is found in bone marrow and trabecular bone, e.g., spongiosa. Mice with a reduction of bone turnover induced by a 4-day treatment of osteoprotegerin reduces [3H]PK11195 binding in the spongiosa (320±128 Bq.mg−1, 499±106 Bq.mg−1 in saline-treated controls). In contrast, mice with an increase in bone turnover caused by a 4-day low calcium diet increases [3H]PK11195 binding in the spongiosa (615±90 Bq.mg−1). Further, our study includes technical feasibility data on [18F]fluoride microPET imaging of rodent bone with altered turnover. Despite [18F]fluoride having high uptake, the in vivo signal differences were small. Using a phantom model, we describe the spillover effect and partial volume loss that affect the quantitative microPET imaging of the small bone structures in experimental mouse models. In summary, we demonstrate the expression of TSPO in small rodent bone tissues, including osteoblasts and osteoclasts. A trend increase in TSPO expression was observed in the spongiosa from low to high bone turnover conditions. However, despite the potential utility of TSPO expression as an in vivo biomarker of bone turnover in experimental rodent models, our small animal PET imaging data using [18F]fluoride show that even under the condition of a good biological signal-to-noise ratio and high tracer uptake, the currently achievable instrument sensitivity and spatial resolution is unlikely to be sufficient to detect subtle differences in small structures, such as mouse bone. PMID:22295097

Urocortin1-induced anorexia is regulated by activation of the serotonin 2C receptor in the brain.

PubMed

Harada, Yumi; Takayama, Kiyoshige; Ro, Shoki; Ochiai, Mitsuko; Noguchi, Masamichi; Iizuka, Seiichi; Hattori, Tomohisa; Yakabi, Koji

2014-01-01

This study was conducted to determine the mechanisms by which serotonin (5-hydroxytryptamine, 5-HT) receptors are involved in the suppression of food intake in a rat stress model and to observe the degree of activation in the areas of the brain involved in feeding. In the stress model, male Sprague-Dawley rats (8 weeks old) were given intracerebroventricular injections of urocortin (UCN) 1. To determine the role of the 5-HT2c receptor (5-HT2cR) in the decreased food intake in UCN1-treated rats, specific 5-HT2cR or 5-HT2b receptor (5-HT2bR) antagonists were administered. Food intake was markedly reduced in UCN1-injected rats compared with phosphate buffered saline treated control rats. Intraperitoneal administration of a 5-HT2cR antagonist, but not a 5-HT2bR antagonist, significantly inhibited the decreased food intake. To assess the involvement of neural activation, we tracked the expression of c-fos mRNA as a neuronal activation marker. Expression of the c-fos mRNA in the arcuate nucleus, ventromedial hypothalamic nucleus (VMH) and rostral ventrolateral medulla (RVLM) in UNC1-injected rats showed significantly higher expression than in the PBS-injected rats. Increased c-fos mRNA was also observed in the paraventricular nucleus (PVN), the nucleus of the solitary tract (NTS), and the amygdala (AMG) after injection of UCN1. Increased 5-HT2cR protein expression was also observed in several areas. However, increased coexpression of 5-HT2cR and c-fos was observed in the PVN, VMH, NTS, RVLM and AMG. Whereas, pro-opiomelanocortin mRNA expression was not changed. In an UNC1-induced stress model, 5-HT2cR expression and activation was found in brain areas involved in feeding control. Copyright © 2013 Elsevier Inc. All rights reserved.

Neuronal injury-induced expression and release of apolipoprotein E in mixed neuron/glia co-cultures: nuclear factor kappaB inhibitors reduce basal and lesion-induced secretion of apolipoprotein E.

PubMed

Petegnief, V; Saura, J; de Gregorio-Rocasolano, N; Paul, S M

2001-01-01

In order to better delineate the intracellular signaling pathways underlying glial apolipoprotein E (apoE) expression and release, we have characterized an in vitro model of induction of glial apoE production induced by neuronal death. Exposure of mixed fetal cortical neuron/glia co-cultures to the neurotoxin N-methyl-D-aspartate results in increased apoE expression and release in a time- and concentration-dependent manner. Increased expression of apoE messenger RNA precedes the increase in intracellular apoE, followed by accumulation of the holoprotein in the culture medium. Neuronal injury induced by N-methyl-D-aspartate is accompanied by a reactive astrogliosis as measured by an increase in glial fibrillary acidic protein messenger RNA and protein at 48 and 72h post-lesion, respectively. A similar microgliosis was observed using the microglial marker ED-1. Neuronal injury-induced glial apoE secretion is attenuated by the nuclear factor kappaB inhibitors, aspirin, Bay 11-7082 and MG-132, suggesting that this transcription factor is involved in both constitutive and induced glial apoE expression. The present data show that up-regulation of apoE is an early event in the glial activation triggered by neurodegeneration in vitro and that activation of nuclear factor kappaB directly or indirectly mediates the increase in apoE expression.

Biological Effects Induced by Specific Advanced Glycation End Products in the Reconstructed Skin Model of Aging

PubMed Central

Pageon, Hervé; Zucchi, Hélène; Dai, Zhenyu; Sell, David R.; Strauch, Christopher M.; Monnier, Vincent M.; Asselineau, Daniel

2015-01-01

Abstract Advanced glycation end products (AGEs) accumulate in the aging skin. To understand the biological effects of individual AGEs, skin reconstructed with collagen selectively enriched with Nɛ-(carboxymethyl)-lysine (CML), Nɛ-(carboxyethyl)-lysine (CEL), methylglyoxal hydroimidazolone (MG-H1), or pentosidine was studied. Immunohistochemistry revealed increased expression of α6 integrin at the dermal epidermal junction by CEL and CML (p<0.01). Laminin 5 was diminished by CEL and MG-H1 (p<0.05). Both CML and CEL induced a robust increase (p<0.01) in procollagen I. In the culture medium, IL-6, VEGF, and MMP1 secretion were significantly decreased (p<0.05) by MG-H1. While both CEL and CML decreased MMP3, only CEL decreased IL-6 and TIMP1, while CML stimulated TIMP1 synthesis significantly (p<0.05). mRNA expression studies using qPCR in the epidermis layer showed that CEL increased type 7 collagen (COL7A1), β1, and α6 integrin, while CML increased only COL7A1 (p<0.05). MG-H1-modified collagen had no effect. Importantly, in the dermis layer, MMP3 mRNA expression was increased by both CML and MG-H1. CML also significantly increased the mRNAs of MMP1, TIMP1, keratinocyte growth factor (KGF), IL-6, and monocyte chemoattractant protein 1 (MCP1) (p<0.05). Mixed effects were present in CEL-rich matrix. Minimally glycoxidized pentosidine-rich collagen suppressed most mRNAs of the genes studied (p<0.05) and decreased VEGF and increased MCP1 protein expression. Taken together, this model of the aging skin suggests that a combination of AGEs tends to counterbalance and thus minimizes the detrimental biological effects of individual AGEs. PMID:26309782

Loss of TIMP3 underlies diabetic nephropathy via FoxO1/STAT1 interplay.

PubMed

Fiorentino, Loredana; Cavalera, Michele; Menini, Stefano; Marchetti, Valentina; Mavilio, Maria; Fabrizi, Marta; Conserva, Francesca; Casagrande, Viviana; Menghini, Rossella; Pontrelli, Paola; Arisi, Ivan; D'Onofrio, Mara; Lauro, Davide; Khokha, Rama; Accili, Domenico; Pugliese, Giuseppe; Gesualdo, Loreto; Lauro, Renato; Federici, Massimo

2013-03-01

ADAM17 and its inhibitor TIMP3 are involved in nephropathy, but their role in diabetic kidney disease (DKD) is unclear. Diabetic Timp3(-/-) mice showed increased albuminuria, increased membrane thickness and mesangial expansion. Microarray profiling uncovered a significant reduction of Foxo1 expression in diabetic Timp3(-/-) mice compared to WT, along with FoxO1 target genes involved in autophagy, while STAT1, a repressor of FoxO1 transcription, was increased. Re-expression of Timp3 in Timp3(-/-) mesangial cells rescued the expression of Foxo1 and its targets, and decreased STAT1 expression to control levels; abolishing STAT1 expression led to a rescue of FoxO1, evoking a role of STAT1 in linking Timp3 deficiency to FoxO1. Studies on kidney biopsies from patients with diabetic nephropathy confirmed a significant reduction in TIMP3, FoxO1 and FoxO1 target genes involved in autophagy compared to controls, while STAT1 expression was strongly increased. Our study suggests that loss of TIMP3 is a hallmark of DKD in human and mouse models and designates TIMP3 as a new possible therapeutic target for diabetic nephropathy. Copyright © 2013 The Authors. Published by John Wiley and Sons, Ltd on behalf of EMBO.

Deficiency of the protein-tyrosine phosphatase DEP-1/PTPRJ promotes matrix metalloproteinase-9 expression in meningioma cells.

PubMed

Petermann, Astrid; Stampnik, Yvonn; Cui, Yan; Morrison, Helen; Pachow, Doreen; Kliese, Nadine; Mawrin, Christian; Böhmer, Frank-D

2015-05-01

Brain-invasive growth of a subset of meningiomas is associated with less favorable prognosis. The molecular mechanisms causing invasiveness are only partially understood, however, the expression of matrix metalloproteinases (MMPs) has been identified as a contributing factor. We have previously found that loss of density enhanced phosphatase-1 (DEP-1, also designated PTPRJ), a transmembrane protein-tyrosine phosphatase, promotes meningioma cell motility and invasive growth in an orthotopic xenotransplantation model. We have now analyzed potential alterations of the expression of genes involved in motility control, caused by DEP-1 loss in meningioma cell lines. DEP-1 depleted cells exhibited increased expression of mRNA encoding MMP-9, and the growth factors EGF and FGF-2. The increase of MMP-9 expression in DEP-1 depleted cells was also readily detectable at the protein level by zymography. MMP-9 upregulation was sensitive to chemical inhibitors of growth factor signal transduction. Conversely, MMP-9 mRNA levels could be stimulated with growth factors (e.g. EGF) and inflammatory cytokines (e.g. TNFα). Increase of MMP-9 expression by DEP-1 depletion, or growth factor/cytokine stimulation qualitatively correlated with increased invasiveness in vitro scored as transmigration through matrigel-coated membranes. The studies suggest induction of MMP-9 expression promoted by DEP-1 deficiency, or potentially by growth factors and inflammatory cytokines, as a mechanism contributing to meningioma brain invasiveness.

Repetitive ischemia increases myocardial dimethylarginine dimethylaminohydrolase 1 expression.

PubMed

Zhang, Ping; Fassett, John T; Zhu, Guangshuo; Li, Jingxin; Hu, Xinli; Xu, Xin; Chen, Yingjie; Bache, Robert J

2017-06-01

Pharmacologic inhibition of nitric oxide production inhibits growth of coronary collateral vessels. Dimethylarginine dimethylaminohydrolase 1 (DDAH1) is the major enzyme that degrades asymmetric dimethylarginine (ADMA), a potent inhibitor of nitric oxide synthase. Here we examined regulation of the ADMA-DDAH1 pathway in a canine model of recurrent myocardial ischemia during the time when coronary collateral growth is known to occur. Under basal conditions, DDAH1 expression was non-uniform across the left ventricular (LV) wall, with expression strongest in the subepicardium. In response to ischemia, DDAH1 expression was up-regulated in the midmyocardium of the ischemic zone, and this was associated with a significant reduction in myocardial interstitial fluid (MIF) ADMA. The decrease in MIF ADMA during ischemia was likely due to increased DDAH1 because myocardial protein arginine N-methyl transferase 1 (PRMT1) and the methylated arginine protein content (the source of ADMA) were unchanged or increased, respectively, at this time. The inflammatory mediators interleukin (IL-1β) and tumor necrosis factor (TNF-α) were also elevated in the midmyocardium where DDAH1 expression was increased. Both of these factors significantly up-regulated DDAH1 expression in cultured human coronary artery endothelial cells. Taken together, these results suggest that inflammatory factors expressed in response to myocardial ischemia contributed to up-regulation of DDAH1, which was responsible for the decrease in MIF ADMA.

The expression of CXCR4 is induced by the luteinizing hormone surge and mediated by progesterone receptors in human preovulatory granulosa cells.

PubMed

Choi, Yohan; Park, Ji Yeon; Wilson, Kalin; Rosewell, Katherine L; Brännström, Mats; Akin, James W; Curry, Thomas E; Jo, Misung

2017-06-01

The chemokine CXC motif ligand 12 (CXCL12) and its cognate receptor, CXCR4, have been implicated in the ovulatory process in various animal models. However, little is known about the expression and regulation of CXCL12 and CXCR4 and their functions during the ovulatory period in the human ovary. In this study, we characterized the expression patterns of CXCL12 and CXCR4 in preovulatory follicles collected before the luteinizing hormone (LH) surge and at defined hours after hCG administration in women with the regular menstrual cycle. The levels of mRNA and protein for CXCR4 were increased in granulosa cells of late ovulatory follicles, whereas CXCL12 expression was constant in follicles throughout the ovulatory period. Both CXCR4 and CXCL12 were localized to a subset of leukocytes around and inside the vasculature of human preovulatory follicles. Using a human granulosa cell culture model, the regulatory mechanisms and functions of CXCL12 and CXCR4 expression were investigated. Human chorionic gonadotropin (hCG) stimulated CXCR4 expression, whereas CXCL12 expression was not affected, mimicking in vivo expression patterns. Both RU486 (progesterone receptor antagonist) and CoCl2 (HIFs activator) blocked the hCG-induced increase in CXCR4 expression, whereas AG1478 (EGFR inhibitor) had no effect. The treatment with CXCL12 had no effect on granulosa cell viability but decreased hCG-stimulated CXCR4 expression. © The Authors 2017. Published by Oxford University Press on behalf of Society for the Study of Reproduction. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

CRE-Mediated Transcription and COX-2 Expression in the Pilocarpine Model of Status Epilepticus

PubMed Central

Lee, Boyoung; Dziema, Heather; Lee, Kyu Hyun; Choi, Yun-Sik; Obrietan, Karl

2007-01-01

Status epilepticus (SE) triggers neuronal death, reactive gliosis and remodeling of synaptic circuitry, thus leading to profound pathological alterations in CNS physiology. These processes are, in part, regulated by the rapid upregulation of both cytotoxic and cytoprotective genes. One pathway that may couple SE to transcriptionally-dependent alterations in CNS physiology is the CREB (cAMP response element-binding protein)/CRE (cAMP response element) cascade. Here, we utilized the pilocarpine model of SE on a mouse strain transgenic for a CRE-reporter construct (β-galactosidase) to begin to characterize how seizure activity regulates the activation state of the CREB/CRE pathway in both glia and neurons of the hippocampus. SE triggered a rapid (4–8 hrs post SE) but transient increase in CRE-mediated gene expression in the neuronal sublayers. In contrast to neurons, SE induced a lasting increase (up to 20 days) in CRE-mediated transcription in both reactive astrocytes and microglia. CRE-mediated gene expression correlated with expression of the pro-inflammatory enzyme cyclooxygenase-2 (COX-2). To examine the role of CREB in SE-induced COX-2 expression, we generated a transgenic mouse strain that expresses A-CREB, a potent repressor of CREB-dependent transcription. In these animals, the capacity of SE to stimulate COX-2 expression was markedly attenuated, indicating that CREB is a key intermediate in SE-induced COX-2 expression. Collectively these data show that SE triggers two waves of CREB-mediated gene expression, a transient wave in neurons and a long-lasting wave in reactive glial cells, and that CREB couples SE to COX-2 expression. PMID:17029965

Effects of hypoxic preconditioning on expression of transcription factor NGFI-A in the rat brain after unavoidable stress in the "learned helplessness" model.

PubMed

Baranova, K A; Rybnikova, E A; Mironova, V I; Samoilov, M O

2010-07-01

We report here our immunocytochemical studies establishing that the development of a depression-like state in rats following unavoidable stress in a "learned helplessness" model is accompanied by stable activation of the expression of transcription factor NGFI-A in the dorsal hippocampus (field CA1) and the magnocellular paraventricular nucleus of the hypothalamus, along with an early wave of post-stress expression, which died down rapidly, in the ventral hippocampus (the dentate gyrus) and a long period of up to five days of high-level expression in the neocortex. In rats subjected to three sessions of preconditioning consisting of moderate hypobaric hypoxia (360 mmHg, 2 h, with intervals of 24 h), which did not form depression in these circumstances, there were significant changes in the dynamics of immunoreactive protein content in the hippocampus, with a stable increase in expression in the ventral hippocampus and only transient and delayed (by five days) expression in field CA1. In the neocortex (layer II), preconditioning eliminated the effects of stress, preventing prolongation of the first wave of NGFI-A expression to five days, while in the magnocellular hypothalamus, conversely, preconditioning stimulated a second (delayed) wave of the expression of this transcription factor. The pattern of NGFI-A expression in the hippocampus, neocortex, and hypothalamus seen in non-preconditioned rats appears to reflect the pathological reaction to aversive stress, which, rather than adaptation, produced depressive disorders. Post-stress modification of the expression of the product of the early gene NGFI-A in the brain induced by hypoxic preconditioning probably plays an important role in increased tolerance to severe psychoemotional stresses and is an important component of antidepressant mechanisms.

Disruption of DNA methylation-dependent long gene repression in Rett syndrome

PubMed Central

Gabel, Harrison W.; Kinde, Benyam Z.; Stroud, Hume; Gilbert, Caitlin S.; Harmin, David A.; Kastan, Nathaniel R.; Hemberg, Martin; Ebert, Daniel H.; Greenberg, Michael E.

2015-01-01

Disruption of the MECP2 gene leads to Rett syndrome (RTT), a severe neurological disorder with features of autism1. MECP2 encodes a methyl-DNA-binding protein2 that has been proposed to function as a transcriptional repressor, but despite numerous studies examining neuronal gene expression in Mecp2 mutants, no clear model has emerged for how MeCP2 regulates transcription3–9. Here we identify a genome-wide length-dependent increase in gene expression in MeCP2 mutant mouse models and human RTT brains. We present evidence that MeCP2 represses gene expression by binding to methylated CA sites within long genes, and that in neurons lacking MeCP2, decreasing the expression of long genes attenuates RTT-associated cellular deficits. In addition, we find that long genes as a population are enriched for neuronal functions and selectively expressed in the brain. These findings suggest that mutations in MeCP2 may cause neurological dysfunction by specifically disrupting long gene expression in the brain. PMID:25762136

Milk with and without lactoferrin can influence intestinal damage in a pig model of malnutrition.

PubMed

Garas, Lydia C; Feltrin, Cristiano; Hamilton, M Kristina; Hagey, Jill V; Murray, James D; Bertolini, Luciana R; Bertolini, Marcelo; Raybould, Helen E; Maga, Elizabeth A

2016-02-01

Malnutrition remains a leading contributor to the morbidity and mortality of children under the age of five worldwide. However, the underlying mechanisms are not well understood necessitating an appropriate animal model to answer fundamental questions and conduct translational research into optimal interventions. One potential intervention is milk from livestock that more closely mimics human milk by increased levels of bioactive components that can promote a healthy intestinal epithelium. We tested the ability of cow milk and milk from transgenic cows expressing human lactoferrin at levels found in human milk (hLF milk) to mitigate the effects of malnutrition at the level of the intestine in a pig model of malnutrition. Weaned pigs (3 weeks old) were fed a protein and calorie restricted diet for five weeks, receiving cow, hLF or no milk supplementation daily from weeks 3-5. After three weeks, the restricted diet induced changes in growth, blood chemistry and intestinal structure including villous atrophy, increased ex vivo permeability and decreased expression of tight junction proteins. Addition of both cow and hLF milk to the diet increased growth rate and calcium and glucose levels while promoting growth of the intestinal epithelium. In the jejunum hLF milk restored intestinal morphology, reduced permeability and increased expression of anti-inflammatory IL-10. Overall, this pig model of malnutrition mimics salient aspects of the human condition and demonstrates that cow milk can stimulate the repair of damage to the intestinal epithelium caused by protein and calorie restriction with hLF milk improving this recovery to a greater extent.

Selective Deletion of the Brain-Specific Isoform of Renin Causes Neurogenic Hypertension.

PubMed

Shinohara, Keisuke; Liu, Xuebo; Morgan, Donald A; Davis, Deborah R; Sequeira-Lopez, Maria Luisa S; Cassell, Martin D; Grobe, Justin L; Rahmouni, Kamal; Sigmund, Curt D

2016-12-01

The renin-angiotensin system (RAS) in the brain is a critical determinant of blood pressure, but the mechanisms regulating RAS activity in the brain remain unclear. Expression of brain renin (renin-b) occurs from an alternative promoter-first exon. The predicted translation product is a nonsecreted enzymatically active renin whose function is unknown. We generated a unique mouse model by selectively ablating the brain-specific isoform of renin (renin-b) while preserving the expression and function of the classical isoform expressed in the kidney (renin-a). Preservation of renal renin was confirmed by measurements of renin gene expression and immunohistochemistry. Surprisingly, renin-b-deficient mice exhibited hypertension, increased sympathetic nerve activity to the kidney and heart, and impaired baroreflex sensitivity. Whereas these mice displayed decreased circulating RAS activity, there was a paradoxical increase in brain RAS activity. Physiologically, renin-b-deficient mice exhibited an exaggerated depressor response to intracerebroventricular administration of losartan, captopril, or aliskiren. At the molecular level, renin-b-deficient mice exhibited increased expression of angiotensin-II type 1 receptor in the paraventricular nucleus, which correlated with an increased renal sympathetic nerve response to leptin, which was dependent on angiotensin-II type 1 receptor activity. Interestingly, despite an ablation of renin-b expression, expression of renin-a was significantly increased in rostral ventrolateral medulla. These data support a new paradigm for the genetic control of RAS activity in the brain by a coordinated regulation of the renin isoforms, with expression of renin-b tonically inhibiting expression of renin-a under baseline conditions. Impairment of this control mechanism causes neurogenic hypertension. © 2016 American Heart Association, Inc.

Abstract knowledge versus direct experience in processing of binomial expressions

PubMed Central

Morgan, Emily; Levy, Roger

2016-01-01

We ask whether word order preferences for binomial expressions of the form A and B (e.g. bread and butter) are driven by abstract linguistic knowledge of ordering constraints referencing the semantic, phonological, and lexical properties of the constituent words, or by prior direct experience with the specific items in questions. Using forced-choice and self-paced reading tasks, we demonstrate that online processing of never-before-seen binomials is influenced by abstract knowledge of ordering constraints, which we estimate with a probabilistic model. In contrast, online processing of highly frequent binomials is primarily driven by direct experience, which we estimate from corpus frequency counts. We propose a trade-off wherein processing of novel expressions relies upon abstract knowledge, while reliance upon direct experience increases with increased exposure to an expression. Our findings support theories of language processing in which both compositional generation and direct, holistic reuse of multi-word expressions play crucial roles. PMID:27776281

Activation of UCPs gene expression in skeletal muscle can be independent on both circulating fatty acids and food intake. Involvement of ROS in a model of mouse cancer cachexia.

PubMed

Busquets, Sílvia; Almendro, Vanessa; Barreiro, Esther; Figueras, Maite; Argilés, Josep M; López-Soriano, Francisco J

2005-01-31

Implantation of a fast growing tumour to mice (Lewis lung carcinoma) resulted in a clear cachectic state characterized by a profound muscle wasting. This was accompanied by a significant increase in both UCP2 and UCP3 gene expression in skeletal muscle and heart. Interestingly, this increase in gene expression was not linked to a rise in circulating fatty acids or in a decrease in food intake, as previously reported in other pathophysiological states. These results question the concept that hyperlipaemia is the only factor controlling UCP gene expression in different pathophysiological conditions. In addition, the present work suggests that UCPs might participate in a counter-regulatory mechanism to lower the production of ROS.

Experimental and Human Evidence for Lipocalin-2 (Neutrophil Gelatinase-Associated Lipocalin [NGAL]) in the Development of Cardiac Hypertrophy and heart failure.

PubMed

Marques, Francine Z; Prestes, Priscilla R; Byars, Sean G; Ritchie, Scott C; Würtz, Peter; Patel, Sheila K; Booth, Scott A; Rana, Indrajeetsinh; Minoda, Yosuke; Berzins, Stuart P; Curl, Claire L; Bell, James R; Wai, Bryan; Srivastava, Piyush M; Kangas, Antti J; Soininen, Pasi; Ruohonen, Saku; Kähönen, Mika; Lehtimäki, Terho; Raitoharju, Emma; Havulinna, Aki; Perola, Markus; Raitakari, Olli; Salomaa, Veikko; Ala-Korpela, Mika; Kettunen, Johannes; McGlynn, Maree; Kelly, Jason; Wlodek, Mary E; Lewandowski, Paul A; Delbridge, Lea M; Burrell, Louise M; Inouye, Michael; Harrap, Stephen B; Charchar, Fadi J

2017-06-14

Cardiac hypertrophy increases the risk of developing heart failure and cardiovascular death. The neutrophil inflammatory protein, lipocalin-2 (LCN2/NGAL), is elevated in certain forms of cardiac hypertrophy and acute heart failure. However, a specific role for LCN2 in predisposition and etiology of hypertrophy and the relevant genetic determinants are unclear. Here, we defined the role of LCN2 in concentric cardiac hypertrophy in terms of pathophysiology, inflammatory expression networks, and genomic determinants. We used 3 experimental models: a polygenic model of cardiac hypertrophy and heart failure, a model of intrauterine growth restriction and Lcn2 -knockout mouse; cultured cardiomyocytes; and 2 human cohorts: 114 type 2 diabetes mellitus patients and 2064 healthy subjects of the YFS (Young Finns Study). In hypertrophic heart rats, cardiac and circulating Lcn2 was significantly overexpressed before, during, and after development of cardiac hypertrophy and heart failure. Lcn2 expression was increased in hypertrophic hearts in a model of intrauterine growth restriction, whereas Lcn2 -knockout mice had smaller hearts. In cultured cardiomyocytes, Lcn2 activated molecular hypertrophic pathways and increased cell size, but reduced proliferation and cell numbers. Increased LCN2 was associated with cardiac hypertrophy and diastolic dysfunction in diabetes mellitus. In the YFS, LCN2 expression was associated with body mass index and cardiac mass and with levels of inflammatory markers. The single-nucleotide polymorphism, rs13297295, located near LCN2 defined a significant cis -eQTL for LCN2 expression. Direct effects of LCN2 on cardiomyocyte size and number and the consistent associations in experimental and human analyses reveal a central role for LCN2 in the ontogeny of cardiac hypertrophy and heart failure. © 2017 The Authors. Published on behalf of the American Heart Association, Inc., by Wiley.

Fatty Acid Oxidation Changes and the Correlation with Oxidative Stress in Different Preeclampsia-Like Mouse Models

PubMed Central

Ding, Xiaoyan; Yang, Zi; Han, Yiwei; Yu, Huan

2014-01-01

Background Long-chain 3-hydroxyacyl-CoA dehydrogenase (LCHAD) expression is decreased in placenta of some cases of preeclampsia (PE) which may result in free fatty acid (FFA) increased. High FFA level will induce oxidative stress, so abnormal long-chain fatty acid-oxidation may participate in the pathogenesis of PE through oxidative stress pathway. Methods PE-like groups were ApoC3 transgenic mice with abnormal fatty acid metabolism, classical PE-like models with injection of Nw-nitro-L-arginine-methyl ester (L-NA) or lipopolysaccharide (LPS) and the antiphospholipid syndrome (APS) mouse model with β2GPI injection (ApoC3+NS, ApoC3+L-NA, L-NA, LPS and β2GPI groups). The control group was wild-type mice with normal saline injection. Except for β2GPI mice, the other mice were subdivided into pre-implantation (Pre) and mid-pregnancy (Mid) subgroups by injection time. Results All PE-like groups showed hypertension and proteinuria except ApoC3+NS mice only showed hypertension. Serum FFA levels increased significantly except in LPS group compared to controls (P<0.05). LCHAD mRNA and protein expression in the liver and placenta was significantly higher for ApoC3+NS, ApoC3+L-NA and β2GPI mice and lower for L-NA mice than controls (P<0.05) but did not differ between LPS mice and controls. P47phox mRNA and protein expression in the liver significantly increased in all PE-like groups except LPS group, while P47phox expression in the placenta only significantly increased in L-NA and β2GPI groups. Conclusions Abnormal long-chain fatty acid-oxidation may play a different role in different PE-like models and in some cases participate in the pathogenesis of PE through oxidative stress pathway. PMID:25302499

Lamellar pro-inflammatory cytokine expression patterns in laminitis at the developmental stage and at the onset of lameness: innate vs. adaptive immune response.

PubMed

Belknap, J K; Giguère, S; Pettigrew, A; Cochran, A M; Van Eps, A W; Pollitt, C C

2007-01-01

Recent research has indicated that inflammation plays a role in the early stages of laminitis and that, similar to organ failure in human sepsis, early inflammatory mechanisms may lead to downstream events resulting in lamellar failure. Characterisation of the type of immune response (i.e. innate vs. adaptive) is essential in order to develop therapeutic strategies to counteract these deleterious events. To quantitate gene expression of pro-inflammatory cytokines known to be important in the innate and adaptive immune response during the early stages of laminitis, using both the black walnut extract (BWE) and oligofructose (OF) models of laminitis. Real-time qPCR was used to assess lamellar mRNA expression of interleukins-1beta, 2, 4, 6, 8, 10, 12 and 18, and tumour necrosis factor alpha and interferon gamma at the developmental stage and at the onset of lameness. Significantly increased lamellar mRNA expression of cytokines important in the innate immune response were present at the developmental stage of the BWE model, and at the onset of acute lameness in both the BWE model and OF model. Of the cytokines characteristic of the Th1 and Th2 arms of the adaptive immune response, a mixed response was noted at the onset of acute lameness in the BWE model, whereas the response was skewed towards a Th1 response at the onset of lameness in the OF model. Lamellar inflammation is characterised by strong innate immune response in the developmental stages of laminitis; and a mixture of innate and adaptive immune responses at the onset of lameness. These results indicate that anti-inflammatory treatment of early stage laminitis (and the horse at risk of laminitis) should include not only therapeutic drugs that address prostanoid activity, but should also address the marked increases in lamellar cytokine expression.

Role of fractalkine/CX3CR1 signaling pathway in the recovery of neurological function after early ischemic stroke in a rat model.

PubMed

Liu, Yan-Zhi; Wang, Chun; Wang, Qian; Lin, Yong-Zhong; Ge, Yu-Song; Li, Dong-Mei; Mao, Geng-Sheng

2017-09-01

This study aims to explore the role of fractalkine/CX3C chemokine receptor 1 (CX3CR1) signaling pathway in the recovery of neurological functioning after an early ischemic stroke in rats. After establishment of permanent middle cerebral artery occlusion (pMCAO) models, 50 rats were divided into blank, sham, model, positive control and CX3CR1 inhibitor groups. Neurological impairment, walking and grip abilities, and cortical and hippocampal infarctions were evaluated by Zea Longa scoring criterion, beam-walking assay and grip strength test, and diffusion-weighted magnetic resonance imaging. qRT-PCR and Western blotting were performed to detect mRNA and protein expressions. ELISA was conducted to measure concentration of sFractalkine (sFkn), interleukin-1β (IL-1β) and TNF-α. The recovery rate of neurological functioning impairment and reduced walking and grip abilities was faster in the positive control and CX3CR1 inhibitor groups than the model group. The model, positive control and CX3CR1 inhibitor groups showed increased mRNA and protein expression of chemokine C-X3-C motif ligand 1 (CX3CL1) and CX3CR1, concentration of sFkn, IL-1β and TNF-α, and size of cortical and cerebral infarctions while decreased expression of NGF and BDNF compared with the blank and sham groups. Compared with the model group, the mRNA and protein expression of CX3CL1 and CX3CR1, concentration of sFkn, IL-1β and TNF-α, and size of cortical and cerebral infarctions decreased while expression of NGF and BDNF increased in the positive control and CX3CR1 inhibitor groups. Thus, the study suggests that inhibition of fractalkine/CX3CR1 signaling pathway promotes the recovery of neurological functioning after the occurrence of an early ischemic stroke. Copyright © 2017 Elsevier Inc. All rights reserved.

[Effect of American Ginseng Capsule on the liver oxidative injury and the Nrf2 protein expression in rats exposed by electromagnetic radiation of frequency of cell phone].

PubMed

Luo, Ya-ping; Ma, Hui-Rong; Chen, Jing-Wei; Li, Jing-Jing; Li, Chun-xiang

2014-05-01

To observe the effect of American Ginseng Capsule (AGC) on the liver oxidative injury and the Nrf2 protein expression in the liver tissue of rats exposed by 900 MHz cell phone electromagnetic radiation. Totally 40 male SD rats were randomly divided into the normal control group, the model group, the Shuifei Jibin Capsule (SJC) group, and the AGC group,10 in each group. Rats in the normal control group were not irradiated. Rats in the rest three groups were exposed by imitated 900 MHz cellular phone for 4 h in 12 consecutive days. Meanwhile, rats in the SJC group and the AGC group were intragastrically administrated with suspension of SJC and AGC (1 mL/200 g body weight) respectively. Normal saline was administered to rats in the normal control group and the model group. The histolomorphological changes of the liver tissue were observed by HE staining. Contents of malonic dialdehyde (MDA), superoxide dismutase (SOD), glutathione (GSH), and glutathione peroxidase (GSH-PX)were detected by colorimetry. The Nrf2 protein expression of hepatocytes was detected by immunohistochemical assay and Western blot. Compared with the normal control group, hepatocyte nucleus was atrophied or partially disappeared, the contents of liver MDA and Nrf2 protein obviously increased (P <0. 05, P <0. 01); contents of liver SOD and GSH decreased (P <0. 05) in the model group. Compared with the model group, karyopyknosis was obviously attenuated and approached to the normal level in the SJC group and the AGC group. The contents of liver MDA and Nrf2 protein expression decreased (P <0. 05), and the contents of liver SOD, GSH, and GSH-PX obviously increased (P < 0.05) in the SJC group. The contents of liver MDA and the Nrf2 protein expression decreased (P < 0.05), and contents of SOD and GSH obviously increased in the AGC group (P <0.01, P <0.05). The electromagnetic radiation induced by 900 MHz cell phone could affect the expression of Nrf2 protein, induce oxidative injury, and induce abnormal morphology of liver cells. SJC and AGC could promote the morphological recovery of the liver cells. Its mechanism might be related to affecting the expression of Nrf2 protein and attenuating oxidative damage of liver cells.

Th 17 Cells and Nesfatin-1 are associated with Spontaneous Abortion in the CBA/j × DBA/2 Mouse Model

PubMed Central

Chung, Yiwa; Kim, Heejeong; Im, Eunji; Kim, Philjae; Yang, Hyunwon

2015-01-01

The pregnancy and abortion process involves a complex mechanism with various immune cells present in the implantation sites and several hormones associated with pregnancy, such as leptin, ghrelin and nesfatin-1. However, the mechanism underlying spontaneous abortion by maternal T helper 17 (Th17) present in the implantation sites and nesfatin-1, which is of anorexigenic hormones, is not fully understood so far. Therefore, the purpose of this study was to examine the possible roles of Th17 cells present in the implantation sites and nesfatin-1 expressed in the uterus on spontaneous abortion using the CBA/j × DBA/2 mouse model. Th17 transcription factor, ROR-γt mRNA expression was significantly increased in the abortion sites compared with the implantation sites of abortion model mice on day 14.5 and 19.5 of pregnancy. In addition, the expression levels of IL-17A mRNA were significantly higher in abortion sites than in implantation sites on day 14.5 and 19.5. Moreover, the nesfatin-1/NUCB2 protein and mRNA levels were increased in abortion sites compared with levels in implantation sites of both normal pregnant and abortion model mice on day 14.5 of pregnancy. Interestingly, nesfatin- 1/NUCB2 serum levels were not changed throughout the whole pregnancy in abortion model mice, but its serum level was dramatically increased on day 14.5, and then rapidly decreased on day 19.5 in normal pregnant mice. In this study, we showed for the first time the expression of nesfatin-1/NUCB2 mRNA and protein in implantation sites during pregnancy. The present results suggest that Th17 cells in the uterus may play an important role in the period of implantation and for maintenance of pregnancy. Furthermore, the present results suggest that Th17 cells in implantation sites may be a key regulator for maintenance of pregnancy and provides evidence that activation of these cells may be regulated by nesfatin-1/NUCB2. Further study is needed to elucidate the role of nesfatin-1 expressed in the uterus during pregnancy. PMID:26973976

Infection of human intestinal epithelial cells with invasive bacteria upregulates apical intercellular adhesion molecule-1 (ICAM)-1) expression and neutrophil adhesion.

PubMed Central

Huang, G T; Eckmann, L; Savidge, T C; Kagnoff, M F

1996-01-01

The acute host response to gastrointestinal infection with invasive bacteria is characterized by an accumulation of neutrophils in the lamina propria, and neutrophil transmigration to the luminal side of the crypts. Intestinal epithelial cells play an important role in the recruitment of inflammatory cells to the site of infection through the secretion of chemokines. However, little is known regarding the expression, by epithelial cells, of molecules that are involved in interactions between the epithelium and neutrophils following bacterial invasion. We report herein that expression of ICAM-1 on human colon epithelial cell lines, and on human enterocytes in an in vivo model system, is upregulated following infection with invasive bacteria. Increased ICAM-1 expression in the early period (4-9 h) after infection appeared to result mainly from a direct interaction between invaded bacteria and host epithelial cells since it co-localized to cells invaded by bacteria, and the release of soluble factors by epithelial cells played only a minor role in mediating increased ICAM-1 expression. Furthermore, ICAM-1 was expressed on the apical side of polarized intestinal epithelial cells, and increased expression was accompanied by increased neutrophil adhesion to these cells. ICAM-1 expression by intestinal epithelial cells following infection with invasive bacteria may function to maintain neutrophils that have transmigrated through the epithelium in close contact with the intestinal epithelium, thereby reducing further invasion of the mucosa by invading pathogens. PMID:8755670

Longitudinal changes in zinc transport kinetics, metallothionein, and zinc transporter expression in a blood-brain barrier model in response to a moderately excessive zinc environment$

PubMed Central

Gauthier, Nicole A.; Karki, Shakun; Olley, Bryony J.; Thomas, W. Kelly

2008-01-01

A blood-brain barrier (BBB) model composed of porcine brain capillary endothelial cells (BCEC) was exposed to a moderately excessive zinc environment (50 µmol Zn/L) in cell culture and longitudinal measurements were made of zinc transport kinetics, ZnT-1 (SLC30A1) expression, and changes in the protein concentration of metallothionein (MT), ZnT-1, ZnT-2 (SLC30A2), and Zip1 (SLC39A1). Zinc release by cells of the BBB model was significantly increased after 12–24 h of exposure, but decreased back to control levels after 48–96 h, as indicated by transport across the BBB from both the ablumenal (brain) and lumenal (blood) directions. Expression of ZnT-1, the zinc export protein, increased 169% within 12 h, but was no longer different from controls after 24 h. Likewise, ZnT-1 protein content increased transiently after 12 h of exposure but returned to control levels by 24 h. Capacity for zinc uptake and retention increased from both the lumenal and ablumenal directions within 12–24 h of exposure and remained elevated. MT and ZnT-2 were elevated within 12 h and remained elevated throughout the study. Zip1 was unchanged by the treatment. The BBB’s response to a moderately high zinc environment was dynamic and involved multiple mechanisms. The initial response was to increase the cell’s capacity to sequester zinc with additional MT and increase zinc export with the ZnT-1 protein. But, the longer term strategy involved increasing ZnT-2 transporters, presumably to sequester zinc into intracellular vesicles as a mechanism to protect the brain and maintain brain zinc homeostasis. PMID:18061429

A composite mouse model of aplastic anemia complicated with iron overload

PubMed Central

Wu, Dijiong; Wen, Xiaowen; Liu, Wenbin; Xu, Linlong; Ye, Baodong; Zhou, Yuhong

2018-01-01

Iron overload is commonly encountered during the course of aplastic anemia (AA), but no composite animal model has been developed yet, which hinders drug research. In the present study, the optimal dosage and duration of intraperitoneal iron dextran injection for the development of an iron overload model in mice were explored. A composite model of AA was successfully established on the principle of immune-mediated bone marrow failure. Liver volume, peripheral hemogram, bone marrow pathology, serum iron, serum ferritin, pathological iron deposition in multiple organs (liver, bone marrow, spleen), liver hepcidin, and bone morphogenetic protein 6 (BMP6), SMAD family member 4 (SMAD4) and transferrin receptor 2 (TfR2) mRNA expression levels were compared among the normal control, AA, iron overload and composite model groups to validate the composite model, and explore the pathogenesis and features of iron overload in this model. The results indicated marked increases in iron deposits, with significantly increased liver/body weight ratios as well as serum iron and ferritin in the iron overload and composite model groups as compared with the normal control and AA groups (P<0.05). There were marked abnormalities in iron regulation gene expression between the AA and composite model groups, as seen by the significant decrease of hepcidin expression in the liver (P<0.01) that paralleled the changes in BMP6, SMAD4, and TfR2. In summary, a composite mouse model with iron overload and AA was successfully established, and AA was indicated to possibly have a critical role in abnormal iron metabolism, which promoted the development of iron deposits. PMID:29434729

A composite mouse model of aplastic anemia complicated with iron overload.

PubMed

Wu, Dijiong; Wen, Xiaowen; Liu, Wenbin; Xu, Linlong; Ye, Baodong; Zhou, Yuhong

2018-02-01

Iron overload is commonly encountered during the course of aplastic anemia (AA), but no composite animal model has been developed yet, which hinders drug research. In the present study, the optimal dosage and duration of intraperitoneal iron dextran injection for the development of an iron overload model in mice were explored. A composite model of AA was successfully established on the principle of immune-mediated bone marrow failure. Liver volume, peripheral hemogram, bone marrow pathology, serum iron, serum ferritin, pathological iron deposition in multiple organs (liver, bone marrow, spleen), liver hepcidin, and bone morphogenetic protein 6 (BMP6), SMAD family member 4 (SMAD4) and transferrin receptor 2 (TfR2) mRNA expression levels were compared among the normal control, AA, iron overload and composite model groups to validate the composite model, and explore the pathogenesis and features of iron overload in this model. The results indicated marked increases in iron deposits, with significantly increased liver/body weight ratios as well as serum iron and ferritin in the iron overload and composite model groups as compared with the normal control and AA groups (P<0.05). There were marked abnormalities in iron regulation gene expression between the AA and composite model groups, as seen by the significant decrease of hepcidin expression in the liver (P<0.01) that paralleled the changes in BMP6, SMAD4, and TfR2. In summary, a composite mouse model with iron overload and AA was successfully established, and AA was indicated to possibly have a critical role in abnormal iron metabolism, which promoted the development of iron deposits.

Genetically engineered cardiac pacemaker: Stem cells transfected with HCN2 gene and myocytes—A model

NASA Astrophysics Data System (ADS)

Kanani, S.; Pumir, A.; Krinsky, V.

2008-01-01

One of the successfully tested methods to design genetically engineered cardiac pacemaker cells consists in transfecting a human mesenchymal stem cell (hMSC) with a HCN2 gene and connecting it to a myocyte. We develop and study a mathematical model, describing a myocyte connected to a hMSC transfected with a HCN2 gene. The cardiac action potential is described both with the simple Beeler Reuter model, as well as with the elaborate dynamic Luo Rudy model. The HCN2 channel is described by fitting electrophysiological records, in the spirit of Hodgkin Huxley. The model shows that oscillations can occur in a pair myocyte-stem cell, that was not observed in the experiments yet. The model predicted that: (1) HCN pacemaker channels can induce oscillations only if the number of expressed I channels is low enough. At too high an expression level of I channels, oscillations cannot be induced, no matter how many pacemaker channels are expressed. (2) At low expression levels of I channels, a large domain of values in the parameter space (n, N) exists, where oscillations should be observed. We denote N the number of expressed pacemaker channels in the stem cell, and n the number of gap junction channels coupling the stem cell and the myocyte. (3) The expression levels of I channels observed in ventricular myocytes, both in the Beeler Reuter and in the dynamic Luo Rudy models are too high to allow to observe oscillations. With expression levels below ˜1/4 of the original value, oscillations can be observed. The main consequence of this work is that in order to obtain oscillations in an experiment with a myocyte-stem cell pair, increasing the values of n, N is unlikely to be helpful, unless the expression level of I has been reduced enough. The model also allows us to explore levels of gene expression not yet achieved in experiments, and could be useful to plan new experiments, aimed at improving the robustness of the oscillations.

Regulation of accumulation and function of myeloid derived suppressor cells in different murine models of hepatocellular carcinoma

PubMed Central

Kapanadze, Tamar; Gamrekelashvili, Jaba; Ma, Chi; Chan, Carmen; Zhao, Fei; Hewitt, Stephen; Zender, Lars; Kapoor, Veena; Felsher, Dean W.; Manns, Michael P.; Korangy, Firouzeh; Greten, Tim F.

2013-01-01

Background and aims Myeloid derived suppressor cells (MDSC) are immature myeloid cells with immunosuppressive activity. They accumulate in tumor-bearing mice and humans with different types of cancer, including hepatocellular carcinoma (HCC). The aim of this study was to examine the biology of MDSC in murine HCC models and to identify a model, which mimics the human disease. Methods: The comparative analysis of MDSC was performed in mice, bearing transplantable, diethylnitrosoamine (DEN)-induced and MYC-expressing HCC at different ages. Results: An accumulation of MDSC was found in mice with HCC irrespectively of the model tested. Transplantable tumors rapidly induced systemic recruitment of MDSC, in contrast to slow-growing DEN-induced or MYC-expressing HCC, where MDSC numbers only increased intra-hepatically in mice with advanced tumors. MDSC derived from mice with subcutaneous tumors were more suppressive than those from mice with DEN-induced HCC. Enhanced expression of genes associated with MDSC generation (GM-CSF, VEGF, IL-6, IL-1β) and migration (MCP-1, KC, S100A8, S100A9) was observed in mice with subcutaneous tumors. In contrast, only KC levels increased in mice with DEN-induced HCC. Both KC and GM-CSF over-expression or anti-KC and anti-GM-CSF treatment controlled MDSC frequency in mice with HCC. Finally, the frequency of MDSC decreased upon successful anti-tumor treatment with sorafenib. Conclusions: Our data indicate that MDSC accumulation is a late event during hepatocarcinogenesis and differs significantly depending on the tumor model studied. PMID:23796475

A quantitative validated model reveals two phases of transcriptional regulation for the gap gene giant in Drosophila.

PubMed

Hoermann, Astrid; Cicin-Sain, Damjan; Jaeger, Johannes

2016-03-15

Understanding eukaryotic transcriptional regulation and its role in development and pattern formation is one of the big challenges in biology today. Most attempts at tackling this problem either focus on the molecular details of transcription factor binding, or aim at genome-wide prediction of expression patterns from sequence through bioinformatics and mathematical modelling. Here we bridge the gap between these two complementary approaches by providing an integrative model of cis-regulatory elements governing the expression of the gap gene giant (gt) in the blastoderm embryo of Drosophila melanogaster. We use a reverse-engineering method, where mathematical models are fit to quantitative spatio-temporal reporter gene expression data to infer the regulatory mechanisms underlying gt expression in its anterior and posterior domains. These models are validated through prediction of gene expression in mutant backgrounds. A detailed analysis of our data and models reveals that gt is regulated by domain-specific CREs at early stages, while a late element drives expression in both the anterior and the posterior domains. Initial gt expression depends exclusively on inputs from maternal factors. Later, gap gene cross-repression and gt auto-activation become increasingly important. We show that auto-regulation creates a positive feedback, which mediates the transition from early to late stages of regulation. We confirm the existence and role of gt auto-activation through targeted mutagenesis of Gt transcription factor binding sites. In summary, our analysis provides a comprehensive picture of spatio-temporal gene regulation by different interacting enhancer elements for an important developmental regulator. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

Pathogenesis pathways of idiopathic pulmonary fibrosis in bleomycin-induced lung injury model in mice.

PubMed

Shi, Keyun; Jiang, Jianzhong; Ma, Tieliang; Xie, Jing; Duan, Lirong; Chen, Ruhua; Song, Ping; Yu, Zhixin; Liu, Chao; Zhu, Qin; Zheng, Jinxu

2014-01-01

Our objective was to investigate the pathogenesis pathways of idiopathic pulmonary fibrosis (IPF). Bleomycin (BLM) induced animal models of experimental lung fibrosis were used. CHIP assay was executed to find the link between Smad3 and IL-31, and the expressions of TGF-β1, Smad3, IL-31 and STAT1 were detected to find whether they were similar with each other. We found that in the early injury or inflammation of the animal model, BLM promoted the development of inflammation, leading to severe pulmonary fibrosis. Then the expression of TGF-β1 and Smad3 increased. Activated Smad3 bound to the IL-31 promoter region, followed by the activation of JAK-STAT pathways. The inhibitor of TGF-β1 receptor decreased the IL-31 expression and knocking-down of IL-31 also decreased the STAT1 expression. We conclude that there is a pathway of pathogenesis in BLM-induced mouse model that involves the TGF-β, IL-31 and JAKs/STATs pathway. Copyright © 2013 Elsevier B.V. All rights reserved.

Expression pattern of cdkl5 during zebrafish early development: implications for use as model for atypical Rett syndrome.

PubMed

Vitorino, Marta; Cunha, Nídia; Conceição, Natércia; Cancela, M Leonor

2018-05-11

Atypical Rett syndrome is a child neurodevelopmental disorder induced by mutations in CDKL5 gene and characterized by a progressive regression in development with loss of purposeful use of the hands, slowed brain and head growth, problems with walking, seizures, and intellectual disability. At the moment, there is no cure for this pathology and little information is available concerning animal models capable of mimicking its phenotypes, thus the development of additional animal models should be of interest to gain more knowledge about the disease. Zebrafish has been used successfully as model organism for many human genetic diseases; however, no information is available concerning the spatial and temporal expression of cdkl5 orthologous in this organism. In the present study, we identified the developmental expression patterns of cdkl5 in zebrafish by quantitative PCR and whole-mount in situ hybridization. cdkl5 is expressed maternally at low levels during the first 24 h of development. After that the expression of the gene increases significantly and it starts to be expressed mainly in the nervous system and in several brain structures, such as telencephalon, mesencephalon and diencephalon. The expression patterns of cdkl5 in zebrafish is in accordance with the tissues known to be affected in humans and associated to symptoms and deficits observed in Rett syndrome patients thus providing the first evidence that zebrafish could be an alternative model to study the molecular pathways of this disease as well as to test possible therapeutic approaches capable of rescuing the phenotype.

Elevation of Il6 is associated with disturbed let-7 biogenesis in a genetic model of depression

PubMed Central

Wei, Y B; Liu, J J; Villaescusa, J C; Åberg, E; Brené, S; Wegener, G; Mathé, A A; Lavebratt, C

2016-01-01

Elevation of the proinflammatory cytokine IL-6 has been implicated in depression; however, the mechanisms remain elusive. MicroRNAs (miRNAs) are small non-coding RNAs that inhibit gene expression post-transcriptionally. The lethal-7 (let-7) miRNA family was suggested to be involved in the inflammation process and IL-6 was shown to be one of its targets. In the present study, we report elevation of Il6 in the prefrontal cortex (PFC) of a genetic rat model of depression, the Flinders Sensitive Line (FSL) compared to the control Flinders Resistant Line. This elevation was associated with an overexpression of LIN28B and downregulation of let-7 miRNAs, the former an RNA-binding protein that selectively represses let-7 synthesis. Also DROSHA, a key enzyme in miRNA biogenesis was downregulated in FSL. Running was previously shown to have an antidepressant-like effect in the FSL rat. We found that running reduced Il6 levels and selectively increased let-7i and miR-98 expression in the PFC of FSL, although there were no differences in LIN28B and DROSHA expression. Pri-let-7i was upregulated in the running FSL group, which associated with increased histone H4 acetylation. In conclusion, the disturbance of let-7 family biogenesis may underlie increased proinflammatory markers in the depressed FSL rats while physical activity could reduce their expression, possibly through regulating primary miRNA expression via epigenetic mechanisms. PMID:27529677

Elevation of Il6 is associated with disturbed let-7 biogenesis in a genetic model of depression.

PubMed

Wei, Y B; Liu, J J; Villaescusa, J C; Åberg, E; Brené, S; Wegener, G; Mathé, A A; Lavebratt, C

2016-08-16

Elevation of the proinflammatory cytokine IL-6 has been implicated in depression; however, the mechanisms remain elusive. MicroRNAs (miRNAs) are small non-coding RNAs that inhibit gene expression post-transcriptionally. The lethal-7 (let-7) miRNA family was suggested to be involved in the inflammation process and IL-6 was shown to be one of its targets. In the present study, we report elevation of Il6 in the prefrontal cortex (PFC) of a genetic rat model of depression, the Flinders Sensitive Line (FSL) compared to the control Flinders Resistant Line. This elevation was associated with an overexpression of LIN28B and downregulation of let-7 miRNAs, the former an RNA-binding protein that selectively represses let-7 synthesis. Also DROSHA, a key enzyme in miRNA biogenesis was downregulated in FSL. Running was previously shown to have an antidepressant-like effect in the FSL rat. We found that running reduced Il6 levels and selectively increased let-7i and miR-98 expression in the PFC of FSL, although there were no differences in LIN28B and DROSHA expression. Pri-let-7i was upregulated in the running FSL group, which associated with increased histone H4 acetylation. In conclusion, the disturbance of let-7 family biogenesis may underlie increased proinflammatory markers in the depressed FSL rats while physical activity could reduce their expression, possibly through regulating primary miRNA expression via epigenetic mechanisms.

Inhibition of GLO1 in Glioblastoma Multiforme Increases DNA-AGEs, Stimulates RAGE Expression, and Inhibits Brain Tumor Growth in Orthotopic Mouse Models.

PubMed

Jandial, Rahul; Neman, Josh; Lim, Punnajit P; Tamae, Daniel; Kowolik, Claudia M; Wuenschell, Gerald E; Shuck, Sarah C; Ciminera, Alexandra K; De Jesus, Luis R; Ouyang, Ching; Chen, Mike Y; Termini, John

2018-01-30

Cancers that exhibit the Warburg effect may elevate expression of glyoxylase 1 (GLO1) to detoxify the toxic glycolytic byproduct methylglyoxal (MG) and inhibit the formation of pro-apoptotic advanced glycation endproducts (AGEs). Inhibition of GLO1 in cancers that up-regulate glycolysis has been proposed as a therapeutic targeting strategy, but this approach has not been evaluated for glioblastoma multiforme (GBM), the most aggressive and difficult to treat malignancy of the brain. Elevated GLO1 expression in GBM was established in patient tumors and cell lines using bioinformatics tools and biochemical approaches. GLO1 inhibition in GBM cell lines and in an orthotopic xenograft GBM mouse model was examined using both small molecule and short hairpin RNA (shRNA) approaches. Inhibition of GLO1 with S -( p -bromobenzyl) glutathione dicyclopentyl ester ( p- BrBzGSH(Cp)₂) increased levels of the DNA-AGE N ²-1-(carboxyethyl)-2'-deoxyguanosine (CEdG), a surrogate biomarker for nuclear MG exposure; substantially elevated expression of the immunoglobulin-like receptor for AGEs (RAGE); and induced apoptosis in GBM cell lines. Targeting GLO1 with shRNA similarly increased CEdG levels and RAGE expression, and was cytotoxic to glioma cells. Mice bearing orthotopic GBM xenografts treated systemically with p -BrBzGSH(Cp)₂ exhibited tumor regression without significant off-target effects suggesting that GLO1 inhibition may have value in the therapeutic management of these drug-resistant tumors.

Inhibition of GLO1 in Glioblastoma Multiforme Increases DNA-AGEs, Stimulates RAGE Expression, and Inhibits Brain Tumor Growth in Orthotopic Mouse Models

PubMed Central

Jandial, Rahul; Neman, Josh; Tamae, Daniel; Kowolik, Claudia M.; Wuenschell, Gerald E.; Ciminera, Alexandra K.; De Jesus, Luis R.; Ouyang, Ching; Chen, Mike Y.

2018-01-01

Cancers that exhibit the Warburg effect may elevate expression of glyoxylase 1 (GLO1) to detoxify the toxic glycolytic byproduct methylglyoxal (MG) and inhibit the formation of pro-apoptotic advanced glycation endproducts (AGEs). Inhibition of GLO1 in cancers that up-regulate glycolysis has been proposed as a therapeutic targeting strategy, but this approach has not been evaluated for glioblastoma multiforme (GBM), the most aggressive and difficult to treat malignancy of the brain. Elevated GLO1 expression in GBM was established in patient tumors and cell lines using bioinformatics tools and biochemical approaches. GLO1 inhibition in GBM cell lines and in an orthotopic xenograft GBM mouse model was examined using both small molecule and short hairpin RNA (shRNA) approaches. Inhibition of GLO1 with S-(p-bromobenzyl) glutathione dicyclopentyl ester (p-BrBzGSH(Cp)2) increased levels of the DNA-AGE N2-1-(carboxyethyl)-2′-deoxyguanosine (CEdG), a surrogate biomarker for nuclear MG exposure; substantially elevated expression of the immunoglobulin-like receptor for AGEs (RAGE); and induced apoptosis in GBM cell lines. Targeting GLO1 with shRNA similarly increased CEdG levels and RAGE expression, and was cytotoxic to glioma cells. Mice bearing orthotopic GBM xenografts treated systemically with p-BrBzGSH(Cp)2 exhibited tumor regression without significant off-target effects suggesting that GLO1 inhibition may have value in the therapeutic management of these drug-resistant tumors. PMID:29385725

The 15-LO-1/15-HETE system promotes angiogenesis by upregulating VEGF in ischemic brains.

PubMed

Chen, Li; Zhu, Yan-Mei; Li, Yu-Nong; Li, Peng-Yan; Wang, Di; Liu, Yu; Qu, You-Yang; Zhu, Da-Ling; Zhu, Yu-Lan

2017-09-01

Angiogenesis promotes neurobehavioral recovery after cerebral ischemic stroke. 15(S)-hydroxyeicosatetraenoic acid (15-HETE) is one of the major metabolites of arachidonic acid by 15-lipoxygenase (15-LO) and stimulates the production of vascular endothelial growth factor (VEGF), thus, inducing autocrine-mediated angiogenesis. The present study aimed to investigate the role of 15-LO/15-HETE system on VEGF expression and angiogenesis in brain ischemia. Rat cerebral arterial vascular endothelial cells were used to set up a cell injury model of oxygen-glucose deprivation and reoxygenation (OGD/R), mimicking a condition of brain ischemia. A mouse model of middle cerebral artery occlusion (MCAO) was established. Oxygen-glucose deprivation increased cellular expression of 15-LO-1 and VEGF. Transfection of 15-LO-1 siRNA depleted cells of 15-LO-1, and sequentially induced downregulation of VEGF expression; while, incubation of 15-HETE increased the expression of VEGF. Incubation of 15-HETE attenuated the reduction in cell viability induced by oxygen-glucose deprivation, and promoted cell migration, while transfection of 15-LO-1 siRNA showed an opposite effect. In animal experiments, the density of microvessels in hypoxic regions of brains was significantly increased after MCAO, while intracerebroventricular delivery of 15-LO-1 siRNA significantly reduced the density of microvessels, and downregulates VEGF expression. The results indicate that the 15-LO-1/15-HETE system promotes angiogenesis in ischemic brains by upregulation of VEGF, representing a potential target for improving neurobehavioral recovery after cerebral ischemic stroke.

Central Nervous System Infection with Borna Disease Virus Causes Kynurenine Pathway Dysregulation and Neurotoxic Quinolinic Acid Production

PubMed Central

Formisano, Simone; Hornig, Mady; Yaddanapudi, Kavitha; Vasishtha, Mansi; Parsons, Loren H.; Briese, Thomas; Lipkin, W. Ian

2017-01-01

ABSTRACT Central nervous system infection of neonatal and adult rats with Borna disease virus (BDV) results in neuronal destruction and behavioral abnormalities with differential immune-mediated involvement. Neuroactive metabolites generated from the kynurenine pathway of tryptophan degradation have been implicated in several human neurodegenerative disorders. Here, we report that brain expression of key enzymes in the kynurenine pathway are significantly, but differentially, altered in neonatal and adult rats with BDV infection. Gene expression analysis of rat brains following neonatal infection showed increased expression of kynurenine amino transferase II (KATII) and kynurenine-3-monooxygenase (KMO) enzymes. Additionally, indoleamine 2,3-dioxygenase (IDO) expression was only modestly increased in a brain region- and time-dependent manner in neonatally infected rats; however, its expression was highly increased in adult infected rats. The most dramatic impact on gene expression was seen for KMO, whose activity promotes the production of neurotoxic quinolinic acid. KMO expression was persistently elevated in brain regions of both newborn and adult BDV-infected rats, with increases reaching up to 86-fold. KMO protein levels were increased in neonatally infected rats and colocalized with neurons, the primary target cells of BDV infection. Furthermore, quinolinic acid was elevated in neonatally infected rat brains. We further demonstrate increased expression of KATII and KMO, but not IDO, in vitro in BDV-infected C6 astroglioma cells. Our results suggest that BDV directly impacts the kynurenine pathway, an effect that may be exacerbated by inflammatory responses in immunocompetent hosts. Thus, experimental models of BDV infection may provide new tools for discriminating virus-mediated from immune-mediated impacts on the kynurenine pathway and their relative contribution to neurodegeneration. IMPORTANCE BDV causes persistent, noncytopathic infection in vitro yet still elicits widespread neurodegeneration of infected neurons in both immunoincompetent and immunocompetent hosts. Here, we show that BDV infection induces expression of key enzymes of the kynurenine pathway in brains of newborn and adult infected rats and cultured astroglioma cells, shunting tryptophan degradation toward the production of neurotoxic quinolinic acid. Thus, our findings newly implicate this metabolic pathway in BDV-induced neurodegeneration. Given the importance of the kynurenine pathway in a wide range of human infections and neurodegenerative and neuropsychiatric disorders, animal models of BDV infection may serve as important tools for contrasting direct viral and indirect antiviral immune-mediated impacts on kynurenine pathway dysregulation and the ensuing neurodevelopmental and neuropathological consequences. PMID:28446679

Central Nervous System Infection with Borna Disease Virus Causes Kynurenine Pathway Dysregulation and Neurotoxic Quinolinic Acid Production.

PubMed

Formisano, Simone; Hornig, Mady; Yaddanapudi, Kavitha; Vasishtha, Mansi; Parsons, Loren H; Briese, Thomas; Lipkin, W Ian; Williams, Brent L

2017-07-15

Central nervous system infection of neonatal and adult rats with Borna disease virus (BDV) results in neuronal destruction and behavioral abnormalities with differential immune-mediated involvement. Neuroactive metabolites generated from the kynurenine pathway of tryptophan degradation have been implicated in several human neurodegenerative disorders. Here, we report that brain expression of key enzymes in the kynurenine pathway are significantly, but differentially, altered in neonatal and adult rats with BDV infection. Gene expression analysis of rat brains following neonatal infection showed increased expression of kynurenine amino transferase II (KATII) and kynurenine-3-monooxygenase (KMO) enzymes. Additionally, indoleamine 2,3-dioxygenase (IDO) expression was only modestly increased in a brain region- and time-dependent manner in neonatally infected rats; however, its expression was highly increased in adult infected rats. The most dramatic impact on gene expression was seen for KMO, whose activity promotes the production of neurotoxic quinolinic acid. KMO expression was persistently elevated in brain regions of both newborn and adult BDV-infected rats, with increases reaching up to 86-fold. KMO protein levels were increased in neonatally infected rats and colocalized with neurons, the primary target cells of BDV infection. Furthermore, quinolinic acid was elevated in neonatally infected rat brains. We further demonstrate increased expression of KATII and KMO, but not IDO, in vitro in BDV-infected C6 astroglioma cells. Our results suggest that BDV directly impacts the kynurenine pathway, an effect that may be exacerbated by inflammatory responses in immunocompetent hosts. Thus, experimental models of BDV infection may provide new tools for discriminating virus-mediated from immune-mediated impacts on the kynurenine pathway and their relative contribution to neurodegeneration. IMPORTANCE BDV causes persistent, noncytopathic infection in vitro yet still elicits widespread neurodegeneration of infected neurons in both immunoincompetent and immunocompetent hosts. Here, we show that BDV infection induces expression of key enzymes of the kynurenine pathway in brains of newborn and adult infected rats and cultured astroglioma cells, shunting tryptophan degradation toward the production of neurotoxic quinolinic acid. Thus, our findings newly implicate this metabolic pathway in BDV-induced neurodegeneration. Given the importance of the kynurenine pathway in a wide range of human infections and neurodegenerative and neuropsychiatric disorders, animal models of BDV infection may serve as important tools for contrasting direct viral and indirect antiviral immune-mediated impacts on kynurenine pathway dysregulation and the ensuing neurodevelopmental and neuropathological consequences. Copyright © 2017 Formisano et al.

Improving RNA-Seq expression estimation by modeling isoform- and exon-specific read sequencing rate.

PubMed

Liu, Xuejun; Shi, Xinxin; Chen, Chunlin; Zhang, Li

2015-10-16

The high-throughput sequencing technology, RNA-Seq, has been widely used to quantify gene and isoform expression in the study of transcriptome in recent years. Accurate expression measurement from the millions or billions of short generated reads is obstructed by difficulties. One is ambiguous mapping of reads to reference transcriptome caused by alternative splicing. This increases the uncertainty in estimating isoform expression. The other is non-uniformity of read distribution along the reference transcriptome due to positional, sequencing, mappability and other undiscovered sources of biases. This violates the uniform assumption of read distribution for many expression calculation approaches, such as the direct RPKM calculation and Poisson-based models. Many methods have been proposed to address these difficulties. Some approaches employ latent variable models to discover the underlying pattern of read sequencing. However, most of these methods make bias correction based on surrounding sequence contents and share the bias models by all genes. They therefore cannot estimate gene- and isoform-specific biases as revealed by recent studies. We propose a latent variable model, NLDMseq, to estimate gene and isoform expression. Our method adopts latent variables to model the unknown isoforms, from which reads originate, and the underlying percentage of multiple spliced variants. The isoform- and exon-specific read sequencing biases are modeled to account for the non-uniformity of read distribution, and are identified by utilizing the replicate information of multiple lanes of a single library run. We employ simulation and real data to verify the performance of our method in terms of accuracy in the calculation of gene and isoform expression. Results show that NLDMseq obtains competitive gene and isoform expression compared to popular alternatives. Finally, the proposed method is applied to the detection of differential expression (DE) to show its usefulness in the downstream analysis. The proposed NLDMseq method provides an approach to accurately estimate gene and isoform expression from RNA-Seq data by modeling the isoform- and exon-specific read sequencing biases. It makes use of a latent variable model to discover the hidden pattern of read sequencing. We have shown that it works well in both simulations and real datasets, and has competitive performance compared to popular methods. The method has been implemented as a freely available software which can be found at https://github.com/PUGEA/NLDMseq.

HSV-2-Driven Increase in the Expression of α4β7 Correlates with Increased Susceptibility to Vaginal SHIVSF162P3 Infection

PubMed Central

Goode, Diana; Truong, Rosaline; Villegas, Guillermo; Calenda, Giulia; Guerra-Perez, Natalia; Piatak, Michael; Lifson, Jeffrey D.; Blanchard, James; Gettie, Agegnehu; Robbiani, Melissa; Martinelli, Elena

2014-01-01

The availability of highly susceptible HIV target cells that can rapidly reach the mucosal lymphoid tissues may increase the chances of an otherwise rare transmission event to occur. Expression of α4β7 is required for trafficking of immune cells to gut inductive sites where HIV can expand and it is expressed at high level on cells particularly susceptible to HIV infection. We hypothesized that HSV-2 modulates the expression of α4β7 and other homing receptors in the vaginal tissue and that this correlates with the increased risk of HIV acquisition in HSV-2 positive individuals. To test this hypothesis we used an in vivo rhesus macaque (RM) model of HSV-2 vaginal infection and a new ex vivo model of macaque vaginal explants. In vivo we found that HSV-2 latently infected RMs appeared to be more susceptible to vaginal SHIVSF162P3 infection, had higher frequency of α4β7 high CD4+ T cells in the vaginal tissue and higher expression of α4β7 and CD11c on vaginal DCs. Similarly, ex vivo HSV-2 infection increased the susceptibility of the vaginal tissue to SHIVSF162P3. HSV-2 infection increased the frequencies of α4β7 high CD4+ T cells and this directly correlated with HSV-2 replication. A higher amount of inflammatory cytokines in vaginal fluids of the HSV-2 infected animals was similar to those found in the supernatants of the infected explants. Remarkably, the HSV-2-driven increase in the frequency of α4β7 high CD4+ T cells directly correlated with SHIV replication in the HSV-2 infected tissues. Our results suggest that the HSV-2-driven increase in availability of CD4+ T cells and DCs that express high levels of α4β7 is associated with the increase in susceptibility to SHIV due to HSV-2. This may persists in absence of HSV-2 shedding. Hence, higher availability of α4β7 positive HIV target cells in the vaginal tissue may constitute a risk factor for HIV transmission. PMID:25521298

HSV-2-driven increase in the expression of α4β7 correlates with increased susceptibility to vaginal SHIV(SF162P3) infection.

PubMed

Goode, Diana; Truong, Rosaline; Villegas, Guillermo; Calenda, Giulia; Guerra-Perez, Natalia; Piatak, Michael; Lifson, Jeffrey D; Blanchard, James; Gettie, Agegnehu; Robbiani, Melissa; Martinelli, Elena

2014-12-01

The availability of highly susceptible HIV target cells that can rapidly reach the mucosal lymphoid tissues may increase the chances of an otherwise rare transmission event to occur. Expression of α4β7 is required for trafficking of immune cells to gut inductive sites where HIV can expand and it is expressed at high level on cells particularly susceptible to HIV infection. We hypothesized that HSV-2 modulates the expression of α4β7 and other homing receptors in the vaginal tissue and that this correlates with the increased risk of HIV acquisition in HSV-2 positive individuals. To test this hypothesis we used an in vivo rhesus macaque (RM) model of HSV-2 vaginal infection and a new ex vivo model of macaque vaginal explants. In vivo we found that HSV-2 latently infected RMs appeared to be more susceptible to vaginal SHIVSF162P3 infection, had higher frequency of α4β7high CD4+ T cells in the vaginal tissue and higher expression of α4β7 and CD11c on vaginal DCs. Similarly, ex vivo HSV-2 infection increased the susceptibility of the vaginal tissue to SHIVSF162P3. HSV-2 infection increased the frequencies of α4β7high CD4+ T cells and this directly correlated with HSV-2 replication. A higher amount of inflammatory cytokines in vaginal fluids of the HSV-2 infected animals was similar to those found in the supernatants of the infected explants. Remarkably, the HSV-2-driven increase in the frequency of α4β7high CD4+ T cells directly correlated with SHIV replication in the HSV-2 infected tissues. Our results suggest that the HSV-2-driven increase in availability of CD4+ T cells and DCs that express high levels of α4β7 is associated with the increase in susceptibility to SHIV due to HSV-2. This may persists in absence of HSV-2 shedding. Hence, higher availability of α4β7 positive HIV target cells in the vaginal tissue may constitute a risk factor for HIV transmission.

Pediatric Crohn disease patients exhibit specific ileal transcriptome and microbiome signature.

PubMed

Haberman, Yael; Tickle, Timothy L; Dexheimer, Phillip J; Kim, Mi-Ok; Tang, Dora; Karns, Rebekah; Baldassano, Robert N; Noe, Joshua D; Rosh, Joel; Markowitz, James; Heyman, Melvin B; Griffiths, Anne M; Crandall, Wallace V; Mack, David R; Baker, Susan S; Huttenhower, Curtis; Keljo, David J; Hyams, Jeffrey S; Kugathasan, Subra; Walters, Thomas D; Aronow, Bruce; Xavier, Ramnik J; Gevers, Dirk; Denson, Lee A

2014-08-01

Interactions between the host and gut microbial community likely contribute to Crohn disease (CD) pathogenesis; however, direct evidence for these interactions at the onset of disease is lacking. Here, we characterized the global pattern of ileal gene expression and the ileal microbial community in 359 treatment-naive pediatric patients with CD, patients with ulcerative colitis (UC), and control individuals. We identified core gene expression profiles and microbial communities in the affected CD ilea that are preserved in the unaffected ilea of patients with colon-only CD but not present in those with UC or control individuals; therefore, this signature is specific to CD and independent of clinical inflammation. An abnormal increase of antimicrobial dual oxidase (DUOX2) expression was detected in association with an expansion of Proteobacteria in both UC and CD, while expression of lipoprotein APOA1 gene was downregulated and associated with CD-specific alterations in Firmicutes. The increased DUOX2 and decreased APOA1 gene expression signature favored oxidative stress and Th1 polarization and was maximally altered in patients with more severe mucosal injury. A regression model that included APOA1 gene expression and microbial abundance more accurately predicted month 6 steroid-free remission than a model using clinical factors alone. These CD-specific host and microbe profiles identify the ileum as the primary inductive site for all forms of CD and may direct prognostic and therapeutic approaches.

The orexin-1 receptor antagonist SB-334867 decreases anxiety-like behavior and c-Fos expression in the hypothalamus of rats exposed to cat odor.

PubMed

Vanderhaven, M W; Cornish, J L; Staples, L G

2015-02-01

Increasing evidence suggests that the orexin system is involved in modulating anxiety, and we have recently shown that cat odor-induced anxiety in rats is attenuated by the orexin receptor antagonist SB-334867. In the current experiment, c-Fos expression was used to map changes in neuronal activation following SB-334867 administration in the cat odor anxiety model. Male Wistar rats were exposed to cat odor with or without SB-334867 pre-treatment (10 mg/kg, i.p.). A naïve control group not exposed to cat odor was also used. Following cat odor exposure, brains were processed for c-Fos expression. Vehicle-treated rats showed an increase in anxiety-like behaviors (increased hiding and decreased approach toward the cat odor), and increased c-Fos expression in the posteroventral medial amygdala (MePV), paraventricular hypothalamus (PVN) and dorsal premammillary nucleus (PMd). In rats pretreated with SB-334867, approach scores increased and c-Fos expression decreased in the PVN and PMd. These results provide both behavioral and neuroanatomical evidence for the attenuation of cat odor-induced anxiety in rats via the orexin system. Crown Copyright © 2014. Published by Elsevier B.V. All rights reserved.

miR-1 is increased in pulmonary hypertension and downregulates Kv1.5 channels in rat pulmonary arteries.

PubMed

Mondejar-Parreño, Gema; Callejo, María; Barreira, Bianca; Morales-Cano, Daniel; Esquivel-Ruiz, Sergio; Moreno, Laura; Cogolludo, Angel; Perez-Vizcaino, Francisco

2018-05-02

■The expression of miR-1 is increased in lungs from the Hyp/Su5416 PAH rat model. ■PASMC from this animal model are more depolarised and show decreased expression and activity of Kv1.5. ■miR-1 directly targets Kv1.5 channels, reduces Kv1.5 activity and induces membrane depolarization. ■Antagomir-1 prevents Kv1.5 channel downregulation and the depolarization induced by hypoxia/Su5416 exposition. Impairment of voltage-dependent potassium channel (Kv) plays a central role in the development of cardiovascular diseases, including pulmonary arterial hypertension (PAH). MicroRNAs (miRNAs) are non-coding RNAs that regulate gene expression by binding to the 3'-UTR region of specific mRNAs. The aim of this study was to analyze the effects of miR-1 on Kv channel function in pulmonary arteries (PA). Kv channel activity was studied in PA from healthy animals transfected with miR-1 or scrambled-miR. Kv currents were studied using the whole-cell configuration of patch-clamp technique. The characterization of the Kv1.5 currents was performed with the selective inhibitor DPO-1. miR-1 expression was increased and Kv1.5 channels were decreased in lungs from a rat model of PAH induced by hypoxia and Su5416. miR-1 transfection increased cell capacitance, reduced Kv1.5 currents and induced membrane depolarization in isolated pulmonary artery smooth muscle cells (PASMCs). Luciferase reporter assay indicated that KCNA5, which encodes Kv1.5 channels, is a direct target gene of miR-1. Incubation of PA with Su5416 and hypoxia (3% O 2 ) increased miR-1 and induced a decline in Kv1.5 currents, which was prevented by antagomiR-1. In conclusion, these data indicate that miR-1 induces PASMC hypertrophy and reduces the activity and expression of Kv channels, suggesting a pathophysiological role in PAH. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

Changes in Ca(2+) channel expression upon differentiation of SN56 cholinergic cells.

PubMed

Kushmerick, C; Romano-Silva, M A; Gomez, M V; Prado, M A

2001-10-19

The SN56 cell line, a fusion of septal neurons and neuroblastoma cells, has been used as a model for central cholinergic neurons. These cells show increased expression of cholinergic neurochemical features upon differentiation, but little is known about how differentiation affects their electrophysiological properties. We examined the changes in Ca(2+) channel expression that occur as these cells undergo morphological differentiation in response to serum withdrawal and exposure to dibutyryl-cAMP. Undifferentiated cells expressed a T-type current with biophysical and pharmacological properties similar, although not identical, to those reported for the current generated by the alpha(1H) (CaV3.2) Ca(2+) channel subunit. Differentiated cells expressed, in addition to this T-type current, high voltage activated currents which were inhibited 38% by the L-type channel antagonist nifedipine (5 microM), 37% by the N-type channel antagonist omega-conotoxin-GVIA (1 microM), and 15% by the P/Q-type channel antagonist omega-agatoxin-IVA (200 nM). Current resistant to these inhibitors accounted for 15% of the high voltage activated current in differentiated SN56 cells. Our data demonstrate that differentiation increases the expression of neuronal type voltage gated Ca(2+) channels in this cell line, and that the channels expressed are comparable to those reported for native basal forebrain cholinergic neurons. This cell line should thus provide a useful model system to study the relationship between calcium currents and cholinergic function and dysfunction.

Inositol-requiring enzyme 1-mediated endoplasmic reticulum stress triggers apoptosis and fibrosis formation in liver cirrhosis rat models.

PubMed

Jiang, Tianpeng; Wang, Lizhou; Li, Xing; Song, Jie; Wu, Xiaoping; Zhou, Shi

2015-04-01

Long‑term and advanced cirrhosis is usually irreversible and often coincides with variceal hemorrhage or development of hepatocellular carcinoma; therefore, liver cirrhosis is a major cause of morbidity and mortality globally. The aim of the present study was to investigate the specific mechanism behind the formation of fibrosis or cirrhosis using rat models of hepatic fibrosis. The cirrhosis model was established by intraperitoneally administering dimethylnitrosamine to the rats. Hematoxylin and eosin staining was performed on the hepatic tissues of the rats to observe the fibrosis or cirrhosis, and western blot analysis was employed to detect α‑smooth muscle actin and desmin protein expression. Flow cytometric analysis was used to examine early and late apoptosis, and the protein and mRNA expression of endoplasmic reticulum (ER) stress-associated unfolded protein response (UPR) pathway proteins and apoptotic proteins [C/EBP homologous protein (CHOP) and caspase‑12] was detected by western blotting and the reverse-transcription polymerase chain reaction, respectively. The results indicated that the cirrhosis model was established successfully and that fibrosis was significantly increased in the cirrhosis model group compared with that in the normal control group. Flow cytometric analysis showed that early and late apoptosis in the cirrhosis model was significantly higher compared with that in the control group. The expression of the UPR pathway protein inositol-requiring enzyme (IRE) 1, as well as the expression of CHOP, was increased significantly in the cirrhotic rat tissues compared with that in the control group tissues (P<0.05). In conclusion, apoptosis was clearly observed in the hepatic tissue of cirrhotic rats, and the apoptosis was caused by activation of the ER stress-mediated IRE1 and CHOP.

Increased H+ efflux is sufficient to induce dysplasia and necessary for viability with oncogene expression

PubMed Central

Grillo-Hill, Bree K; Choi, Changhoon; Jimenez-Vidal, Maite; Barber, Diane L

2015-01-01

Intracellular pH (pHi) dynamics is increasingly recognized as an important regulator of a range of normal and pathological cell behaviors. Notably, increased pHi is now acknowledged as a conserved characteristic of cancers and in cell models is confirmed to increase proliferation and migration as well as limit apoptosis. However, the significance of increased pHi for cancer in vivo remains unresolved. Using Drosophila melanogaster, we show that increased pHi is sufficient to induce dysplasia in the absence of other transforming cues and potentiates growth and invasion with oncogenic Ras. Using a genetically encoded biosensor we also confirm increased pHi in situ. Moreover, in Drosophila models and clonal human mammary cells we show that limiting H+ efflux with oncogenic Raf or Ras induces acidosis and synthetic lethality. Further, we show lethality in invasive primary tumor cell lines with inhibiting H+ efflux. Synthetic lethality with reduced H+ efflux and activated oncogene expression could be exploited therapeutically to restrain cancer progression while limiting off-target effects. DOI: http://dx.doi.org/10.7554/eLife.03270.001 PMID:25793441

Experimental and in silico modelling analyses of the gene expression pathway for recombinant antibody and by-product production in NS0 cell lines.

PubMed

Mead, Emma J; Chiverton, Lesley M; Spurgeon, Sarah K; Martin, Elaine B; Montague, Gary A; Smales, C Mark; von der Haar, Tobias

2012-01-01

Monoclonal antibodies are commercially important, high value biotherapeutic drugs used in the treatment of a variety of diseases. These complex molecules consist of two heavy chain and two light chain polypeptides covalently linked by disulphide bonds. They are usually expressed as recombinant proteins from cultured mammalian cells, which are capable of correctly modifying, folding and assembling the polypeptide chains into the native quaternary structure. Such recombinant cell lines often vary in the amounts of product produced and in the heterogeneity of the secreted products. The biological mechanisms of this variation are not fully defined. Here we have utilised experimental and modelling strategies to characterise and define the biology underpinning product heterogeneity in cell lines exhibiting varying antibody expression levels, and then experimentally validated these models. In undertaking these studies we applied and validated biochemical (rate-constant based) and engineering (nonlinear) models of antibody expression to experimental data from four NS0 cell lines with different IgG4 secretion rates. The models predict that export of the full antibody and its fragments are intrinsically linked, and cannot therefore be manipulated individually at the level of the secretory machinery. Instead, the models highlight strategies for the manipulation at the precursor species level to increase recombinant protein yields in both high and low producing cell lines. The models also highlight cell line specific limitations in the antibody expression pathway.

[Effects of the scalp acupuncture at baihui (DU20) through qubin (GB7) on the expressions of GDNF VEGF in the brain tissue of rats with acute intracerebral hemorrhage].

PubMed

Zhang, Guo-Wei; Zou, Wei; Liu, Fang

2012-09-01

To observe the effects of the scalp acupuncture at Baihui (DU20) through Qubin (GB7) on the expressions of glial cell line-derived neurotrophic factor (GDNF) and vascular endothelial growth factor (VEGF) in the brain tissue of rats with acute intracerebral hemorrhage (AICH). Totally 150 healthy male Wistar rats were used to prepare the intracerebral hemorrhage (ICH) model. They were randomly divided into the model group, the acupuncture group, and the Western medicine group, 50 in each group. Rats in each group were then randomly divided into five subgroups, i. e., 6 h, day 1, day 2, day 3, and day 7, 10 in each subgroup. Another 10 normal rats were recruited as the blank control group. Rats in the acupuncture group were acupunctured at Baihui (DU20) through Qubin (GB7) on the lesion side. Rats in the Western medicine group were administered with aniracetam diluent 1 mL by gastrogavage, 3 times daily. Rats in the model group were bound the same way as those in the acupuncture group 30 min daily, and then administered with normal saline 1 mL by gastrogavage, 3 times daily. The expressions of GDNF and VEGF in the brain tissue of the rats in each group were detected using in situ hybridization and immunohistochemical methods. Compared with the blank control group, the number of GDNF positive cells in the model group increased at 6 h-3 days, and the number of VEGF positive cells in the model group increased at each time point, showing statistical difference (P<0.01). The numbers of GDNF positive cells increased in the acupuncture group at each time point, showing statistical difference when compared with those of the model group and the Western medicine group (P<0.01). There was no statistical difference between the model group and the Western medicine group at each time point (P>0.05). Compared with the model group, the number of VEGF positive cells in the acupuncture group decreased at 6 h -3 days. The VEGF positive cells increased at day 7 and were higher than that of the Western medicine group, showing statistical difference (P<0.01). The scalp acupuncture at Baihui (DU20) through Qubin (GB7) played the role of nerves remodeling in AICH possibly through promoting the expression of endogenous GDNF and early inhibiting the expression of VEGF in the AICH. This method possibly had favorable bidirectional regulation effects in AICH. Its therapeutic effects were superior to Aniracetam.

Differential transcriptional regulation by alternatively designed mechanisms: A mathematical modeling approach.

PubMed

Yildirim, Necmettin; Aktas, Mehmet Emin; Ozcan, Seyma Nur; Akbas, Esra; Ay, Ahmet

2017-01-01

Cells maintain cellular homeostasis employing different regulatory mechanisms to respond external stimuli. We study two groups of signal-dependent transcriptional regulatory mechanisms. In the first group, we assume that repressor and activator proteins compete for binding to the same regulatory site on DNA (competitive mechanisms). In the second group, they can bind to different regulatory regions in a noncompetitive fashion (noncompetitive mechanisms). For both competitive and noncompetitive mechanisms, we studied the gene expression dynamics by increasing the repressor or decreasing the activator abundance (inhibition mechanisms), or by decreasing the repressor or increasing the activator abundance (activation mechanisms). We employed delay differential equation models. Our simulation results show that the competitive and noncompetitive inhibition mechanisms exhibit comparable repression effectiveness. However, response time is fastest in the noncompetitive inhibition mechanism due to increased repressor abundance, and slowest in the competitive inhibition mechanism by increased repressor level. The competitive and noncompetitive inhibition mechanisms through decreased activator abundance show comparable and moderate response times, while the competitive and noncompetitive activation mechanisms by increased activator protein level display more effective and faster response. Our study exemplifies the importance of mathematical modeling and computer simulation in the analysis of gene expression dynamics.

LocExpress: a web server for efficiently estimating expression of novel transcripts.

PubMed

Hou, Mei; Tian, Feng; Jiang, Shuai; Kong, Lei; Yang, Dechang; Gao, Ge

2016-12-22

The temporal and spatial-specific expression pattern of a transcript in multiple tissues and cell types can indicate key clues about its function. While several gene atlas available online as pre-computed databases for known gene models, it's still challenging to get expression profile for previously uncharacterized (i.e. novel) transcripts efficiently. Here we developed LocExpress, a web server for efficiently estimating expression of novel transcripts across multiple tissues and cell types in human (20 normal tissues/cells types and 14 cell lines) as well as in mouse (24 normal tissues/cell types and nine cell lines). As a wrapper to RNA-Seq quantification algorithm, LocExpress efficiently reduces the time cost by making abundance estimation calls increasingly within the minimum spanning bundle region of input transcripts. For a given novel gene model, such local context-oriented strategy allows LocExpress to estimate its FPKMs in hundreds of samples within minutes on a standard Linux box, making an online web server possible. To the best of our knowledge, LocExpress is the only web server to provide nearly real-time expression estimation for novel transcripts in common tissues and cell types. The server is publicly available at http://loc-express.cbi.pku.edu.cn .

Effects of prenatal low protein and postnatal high fat diets on visceral adipose tissue macrophage phenotypes and IL-6 expression in Sprague Dawley rat offspring

USDA-ARS?s Scientific Manuscript database

Adipose tissue macrophages (ATM) are implicated in adipose tissue inflammation and obesity-related insulin resistance. Maternal low protein models result in fetal programming of obesity. However, it is not known whether maternal undernutrition increases ATM phenotypic expression in F1 offspring. Us...

Changes in dopamine transporter expression in the midbrain following traumatic brain injury: an immunohistochemical and in situ hybridization study in a mouse model.

PubMed

Shimada, Ryo; Abe, Keiichi; Furutani, Rui; Kibayashi, Kazuhiko

2014-03-01

An association has been suggested between trauma and neurological degenerative diseases. Magnetic resonance imaging has revealed that traumatic brain injury (TBI) can cause primary lesions in the midbrain including the substantia nigra (SN). Dopamine transporter (DAT) is mainly expressed in the SN, ventral tegmental area (VTA), and retrorubral field (RRF) of the ventral midbrain. Previous western blot studies have examined DAT levels in the rat frontal cortex and striatum after a controlled cortical impact (CCI); however, no study has comprehensively examined DAT expression in the midbrain following TBI in an animal model. We used immunohistochemistry and in situ hybridization to examine the time-dependent changes in the expression of DAT in the midbrain during the first 14 days after TBI in a mouse CCI model. The expression of DAT protein in the RRF on the side ipsilateral to the site of injury decreased in 14 days after injury. Dopamine transporter mRNA expression in the RRF on the ipsilateral side decreased in 1, 7, and 14 days and increased in 4 days after injury. These findings indicated that TBI induced changes in DAT expression in the RRF. Because the DAT pumps dopamine (DA) out of the synapse back into the cytosol and maintains DA homeostasis, the decreased expression of DAT after TBI may result in decreased DA neurotransmission in the brain.

Upregulation of fibroblast growth factor receptor 2 and 3 in the late stages of fetal lung development in the nitrofen rat model.

PubMed

Friedmacher, Florian; Doi, Takashi; Gosemann, Jan-Hendrik; Fujiwara, Naho; Kutasy, Balazs; Puri, Prem

2012-02-01

Nitrofen model of congenital diaphragmatic hernia (CDH) has been widely used to investigate the pathogenesis of pulmonary hypoplasia (PH). Fibroblast growth factor (FGF) signaling pathway plays a fundamental role in fetal lung development. FGF7 and FGF10, which are critical for lung morphogenesis, have been reported to be downregulated in nitrofen-induced PH. FGF signaling is mediated by a family of four single transmembrane receptors, FGFR1-4. FGFR2 and FGFR3 have been shown to be expressed predominantly in the late stages of developing lungs. In addition, the upregulation of FGFR2 gene expression has been associated with severe defects in lung development and resulted in arrested alveologenesis similar to PH seen in the nitrofen model. Furthermore, FGFR3(-/-)FGFR4(-/-) double mutants showed thinner mesenchyme and larger air spaces. We designed this study to test the hypothesis that FGFR gene expression is upregulated in the late stages of lung development in the nitrofen CDH model. Pregnant rats were exposed to either olive oil or nitrofen on day 9 of gestation (D9). Cesarean section was performed and fetuses were harvested on D18 and D21. Fetal lungs were divided into three groups: control, nitrofen without CDH [CDH(-)], and nitrofen with CDH [CDH(+)] (n = 24 at each time-point). Pulmonary gene expression levels of FGFR1-4 were analyzed by real-time RT-PCR. Immunohistochemistry was also performed to evaluate protein expression/distribution at each time-point. The relative messenger RNA expression levels of pulmonary FGFR2 and FGFR3 on D21 were significantly increased in CDH(-) (6.38 ± 1.93 and 7.84 ± 2.86, respectively) and CDH(+) (7.09 ± 2.50 and 7.25 ± 3.43, respectively) compared to controls (P < 0.05 and P < 0.01, respectively), whereas no significant alteration was observed on D18. There were no differences in FGFR1 and FGFR4 expression at both time-points. Increased immunoreactivity of FGFR2 and FGFR3, mainly in the distal epithelium and mesenchyme, was observed in the nitrofen-induced hypoplastic lungs on D21 compared to controls. Upregulation of FGFR2 and FGFR3 pulmonary gene expression in the late stages of fetal lung development may disrupt FGFR-mediated alveologenesis resulting in PH in the CDH model.

A Pathway Based Classification Method for Analyzing Gene Expression for Alzheimer's Disease Diagnosis.

PubMed

Voyle, Nicola; Keohane, Aoife; Newhouse, Stephen; Lunnon, Katie; Johnston, Caroline; Soininen, Hilkka; Kloszewska, Iwona; Mecocci, Patrizia; Tsolaki, Magda; Vellas, Bruno; Lovestone, Simon; Hodges, Angela; Kiddle, Steven; Dobson, Richard Jb

2016-01-01

Recent studies indicate that gene expression levels in blood may be able to differentiate subjects with Alzheimer's disease (AD) from normal elderly controls and mild cognitively impaired (MCI) subjects. However, there is limited replicability at the single marker level. A pathway-based interpretation of gene expression may prove more robust. This study aimed to investigate whether a case/control classification model built on pathway level data was more robust than a gene level model and may consequently perform better in test data. The study used two batches of gene expression data from the AddNeuroMed (ANM) and Dementia Case Registry (DCR) cohorts. Our study used Illumina Human HT-12 Expression BeadChips to collect gene expression from blood samples. Random forest modeling with recursive feature elimination was used to predict case/control status. Age and APOE ɛ4 status were used as covariates for all analysis. Gene and pathway level models performed similarly to each other and to a model based on demographic information only. Any potential increase in concordance from the novel pathway level approach used here has not lead to a greater predictive ability in these datasets. However, we have only tested one method for creating pathway level scores. Further, we have been able to benchmark pathways against genes in datasets that had been extensively harmonized. Further work should focus on the use of alternative methods for creating pathway level scores, in particular those that incorporate pathway topology, and the use of an endophenotype based approach.

Reduced expression of brain cannabinoid receptor 1 (Cnr1) is coupled with an increased complementary micro-RNA (miR-26b) in a mouse model of fetal alcohol spectrum disorders

PubMed Central

2013-01-01

Background Prenatal alcohol exposure is known to result in fetal alcohol spectrum disorders, a continuum of physiological, behavioural, and cognitive phenotypes that include increased risk for anxiety and learning-associated disorders. Prenatal alcohol exposure results in life-long disorders that may manifest in part through the induction of long-term gene expression changes, potentially maintained through epigenetic mechanisms. Findings Here we report a decrease in the expression of Canabinoid receptor 1 (Cnr1) and an increase in the expression of the regulatory microRNA miR-26b in the brains of adult mice exposed to ethanol during neurodevelopment. Furthermore, we show that miR-26b has significant complementarity to the 3’-UTR of the Cnr1 transcript, giving it the potential to bind and reduce the level of Cnr1 expression. Conclusions These findings elucidate a mechanism through which some genes show long-term altered expression following prenatal alcohol exposure, leading to persistent alterations to cognitive function and behavioural phenotypes observed in fetal alcohol spectrum disorders. PMID:23915435

Increased receptor for advanced glycation end product expression in the human alcoholic prefrontal cortex is linked to adolescent drinking.

PubMed

Vetreno, Ryan P; Qin, Liya; Crews, Fulton T

2013-11-01

Adolescence is characterized behaviorally by increased impulsivity and risk-taking that declines in parallel with maturation of the prefrontal cortex and executive function. In the brain, the receptor for advanced glycation end products (RAGE) is critically involved in neurodevelopment and neuropathology. In humans, the risk of alcoholism is greatly increased in those who begin drinking between 13 and 15years of age, and adolescents binge drink more than any other age group. We have previously found that alcoholism is associated with increased expression of neuroimmune genes. This manuscript tested the hypothesis that adolescent binge drinking upregulates RAGE and Toll-like receptor (TLR) 4 as well as their endogenous agonist, high-mobility group box 1 (HMGB1). Immunohistochemistry, Western blot, and mRNA analyses found that RAGE expression was increased in the human post-mortem alcoholic orbitofrontal cortex (OFC). Further, an earlier age of drinking onset correlated with increased expression of RAGE, TLR4, and HMGB1. To determine if alcohol contributed to these changes, we used an adolescent binge ethanol model in rats (5.0g/kg, i.g., 2-day on/2-day off from postnatal day [P] 25 to P55) and assessed neuroimmune gene expression. We found an age-associated decline of RAGE expression from late adolescence (P56) to young adulthood (P80). Adolescent intermittent ethanol exposure did not alter RAGE expression at P56, but increased RAGE in the young adult PFC (P80). Adolescent intermittent ethanol exposure also increased TLR4 and HMGB1 expression at P56 that persisted into young adulthood (P80). Assessment of young adult frontal cortex mRNA (RT-PCR) found increased expression of proinflammatory cytokines, oxidases, and neuroimmune agonists at P80, 25days after ethanol treatment. Together, these human and animal data support the hypothesis that an early age of drinking onset upregulates RAGE/TLR4-HMGB1 and other neuroimmune genes that persist into young adulthood and could contribute to risk of alcoholism or other brain diseases associated with neuroinflammation. © 2013.

Expression of heat shock protein 47 is increased in remnant kidney and correlates with disease progression

PubMed Central

SUNAMOTO, MASAAKI; KUZE, KOGO; IEHARA, NORIYUKI; TAKEOKA, HIROYA; NAGATA, KAZUHIRO; KITA, TORU; DOI, TOSHIO

1998-01-01

Glomerulosclerosis is characterized by accumulation of the mesangial extracellular matrix, including type I and IV collagen. The processing for the collagens in the glomeruli may play a critical role for development of glomerulosclerosis. We examined the expression of heat shock protein 47 (HSP47), a collagen-binding molecular chaperone in the progresive glomerulosclerosis model. Subtotally nephrectomized rats, unlike sham-operated rats, developed focal and segmental glomerulosclerosis. Immunological staining demonstrated an increased expression of HSP47 which paralleled the expression of type I and IV collagen in the glomeruli of the nephrectomized rats as the glomerulosclerosis developed. The mRNA levels encoding type I and type IV collagen and HSP47 were increased 3.4 fold, 3.6 fold and 2.8 fold, respectively, at week 7 after nephrectomy. By in situ hybridization, the expression of HSP47 mRNA was determined to be localized to the glomeruli with segmental sclerosis. These results suggest that HSP47 may play a central role in the process of extracellular matrix accumulation during the development of glomerulosclerosis. PMID:9741355

Employing conservation of co-expression to improve functional inference

PubMed Central

Daub, Carsten O; Sonnhammer, Erik LL

2008-01-01

Background Observing co-expression between genes suggests that they are functionally coupled. Co-expression of orthologous gene pairs across species may improve function prediction beyond the level achieved in a single species. Results We used orthology between genes of the three different species S. cerevisiae, D. melanogaster, and C. elegans to combine co-expression across two species at a time. This led to increased function prediction accuracy when we incorporated expression data from either of the other two species and even further increased when conservation across both of the two other species was considered at the same time. Employing the conservation across species to incorporate abundant model organism data for the prediction of protein interactions in poorly characterized species constitutes a very powerful annotation method. Conclusion To be able to employ the most suitable co-expression distance measure for our analysis, we evaluated the ability of four popular gene co-expression distance measures to detect biologically relevant interactions between pairs of genes. For the expression datasets employed in our co-expression conservation analysis above, we used the GO and the KEGG PATHWAY databases as gold standards. While the differences between distance measures were small, Spearman correlation showed to give most robust results. PMID:18808668

Chronic Exercise Reduces CETP and Mesterolone Treatment Counteracts Exercise Benefits on Plasma Lipoproteins Profile: Studies in Transgenic Mice.

PubMed

Casquero, Andrea Camargo; Berti, Jairo Augusto; Teixeira, Laura Lauand Sampaio; de Oliveira, Helena Coutinho Franco

2017-12-01

Regular exercise and anabolic androgenic steroids have opposing effects on the plasma lipoprotein profile and risk of cardio-metabolic diseases in humans. Studies in humans and animal models show conflicting results. Here, we used a mice model genetically modified to mimic human lipoprotein profile and metabolism. They under-express the endogenous LDL receptor gene (R1) and express a human transgene encoding the cholesteryl ester transfer protein (CETP), normally absent in mice. The present study was designed to evaluate the independent and interactive effects of testosterone supplementation, exercise training and CETP expression on the plasma lipoprotein profile and CETP activity. CETP/R1 and R1 mice were submitted to a 6-week swimming training and mesterolone (MEST) supplementation in the last 3 weeks. MEST treatment increased markedly LDL levels (40%) in sedentary CETP/R1 mice and reduced HDL levels in exercised R1 mice (18%). A multifactorial ANOVA revealed the independent effects of each factor, as follows. CETP expression reduced HDL (21%) and increased non-HDL (15%) fractions. MEST treatment increased the VLDL concentrations (42%) regardless of other interventions. Exercise training reduced triacylglycerol (25%) and free fatty acids (20%), increased both LDL and HDL (25-33%), and reduced CETP (19%) plasma levels. Significant factor interactions showed that the increase in HDL induced by exercise is explained by reducing CETP activity and that MEST blunted the exercise-induced elevation of HDL-cholesterol. These results reinforce the positive metabolic effects of exercise, resolved a controversy about CETP response to exercise and evidenced MEST potency to counteract specific exercise benefits.

Fluoxetine prevents the development of depressive-like behavior in a mouse model of cancer related fatigue.

PubMed

Norden, Diana M; Devine, Raymond; Bicer, Sabahattin; Jing, Runfeng; Reiser, Peter J; Wold, Loren E; Godbout, Jonathan P; McCarthy, Donna O

2015-03-01

Cancer patients frequently suffer from fatigue, a complex syndrome associated with tiredness and depressed mood. Cancer-related fatigue (CRF) can be present at the time of diagnosis, escalates during treatment, and can persist for years after treatment. CRF negatively influences quality of life, limits functional independence, and is associated with decreased survival in patients with incurable disease. We have previously shown that increased pro-inflammatory cytokine expression in the brain contributes to depressive- and fatigue-like behaviors in a mouse model of CRF. Inflammatory cytokines increase the activity of indoleamine 2,3-dioxygenase (IDO) and kynurenine 3-monooxygenase (KMO), which competitively reduce serotonin synthesis. Reduced serotonin availability in the brain and increased production of alternative neuroactive metabolites of tryptophan are thought to contribute to the development of depression and fatigue. The purpose of this study was to determine the effects of fluoxetine, a selective serotonin reuptake inhibitor (SSRI), on brain cytokines and behavioral measures of fatigue and depression in tumor-bearing mice. Here we show that tumor growth increased brain expression of pro-inflammatory cytokines and KMO. Treatment with fluoxetine had no effect on tumor growth, muscle wasting, fatigue behavior, or cytokine expression in the brain. Fluoxetine, however, reduced depressive-like behaviors in tumor bearing mice. In conclusion, our data confirm that increased brain expression of pro-inflammatory cytokines is associated with tumor-induced fatigue- and depressive-like behaviors. However, it is possible to separate the effects of tumor growth on mood and fatigue-like behaviors using SSRIs such as fluoxetine. Copyright © 2014 Elsevier Inc. All rights reserved.

Upregulation of the sodium channel NaVβ4 subunit and its contributions to mechanical hypersensitivity and neuronal hyperexcitability in a rat model of radicular pain induced by local DRG inflammation

PubMed Central

Xie, Wenrui; Tan, Zhi-Yong; Barbosa, Cindy; Strong, Judith A.; Cummins, Theodore R.; Zhang, Jun-Ming

2016-01-01

High frequency spontaneous firing in myelinated sensory neurons plays a key role in initiating pain behaviors in several different models, including the radicular pain model in which the rat lumbar dorsal root ganglia (DRG) are locally inflamed. The sodium channel isoform NaV1.6 contributes to pain behaviors and spontaneous activity in this model. Among all the isoforms in adult DRG, NaV1.6 is the main carrier of TTX-sensitive resurgent Na currents that allow high-frequency firing. Resurgent currents flow after a depolarization or action potential, as a blocking particle exits the pore. In most neurons the regulatory β4 subunit is potentially the endogenous blocker. We used in vivo siRNA mediated knockdown of NaVβ4 to examine its role in the DRG inflammation model. NaVβ4 but not control siRNA almost completely blocked mechanical hypersensitivity induced by DRG inflammation. Microelectrode recordings in isolated whole DRGs showed that NaVβ4 siRNA blocked the inflammation-induced increase in spontaneous activity of Aβ neurons, and reduced repetitive firing and other measures of excitability. NaVβ4 was preferentially expressed in larger diameter cells; DRG inflammation increased its expression and this was reversed by NaVβ4 siRNA, based on immunohistochemistry and Western blotting. NaVβ4 siRNA also reduced immunohistochemical NaV1.6 expression. Patch clamp recordings of TTX-sensitive Na currents in acutely cultured medium diameter DRG neurons showed that DRG inflammation increased transient and especially resurgent current; effects blocked by NaVβ4 siRNA. NaVβ4 may represent a more specific target for pain conditions that depend on myelinated neurons expressing NaV1.6. PMID:26785322

Mouse repeated electroconvulsive seizure (ECS) does not reverse social stress effects but does induce behavioral and hippocampal changes relevant to electroconvulsive therapy (ECT) side-effects in the treatment of depression

PubMed Central

Sigrist, Hannes; Seifritz, Erich; Fikse, Lianne; Bosker, Fokko J.; Schoevers, Robert A.; Klein, Hans C.

2017-01-01

Electroconvulsive therapy (ECT) is an effective treatment for depression, but can have negative side effects including amnesia. The mechanisms of action underlying both the antidepressant and side effects of ECT are not well understood. An equivalent manipulation that is conducted in experimental animals is electroconvulsive seizure (ECS). Rodent studies have provided valuable insights into potential mechanisms underlying the antidepressant and side effects of ECT. However, relatively few studies have investigated the effects of ECS in animal models with a depression-relevant manipulation such as chronic stress. In the present study, mice were first exposed to chronic social stress (CSS) or a control procedure for 15 days followed by ECS or a sham procedure for 10 days. Behavioral effects were investigated using an auditory fear conditioning (learning) and expression (memory) test and a treadmill-running fatigue test. Thereafter, immunohistochemistry was conducted on brain material using the microglial marker Iba-1 and the cholinergic fibre marker ChAT. CSS did not increase fear learning and memory in the present experimental design; in both the control and CSS mice ECS reduced fear learning and fear memory expression. CSS induced the expected fatigue-like effect in the treadmill-running test; ECS induced increased fatigue in CSS and control mice. In CSS and control mice ECS induced inflammation in hippocampus in terms of increased expression of Iba-1 in radiatum of CA1 and CA3. CSS and ECS both reduced acetylcholine function in hippocampus as indicated by decreased expression of ChAT in several hippocampal sub-regions. Therefore, CSS increased fatigue and reduced hippocampal ChAT activity and, rather than reversing these effects, a repeated ECS regimen resulted in impaired fear learning-memory, increased fatigue, increased hippocampal Iba-1 expression, and decreased hippocampal ChAT expression. As such, the current model does not provide insights into the mechanism of ECT antidepressant function but does provide evidence for pathophysiological mechanisms that might contribute to important ECT side-effects. PMID:28910337

Mouse repeated electroconvulsive seizure (ECS) does not reverse social stress effects but does induce behavioral and hippocampal changes relevant to electroconvulsive therapy (ECT) side-effects in the treatment of depression.

PubMed

van Buel, Erin M; Sigrist, Hannes; Seifritz, Erich; Fikse, Lianne; Bosker, Fokko J; Schoevers, Robert A; Klein, Hans C; Pryce, Christopher R; Eisel, Ulrich Lm

2017-01-01

Electroconvulsive therapy (ECT) is an effective treatment for depression, but can have negative side effects including amnesia. The mechanisms of action underlying both the antidepressant and side effects of ECT are not well understood. An equivalent manipulation that is conducted in experimental animals is electroconvulsive seizure (ECS). Rodent studies have provided valuable insights into potential mechanisms underlying the antidepressant and side effects of ECT. However, relatively few studies have investigated the effects of ECS in animal models with a depression-relevant manipulation such as chronic stress. In the present study, mice were first exposed to chronic social stress (CSS) or a control procedure for 15 days followed by ECS or a sham procedure for 10 days. Behavioral effects were investigated using an auditory fear conditioning (learning) and expression (memory) test and a treadmill-running fatigue test. Thereafter, immunohistochemistry was conducted on brain material using the microglial marker Iba-1 and the cholinergic fibre marker ChAT. CSS did not increase fear learning and memory in the present experimental design; in both the control and CSS mice ECS reduced fear learning and fear memory expression. CSS induced the expected fatigue-like effect in the treadmill-running test; ECS induced increased fatigue in CSS and control mice. In CSS and control mice ECS induced inflammation in hippocampus in terms of increased expression of Iba-1 in radiatum of CA1 and CA3. CSS and ECS both reduced acetylcholine function in hippocampus as indicated by decreased expression of ChAT in several hippocampal sub-regions. Therefore, CSS increased fatigue and reduced hippocampal ChAT activity and, rather than reversing these effects, a repeated ECS regimen resulted in impaired fear learning-memory, increased fatigue, increased hippocampal Iba-1 expression, and decreased hippocampal ChAT expression. As such, the current model does not provide insights into the mechanism of ECT antidepressant function but does provide evidence for pathophysiological mechanisms that might contribute to important ECT side-effects.

White adipose tissue IFN-γ expression and signalling along the progression of rodent cancer cachexia.

PubMed

Yamashita, Alex Shimura; das Neves, Rodrigo Xavier; Rosa-Neto, José Cesar; Lira, Fábio Dos Santos; Batista, Miguel Luís; Alcantara, Paulo Sérgio; Otoch, José Pinhata; Seelaender, Marília

2017-01-01

Cachexia is associated with increased morbidity and mortality in cancer. The White adipose tissue (WAT) synthesizes and releases several pro-inflammatory cytokines that play a role in cancer cachexia-related systemic inflammation. IFN-γ is a pleiotropic cytokine that regulates several immune and metabolic functions. To assess whether IFN-γ signalling in different WAT pads is modified along cancer-cachexia progression, we evaluated IFN-γ receptors expression (IFNGR1 and IFNGR2) and IFN-γ protein expression in a rodent model of cachexia (7, 10, and 14days after tumour implantation). IFN-γ protein expression was heterogeneously modulated in WAT, with increases in the mesenteric pad and decreased levels in the retroperitoneal depot along cachexia progression. Ifngr1 was up-regulated 7days after tumour cell injection in mesenteric and epididymal WAT, but the retroperitoneal depot showed reduced Ifngr1 gene expression. Ifngr2 gene expression was increased 7 and 14days after tumour inoculation in mesenteric WAT. The results provide evidence that changes in IFN-γ expression and signalling may be perceived at stages preceding refractory cachexia, and therefore, might be employed as a means to assess the early stage of the syndrome. Copyright © 2016 Elsevier Ltd. All rights reserved.

Stimulation of glycerol kinase in grass carp preadipocytes by EPA.

PubMed

Lei, Caixia; Tian, Jingjing; Ji, Hong

2017-06-01

This study was conducted to assess the effect of eicosapentaenoic acid (EPA) on grass carp preadipocyte glycerol kinase (GyK) expression, as well as to explore the mechanism. Here, we cloned partial sequence of grass carp GyK gene and analyzed its tissue distribution. The result showed that GyK gene expressed most in the liver, followed by adipose tissue and the kidney. Besides, 400 μM oleic acid (18:1n-9, OA) was used to establish a hypertrophic preadipocyte model. GyK gene expression and enzyme activity were significantly enhanced after model cells were treated with 100 μM eicosapentaenoic acid (20:5n-3, EPA) for 6, 12, and 24 h. Meanwhile, peroxisome proliferative-activated receptor (PPAR)γ, adipose triglyceride lipase (ATGL), and the two isoforms of grass carp HSL gene were first identified by Sun et al (2016), and they defined the two isoforms as HSLa and HSLb. Therefore, maybe HSLa and HSLb are appropriate.. The content of triglyceride was dramatically increased by EPA treatment for 24 h. Further, a competitive ATGL antagonist, HY-15859, attenuated the increase in GyK induced by EPA at 12 h. Surprisingly, the enhanced lipolysis and PPARγ gene expression induced by serum deprivation were paralleled by an increase in GyK gene expression, whereas a stabilization in GyK enzyme activity. Other fatty acids, including docosahexaenoic acid, alpha-linolenic acid, linoleic acid, and OA also promoted GyK gene expression. Moreover, an irreversible PPARγ antagonist, GW9662, was used to investigate the role of PPARγ in GyK induction. Data showed that GW9662 abolished the induction of GyK by EPA at 12 h. Together, these data suggested that EPA elevated grass carp preadipocytes GyK expression. ATGL and PPARγ contributed to the induction of GyK. PPARγ may be a key regulator in response to GyK expression induced by EPA.

Vitamin K2 alleviates type 2 diabetes in rats by induction of osteocalcin gene expression.

PubMed

Hussein, Atef G; Mohamed, Randa H; Shalaby, Sally M; Abd El Motteleb, Dalia M

2018-03-01

The biological mechanisms behind the association between vitamin K (Vit K) and glucose metabolism are uncertain. We aimed to analyze the expression of insulin 1 (Ins 1), insulin 2 (Ins 2) and cyclin D2, the expression of adiponectin and UCP-1 . In addition, we aimed to estimate the doses of Vit K2 able to affect various aspects of glucose and energy metabolism in type 2 diabetes. Thirty adult male rats were allocated equally into five groups: control group, diabetes mellitus group, and groups 3, 4, and 5, which received Vit K 2 at three daily dose levels (10, 15, and 30 mg/kg, respectively) for 8 wk. At the end of the study, blood samples were collected to quantify total osteocalcin, fasting plasma glucose, fasting insulin, and relevant variables. The expression of OC, Ins 1, Ins 2, cyclin D2, adiponectin, UCP-1 genes was analyzed by real-time polymerase chain reaction. After administration of Vit K 2 , a dose-dependent decrease in fasting plasma glucose, hemoglobin A1c and homeostatic model assessment method insulin resistance, and a dose-dependent increase in fasting insulin and homeostatic model assessment method β cell function levels, when compared with diabetes mellitus rats, were detected. There was significant upregulation of OC, Ins 1, Ins 2, or cyclin D2 gene expression in the three treated groups in a dose-dependent manner when compared with the diabetic rats. However, expression of adiponectin and UCP-1 were significantly increased at the highest dose (30 mg/kg daily) only. Vit K 2 administration could improve glycemic status in type 2 diabetic rats by induction of OC gene expression. Osteocalcin could increase β-cell proliferation, energy expenditure, and adiponectin expression. Different concentrations of Vit K 2 were required to affect glucose metabolism and insulin sensitivity. Copyright © 2017 Elsevier Inc. All rights reserved.

Regulation of Facilitative Glucose Transporters and AKT/MAPK/PRKAA Signaling via Estradiol and Progesterone in the Mouse Uterine Epithelium1

PubMed Central

Kim, Sung Tae; Moley, Kelle H.

2009-01-01

Adequate uterine glucose metabolism is an essential part of embryo implantation and the development of an adequate utero-fetal environment. However, expression of facilitative glucose transporters (GLUTs [solute transporter family SLC2A]) and AKT/MAPK/PRKAA (PRKAA) signaling has not been described in the mouse uterine cells, to our knowledge. The objective of this study was to determine the hormonal regulation of SLC2A protein expression and AKT/MAPK/PRKAA signaling in the mouse uterine epithelial cells during estrous cycles and peri-implantation periods. SLC2As 1, 4, 8, and 9B were highly expressed in the luminal and glandular epithelia of estrous stage. In metestrous and diestrous stages, expression of SLC2As 1, 4, 8, and 9B was lower than that in proestrous stage. Levels of activated phospho-AKT (p-AKT), p-MAPK3, and p-MAPK1 also varied during the estrous cycle. Estrogen and progesterone injection in an ovariectomized mouse (delayed implantation model) resulted in a decrease and an increase, respectively, in expression of GLUTs in the luminal epithelial cells of the uterus. The expression of SLC2A1, SLC2A8, SLC2A9B, p-AKT, p-MAPK3/1, and p-PRKAA was increased in the decidual region of the implantation sites and was significantly increased in the uterus of activated implantation. Using an artificial decidualization mouse model, it was also demonstrated that expression of the same GLUTs, p-MAPK3/1, and p-PRKAA was dramatically higher in the decidualized uteri than that in the control uteri. These results suggest that steroid hormones regulate expression of uterine epithelial GLUTs possibly through AKT/MAPK/PRKAA signaling pathways and that glucose utilization may have an important role in decidualization and possibly in the maintenance of pregnancy. PMID:19208550

High levels of the extracellular matrix proteoglycan decorin are associated with inhibition of testicular function

PubMed Central

Adam, Marion; Urbanski, Henryk F.; Garyfallou, Vasilios T.; Welsch, Ulrich; Köhn, Frank M.; Schwarzer, J. Ullrich; Strauss, Leena; Poutanen, Matti; Mayerhofer, Artur

2011-01-01

Decorin (DCN), a component of the extracellular matrix of the peritubular wall and the interstitial areas of the human testis, can interact with growth factor (GF) signaling, thereby blocking downstream actions of GFs. In the present study the expression and regulation of DCN using both human testes and two experimental animal models, namely the rhesus monkey and mouse, were examined. DCN protein was present in peritubular and interstitial areas of adult human and monkey testes, while it was almost undetectable in adult wild-type mice. Interestingly, the levels and sites of testicular DCN expression in the monkeys were inversely correlated with testicular maturation markers. A strong DCN expression associated with the abundant connective tissue of the interstitial areas in the postnatal through prepubertal phases was observed. In adult and old monkeys the DCN pattern was similar to the one in normal human testes, presenting strong expression at the peritubular region. In the testes of both infertile men and in a mouse model of inflammation associated infertility (aromatase-overexpressing transgenic mice), the fibrotic changes and increased numbers of Tumor necrosis factor (TNF)-α-producing immune cells were shown to be associated with increased production of DCN. Furthermore, studies with human testicular peritubular cells isolated from fibrotic testis indicated that TNF-α significantly increased DCN production. The data, thus, show that an increased DCN level is associated with impaired testicular function, supporting our hypothesis that DCN interferes with paracrine signaling of the testis in health and disease. PMID:22413766

Binge alcohol alters PNPLA3 levels in liver through epigenetic mechanism involving histone H3 acetylation.

PubMed

Restrepo, Ricardo J; Lim, Robert W; Korthuis, Ronald J; Shukla, Shivendra D

2017-05-01

The human PNPLA3 (patatin-like phospholipase domain-containing 3) gene codes for a protein which is highly expressed in adipose tissue and liver, and is implicated in lipid homeostasis. While PNPLA3 protein contains regions homologous to functional lipolytic proteins, the regulation of its tissue expression is reflective of lipogenic genes. A naturally occurring genetic variant of PNPLA3 in humans has been linked to increased susceptibility to alcoholic liver disease. We have examined the modulatory effect of alcohol on PNPLA3 protein and mRNA expression as well as the association of its gene promoter with acetylated histone H3K9 by chromatin immunoprecipitation (ChIP) assay in rat hepatocytes in vitro, and in vivo in mouse and rat models of acute binge, chronic, and chronic followed by acute binge ethanol administration. Protein expression of PNPLA3 was significantly increased by alcohol in all three models used. PNPLA3 mRNA also increased, albeit to a varying degree. ChIP assay using H3AcK9 antibody showed increased association with the promoter of PNPLA3 in hepatocytes and in mouse liver. This was less evident in rat livers in vivo except under chronic treatment. It is concluded for the first time that histone acetylation plays a role in the modulation of PNPLA3 levels in the liver exposed to binge ethanol both in vitro and in vivo. Copyright © 2017 Elsevier Inc. All rights reserved.

Binge alcohol alters PNPLA3 levels in liver through epigenetic mechanism involving histone H3 acetylation

PubMed Central

Restrepo, Ricardo J.; Lim, Robert W.; Korthuis, Ronald J.; Shukla, Shivendra D.

2017-01-01

The human PNPLA3 (patatin-like phospholipase domain-containing 3) gene codes for a protein which is highly expressed in adipose tissue and liver, and is implicated in lipid homeostasis. While PNPLA3 protein contains regions homologous to functional lipolytic proteins, the regulation of its tissue expression is reflective of lipogenic genes. A naturally occurring genetic variant of PNPLA3 in humans has been linked to increased susceptibility to alcoholic liver disease. We have examined the modulatory effect of alcohol on PNPLA3 protein and mRNA expression as well as the association of its gene promoter with acetylated histone H3K9 by chromatin immunoprecipitation (ChIP) assay in rat hepatocytes in vitro, and in vivo in mouse and rat models of acute binge, chronic, and chronic followed by acute binge ethanol administration. Protein expression of PNPLA3 was significantly increased by alcohol in all three models used. PNPLA3 mRNA also increased, albeit to a varying degree. ChIP assay using H3AcK9 antibody showed increased association with the promoter of PNPLA3 in hepatocytes and in mouse liver. This was less evident in rat livers in vivo except under chronic treatment. It is concluded for the first time that histone acetylation plays a role in the modulation of PNPLA3 levels in the liver exposed to binge ethanol both in vitro and in vivo. PMID:28433418

Aromatase Blockade Is Associated With Increased Mortality in Acute Illness in Male Mice.

PubMed

Connerney, Jeannette J; Spratt, Daniel I

2017-09-01

The increase in circulating estrogen levels with acute illness in humans is accompanied by increased aromatase expression in adipose tissue and increased peripheral aromatization of estrogens to androgens. Animal studies indicate that estrogen may be beneficial in acute illness. We hypothesized that blockade of aromatase in acute illness would decrease survival. Prospective sham controlled. Maine Medical Center Research Institute animal facility. Six- to 8-week-old male black 6 mice. Mice underwent cecal ligation and puncture (CLP) to induce acute illness and were administered letrozole to block aromatase or saline. Mice undergoing sham surgery with or without letrozole served as controls. Adipose and cardiovascular tissue was harvested for preliminary evaluation of aromatase expression. Survival was the main outcome measurement. Evidence for aromatase expression in tissue samples was assessed using western blot and/or immunohistochemistry. With aromatase blockade, survival in CLP mice was decreased ( P = 0.04). The presence of aromatase in adipose tissue was observed by western blot in CLP but not control mice. Similarly, the presence of aromatase was observed in cardiac tissue of CLP but not in control mice. The decreased survival during sepsis with aromatase blockade suggests that this response to acute illness may be important both physiologically and clinically. The preliminary observation of aromatase expression in adipose and cardiovascular tissue during acute illness in this mouse model indicates that this model has parallels to human physiology and may be useful for further studying the aromatase response to acute illness.

[Intramuscular injection of lentivirus-mediated EPAS1 gene improves hind limb ischemia and its mechanism in a rat model of peripheral artery vascular disease].

PubMed

Wang, Zhihong; Gu, Hongbin; Yang, Fan; Xie, Huajie; Sheng, Lei; Li, Mingfei

2017-11-01

Objective To investigate the effect of over-expressed endothelial Per-Arnt-Sim domain protein 1 (EPAS1) on peripheral arterial disease (PAD) in a rat model. Methods PAD rat model was established by external iliac artery ligation followed by lentivirus-mediated EPAS1 gene injection into rat right adductor magnus. The models were evaluated by quantitative analysis of gait disturbance. The changes of blood flow in the posterior extremity of the rats were detected using laser Doppler. The expressions of EPAS1, hepatocyte growth factor (HGF), basic fibroblast growth factor (bFGF), and vascular endothelial growth factor (VEGF) mRNAs were tested by real-time quantitative PCR. The expression of α-smooth muscle actin (αSMA) was detected by immunohistochemical staining. Results Compared with lenti-EGFP group, rat hind limb function and circulation got recovered obviously 7 days after lenti-EPAS1 injection. The mRNA expressions of EPAS1, HGF, bFGF, and VEGF were up-regulated in the lenti-EPAS1-treated sites.The expression of αSMA showed an obvious increase in the lenti-EPAS1-treated muscles. Conclusion Over-expressed lenti-EPAS1 can promote angiogenesis via the up-regulation of EPAS1-related angiogenic factors in the muscles of the affected hind limb and reduce gait disturbance.

RNA Sequencing Identifies Novel Translational Biomarkers of Kidney Fibrosis

PubMed Central

Craciun, Florin L.; Bijol, Vanesa; Ajay, Amrendra K.; Rao, Poornima; Kumar, Ramya K.; Hutchinson, John; Hofmann, Oliver; Joshi, Nikita; Luyendyk, James P.; Kusebauch, Ulrike; Moss, Christopher L.; Srivastava, Anand; Himmelfarb, Jonathan; Waikar, Sushrut S.; Moritz, Robert L.

2016-01-01

CKD is the gradual, asymptomatic loss of kidney function, but current tests only identify CKD when significant loss has already happened. Several potential biomarkers of CKD have been reported, but none have been approved for preclinical or clinical use. Using RNA sequencing in a mouse model of folic acid-induced nephropathy, we identified ten genes that track kidney fibrosis development, the common pathologic finding in patients with CKD. The gene expression of all ten candidates was confirmed to be significantly higher (approximately ten- to 150-fold) in three well established, mechanistically distinct mouse models of kidney fibrosis than in models of nonfibrotic AKI. Protein expression of these genes was also high in the folic acid model and in patients with biopsy-proven kidney fibrosis. mRNA expression of the ten genes increased with increasing severity of kidney fibrosis, decreased in response to therapeutic intervention, and increased only modestly (approximately two- to five-fold) with liver fibrosis in mice and humans, demonstrating specificity for kidney fibrosis. Using targeted selected reaction monitoring mass spectrometry, we detected three of the ten candidates in human urine: cadherin 11 (CDH11), macrophage mannose receptor C1 (MRC1), and phospholipid transfer protein (PLTP). Furthermore, urinary levels of each of these three proteins distinguished patients with CKD (n=53) from healthy individuals (n=53; P<0.05). In summary, we report the identification of urinary CDH11, MRC1, and PLTP as novel noninvasive biomarkers of CKD. PMID:26449608

Histone deacetylation contributes to low extracellular superoxide dismutase expression in human idiopathic pulmonary arterial hypertension

PubMed Central

Woods, Crystal; Stearman, Robert S.; Venkataraman, Sujatha; Ferguson, Bradley S.; Swain, Kalin; Bowler, Russell P.; Geraci, Mark W.; Ihida-Stansbury, Kaori; Stenmark, Kurt R.; McKinsey, Timothy A.; Domann, Frederick E.

2016-01-01

Epigenetic mechanisms, including DNA methylation and histone acetylation, regulate gene expression in idiopathic pulmonary arterial hypertension (IPAH). These mechanisms can modulate expression of extracellular superoxide dismutase (SOD3 or EC-SOD), a key vascular antioxidant enzyme, and loss of vascular SOD3 worsens outcomes in animal models of pulmonary arterial hypertension. We hypothesized that SOD3 gene expression is decreased in patients with IPAH due to aberrant DNA methylation and/or histone deacetylation. We used lung tissue and pulmonary artery smooth muscle cells (PASMC) from subjects with IPAH at transplantation and from failed donors (FD). Lung SOD3 mRNA expression and activity was decreased in IPAH vs. FD. In contrast, mitochondrial SOD (Mn-SOD or SOD2) protein expression was unchanged and intracellular SOD activity was unchanged. Using bisulfite sequencing in genomic lung or PASMC DNA, we found the methylation status of the SOD3 promoter was similar between FD and IPAH. Furthermore, treatment with 5-aza-2′-deoxycytidine did not increase PASMC SOD3 mRNA, suggesting DNA methylation was not responsible for PASMC SOD3 expression. Though total histone deacetylase (HDAC) activity, histone acetyltransferase (HAT) activity, acetylated histones, and acetylated SP1 were similar between IPAH and FD, treatment with two selective class I HDAC inhibitors increased SOD3 only in IPAH PASMC. Class I HDAC3 siRNA also increased SOD3 expression. Trichostatin A, a pan-HDAC inhibitor, decreased proliferation in IPAH, but not in FD PASMC. These data indicate that histone deacetylation, specifically via class I HDAC3, decreases SOD3 expression in PASMC and HDAC inhibitors may protect IPAH in part by increasing PASMC SOD3 expression. PMID:27233998

Valsartan Upregulates Kir2.1 in Rats Suffering from Myocardial Infarction via Casein Kinase 2.

PubMed

Li, Xinran; Hu, Hesheng; Wang, Ye; Xue, Mei; Li, Xiaolu; Cheng, Wenjuan; Xuan, Yongli; Yin, Jie; Yang, Na; Yan, Suhua

2015-06-01

Myocardial infarction (MI) results in an increased susceptibility to ventricular arrhythmias, due in part to decreased inward-rectifier K+ current (IK1), which is mediated primarily by the Kir2.1 protein. The use of renin-angiotensin-aldosterone system antagonists is associated with a reduced incidence of ventricular arrhythmias. Casein kinase 2 (CK2) binds and phosphorylates SP1, a transcription factor of KCNJ2 that encodes Kir2.1. Whether valsartan represses CK2 activation to ameliorate IK1 remodeling following MI remains unclear. Wistar rats suffering from MI received either valsartan or saline for 7 days. The protein levels of CK2 and Kir2.1 were each detected via a Western blot analysis. The mRNA levels of CK2 and Kir2.1 were each examined via quantitative real-time PCR. CK2 expression was higher at the infarct border; and was accompanied by a depressed IK1/Kir2.1 protein level. Additionally, CK2 overexpression suppressed KCNJ2/Kir2.1 expression. By contrast, CK2 inhibition enhanced KCNJ2/Kir2.1 expression, establishing that CK2 regulates KCNJ2 expression. Among the rats suffering from MI, valsartan reduced CK2 expression and increased Kir2.1 expression compared with the rats that received saline treatment. In vitro, hypoxia increased CK2 expression and valsartan inhibited CK2 expression. The over-expression of CK2 in cells treated with valsartan abrogated its beneficial effect on KCNJ2/Kir2.1. AT1 receptor antagonist valsartan reduces CK2 activation, increases Kir2.1 expression and thereby ameliorates IK1 remodeling after MI in the rat model.

Different Expression Levels of Human Mutant Ubiquitin B+1 (UBB+1) Can Modify Chronological Lifespan or Stress Resistance of Saccharomyces cerevisiae

PubMed Central

Muñoz-Arellano, Ana Joyce; Chen, Xin; Molt, Andrea; Meza, Eugenio; Petranovic, Dina

2018-01-01

The ubiquitin-proteasome system (UPS) is the main pathway responsible for the degradation of misfolded proteins, and its dysregulation has been implicated in several neurodegenerative diseases, including Alzheimer’s disease (AD). UBB+1, a mutant variant of ubiquitin B, was found to accumulate in neurons of AD patients and it has been linked to UPS dysfunction and neuronal death. Using the yeast Saccharomyces cerevisiae as a model system, we constitutively expressed UBB+1 to evaluate its effects on proteasome function and cell death, particularly under conditions of chronological aging. We showed that the expression of UBB+1 caused inhibition of the three proteasomal proteolytic activities (caspase-like (β1), trypsin-like (β2) and chymotrypsin-like (β5) activities) in yeast. Interestingly, this inhibition did not alter cell viability of growing cells. Moreover, we showed that cells expressing UBB+1 at lower level displayed an increased capacity to degrade induced misfolded proteins. When we evaluated cells during chronological aging, UBB+1 expression at lower level, prevented cells to accumulate reactive oxygen species (ROS) and avert apoptosis, dramatically increasing yeast life span. Since proteasome inhibition by UBB+1 has previously been shown to induce chaperone expression and thus protect against stress, we evaluated our UBB+1 model under heat shock and oxidative stress. Higher expression of UBB+1 caused thermotolerance in yeast due to induction of chaperones, which occurred to a lesser extent at lower expression level of UBB+1 (where we observed the phenotype of extended life span). Altering UPS capacity by differential expression of UBB+1 protects cells against several stresses during chronological aging. This system can be valuable to study the effects of UBB+1 on misfolded proteins involved in neurodegeneration and aging.

Drug resistance is conferred on the model yeast Saccharomyces cerevisiae by expression of full-length melanoma-associated human ATP-binding cassette transporter ABCB5.

PubMed

Keniya, Mikhail V; Holmes, Ann R; Niimi, Masakazu; Lamping, Erwin; Gillet, Jean-Pierre; Gottesman, Michael M; Cannon, Richard D

2014-10-06

ABCB5, an ATP-binding cassette (ABC) transporter, is highly expressed in melanoma cells, and may contribute to the extreme resistance of melanomas to chemotherapy by efflux of anti-cancer drugs. Our goal was to determine whether we could functionally express human ABCB5 in the model yeast Saccharomyces cerevisiae, in order to demonstrate an efflux function for ABCB5 in the absence of background pump activity from other human transporters. Heterologous expression would also facilitate drug discovery for this important target. DNAs encoding ABCB5 sequences were cloned into the chromosomal PDR5 locus of a S. cerevisiae strain in which seven endogenous ABC transporters have been deleted. Protein expression in the yeast cells was monitored by immunodetection using both a specific anti-ABCB5 antibody and a cross-reactive anti-ABCB1 antibody. ABCB5 function in recombinant yeast cells was measured by determining whether the cells possessed increased resistance to known pump substrates, compared to the host yeast strain, in assays of yeast growth. Three ABCB5 constructs were made in yeast. One was derived from the ABCB5-β mRNA, which is highly expressed in human tissues but is a truncation of a canonical full-size ABC transporter. Two constructs contained full-length ABCB5 sequences: either a native sequence from cDNA or a synthetic sequence codon-harmonized for S. cerevisiae. Expression of all three constructs in yeast was confirmed by immunodetection. Expression of the codon-harmonized full-length ABCB5 DNA conferred increased resistance, relative to the host yeast strain, to the putative substrates rhodamine 123, daunorubicin, tetramethylrhodamine, FK506, or clorgyline. We conclude that full-length ABCB5 can be functionally expressed in S. cerevisiae and confers drug resistance.

Metabolic regulation of ghrelin O-acyl transferase (GOAT) expression in the mouse hypothalamus, pituitary, and stomach.

PubMed

Gahete, Manuel D; Córdoba-Chacón, Jose; Salvatori, Roberto; Castaño, Justo P; Kineman, Rhonda D; Luque, Raul M

2010-04-12

Ghrelin acts as an endocrine link connecting physiological processes regulating food intake, body composition, growth, and energy balance. Ghrelin is the only peptide known to undergo octanoylation. The enzyme mediating this process, ghrelin O-acyltransferase (GOAT), is expressed in the gastrointestinal tract (GI; primary source of circulating ghrelin) as well as other tissues. The present study demonstrates that stomach GOAT mRNA levels correlate with circulating acylated-ghrelin levels in fasted and diet-induced obese mice. In addition, GOAT was found to be expressed in both the pituitary and hypothalamus (two target tissues of ghrelin's actions), and regulated in response to metabolic status. Using primary pituitary cell cultures as a model system to study the regulation of GOAT expression, we found that acylated-ghrelin, but not desacyl-ghrelin, increased GOAT expression. In addition, growth-hormone-releasing hormone (GHRH) and leptin increased, while somatostatin (SST) decreased GOAT expression. The physiologic relevance of these later results is supported by the observation that pituitary GOAT expression in mice lacking GHRH, SST and leptin showed opposite changes to those observed after in vitro treatment with the corresponding peptides. Therefore, it seems plausible that these hormones directly contribute to the regulation of pituitary GOAT. Interestingly, in all the models studied, pituitary GOAT expression paralleled changes in the expression of a dominant spliced-variant of ghrelin (In2-ghrelin) and therefore this transcript may be a primary substrate for pituitary GOAT. Collectively, these observations support the notion that the GI tract is not the only source of acylated-ghrelin, but in fact locally produced des-acylated-ghrelin could be converted to acylated-ghrelin within target tissues by locally active GOAT, to mediate its tissue-specific effects.

Spermine oxidase maintains basal skeletal muscle gene expression and fiber size and is strongly repressed by conditions that cause skeletal muscle atrophy

PubMed Central

Bongers, Kale S.; Fox, Daniel K.; Kunkel, Steven D.; Stebounova, Larissa V.; Murry, Daryl J.; Pufall, Miles A.; Ebert, Scott M.; Dyle, Michael C.; Bullard, Steven A.; Dierdorff, Jason M.

2014-01-01

Skeletal muscle atrophy is a common and debilitating condition that remains poorly understood at the molecular level. To better understand the mechanisms of muscle atrophy, we used mouse models to search for a skeletal muscle protein that helps to maintain muscle mass and is specifically lost during muscle atrophy. We discovered that diverse causes of muscle atrophy (limb immobilization, fasting, muscle denervation, and aging) strongly reduced expression of the enzyme spermine oxidase. Importantly, a reduction in spermine oxidase was sufficient to induce muscle fiber atrophy. Conversely, forced expression of spermine oxidase increased muscle fiber size in multiple models of muscle atrophy (immobilization, fasting, and denervation). Interestingly, the reduction of spermine oxidase during muscle atrophy was mediated by p21, a protein that is highly induced during muscle atrophy and actively promotes muscle atrophy. In addition, we found that spermine oxidase decreased skeletal muscle mRNAs that promote muscle atrophy (e.g., myogenin) and increased mRNAs that help to maintain muscle mass (e.g., mitofusin-2). Thus, in healthy skeletal muscle, a relatively low level of p21 permits expression of spermine oxidase, which helps to maintain basal muscle gene expression and fiber size; conversely, during conditions that cause muscle atrophy, p21 expression rises, leading to reduced spermine oxidase expression, disruption of basal muscle gene expression, and muscle fiber atrophy. Collectively, these results identify spermine oxidase as an important positive regulator of muscle gene expression and fiber size, and elucidate p21-mediated repression of spermine oxidase as a key step in the pathogenesis of skeletal muscle atrophy. PMID:25406264

Association between renal iron accumulation and renal interstitial fibrosis in a rat model of chronic kidney disease.

PubMed

Naito, Yoshiro; Fujii, Aya; Sawada, Hisashi; Oboshi, Makiko; Iwasaku, Toshihiro; Okuhara, Yoshitaka; Morisawa, Daisuke; Eguchi, Akiyo; Hirotani, Shinichi; Masuyama, Tohru

2015-07-01

Iron accumulation is associated with the pathophysiology of chronic kidney disease (CKD). Renal fibrosis is a final common feature that contributes to the progression of CKD; however, little is known about the association between renal iron accumulation and renal interstitial fibrosis in CKD. Here we investigate the effects of iron chelation on renal interstitial fibrosis in a rat model of CKD. CKD was induced by 5/6 nephrectomy in Sprague-Dawley rats. At 8 weeks after operation, 5/6 nephrectomized rats were administered an oral iron chelator, deferasirox (DFX), in chow for 8 weeks. Other CKD rats were given a normal diet. Sham-operative rats given a normal diet served as a control. CKD rats exhibited hypertension, glomerulosclerosis and renal interstitial fibrosis. Iron chelation with DFX did not change hypertension and glomerulosclerosis; however, renal interstitial fibrosis was attenuated in CKD rats. Consistent with these findings, renal gene expression of collagen type III and transforming growth factor-β was increased in CKD rats compared with the controls, while iron chelation suppressed these increments. In addition, a decrease in vimentin along an increase in E-cadherin in renal gene expression was observed in CKD rats with iron chelation. CKD rats also showed increased CD68-positive cells in the kidney, whereas its increase was attenuated by iron deprivation. Similarly, increased renal gene expression of CD68, tumor necrosis factor-α and monocyte chemoattractant protein-1 was suppressed in CKD rats with iron chelation. Renal iron accumulation seems to be associated with renal interstitial fibrosis in a rat model of CKD.

Protective Effects and Mechanism of Meretrix meretrix Oligopeptides against Nonalcoholic Fatty Liver Disease

PubMed Central

Huang, Fangfang; Zhao, Shasha; Yu, Fangmiao; Yang, Zuisu; Ding, Guofang

2017-01-01

Meretrix meretrix oligopeptides (MMO) derived from shellfish have important medicinal properties. We previously obtained MMO from alcalase by hydrolysis processes. Here we examine the protective effects of MMO against nonalcoholic fatty liver disease (NAFLD) and explored the underlying mechanism. Human Chang liver cells were used in our experiments after exposure to palmitic acid at a final concentration of 15 μg/mL for 48 h to induce an overload of fatty acid as NAFLD model cells. Treatment with MMO for 24 h increased the viability of the NAFLD model cells by inhibiting apoptosis. MMO alleviated oxidative stress in the NAFLD model cells by preserving reactive oxygen species activity and increasing malondialdehyde and superoxide dismutase activity. MMO improved mitochondrial dysfunction by decreasing the mitochondrial membrane potential and increasing the activities of Na+/K+-ATPase and Ca2+/Mg2+-ATPase. In addition, MMO inhibited the activation of cell death-related pathways, based on reduced p-JNK, Bax expression, tumor necrosis factor-α, caspase-9, and caspase-3 activity in the NAFLD model cells, and Bcl-2 expression was enhanced in the NAFLD model cells compared with the control group. These findings indicate that MMO have antioxidant and anti-apoptotic effects on NAFLD model cells and may thus exert protective effects against NAFLD. PMID:28216552

Carcinogen-induced mdr overexpression is associated with xenobiotic resistance in rat preneoplastic liver nodules and hepatocellular carcinomas.

PubMed

Fairchild, C R; Ivy, S P; Rushmore, T; Lee, G; Koo, P; Goldsmith, M E; Myers, C E; Farber, E; Cowan, K H

1987-11-01

We have previously reported the isolation of a human breast cancer cell line resistant to doxorubicin (adriamycin; AdrR MCF-7 cells) that has also developed the phenotype of multidrug resistance (MDR). MDR in this cell line is associated with increased expression of mdr (P glycoprotein) gene sequences. The development of MDR in AdrR MCF-7 cells is also associated with changes in the expression of several phase I and phase II drug-detoxifying enzymes. These changes are remarkably similar to those associated with development of xenobiotic resistance in rat hyperplastic liver nodules, a well-studied model system of chemical carcinogenesis. Using an mdr-encoded cDNA sequence isolated from AdrR MCF-7 cells, we have examined the expression of mdr sequences in rat livers under a variety of experimental conditions. The expression of mdr increased 3-fold in regenerating liver. It was also elevated (3- to 12-fold) in several different samples of rat hyperplastic nodules and in four of five hepatomas that developed in this system. This suggests that overexpression of mdr, a gene previously associated with resistance to antineoplastic agents, may also be involved in the development of resistance to xenobiotics in rat hyperplastic nodules. In addition, although the acute administration of 2-acetylaminofluorene induced an 8-fold increase in hepatic mdr-encoded RNA, performance of a partial hepatectomy either before or after administration of 2-acetylaminofluorene resulted in a greater than 80-fold increase in mdr gene expression over that in normal untreated livers. This represents an important in vivo model system in which to study the acute regulation of this drug resistance gene.

Serum response factor: positive and negative regulation of an epithelial gene expression network in the destrin mutant cornea

PubMed Central

Kawakami-Schulz, Sharolyn V.; Verdoni, Angela M.; Sattler, Shannon G.; Jessen, Erik; Kao, Winston W.-Y.; Ikeda, Akihiro

2014-01-01

Increased angiogenesis, inflammation, and proliferation are hallmarks of diseased tissues, and in vivo models of these disease phenotypes can provide insight into disease pathology. Dstncorn1 mice, deficient for the actin depolymerizing factor destrin (DSTN), display an increase of serum response factor (SRF) that results in epithelial hyperproliferation, inflammation, and neovascularization in the cornea. Previous work demonstrated that conditional ablation of Srf from the corneal epithelium of Dstncorn1 mice returns the cornea to a wild-type (WT) like state. This result implicated SRF as a major regulator of genes that contributes to abnormal phenotypes in Dstncorn1 cornea. The purpose of this study is to identify gene networks that are affected by increased expression of Srf in the Dstncorn1 cornea. Microarray analysis led to characterization of gene expression changes that occur when conditional knockout of Srf rescues mutant phenotypes in the cornea of Dstncorn1 mice. Comparison of gene expression values from WT, Dstncorn1 mutant, and Dstncorn1 rescued cornea identified >400 differentially expressed genes that are downstream from SRF. Srf ablation had a significant effect on genes associated with epithelial cell-cell junctions and regulation of actin dynamics. The majority of genes affected by SRF are downregulated in the Dstncorn1 mutant cornea, suggesting that increased SRF negatively affects transcription of SRF gene targets. ChIP-seq analysis on Dstncorn1 mutant and WT tissue revealed that, despite being present in higher abundance, SRF binding is significantly decreased in the Dstncorn1 mutant cornea. This study uses a unique model combining genetic and genomic approaches to identify genes that are regulated by SRF. These findings expand current understanding of the role of SRF in both normal and abnormal tissue homeostasis. PMID:24550211

Effect of adipose-derived mesenchymal stromal cells on tendon healing in aging and estrogen deficiency: an in vitro co-culture model.

PubMed

Veronesi, Francesca; Della Bella, Elena; Torricelli, Paola; Pagani, Stefania; Fini, Milena

2015-11-01

Aging and estrogen deficiency play a pivotal role in reducing tenocyte proliferation, collagen turnover and extracellular matrix remodeling. Mesenchymal stromal cells are being studied as an alternative for tendon regeneration, but little is known about the molecular events of adipose-derived mesenchymal stromal cells (ADSCs) on tenocytes in tendons compromised by aging and estrogen deficiency. The present in vitro study aims to compare the potential therapeutic effects of ADSCs, harvested from healthy young (sham) and aged estrogen-deficient (OVX) subjects, for tendon healing. An indirect co-culture system was set up with ADSCs, isolated from OVX or sham rats, and tenocytes from OVX rats. Cell proliferation, healing rate and gene expression were evaluated in both a standard culture condition and a microwound-healing model. It was observed that tenocyte proliferation, healing rate and collagen expression improved after the addition of sham ADSCs in both culture situations. OVX ADSCs also increased tenocyte proliferation and healing rate but less compared with sham ADSCs. Decorin and Tenascin C expression increased in the presence of OVX ADSCs. Findings suggest that ADSCs might be a promising treatment for tendon regeneration in advanced age and estrogen deficiency. However, some differences between allogenic and autologous cells were found and should be investigated in further in vivo studies. It appears that allogenic ADSCs improve tenocyte proliferation, collagen expression and the healing rate more than autologous cells. Autologous cells increase collagen expression only in the absence of an injury and increase Decorin and Tenascin C more than allogenic cells. Copyright © 2015 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

Nerve-dependent changes in skeletal muscle myosin heavy chain after experimental denervation and cross-reinnervation and in a demyelinating mouse model of Charcot-Marie-Tooth disease type 1A.

PubMed

Maggs, Alison M; Huxley, Clare; Hughes, Simon M

2008-12-01

Innervation regulates the contractile properties of vertebrate muscle fibers, in part through the effect of electrical activity on expression of distinct myosins. Herein we analyze the role of innervation in regulating the accumulation of the general, maturational, and adult forms of rodent slow myosin heavy chain (MyHC) that are defined by the presence of distinct antigenic epitopes. Denervation increases the number of fibers that express general slow MyHC, but it decreases the adult slow MyHC epitope. Cross-reinnervation of slow muscle by a fast nerve leads to an increase in the number of fibers that express fast MyHC. In both cases, there is an increase in the number of fibers that express slow and fast IIA MyHCs, but without the adult slow MyHC epitope. The data suggest that innervation is required for maturation and maintenance of diversity of both slow and fast fibers. The sequence of slow MyHC epitope transitions is a useful biomarker, and it may play a significant role during nerve-dependent changes in muscle fiber function. We applied this detailed muscle analysis to a transgenic mouse model of human motor and sensory neuropathy IA, also known as Charcot-Marie-Tooth disease type 1A (CMT1A), in which electrical conduction in some motor nerves is poor due to demyelination. The mice display atrophy of some muscle fibers and changes in slow and fast MyHC epitope expression, suggestive of a progressive increase in innervation of muscle fibers by fast motor neurons, even at early stages. The potential role of these early changes in disease pathogenesis is assessed.

Upregulation of aldolase B and overproduction of methylglyoxal in vascular tissues from rats with metabolic syndrome.

PubMed

Liu, Jianghai; Wang, Rui; Desai, Kaushik; Wu, Lingyun

2011-12-01

Methylglyoxal (MG) overproduction has been reported in metabolic syndrome with hyperglycaemia (diabetes) or without hyperglycaemia (hypertension), and the underlying mechanism was investigated. Contributions of different pathways or enzymes to MG formation were evaluated in aorta or cultured vascular smooth muscle cells (VSMCs). In all four animal models of metabolic syndrome, i.e. chronically fructose-fed hypertensive Sprague-Dawley rats, spontaneously hypertensive rats, obese non-diabetic Zucker rats, and diabetic Zucker rats, serum and aortic MG and fructose levels were increased, and the expression of GLUT5 (transporting fructose) and aldolase B (converting fructose to MG) in aorta were up-regulated. Aortic expressions of aldolase A, semicarbazide-sensitive amine oxidase (SSAO), and cytochrome P450 2E1 (CYP 2E1), accounting for MG formation during glycolysis, protein, and lipid metabolism, respectively, was unchanged/reduced. Fructose (25 mM) treatment of VSMCs up-regulated the expression of GLUT5 and aldolase B and accelerated MG formation. Insulin (100 nM) increased GLUT5 expression and augmented fructose-increased cellular fructose accumulation and MG formation. Glucose (25 mM) treatment activated the polyol pathway and enhanced fructose formation, leading to aldolase B upregulation and MG overproduction. Inhibition of the polyol pathway reduced the glucose-increased aldolase B expression and MG generation. The excess formation of MG in under these conditions was eliminated by knock-down of aldolase B, but not by knock-down of aldolase A or inhibition of SSAO or CYP 2E1. Upregulation of aldolase B by accumulated fructose is a common mechanism for MG overproduction in VSMCs and aorta in different models of metabolic syndrome.

Analysis of molecular mechanisms of 5-fluorouracil-induced steatosis and inflammation in vitro and in mice

PubMed Central

Freese, Kim; Schiergens, Tobias S.; Kuecuekoktay, Fulya Suzan; Teufel, Andreas; Thasler, Wolfgang E.; Müller, Martina; Bosserhoff, Anja K.; Hellerbrand, Claus

2017-01-01

Chemotherapy-associated steatohepatitis is attracting increasing attention because it heralds an increased risk of morbidity and mortality in patients undergoing surgery because of liver metastases. The aim of this study was to develop in vitro and in vivo models to analyze the pathogenesis of 5-fluorouracil (5-FU)-induced steatohepatitis. Therefore, primary human hepatocytes and HepG2 hepatoma cells were incubated with 5-FU at non-toxic concentrations up to 24 h. Furthermore, hepatic tissue of C57BL/6N mice was analyzed 24 h after application of a single 5-FU dose (200 mg/kg body weight). In vitro, incubation with 5-FU induced a significant increase of hepatocellular triglyceride levels. This was paralleled by an impairment of mitochondrial function and a dose- and time-dependently increased expression of fatty acid acyl-CoA oxidase 1 (ACOX1), which catalyzes the initial step for peroxisomal β-oxidation. The latter is known to generate reactive oxygen species, and consequently, expression of the antioxidant enzyme heme oxygenase 1 (HMOX1) was significantly upregulated in 5-FU-treated cells, indicative for oxidative stress. Furthermore, 5-FU significantly induced c-Jun N-terminal kinase (JNK) activation and the expression of pro-inflammatory genes IL-8 and ICAM-1. Also in vivo, 5-FU significantly induced hepatic ACOX1 and HMOX1 expression as well as JNK-activation, pro-inflammatory gene expression and immune cell infiltration. In summary, we identified molecular mechanisms by which 5-FU induces hepatocellular lipid accumulation and inflammation. Our newly developed models can be used to gain further insight into the pathogenesis of 5-FU-induced steatohepatitis and to develop therapeutic strategies to inhibit its development and progression. PMID:28055957

Defective Hematopoietic Stem Cell and Lymphoid Progenitor Development in the Ts65Dn Mouse Model of Down Syndrome: Potential Role of Oxidative Stress

PubMed Central

Lorenzo, Laureanne Pilar E.; Chen, Haiyan; Shatynski, Kristen E.; Clark, Sarah; Yuan, Rong; Harrison, David E.; Yarowsky, Paul J.

2011-01-01

Abstract Aims Down Syndrome (DS), a genetic disease caused by a triplication of chromosome 21, is characterized by increased markers of oxidative stress. In addition to cognitive defects, patients with DS also display hematologic disorders and increased incidence of infections and leukemia. Using the Ts65Dn mouse model of DS, the goal of this study was to examine hematopoietic stem and lymphoid progenitor cell function in DS. Results Analysis of hematopoietic progenitor populations showed that Ts65Dn mice possessed fewer functional hematopoietic stem cells and a significantly decreased percentage of bone marrow lymphoid progenitors. Increased reactive oxygen species and markers of oxidative stress were detected in hematopoietic stem cell populations and were associated with a loss of quiescence. Bone marrow progenitor populations expressed diminished levels of the IL-7Rα chain, which was associated with decreased proliferation and increased apoptosis. Modulating oxidative stress in vitro suggested that oxidative stress selectively leads to decreased IL-7Rα expression, and inhibits the survival of IL-7Rα-expressing hematopoietic progenitors, potentially linking increased reactive oxygen species and immunopathology. Innovation The study results identify a link between oxidative stress and diminished IL-7Rα expression and function. Further, the data suggest that this decrease in IL-7Rα is associated with defective hematopoietic development in Down Syndrome. Conclusion The data suggest that hematopoietic stem and lymphoid progenitor cell defects underlie immune dysfunction in DS and that increased oxidative stress and reduced cytokine signaling may alter hematologic development in Ts65Dn mice. Antioxid. Redox Signal. 15, 2083–2094. PMID:21504363

Intervertebral disc degeneration-induced expression of pain-related molecules: glial cell-derived neurotropic factor as a key factor.

PubMed

Jung, Woon-Won; Kim, Hyun-Sook; Shon, Jong-Ryeul; Lee, Min; Lee, Sang-Heon; Sul, Donggeun; Na, Heung Sik; Kim, Joo Han; Kim, Byung-Jo

2011-10-01

Discogenic low back pain has been shown to develop into chronic intractable pain due to an unknown pathogenesis. To study the mechanism of discogenic pain, we analyzed the serial expression of pain-related molecules in the dorsal root ganglia (DRG) and thalamus using a newly developed rat model of disc degeneration. Ten microliters of complete Freund's adjuvant was injected into the L5-6 disc of male Sprague-Dawley rats for 10 minutes using a 26-gauge needle. Using a behavioral test, rats with significant pain were selected and subsequently serial gene expression of pain-related molecules in the DRG and the thalamus was analyzed by reverse transcriptase polymerase chain reaction. The expression of tumor necrosis factor-α and interleukin-1β significantly increased at 4 and 8 weeks in the DRG of rats with pain. Furthermore, interleukin-6 was significantly increased at 4 weeks in the DRG; however, these cytokines did not show a significant change in the thalamus. Calcitonin gene-related peptide and substance P were significantly increased in DRG at 4 and 8 weeks and in the thalamus at 2 and 4 weeks. The level of nerve growth factor-β did not significantly increase in the DRG or thalamus, whereas glial cell line-derived neurotropic factor (GDNF) was significantly increased at 2 weeks and was sustained through 8 weeks in both the DRG and thalamus. The disc degeneration rat model described herein led to significant pain of a chronic nature. The gradual and persistent increase of GDNF in both the thalamus and DRG suggests that GDNF might be a key factor in the development of intractable, chronic discogenic pain.

Fatty Acid Synthase Mediates the Epithelial-Mesenchymal Transition of Breast Cancer Cells

PubMed Central

Li, Junqin; Dong, Lihua; Wei, Dapeng; Wang, Xiaodong; Zhang, Shuo; Li, Hua

2014-01-01

This study aimed to investigate the role of fatty acid synthase (FASN) in the epithelial-mesenchymal transition (EMT) of breast cancer cells. MCF-7 cells and MCF-7 cells overexpressing mitogen-activated protein kinase 5 (MCF-7-MEK5) were used in this study. MCF-7-MEK5 cells showed stable EMT characterized by increased vimentin and decreased E-cadherin expression. An In vivo animal model was established using the orthotopic injection of MCF-7 or MCF-7-MEK5 cells. Real-time quantitative PCR and western blotting were used to detect the expression levels of FASN and its downstream proteins liver fatty acid-binding protein (L-FABP) and VEGF/VEGFR-2 in both in vitro and in vivo models (nude mouse tumor tissues). In MCF-7-MEK5 cells, significantly increased expression of FASN was associated with increased levels of L-FABP and VEGF/VEGFR-2. Cerulenin inhibited MCF-7-MEK5 cell migration and EMT, and reduced FASN expression and down-stream proteins L-FABP, VEGF, and VEGFR-2. MCF-7-MEK5 cells showed higher sensitivity to Cerulenin than MCF-7 cells. Immunofluorescence revealed an increase of co-localization of FASN with VEGF on the cell membrane and with L-FABP within MCF-7-MEK5 cells. Immunohistochemistry further showed that increased percentage of FASN-positive cells in the tumor tissue was associated with increased percentages of L-FABP- and VEGF-positive cells and the Cerulenin treatment could reverse the effect. Altogether, our results suggest that FASN is essential to EMT possibly through regulating L-FABP, VEGF and VEGFR-2. This study provides a theoretical basis and potential strategy for effective suppression of malignant cells with EMT. PMID:24520215

Myrtol ameliorates cartilage lesions in an osteoarthritis rat model.

PubMed

Ying, Binbin; Maimaiti, Abudu Kelimujiang; Song, Donghui; Zhu, Songsong

2015-01-01

The aim of this study is to evaluate the effects of myrtol standardized on cartilage lesions in osteoarthritis (OA) rats. Fifty-six healthy Sprague-Dawley rats were randomly divided into sham group (13 rats) and OA model group (43 rats) with interior meniscus excision. Then serum estradiol (E2) and glycosaminoglycan (GAG) content in cartilage tissue were measured by radioimmunoassay and toluidine blue staining, respectively. After that, the model rats were randomly divided into low dose myrtol (LDM) group, middle dose myrtol (MDM) group and high dose myrtol (HDM) group (10 rats in each group) with treatment of 450 mg/kg, 300 mg/kg and 150 mg/kg myrtol, respectively. Then, Mankin scores were used to evaluate lesion extent of knee joint cartilage. Expression of tumor necrosis factor α (TNF-α), transforming growth factor β1 (TGF-β1), interleukin (IL)-6, Bax and Bcl-2 were investigated using PCR gel electrophoresis method. Mankin cores were lower in sham group and myrtol group than in model group. There were statistically significant differences (P < 0.01) between sham group and model group in expression of TNF-α, TGF-β1, IL-6, Bax and Bcl-2 in the cartilage tissue. Myrtol significantly reduced the expression of TNF-α, IL-6 and Bax, and increased the expression of TGF-β1 and Bcl-2 in myrtol group, comparing with those in model group (P < 0.01). Myrtol could down-regulate the expression of TNF-α, IL-6 and Bax, and up-regulate the expression of TGF-β1 and Bcl-2. Myrtol standardized is a promising drug to ameliorate knee cartilage lesions in the OA rat model.

The Bile Acid Receptor GPBAR-1 (TGR5) Modulates Integrity of Intestinal Barrier and Immune Response to Experimental Colitis

PubMed Central

Cipriani, Sabrina; Mencarelli, Andrea; Chini, Maria Giovanna; Distrutti, Eleonora; Renga, Barbara; Bifulco, Giuseppe; Baldelli, Franco; Donini, Annibale; Fiorucci, Stefano

2011-01-01

Background GP-BAR1, a member G protein coupled receptor superfamily, is a cell surface bile acid-activated receptor highly expressed in the ileum and colon. In monocytes, ligation of GP-BAR1 by secondary bile acids results in a cAMP-dependent attenuation of cytokine generation. Aims To investigate the role GP-BAR1 in regulating intestinal homeostasis and inflammation-driven immune dysfunction in rodent models of colitis. Methods Colitis was induced in wild type and GP-BAR1−/− mice by DSS and TNBS administration. Potential GP-BAR1 agonists were identified by in silico screening and computational docking studies. Results GP-BAR1−/− mice develop an abnormal morphology of colonic mucous cells and an altered molecular architecture of epithelial tight junctions with increased expression and abnormal subcellular distribution of zonulin 1 resulting in increased intestinal permeability and susceptibility to develop severe colitis in response to DSS at early stage of life. By in silico screening and docking studies we identified ciprofloxacin as a GP-BAR1 ligand. In monocytes, ciprofloxacin increases cAMP concentrations and attenuates TNFα release induced by TLR4 ligation in a GP-BAR1 dependent manner. Treating mice rendered colitic by TNBS with ciprofloxacin and oleanolic acid, a well characterized GP-BAR1 ligand, abrogates signs and symptoms of colitis. Colonic expression of GP-BAR1 mRNA increases in rodent models of colitis and tissues from Crohn's disease patients. Flow cytometry analysis demonstrates that ≈90% of CD14+ cells isolated from the lamina propria of TNBS-treated mice stained positively for GP-BAR1. Conclusions GP-BAR1 regulates intestinal barrier structure. Its expression increases in rodent models of colitis and Crohn's disease. Ciprofloxacin is a GP-BAR1 ligand. PMID:22046243

S100A8/A9 mRNA induction in an ex vivo model of endotoxin tolerance: roles of IL-10 and IFNγ.

PubMed

Fontaine, Mathieu; Planel, Séverine; Peronnet, Estelle; Turrel-Davin, Fanny; Piriou, Vincent; Pachot, Alexandre; Monneret, Guillaume; Lepape, Alain; Venet, Fabienne

2014-01-01

Septic syndromes are the leading cause of death in intensive care units. They are characterized by the development of immune dysfunctions such as endotoxin tolerance (ET), whose intensity and duration are associated with increased risk of nosocomial infections and mortality. Alarmins S100A8 and S100A9 have been shown to be increased after septic shock. Importantly, a delayed S100A9 mRNA increase predicts hospital-acquired infection in patients. The aim of this study was to investigate the regulation of S100A8 and S100A9 mRNA expression in an ex vivo model of ET. ET was reproduced ex vivo by priming healthy peripheral blood mononuclear cells (number of donors  = 9 to 10) with low-dose endotoxin (2 ng/ml) before stimulation with high dose endotoxin (100 ng/ml). S100A8 and S100A9 mRNA levels were measured by quantitative real-time polymerase chain reactions. ET was established by observing decreased TNFα and increased IL-10 transcriptomic responses to two subsequent endotoxin challenges. Interestingly, ET was associated with increased S100A8 and S100A9 mRNA expression ex vivo. We showed that IL-10 played a role in this process, since S100A8 and S100A9 mRNA increases were significantly abrogated by IL-10 blockade in the model. Conversely, treatment with rIFN-γ, a pro-inflammatory and immunostimulating molecule known to block ET induction, was able to restore normal S100A8 and S100A9 mRNA in this model. In this ex vivo model, we observed that S100A8 and S100A9 mRNA expression was significantly increased during ET. This reproduced ex vivo the observations we had previously made in septic shock patients. Interestingly, IL-10 blockade and rIFN-γ treatment partially abrogated S100A8/A9 mRNA increases in this model. Pending confirmation in larger, independent clinical studies, these preliminary results suggest that S100A8 and S100A9 mRNA levels might be used as surrogate markers of ET and as stratification tools for personalized immunotherapy in septic shock patients.

Clostridium butyricum exerts a neuroprotective effect in a mouse model of traumatic brain injury via the gut-brain axis.

PubMed

Li, H; Sun, J; Du, J; Wang, F; Fang, R; Yu, C; Xiong, J; Chen, W; Lu, Z; Liu, J

2018-05-01

Traumatic brain injury (TBI) is a common occurrence following gastrointestinal dysfunction. Recently, more and more attentions are being focused on gut microbiota in brain and behavior. Glucagon-like peptide-1 (GLP-1) is considered as a mediator that links the gut-brain axis. The aim of this study was to explore the neuroprotective effects of Clostridium butyricum (Cb) on brain damage in a mouse model of TBI. Male C57BL/6 mice were subjected to a model of TBI-induced by weight-drop impact head injury and were treated intragastrically with Cb. The cognitive deficits, brain water content, neuronal death, and blood-brain barrier (BBB) permeability were evaluated. The expression of tight junction (TJ) proteins, Bcl-2, Bax, GLP-1 receptor (GLP-1R), and phosphorylation of Akt (p-Akt) in the brain were also measured. Moreover, the intestinal barrier permeability, the expression of TJ protein and GLP-1, and IL-6 level in the intestine were detected. Cb treatment significantly improved neurological dysfunction, brain edema, neurodegeneration, and BBB impairment. Meanwhile, Cb treatment also significantly increased the expression of TJ proteins (occludin and zonula occluden-1), p-Akt and Bcl-2, but decreased expression of Bax. Moreover, Cb treatment exhibited more prominent effects on decreasing the levels of plasma d-lactate and colonic IL-6, upregulating expression of Occludin, and protecting intestinal barrier integrity. Furthermore, Cb-treated mice showed increased the secretion of intestinal GLP-1 and upregulated expression of cerebral GLP-1R. Our findings demonstrated the neuroprotective effect of Cb in TBI mice and the involved mechanisms were partially attributed to the elevating GLP-1 secretion through the gut-brain axis. © 2017 John Wiley & Sons Ltd.

Impaired anti-fibrotic effect of bone marrow-derived mesenchymal stem cell in a mouse model of pulmonary paracoccidioidomycosis

PubMed Central

Arango, Julián Camilo; Puerta-Arias, Juan David; Pino-Tamayo, Paula Andrea; Salazar-Peláez, Lina María; Rojas, Mauricio

2017-01-01

Bone marrow-derived mesenchymal stem cells (BMMSCs) have been consider as a promising therapy in fibrotic diseases. Experimental models suggest that BMMSCs may be used as an alternative therapy to treat chemical- or physical-induced pulmonary fibrosis. We investigated the anti-fibrotic potential of BMMSCs in an experimental model of lung fibrosis by infection with Paracoccidioides brasiliensis. BMMSCs were isolated and purified from BALB/c mice using standardized methods. BALB/c male mice were inoculated by intranasal infection of 1.5x106 P. brasiliensis yeasts. Then, 1x106 BMMSCs were administered intra venous at 8th week post-infection (p.i.). An additional group of mice was treated with itraconazole (ITC) two weeks before BMMSCs administration. Animals were sacrificed at 12th week p.i. Histopathological examination, fibrocytes counts, soluble collagen and fibrosis-related genes expression in lungs were evaluated. Additionally, human fibroblasts were treated with homogenized lung supernatants (HLS) to determine induction of collagen expression. Histological analysis showed an increase of granulomatous inflammatory areas in BMMSCs-treated mice. A significant increase of fibrocytes count, soluble collagen and collagen-3α1, TGF-β3, MMP-8 and MMP-15 genes expression were also observed in those mice. Interestingly, when combined therapy BMMSCs/ITC was used there is a decrease of TIMP-1 and MMP-13 gene expression in infected mice. Finally, human fibroblasts stimulated with HLS from infected and BMMSCs-transplanted mice showed a higher expression of collagen I. In conclusion, our findings indicate that late infusion of BMMSCs into mice infected with P. brasiliensis does not have any anti-fibrotic effect; possibly because their interaction with the fungus promotes collagen expression and tissue remodeling. PMID:29040281

Effect of Traumatic Brain Injury, Erythropoietin, and Anakinra on Hepatic Metabolizing Enzymes and Transporters in an Experimental Rat Model.

PubMed

Anderson, Gail D; Peterson, Todd C; Vonder Haar, Cole; Farin, Fred M; Bammler, Theo K; MacDonald, James W; Kantor, Eric D; Hoane, Michael R

2015-09-01

In contrast to considerable data demonstrating a decrease in cytochrome P450 (CYP) activity in inflammation and infection, clinically, traumatic brain injury (TBI) results in an increase in CYP and UDP glucuronosyltransferase (UGT) activity. The objective of this study was to determine the effects of TBI alone and with treatment with erythropoietin (EPO) or anakinra on the gene expression of hepatic inflammatory proteins, drug-metabolizing enzymes, and transporters in a cortical contusion impact (CCI) injury model. Microarray-based transcriptional profiling was used to determine the effect on gene expression at 24 h, 72 h, and 7 days post-CCI. Plasma cytokine and liver protein concentrations of CYP2D4, CYP3A1, EPHX1, and UGT2B7 were determined. There was no effect of TBI, TBI + EPO, or TBI + anakinra on gene expression of the inflammatory factors shown to be associated with decreased expression of hepatic metabolic enzymes in models of infection and inflammation. IL-6 plasma concentrations were increased in TBI animals and decreased with EPO and anakinra treatment. There was no significant effect of TBI and/or anakinra on gene expression of enzymes or transporters known to be involved in drug disposition. TBI + EPO treatment decreased the gene expression of Cyp2d4 at 72 h with a corresponding decrease in CYP2D4 protein at 72 h and 7 days. CYP3A1 protein was decreased at 24 h. In conclusion, EPO treatment may result in a significant decrease in the metabolism of Cyp-metabolized drugs. In contrast to clinical TBI, there was not a significant effect of experimental TBI on CYP or UGT metabolic enzymes.

Deoxycholic acid promotes development of gastroesophageal reflux disease and Barrett's oesophagus by modulating integrin-αv trafficking.

PubMed

Prichard, David O; Byrne, Anne Marie; Murphy, James O; Reynolds, John V; O'Sullivan, Jacintha; Feighery, Ronan; Doyle, Brendan; Eldin, Osama Sharaf; Finn, Stephen P; Maguire, Aoife; Duff, Deirdre; Kelleher, Dermot P; Long, Aideen

2017-12-01

The fundamental mechanisms underlying erosive oesophagitis and subsequent development of Barrett's oesophagus (BO) are poorly understood. Here, we investigated the contribution of specific components of the gastric refluxate on adhesion molecules involved in epithelial barrier maintenance. Cell line models of squamous epithelium (HET-1A) and BO (QH) were used to examine the effects of bile acids on cell adhesion to extracellular matrix proteins (Collagen, laminin, vitronectin, fibronectin) and expression of integrin ligands (α 3 , α 4, α 5 , α 6 and α ν ). Experimental findings were validated in human explant oesophageal biopsies, a rat model of gastroesophageal reflux disease (GORD) and in patient tissue microarrays. The bile acid deoxycholic acid (DCA) specifically reduced adhesion of HET-1A cells to vitronectin and reduced cell-surface expression of integrin-α ν via effects on endocytic recycling processes. Increased expression of integrin-α v was observed in ulcerated tissue in a rat model of GORD and in oesophagitis and Barrett's intestinal metaplasia patient tissue compared to normal squamous epithelium. Increased expression of integrin-α ν was observed in QH BO cells compared to HET-1A cells. QH cells were resistant to DCA-mediated loss of adhesion and reduction in cell-surface expression of integrin-α ν . We demonstrated that a specific component of the gastric refluxate, DCA, affects the epithelial barrier through modulation of integrin α ν expression, providing a novel mechanism for bile acid-mediated erosion of oesophageal squamous epithelium and promotion of BO. Strategies aimed at preventing bile acid-mediated erosion should be considered in the clinical management of patients with GORD. © 2017 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.

Complexity reduction of biochemical rate expressions.

PubMed

Schmidt, Henning; Madsen, Mads F; Danø, Sune; Cedersund, Gunnar

2008-03-15

The current trend in dynamical modelling of biochemical systems is to construct more and more mechanistically detailed and thus complex models. The complexity is reflected in the number of dynamic state variables and parameters, as well as in the complexity of the kinetic rate expressions. However, a greater level of complexity, or level of detail, does not necessarily imply better models, or a better understanding of the underlying processes. Data often does not contain enough information to discriminate between different model hypotheses, and such overparameterization makes it hard to establish the validity of the various parts of the model. Consequently, there is an increasing demand for model reduction methods. We present a new reduction method that reduces complex rational rate expressions, such as those often used to describe enzymatic reactions. The method is a novel term-based identifiability analysis, which is easy to use and allows for user-specified reductions of individual rate expressions in complete models. The method is one of the first methods to meet the classical engineering objective of improved parameter identifiability without losing the systems biology demand of preserved biochemical interpretation. The method has been implemented in the Systems Biology Toolbox 2 for MATLAB, which is freely available from http://www.sbtoolbox2.org. The Supplementary Material contains scripts that show how to use it by applying the method to the example models, discussed in this article.

Generation of SNCA Cell Models Using Zinc Finger Nuclease (ZFN) Technology for Efficient High-Throughput Drug Screening.

PubMed

Dansithong, Warunee; Paul, Sharan; Scoles, Daniel R; Pulst, Stefan M; Huynh, Duong P

2015-01-01

Parkinson's disease (PD) is a progressive neurodegenerative disorder caused by loss of dopaminergic neurons of the substantia nigra. The hallmark of PD is the appearance of neuronal protein aggregations known as Lewy bodies and Lewy neurites, of which α-synuclein forms a major component. Familial PD is rare and is associated with missense mutations of the SNCA gene or increases in gene copy number resulting in SNCA overexpression. This suggests that lowering SNCA expression could be therapeutic for PD. Supporting this hypothesis, SNCA reduction was neuroprotective in cell line and rodent PD models. We developed novel cell lines expressing SNCA fused to the reporter genes luciferase (luc) or GFP with the objective to enable high-throughput compound screening (HTS) for small molecules that can lower SNCA expression. Because SNCA expression is likely regulated by far-upstream elements (including the NACP-REP1 located at 8852 bp upstream of the transcription site), we employed zinc finger nuclease (ZFN) genome editing to insert reporter genes in-frame downstream of the SNCA gene in order to retain native SNCA expression control. This ensured full retention of known and unknown up- and downstream genetic elements controlling SNCA expression. Treatment of cells with the histone deacetylase inhibitor valproic acid (VPA) resulted in significantly increased SNCA-luc and SNCA-GFP expression supporting the use of our cell lines for identifying small molecules altering complex modes of expression control. Cells expressing SNCA-luc treated with a luciferase inhibitor or SNCA siRNA resulted in Z'-scores ≥ 0.75, suggesting the suitability of these cell lines for use in HTS. This study presents a novel use of genome editing for the creation of cell lines expressing α-synuclein fusion constructs entirely under native expression control. These cell lines are well suited for HTS for compounds that lower SNCA expression directly or by acting at long-range sites to the SNCA promoter and 5'-UTR.

[Protective mechanism on the vascular pathological process in diabetes mellitus rats by Rheum officeinale].

PubMed

Tian, Feng-Sheng; Li, Zhen-Bin; Wang, Yuan-Song; Su, Xiu-Hai; Li, Wen-Dong; Wang, Xiao-Yun

2008-03-01

To explore the protective mechanism of officeihale on the vascular pathological process in diabetes mellitus (DM) rats. After the DM rat model was established, 24 DM rats were randomly divided into model group (12 DM rats) and Rheum officeinale group (12 DM rats). Rheum officeinale was orally given in 10 g kg(-1) per day, and the other two groups were given equal pure water. 8 weeks later, blood samples were collected to determine the level of nitric oxide (NO) and endothelin-1 (ET-1). Thoracic aortic rings was prepared to observe the inhibiting effect of Ach with different concentration on contraction caused by NE. Another part of aorta was made to observe the expression of ICAM-1 and VCAM-1 by method of SP immunohistochemistry staining, Rheum officeinale group obviously decreased the level of ET-1 and increased the NO compared with model group (P <0.05). The expression of ICAM-1 and VCAM-1 could be obviously inhibited in Rheum officeinale group compared with model group. (P <0.05). Rheum officeinale could decrease the level of ET-1 with increased the NO in diabetes rats, and inhibit the expression of ICAM-1 and VCAM-1, which may be mechanisms of protecting the endothelium of vessel in diabetes rats.

Development of Transgenic Cloned Pig Models of Skin Inflammation by DNA Transposon-Directed Ectopic Expression of Human β1 and α2 Integrin

PubMed Central

Staunstrup, Nicklas Heine; Madsen, Johannes; Primo, Maria Nascimento; Li, Juan; Liu, Ying; Kragh, Peter M.; Li, Rong; Schmidt, Mette; Purup, Stig; Dagnæs-Hansen, Frederik; Svensson, Lars; Petersen, Thomas K.; Callesen, Henrik; Bolund, Lars; Mikkelsen, Jacob Giehm

2012-01-01

Integrins constitute a superfamily of transmembrane signaling receptors that play pivotal roles in cutaneous homeostasis by modulating cell growth and differentiation as well as inflammatory responses in the skin. Subrabasal expression of integrins α2 and/or β1 entails hyperproliferation and aberrant differentiation of keratinocytes and leads to dermal and epidermal influx of activated T-cells. The anatomical and physiological similarities between porcine and human skin make the pig a suitable model for human skin diseases. In efforts to generate a porcine model of cutaneous inflammation, we employed the Sleeping Beauty DNA transposon system for production of transgenic cloned Göttingen minipigs expressing human β1 or α2 integrin under the control of a promoter specific for subrabasal keratinocytes. Using pools of transgenic donor fibroblasts, cloning by somatic cell nuclear transfer was utilized to produce reconstructed embryos that were subsequently transferred to surrogate sows. The resulting pigs were all transgenic and harbored from one to six transgene integrants. Molecular analyses on skin biopsies and cultured keratinocytes showed ectopic expression of the human integrins and localization within the keratinocyte plasma membrane. Markers of perturbed skin homeostasis, including activation of the MAPK pathway, increased expression of the pro-inflammatory cytokine IL-1α, and enhanced expression of the transcription factor c-Fos, were identified in keratinocytes from β1 and α2 integrin-transgenic minipigs, suggesting the induction of a chronic inflammatory phenotype in the skin. Notably, cellular dysregulation obtained by overexpression of either β1 or α2 integrin occurred through different cellular signaling pathways. Our findings mark the creation of the first cloned pig models with molecular markers of skin inflammation. Despite the absence of an overt psoriatic phenotype, these animals may possess increased susceptibility to severe skin damage-induced inflammation and should be of great potential in studies aiming at the development and refinement of topical therapies for cutaneous inflammation including psoriasis. PMID:22590584

Different profiles of neuroendocrine cell differentiation evolve in the PC-310 human prostate cancer model during long-term androgen deprivation.

PubMed

Jongsma, Johan; Oomen, Monique H; Noordzij, Marinus A; Van Weerden, Wytske M; Martens, Gerard J M; van der Kwast, Theodorus H; Schröder, Fritz H; van Steenbrugge, Gert J

2002-03-01

Neuroendocrine (NE) cells are androgen-independent cells and secrete growth-modulating peptide hormones via a regulated secretory pathway (RSP). We studied NE differentiation after long-term androgen withdrawal in the androgen-dependent human prostate cancer xenograft PC-310. Tumor-bearing nude mice were killed at 0, 2, 5, 7, 14, 21, 47, 84, and 154 days after castration. The half-life of the PC-310 tumor was 10 days, with a stable residual tumor volume of 30--40% after 21 days and longer periods of androgen deprivation. Proliferative activity and prostate-specific antigen serum levels decreased to zero after castration, whereas cell-cycle arrest was manifested by increased p27(kip1) expression. A temporary downregulation of androgen receptor (AR) expression was noted after androgen deprivation. The expression of chromogranin A, secretogranin III, and secretogranin V (7B2) increased 5 days after castration and later. Subsequently, pro-hormone convertase 1 and peptidyl alpha--amidating monooxygenase as well as vascular endothelial growth factor were expressed from 7 days after castration on. Finally, such growth factors as gastrin-releasing peptide and serotonin were expressed in a small part of the NE cells 21 days after castration, but strong expression was induced late during androgen deprivation, that is, 84 and 154 days after castration, respectively. Androgen deprivation of the NE-differentiated PC-310 model induced the formation of NE-differentiated AR(minus sign) and non-NE AR(+) tumor residues. The NE-differentiated cells actively produced growth factors via an RSP that may lead to hormone-refractory disease. The dormant non-NE AR(+) tumor cells were shown to remain androgen sensitive even after long-term androgen deprivation. In the PC-310 xenograft, time-dependent NE differentiation and subsequent maturation were induced after androgen depletion. The androgen-dependent PC-310 xenograft model constitutes an excellent model for studying the role of NE cells in the progression of clinical prostate cancer. Copyright 2002 Wiley-Liss, Inc.

NR2B Expression in Rat DRG Is Differentially Regulated Following Peripheral Nerve Injuries That Lead to Transient or Sustained Stimuli-Evoked Hypersensitivity

PubMed Central

Norcini, Monica; Sideris, Alexandra; Adler, Samantha M.; Hernandez, Lourdes A. M.; Zhang, Jin; Blanck, Thomas J. J.; Recio-Pinto, Esperanza

2016-01-01

Following injury, primary sensory neurons undergo changes that drive central sensitization and contribute to the maintenance of persistent hypersensitivity. NR2B expression in the dorsal root ganglia (DRG) has not been previously examined in neuropathic pain models. Here, we investigated if changes in NR2B expression within the DRG are associated with hypersensitivities that result from peripheral nerve injuries. This was done by comparing the NR2B expression in the DRG derived from two modalities of the spared nerve injury (SNI) model, since each variant produces different neuropathic pain phenotypes. Using the electronic von Frey to stimulate the spared and non-spared regions of the hindpaws, we demonstrated that sural-SNI animals develop sustained neuropathic pain in both regions while the tibial-SNI animals recover. NR2B expression was measured at Day 23 and Day 86 post-injury. At Day 23 and 86 post-injury, sural-SNI animals display strong hypersensitivity, whereas tibial-SNI animals display 50 and 100% recovery from post-injury-induced hypersensitivity, respectively. In tibial-SNI at Day 86, but not at Day 23 the perinuclear region of the neuronal somata displayed an increase in NR2B protein. This retention of NR2B protein within the perinuclear region, which will render them non-functional, correlates with the recovery observed in tibial-SNI. In sural-SNI at Day 86, DRG displayed an increase in NR2B mRNA which correlates with the development of sustained hypersensitivity in this model. The increase in NR2B mRNA was not associated with an increase in NR2B protein within the neuronal somata. The latter may result from a decrease in kinesin Kif17, since Kif17 mediates NR2B transport to the soma’s plasma membrane. In both SNIs, microglia/macrophages showed a transient increase in NR2B protein detected at Day 23 but not at Day 86, which correlates with the initial post-injury induced hypersensitivity in both SNIs. In tibial-SNI at Day 86, but not at Day 23, satellite glia cells (SGCs) displayed an increase in NR2B protein. This study is the first to characterize of cell-specific changes in NR2B expression within the DRG following peripheral nerve injury. We discuss how the observed NR2B changes in DRG can contribute to the different neuropathic pain phenotypes displayed by each SNI variant. PMID:27803647

NR2B Expression in Rat DRG Is Differentially Regulated Following Peripheral Nerve Injuries That Lead to Transient or Sustained Stimuli-Evoked Hypersensitivity.

PubMed

Norcini, Monica; Sideris, Alexandra; Adler, Samantha M; Hernandez, Lourdes A M; Zhang, Jin; Blanck, Thomas J J; Recio-Pinto, Esperanza

2016-01-01

Following injury, primary sensory neurons undergo changes that drive central sensitization and contribute to the maintenance of persistent hypersensitivity. NR2B expression in the dorsal root ganglia (DRG) has not been previously examined in neuropathic pain models. Here, we investigated if changes in NR2B expression within the DRG are associated with hypersensitivities that result from peripheral nerve injuries. This was done by comparing the NR2B expression in the DRG derived from two modalities of the spared nerve injury (SNI) model, since each variant produces different neuropathic pain phenotypes. Using the electronic von Frey to stimulate the spared and non-spared regions of the hindpaws, we demonstrated that sural-SNI animals develop sustained neuropathic pain in both regions while the tibial-SNI animals recover. NR2B expression was measured at Day 23 and Day 86 post-injury. At Day 23 and 86 post-injury, sural-SNI animals display strong hypersensitivity, whereas tibial-SNI animals display 50 and 100% recovery from post-injury-induced hypersensitivity, respectively. In tibial-SNI at Day 86, but not at Day 23 the perinuclear region of the neuronal somata displayed an increase in NR2B protein. This retention of NR2B protein within the perinuclear region, which will render them non-functional, correlates with the recovery observed in tibial-SNI. In sural-SNI at Day 86, DRG displayed an increase in NR2B mRNA which correlates with the development of sustained hypersensitivity in this model. The increase in NR2B mRNA was not associated with an increase in NR2B protein within the neuronal somata. The latter may result from a decrease in kinesin Kif17, since Kif17 mediates NR2B transport to the soma's plasma membrane. In both SNIs, microglia/macrophages showed a transient increase in NR2B protein detected at Day 23 but not at Day 86, which correlates with the initial post-injury induced hypersensitivity in both SNIs. In tibial-SNI at Day 86, but not at Day 23, satellite glia cells (SGCs) displayed an increase in NR2B protein. This study is the first to characterize of cell-specific changes in NR2B expression within the DRG following peripheral nerve injury. We discuss how the observed NR2B changes in DRG can contribute to the different neuropathic pain phenotypes displayed by each SNI variant.

SCD1 activity in muscle increases triglyceride PUFA content, exercise capacity, and PPARδ expression in mice[S

PubMed Central

Rogowski, Michael P.; Flowers, Matthew T.; Stamatikos, Alexis D.; Ntambi, James M.; Paton, Chad M.

2013-01-01

Stearoyl-CoA desaturase (SCD)1 converts saturated fatty acids into monounsaturated fatty acids. Using muscle overexpression, we sought to determine the role of SCD1 expression in glucose and lipid metabolism and its effects on exercise capacity in mice. Wild-type C57Bl/6 (WT) and SCD1 muscle transgenic (SCD1-Tg) mice were generated, and expression of the SCD1 transgene was restricted to skeletal muscle. SCD1 overexpression was associated with increased triglyceride (TG) content. The fatty acid composition of the muscle revealed a significant increase in polyunsaturated fatty acid (PUFA) content of TG, including linoleate (18:2n6). Untrained SCD1-Tg mice also displayed significantly increased treadmill exercise capacity (WT = 6.6 ± 3 min, Tg = 71.9 ± 9.5 min; P = 0.0009). SCD1-Tg mice had decreased fasting plasma glucose, glucose transporter (GLUT)1 mRNA, fatty acid oxidation, mitochondrial content, and increased peroxisome proliferator-activated receptor (PPAR)δ and Pgc-1 protein expression in skeletal muscle. In vitro studies in C2C12 myocytes revealed that linoleate (18:2n6) and not oleate (18:1n9) caused a 3-fold increase in PPARδ and a 9-fold increase in CPT-1b with a subsequent increase in fat oxidation. The present model suggests that increasing delta-9 desaturase activity of muscle increases metabolic function, exercise capacity, and lipid oxidation likely through increased PUFA content, which increases PPARδ expression and activity. However, the mechanism of action that results in increased PUFA content of SCD1-Tg mice remains to be elucidated. PMID:23918045

Cigarette smoking causes epigenetic changes associated with cardiorenal fibrosis

PubMed Central

Haller, Steven T.; Fan, Xiaoming; Xie, Jeffrey X.; Kennedy, David J.; Liu, Jiang; Yan, Yanling; Hernandez, Dawn-Alita; Mathew, Denzil P.; Cooper, Christopher J.; Shapiro, Joseph I.; Tian, Jiang

2016-01-01

Clinical studies indicate that smoking combustible cigarettes promotes progression of renal and cardiac injury, leading to functional decline in the setting of chronic kidney disease (CKD). However, basic studies using in vivo small animal models that mimic clinical pathology of CKD are lacking. To address this issue, we evaluated renal and cardiac injury progression and functional changes induced by 4 wk of daily combustible cigarette smoke exposure in the 5/6th partial nephrectomy (PNx) CKD model. Molecular evaluations revealed that cigarette smoke significantly (P < 0.05) decreased renal and cardiac expression of the antifibrotic microRNA miR-29b-3 and increased expression of molecular fibrosis markers. In terms of cardiac and renal organ structure and function, exposure to cigarette smoke led to significantly increased systolic blood pressure, cardiac hypertrophy, cardiac and renal fibrosis, and decreased renal function. These data indicate that decreased expression of miR-29b-3p is a novel mechanism wherein cigarette smoke promotes accelerated cardiac and renal tissue injury in CKD. (155 words) PMID:27789733

Expression and function of survivin in canine osteosarcoma.

PubMed

Shoeneman, Jenette K; Ehrhart, E J; Eickhoff, Jens C; Charles, J B; Powers, Barbara E; Thamm, Douglas H

2012-01-01

Osteosarcoma has a high mortality rate and remains in need of more effective therapeutic approaches. Survivin is an inhibitor of apoptosis family member protein that blocks apoptosis and drives proliferation in human cancer cells where it is commonly elevated. In this study, we illustrate the superiority of a canine osteosarcoma model as a translational tool for evaluating survivin-directed therapies, owing to the striking similarities in gross and microscopic appearance, biologic behavior, gene expression, and signaling pathway alterations. Elevated survivin expression in primary canine osteosarcoma tissue correlated with increased histologic grade and mitotic index and a decreased disease-free interval (DFI). Survivin attenuation in canine osteosarcoma cells inhibited cell-cycle progression, increased apoptosis, mitotic arrest, and chemosensitivity, and cooperated with chemotherapy to significantly improve in vivo tumor control. Our findings illustrate the utility of a canine system to more accurately model human osteosarcoma and strongly suggest that survivin-directed therapies might be highly effective in its treatment. ©2011 AACR.

Antidepressant effects of abscisic acid mediated by the downregulation of corticotrophin-releasing hormone gene expression in rats.

PubMed

Qi, Cong-Cong; Zhang, Zhi; Fang, Hui; Liu, Ji; Zhou, Nan; Ge, Jin-Fang; Chen, Fang-Han; Xiang, Cheng-Bin; Zhou, Jiang-Ning

2014-10-31

Corticotrophin-releasing hormone (CRH) is considered to be the central driving force of the hypothalamic-pituitary-adrenal axis, which plays a key role in the stress response and depression. Clinical reports have suggested that excess retinoic acid (RA) is associated with depression. Abscisic acid (ABA) and RA are direct derivatives of carotenoids and share a similar molecular structure. Here, we proposed that ABA also plays a role in the regulation of CRH activity sharing with the RA signaling pathway. [3H]-ABA radioimmunoassay demonstrated that the hypothalamus of rats shows the highest concentration of ABA compared with the cortex and the hippocampus under basal conditions. Under acute stress, ABA concentrations increased in the serum, but decreased in the hypothalamus and were accompanied by increased corticosterone in the serum and c-fos expression in the hypothalamus. Moreover, chronic ABA administration increased sucrose intake and decreased the mRNA expression of CRH and retinoic acid receptor alpha (RARα) in the hypothalamus of rats. Furthermore, ABA improved the symptom of chronic unpredictable mild stress in model rats, as indicated by increased sucrose intake, increased swimming in the forced swim test, and reduced mRNA expression of CRH and RARα in the rat hypothalamus. In vitro, CRH expression decreased after ABA treatment across different neural cells. In BE(2)-C cells, ABA inhibited a series of retinoid receptor expression, including RARα, a receptor that could facilitate CRH expression directly. These results suggest that ABA may play a role in the pathogenesis of depression by downregulating CRH mRNA expression shared with the RA signaling pathway. © The Author 2014. Published by Oxford University Press on behalf of CINP.

A randomized clinical trial of the effects of supplemental calcium and vitamin D3 on markers of their metabolism in normal mucosa of colorectal adenoma patients.

PubMed

Ahearn, Thomas U; McCullough, Marjorie L; Flanders, W Dana; Long, Qi; Sidelnikov, Eduard; Fedirko, Veronika; Daniel, Carrie R; Rutherford, Robin E; Shaukat, Aasma; Bostick, Roberd M

2011-01-15

In cancer cell lines and rodent models, calcium and vitamin D favorably modulate cell proliferation, differentiation, and apoptosis in colonic epithelia. These effects may be modulated by local expression of the calcium receptor (CaR), the vitamin D receptor (VDR), and the P450 cytochromes, CYP27B1 and CYP24A1; however, they have yet to be investigated in humans. To address this gap, we conducted a randomized, double-blinded, placebo-controlled 2×2 factorial clinical trial. Patients with at least one pathology-confirmed colorectal adenoma were treated with 2 g/d elemental calcium and/or 800 IU/d vitamin D3 versus placebo over 6 months (n=92; 23 per group). CaR, VDR, CYP27B1, and CYP24A1 expression and distribution in biopsies of normal appearing rectal mucosa were detected by standardized, automated immunohistochemistry and quantified by image analysis. In the calcium-supplemented group, CaR expression increased 27% (P=0.03) and CYP24A1 expression decreased 21% (P=0.79). In the vitamin D3-supplemented group, CaR expression increased 39% (P=0.01) and CYP27B1 expression increased 159% (P=0.06). In patients supplemented with both calcium and vitamin D3, VDR expression increased 19% (P=0.13) and CaR expression increased 24% (P=0.05). These results provide mechanistic support for further investigation of calcium and vitamin D3 as chemopreventive agents against colorectal neoplasms, and CaR, VDR, CYP27B1, and CYP24A1 as modifiable, preneoplastic risk biomarkers for colorectal neoplasms. © 2010 AACR.

Expression of Wnt and Notch signaling pathways in inflammatory bowel disease treated with mesenchymal stem cell transplantation: evaluation in a rat model.

PubMed

Xing, Yanfen; Chen, Xiaojie; Cao, Yanwen; Huang, Jianyun; Xie, Xuhong; Wei, Yaming

2015-05-22

The purpose of this study was to investigate the expression of Wnt and Notch signaling pathway-related genes in inflammatory bowel disease (IBD) treated with mesenchymal stem cell transplantation (MSCT). TNBS (2,4,6-trinitrobenzene sulfonic acid) was used to establish IBD in a rat model. Mesenchymal stem cells (MSCs) were transplanted via tail vein transfusion. Saline water was used in a control group. The expression of Wnt and Notch main signaling molecules was screened by gene chips and verified by quantitative reverse transcription-polymerase chain reaction in the IBD rat model on day 14 and day 28 after transplantation. The IBD rat models were successfully established and MSCs were transplanted into those models. Genome-wide expression profile chips identified a total of 388 differentially expressive genes, of which 191 were upregulated and 197 were downregulated in the MSC-transplanted group in comparison with the IBD control group. Real-time quantitative polymerase chain reaction results showed that the level of Olfm4 mRNA expression in the IBD group (2.54±0.20) was significantly increased compared with the MSCT group (1.39±0.54) and the normal group (1.62±0.25) (P <0.05). The Wnt3a mRNA was more highly expressed in IBD rats (2.92±0.94) and decreased in MSCT rats (0.17±0.63, P <0.05). The expression of GSK-3β mRNA was decreased in the setting of inflammation (0.65±0.04 versus 1.00±0.01 in normal group, P <0.05) but returned to normal levels after MSCT (0.81±0.17). The expression of β-catenin was observed to increase in IBD tissues (1.76±0.44) compared with normal tissues (1.00±0.01, P <0.05), but no difference was found in the MSCT group (1.12±0.36). Wnt11 declined at 14 days and returned to normal levels at 28 days in the IBD group; in comparison, a significantly lower expression was found in MSCT rats. There were no differences in the expression of Fzd3, c-myc, TCF4, and Wnt5a in inflammation, but all of those genes declined after MSCT treatment. The canonical Wnt and Notch signaling pathways are activated in IBD and may be suppressed by stem cell transplantation to differentiate into intestinal epithelium after MSCT. Moreover, the non-canonical Wnt signaling may be inhibited by canonical Wnt signaling in the setting of inflammation and may also be suppressed by MSCT.

Transcriptional bursting explains the noise–versus–mean relationship in mRNA and protein levels

DOE PAGES

Dar, Roy; Shaffer, Sydney M.; Singh, Abhyudai; ...

2016-07-28

Recent analysis demonstrates that the HIV-1 Long Terminal Repeat (HIV LTR) promoter exhibits a range of possible transcriptional burst sizes and frequencies for any mean-expression level. However, these results have also been interpreted as demonstrating that cell-tocell expression variability (noise) and mean are uncorrelated, a significant deviation from previous results. Here, we re-examine the available mRNA and protein abundance data for the HIV LTR and find that noise in mRNA and protein expression scales inversely with the mean along analytically predicted transcriptional burst-size manifolds. We then experimentally perturb transcriptional activity to test a prediction of the multiple burst-size model: thatmore » increasing burst frequency will cause mRNA noise to decrease along given burst-size lines as mRNA levels increase. In conclusion, the data show that mRNA and protein noise decrease as mean expression increases, supporting the canonical inverse correlation between noise and mean.« less

Nandrolone reduces activation of Notch signaling in denervated muscle associated with increased Numb expression.

PubMed

Liu, Xin-Hua; Yao, Shen; Qiao, Rui-Fang; Levine, Alice C; Kirschenbaum, Alexander; Pan, Jiangping; Wu, Yong; Qin, Weiping; Bauman, William A; Cardozo, Christopher P

2011-10-14

Nandrolone, an anabolic steroid, slows denervation-atrophy in rat muscle. The molecular mechanisms responsible for this effect are not well understood. Androgens and anabolic steroids activate Notch signaling in animal models of aging and thereby mitigate sarcopenia. To explore the molecular mechanisms by which nandrolone prevents denervation-atrophy, we investigated the effects of nandrolone on Notch signaling in denervated rat gastrocnemius muscle. Denervation significantly increased Notch activity reflected by elevated levels of nuclear Notch intracellular domain (NICD) and expression of Hey1 (a Notch target gene). Activation was greatest at 7 and 35 days after denervation but remained present at 56 days after denervation. Activation of Notch in denervated muscle was prevented by nandrolone associated with upregulated expression of Numb mRNA and protein. These data demonstrate that denervation activates Notch signaling, and that nandrolone abrogates this response associated with increased expression of Numb, suggesting a potential mechanism by which nandrolone reduces denervation-atrophy. Copyright © 2011. Published by Elsevier Inc.

Rhesus monkey model of liver disease reflecting clinical disease progression and hepatic gene expression analysis

PubMed Central

Wang, Hong; Tan, Tao; Wang, Junfeng; Niu, Yuyu; Yan, Yaping; Guo, Xiangyu; Kang, Yu; Duan, Yanchao; Chang, Shaohui; Liao, Jianpeng; Si, Chenyang; Ji, Weizhi; Si, Wei

2015-01-01

Alcoholic liver disease (ALD) is a significant public health issue with heavy medical and economic burdens. The aetiology of ALD is not yet completely understood. The development of drugs and therapies for ALD is hampered by a lack of suitable animal models that replicate both the histological and metabolic features of human ALD. Here, we characterize a rhesus monkey model of alcohol-induced liver steatosis and hepatic fibrosis that is compatible with the clinical progression of the biochemistry and pathology in humans with ALD. Microarray analysis of hepatic gene expression was conducted to identify potential molecular signatures of ALD progression. The up-regulation of expression of hepatic genes related to liver steatosis (CPT1A, FASN, LEPR, RXRA, IGFBP1, PPARGC1A and SLC2A4) was detected in our rhesus model, as was the down-regulation of such genes (CYP7A1, HMGCR, GCK and PNPLA3) and the up-regulation of expression of hepatic genes related to liver cancer (E2F1, OPCML, FZD7, IGFBP1 and LEF1). Our results demonstrate that this ALD model reflects the clinical disease progression and hepatic gene expression observed in humans. These findings will be useful for increasing the understanding of ALD pathogenesis and will benefit the development of new therapeutic procedures and pharmacological reagents for treating ALD. PMID:26442469

Bi-directional transition nets

NASA Astrophysics Data System (ADS)

Staines, Anthony Spiteri

2017-06-01

Ordinary Petri nets are forward directed transition systems. Modern transition systems events and event flows are reversible. Hence modeling structures that reflect this are important. The creation of a bi-directional Petri net extends the modeling power of Petri nets. This work presents the successful implementation of a bi-directional transition net. Some toy examples in comparison to Petri nets are given showing the increased modeling power in a compacted form. The results show some interesting findings on how the expressive power of these structures has been increased.

Moderate Exercise Mitigates the Detrimental Effects of Aging on Tendon Stem Cells.

PubMed

Zhang, Jianying; Wang, James H-C

2015-01-01

Aging is known to cause tendon degeneration whereas moderate exercise imparts beneficial effects on tendons. Since stem cells play a vital role in maintaining tissue integrity, in this study we aimed to define the effects of aging and moderate exercise on tendon stem/progenitor cells (TSCs) using in vitro and in vivo models. TSCs derived from aging mice (9 and 24 months) proliferated significantly slower than TSCs obtained from young mice (2.5 and 5 months). In addition, expression of the stem cell markers Oct-4, nucleostemin (NS), Sca-1 and SSEA-1 in TSCs decreased in an age-dependent manner. Interestingly, moderate mechanical stretching (4%) of aging TSCs in vitro significantly increased the expression of the stem cell marker, NS, but 8% stretching decreased NS expression. Similarly, 4% mechanical stretching increased the expression of Nanog, another stem cell marker, and the tenocyte-related genes, collagen I and tenomodulin. However, 8% stretching increased expression of the non-tenocyte-related genes, LPL, Sox-9 and Runx-2, while 4% stretching had minimal effects on the expression of these genes. In the in vivo study, moderate treadmill running (MTR) of aging mice (9 months) resulted in the increased proliferation rate of aging TSCs in culture, decreased lipid deposition, proteoglycan accumulation and calcification, and increased the expression of NS in the patellar tendons. These findings indicate that while aging impairs the proliferative ability of TSCs and reduces their stemness, moderate exercise can mitigate the deleterious effects of aging on TSCs and therefore may be responsible for decreased aging-induced tendon degeneration.

Na+/H+ exchanger isoform 1-induced osteopontin expression facilitates cardiac hypertrophy through p90 ribosomal S6 kinase.

PubMed

Abdulrahman, Nabeel; Jaspard-Vinassa, Beatrice; Fliegel, Larry; Jabeen, Aayesha; Riaz, Sadaf; Gadeau, Alain-Pierre; Mraiche, Fatima

2018-05-01

Cardiovascular diseases are the leading cause of death worldwide. One in three cases of heart failure is due to dilated cardiomyopathy. The Na + /H + exchanger isoform 1 (NHE1), a multifunctional protein and the key pH regulator in the heart, has been demonstrated to be increased in this condition. We have previously demonstrated that elevated NHE1 activity induced cardiac hypertrophy in vivo. Furthermore, the overexpression of active NHE1 elicited modulation of gene expression in cardiomyocytes including an upregulation of myocardial osteopontin (OPN) expression. To determine the role of OPN in inducing NHE1-mediated cardiomyocyte hypertrophy, double transgenic mice expressing active NHE1 and OPN knockout were generated and assessed by echocardiography and the cardiac phenotype. Our studies showed that hearts expressing active NHE1 exhibited cardiac remodeling indicated by increased systolic and diastolic left ventricular internal diameter and increased ventricular volume. Moreover, these hearts demonstrated impaired function with decreased fractional shortening and ejection fraction. Atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP) mRNA was upregulated, and there was an increase in heart cell cross-sectional area confirming the cardiac hypertrophic effect. Moreover, NHE1 transgenic mice also showed increased collagen deposition, upregulation of CD44 and phosphorylation of p90 ribosomal s6 kinase (RSK), effects that were regressed in OPN knockout mice. In conclusion, we developed an interesting comparative model of active NHE1 transgenic mouse lines which express a dilated hypertrophic phenotype expressing CD44 and phosphorylated RSK, effects which were regressed in absence of OPN.

Antiallodynic Effects of Bee Venom in an Animal Model of Complex Regional Pain Syndrome Type 1 (CRPS-I).

PubMed

Lee, Sung Hyun; Lee, Jae Min; Kim, Yun Hong; Choi, Jung Hyun; Jeon, Seung Hwan; Kim, Dong Kyu; Jeong, Hyeon Do; Lee, You Jung; Park, Hue Jung

2017-09-15

Neuropathic pain in a chronic post-ischaemic pain (CPIP) model mimics the symptoms of complex regional pain syndrome type I (CRPS I). The administration of bee venom (BV) has been utilized in Eastern medicine to treat chronic inflammatory diseases accompanying pain. However, the analgesic effect of BV in a CPIP model remains unknown. The application of a tight-fitting O-ring around the left ankle for a period of 3 h generated CPIP in C57/Bl6 male adult mice. BV (1 mg/kg ; 1, 2, and 3 times) was administered into the SC layer of the hind paw, and the antiallodynic effects were investigated using the von Frey test and by measuring the expression of neurokinin type 1 (NK-1) receptors in dorsal root ganglia (DRG). The administration of BV dose-dependently reduced the pain withdrawal threshold to mechanical stimuli compared with the pre-administration value and with that of the control group. After the development of the CPIP model, the expression of NK-1 receptors in DRG increased and then decreased following the administration of BV. SC administration of BV results in the attenuation of allodynia in a mouse model of CPIP. The antiallodynic effect was objectively proven through a reduction in the increased expression of NK-1 receptors in DRG.

Cocaine and sigma-1 receptors modulate HIV infection, chemokine receptors, and the HPA axis in the huPBL-SCID model.

PubMed

Roth, Michael D; Whittaker, Katherine M; Choi, Ruth; Tashkin, Donald P; Baldwin, Gayle Cocita

2005-12-01

Cocaine is associated with an increased risk for, and progression of, clinical disease associated with human immunodeficiency virus (HIV) infection. A human xenograft model, in which human peripheral blood mononuclear cells were implanted into severe combined immunodeficiency mice (huPBL-SCID) and infected with a HIV reporter virus, was used to investigate the biological interactions between cocaine and HIV infection. Systemic administration of cocaine (5 mg/kg/d) significantly increased the percentage of HIV-infected PBL (two- to threefold) and viral load (100- to 300-fold) in huPBL-SCID mice. Despite the capacity for cocaine to increase corticosterone and adrenocorticotropic hormone levels in control mice, the hypothalamic-pituitary-adrenal axis was suppressed in HIV-infected animals, and corticosterone levels were further decreased when animals were exposed to HIV and cocaine. Activating huPBL in vitro in the presence of 10(-8) M cocaine increased expression of CC chemokine receptor 5 (CCR5) and CXC chemokine receptor 4 (CXCR4) coreceptors. Expression of CCR5 was also increased at early time-points in the huPBL-SCID model following systemic exposure to cocaine (54.1+/-9.4% increase over control, P<0.01). This effect preceded the boost in viral infection and waned as HIV infection progressed. Cocaine has been shown to mediate immunosuppressive effects by activating sigma-1 receptors in immune cells in vitro and in vivo. Consistent with these reports, a selective sigma-1 antagonist, BD1047, blocked the effects of cocaine on HIV replication in the huPBL-SCID mouse. Our results suggest that systemic exposure to cocaine can enhance HIV infection in vivo by activating sigma-1 receptors and by modulating the expression of HIV coreceptors.

Robust Linear Models for Cis-eQTL Analysis.

PubMed

Rantalainen, Mattias; Lindgren, Cecilia M; Holmes, Christopher C

2015-01-01

Expression Quantitative Trait Loci (eQTL) analysis enables characterisation of functional genetic variation influencing expression levels of individual genes. In outbread populations, including humans, eQTLs are commonly analysed using the conventional linear model, adjusting for relevant covariates, assuming an allelic dosage model and a Gaussian error term. However, gene expression data generally have noise that induces heavy-tailed errors relative to the Gaussian distribution and often include atypical observations, or outliers. Such departures from modelling assumptions can lead to an increased rate of type II errors (false negatives), and to some extent also type I errors (false positives). Careful model checking can reduce the risk of type-I errors but often not type II errors, since it is generally too time-consuming to carefully check all models with a non-significant effect in large-scale and genome-wide studies. Here we propose the application of a robust linear model for eQTL analysis to reduce adverse effects of deviations from the assumption of Gaussian residuals. We present results from a simulation study as well as results from the analysis of real eQTL data sets. Our findings suggest that in many situations robust models have the potential to provide more reliable eQTL results compared to conventional linear models, particularly in respect to reducing type II errors due to non-Gaussian noise. Post-genomic data, such as that generated in genome-wide eQTL studies, are often noisy and frequently contain atypical observations. Robust statistical models have the potential to provide more reliable results and increased statistical power under non-Gaussian conditions. The results presented here suggest that robust models should be considered routinely alongside other commonly used methodologies for eQTL analysis.

The expression of 11β-hydroxysteroid dehydrogenase type 1 is increased in experimental periodontitis in rats.

PubMed

Nakata, Takaya; Umeda, Makoto; Masuzaki, Hiroaki; Sawai, Hirofumi

2016-10-03

The involvement of 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1), which converts inactive glucocorticoids into active glucocorticoids intracellularly, in metabolic diseases and chronic inflammatory diseases has been elucidated. We recently reported that an increase in 11β-HSD1 expression was associated with chronic periodontitis in humans irrespective of obesity. To further clarify the role of 11β-HSD1 in chronic periodontitis, the expression of 11β-HSD1 was investigated in experimental periodontitis model in rats. Experimental periodontitis was induced by silk ligature of left maxillary second molars of 7-week-old male Wistar rats, and periodontal tissues were collected at day 3. The expression of 11β-HSD1, 11β-HSD2, and TNFα mRNA was examined using real time reverse transcription-polymerase chain reaction. The expression of TNFα was used as an indicator of inflammation. Thus, the rats in which the levels of TNFα mRNA were increased in the ligature-induced periodontitis compared with the control were analysed. The findings demonstrated that the expression of 11β-HSD1 mRNA was significantly increased in experimental periodontitis compared with the control. The increase in the levels of 11β-HSD1 mRNA in the ligature-induced periodontitis compared with the control was positively correlated with that of TNFα mRNA. On the other hand, the expression of 11β-HSD2 mRNA, which inactivates glucocorticoids, was slightly decreased in experimental periodontitis. Therefore, the ratio of 11β-HSD1 versus 11β-HSD2 mRNA was significantly higher in experimental periodontitis than in the control. These results suggest that the increased expression of 11β-HSD1, which would result in the increased levels of intracellular glucocorticoids, may play a role in the pathophysiology of experimental periodontitis.

Fructose-induced increases in expression of intestinal fructolytic and gluconeogenic genes are regulated by GLUT5 and KHK

PubMed Central

Patel, Chirag; Douard, Veronique; Yu, Shiyan; Tharabenjasin, Phuntila; Gao, Nan

2015-01-01

Marked increases in fructose consumption have been tightly linked to metabolic diseases. One-third of ingested fructose is metabolized in the small intestine, but the underlying mechanisms regulating expression of fructose-metabolizing enzymes are not known. We used genetic mouse models to test the hypothesis that fructose absorption via glucose transporter protein, member 5 (GLUT5), metabolism via ketohexokinase (KHK), as well as GLUT5 trafficking to the apical membrane via the Ras-related protein in brain 11a (Rab11a)-dependent endosomes are required for the regulation of intestinal fructolytic and gluconeogenic enzymes. Fructose feeding increased the intestinal mRNA and protein expression of these enzymes in the small intestine of adult wild-type (WT) mice compared with those gavage fed with lysine or glucose. Fructose did not increase expression of these enzymes in the GLUT5 knockout (KO) mice. Blocking intracellular fructose metabolism by KHK ablation also prevented fructose-induced upregulation. Glycolytic hexokinase I expression was similar between WT and GLUT5- or KHK-KO mice and did not vary with feeding solution. Gavage feeding with the fructose-specific metabolite glyceraldehyde did not increase enzyme expression, suggesting that signaling occurs before the hydrolysis of fructose to three-carbon compounds. Impeding GLUT5 trafficking to the apical membrane using intestinal epithelial cell-specific Rab11a-KO mice impaired fructose-induced upregulation. KHK expression was uniformly distributed along the villus but was localized mainly in the basal region of the cytosol of enterocytes. The feedforward upregulation of fructolytic and gluconeogenic enzymes specifically requires GLUT5 and KHK and may proactively enhance the intestine's ability to process anticipated increases in dietary fructose concentrations. PMID:26084694

Protection by sulforaphane from type 1 diabetes-induced testicular apoptosis is associated with the up-regulation of Nrf2 expression and function.

PubMed

Jiang, Xin; Bai, Yang; Zhang, Zhiguo; Xin, Ying; Cai, Lu

2014-09-01

Diabetes-induced testicular apoptosis is predominantly due to increased oxidative stress. The nuclear factor-erythroid 2-related factor 2 (Nrf2), as a master transcription factor in controlling anti-oxidative systems, is able to be induced by sulforaphane (SFN). To examine whether SFN prevents testicular apoptosis, type 1 diabetic mouse model was induced with multiple low-dose streptozotocin. Diabetic and age-matched control mice were treated with and without SFN at 0.5mg/kg daily in five days of each week for 3months and then kept until 6months. Diabetes significantly increased testicular apoptosis that was associated with endoplasmic reticulum stress and mitochondrial cell death pathways, shown by the increased expression of C/EBP homologous protein (CHOP), cleaved caspase-12, Bax to Bcl2 expression ratio, and cleaved caspase-3. Diabetes also significantly increased testicular oxidative damage, inflammation and fibrosis, and decreased germ cell proliferation. All these diabetic effects were significantly prevented by SFN treatment for the first 3months, and the protective effect could be sustained at 3months after SFN treatment. SFN was able to up-regulate Nrf2 expression and function. The latter was reflected by the increased phosphorylation of Nrf2 at Ser40 and expression of Nrf2 downstream antioxidants at mRNA and protein levels. These results suggest that type 1 diabetes significantly induced testicular apoptosis and damage along with increasing oxidative stress and cell death and suppressing Nrf2 expression and function. SFN is able to prevent testicular oxidative damage and apoptosis in type 1 diabetes mice, which may be associated with the preservation of testicular Nrf2 expression and function under diabetic condition. Copyright © 2014 Elsevier Inc. All rights reserved.

Macrophage migration inhibitory factor induces epithelial to mesenchymal transition, enhances tumor aggressiveness and predicts clinical outcome in resected pancreatic ductal adenocarcinoma.

PubMed

Funamizu, Naotake; Hu, Chaoxin; Lacy, Curtis; Schetter, Aaron; Zhang, Geng; He, Peijun; Gaedcke, Jochen; Ghadimi, Michael B; Ried, Thomas; Yfantis, Harris G; Lee, Dong H; Subleski, Jeffrey; Chan, Tim; Weiss, Jonathan M; Back, Timothy C; Yanaga, Katsuhiko; Hanna, Nader; Alexander, H Richard; Maitra, Anirban; Hussain, S Perwez

2013-02-15

MIF is a proinflammatory cytokine and is implicated in cancer. A higher MIF level is found in many human cancer and cancer-prone inflammatory diseases, including chronic pancreatitis and pancreatic cancer. We tested the hypothesis that MIF contributes to pancreatic cancer aggressiveness and predicts disease outcome in resected cases. Consistent with our hypothesis we found that an elevated MIF mRNA expression in tumors was significantly associated with poor outcome in resected cases. Multivariate Cox-regression analysis further showed that MIF is independently associated with patients' survival (HR = 2.26, 95% CI = 1.17-4.37, p = 0.015). Mechanistic analyses revealed that MIF overexpression decreased E-cadherin and increased vimentin mRNA and protein levels in pancreatic cancer cell lines, consistent with the features of epithelial-to-mesenchymal transition (EMT). Furthermore, MIF-overexpression significantly increased ZEB1/2 and decreased miR-200b expression, while shRNA-mediated inhibition of MIF increased E-cadherin and miR-200b expression, and reduced the expression of ZEB1/2 in Panc1 cells. Re-expression of miR-200b in MIF overexpressing cells restored the epithelial characteristics, as indicated by an increase in E-cadherin and decrease in ZEB1/2 and vimentin expression. A reduced sensitivity to the chemotherapeutic drug, gemcitabine, occurred in MIF-overexpressing cells. Indicative of an increased malignant potential, MIF over-expressing cells showed significant increase in their invasion ability in vitro, and tumor growth and metastasis in an orthotopic xenograft mouse model. These results support a role of MIF in disease aggressiveness, indicating its potential usefulness as a candidate target for designing improved treatment in pancreatic cancer. Copyright © 2012 UICC.

Developmental programming: prenatal testosterone excess disrupts anti-Müllerian hormone expression in preantral and antral follicles.

PubMed

Veiga-Lopez, Almudena; Ye, Wen; Padmanabhan, Vasantha

2012-03-01

To investigate the impact of prenatal T excess on the expression of key ovarian regulators implicated in follicular recruitment and persistence using a large animal model of polycystic ovarian syndrome (PCOS). Interventional, animal model study. Academic research unit. A total of 25 female fetuses, 14 prepubertal female, and 24 adult female Suffolk sheep. Prenatal T treatment. Immunohistochemical determination of expression of anti-Müllerian hormone (AMH), kit ligand, and growth differentiation factor 9 (GDF9) in fetal, prepubertal, and adult ovarian tissues. Prenatal T treatment reduced the AMH protein expression in granulosa cells (GC) of preantral follicles and increased its expression in antral follicles compared with age-matched adult controls. These differences were not evident in prepubertal animals. Protein expression of GDF9 and kit ligand was not altered at any of the developmental time points studied. Prenatal T exposure is associated with changes in AMH expression in preantral and antral follicles in adult ovaries, similar to findings in women with PCOS. These findings indicate that abnormal folliculogenesis in PCOS may be at least in part mediated by changes in AMH expression. Copyright © 2012 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

Novel Dedifferentiated Liposarcoma Xenograft Models Reveal PTEN Down-Regulation as a Malignant Signature and Response to PI3K Pathway Inhibition

PubMed Central

Smith, Kathleen B.; Tran, Linh M.; Tam, Brenna M.; Shurell, Elizabeth M.; Li, Yunfeng; Braas, Daniel; Tap, William D.; Christofk, Heather R.; Dry, Sarah M.; Eilber, Fritz C.; Wu, Hong

2014-01-01

Liposarcoma is a type of soft tissue sarcoma that exhibits poor survival and a high recurrence rate. Treatment is generally limited to surgery and radiation, which emphasizes the need for better understanding of this disease. Because very few in vivo and in vitro models can reproducibly recapitulate the human disease, we generated several xenograft models from surgically resected human dedifferentiated liposarcoma. All xenografts recapitulated morphological and gene expression characteristics of the patient tumors after continuous in vivo passages. Importantly, xenograftability was directly correlated with disease-specific survival of liposarcoma patients. Thus, the ability for the tumor of a patient to engraft may help identify those patients who will benefit from more aggressive treatment regimens. Gene expression analyses highlighted the association between xenograftability and a unique gene expression signature, including down-regulated PTEN tumor-suppressor gene expression and a progenitor-like phenotype. When treated with the PI3K/AKT/mTOR pathway inhibitor rapamycin alone or in combination with the multikinase inhibitor sorafenib, all xenografts responded with increased lipid content and a more differentiated gene expression profile. These human xenograft models may facilitate liposarcoma research and accelerate the generation of readily translatable preclinical data that could ultimately influence patient care. PMID:23416162

Potential Influence of Metro on Bus: A Case Study

NASA Astrophysics Data System (ADS)

Selvakumar, M.; Abishek Reddy, M.; Sathish, V.; Venkatesh, R.

2018-03-01

A modal shift occurs when one mode of transport has a comparative advantage in a similar market over another. The present work concerns with the development of modal shift model for urban travel in Chennai, India. The modal shift model was calibrated using binary logit technique and validated using hold-out sample method. The validated model was used to predict the probability of shift in selected corridor. The recent introduction of metro rail in Chennai has lead to an increasing competition among public transport modes. To study the influence of metro on bus transport, a Stated Preference (SP) survey was conducted among express bus travellers. Using the SP survey data, a modal shift model was calibrated to estimate the plausible shift from bus to metro. Results indicate that variables like fare-difference, age, and income play an important role in the shift behaviour. When metro fare increases with respect to express bus fare, bus passengers are less willing to use metro and vice-versa.

Potential Influence of Metro on Bus: A Case Study

NASA Astrophysics Data System (ADS)

Selvakumar, M.; Abishek Reddy, M.; Sathish, V.; Venkatesh, R.

2018-06-01

A modal shift occurs when one mode of transport has a comparative advantage in a similar market over another. The present work concerns with the development of modal shift model for urban travel in Chennai, India. The modal shift model was calibrated using binary logit technique and validated using hold-out sample method. The validated model was used to predict the probability of shift in selected corridor. The recent introduction of metro rail in Chennai has lead to an increasing competition among public transport modes. To study the influence of metro on bus transport, a Stated Preference (SP) survey was conducted among express bus travellers. Using the SP survey data, a modal shift model was calibrated to estimate the plausible shift from bus to metro. Results indicate that variables like fare- difference, age, and income play an important role in the shift behaviour. When metro fare increases with respect to express bus fare, bus passengers are less willing to use metro and vice-versa.

2-Deoxy-D-glucose treatment induces ketogenesis, sustains mitochondrial function, and reduces pathology in female mouse model of Alzheimer's disease.

PubMed

Yao, Jia; Chen, Shuhua; Mao, Zisu; Cadenas, Enrique; Brinton, Roberta Diaz

2011-01-01

Previously, we demonstrated that mitochondrial bioenergetic deficits preceded Alzheimer's disease (AD) pathology in the female triple-transgenic AD (3xTgAD) mouse model. In parallel, 3xTgAD mice exhibited elevated expression of ketogenic markers, indicating a compensatory mechanism for energy production in brain. This compensatory response to generate an alternative fuel source was temporary and diminished with disease progression. To determine whether this compensatory alternative fuel system could be sustained, we investigated the impact of 2-deoxy-D-glucose (2-DG), a compound known to induce ketogenesis, on bioenergetic function and AD pathology burden in brain. 6-month-old female 3xTgAD mice were fed either a regular diet (AIN-93G) or a diet containing 0.04% 2-DG for 7 weeks. 2-DG diet significantly increased serum ketone body level and brain expression of enzymes required for ketone body metabolism. The 2-DG-induced maintenance of mitochondrial bioenergetics was paralleled by simultaneous reduction in oxidative stress. Further, 2-DG treated mice exhibited a significant reduction of both amyloid precursor protein (APP) and amyloid beta (Aβ) oligomers, which was paralleled by significantly increased α-secretase and decreased γ-secretase expression, indicating that 2-DG induced a shift towards a non-amyloidogenic pathway. In addition, 2-DG increased expression of genes involved in Aβ clearance pathways, degradation, sequestering, and transport. Concomitant with increased bioenergetic capacity and reduced β-amyloid burden, 2-DG significantly increased expression of neurotrophic growth factors, BDNF and NGF. Results of these analyses demonstrate that dietary 2-DG treatment increased ketogenesis and ketone metabolism, enhanced mitochondrial bioenergetic capacity, reduced β-amyloid generation and increased mechanisms of β-amyloid clearance. Further, these data link bioenergetic capacity with β-amyloid generation and demonstrate that β-amyloid burden was dynamic and reversible, as 2-DG reduced activation of the amyloidogenic pathway and increased mechanisms of β-amyloid clearance. Collectively, these data provide preclinical evidence for dietary 2-DG as a disease-modifying intervention to delay progression of bioenergetic deficits in brain and associated β-amyloid burden.

Expression of 11beta-hydroxysteroid dehydrogenase 1 and 2 in subcutaneous adipose tissue of lean and obese women with and without polycystic ovary syndrome.

PubMed

Svendsen, P F; Madsbad, S; Nilas, L; Paulsen, S K; Pedersen, S B

2009-11-01

To investigate the expression of 11beta-hydroxysteroid dehydrogenase (11beta-HSD) type 1 and 2 and hexose-6-phosphate dehydrogenase (H6PDH) mRNA in subcutaneous abdominal tissue from lean and obese women with and without polycystic ovary syndrome (PCOS), and to investigate the association between these enzymes and different measures of insulin sensitivity. Cross-sectional study. A total of 60 women, 36 women with PCOS, 17 lean (lean PCOS, LP) and 19 obese (obese PCOS, OP) and 24 age- and weight-matched control women, 8 lean (lean controls, LC) and 16 obese (obese controls, OC). Subcutaneous adipose tissue was collected from the abdomen. Peripheral insulin sensitivity was assessed by the euglycemic hyperinsulinemic clamp and determined as glucose disposal rate and insulin sensitivity index. Whole-body insulin sensitivity was calculated using homeostasis model assessment insulin resistance index. Body composition was evaluated by dual X-ray absorptiometry. Adipose mRNA expression of leptin and adiponectin were determined by real-time PCR. Polycystic ovary syndrome (P<0.05) and obesity (P<0.05) were independently associated with increased expression of 11beta-HSD1 mRNA. The subgroups LP and OC had increased 11beta-HSD1 and 11beta-HSD2 mRNA expression compared with LC (P<0.05, P<0.05). There were no effects of PCOS or obesity on11beta-HSD2 or H6PDH mRNA expression. Decreased peripheral insulin sensitivity (P<0.001) and increased upper body fat distribution (P<0.01) were associated with increased expression of 11beta-HSD1, but neither 11beta-HSD2 nor H6PDH. Polycystic ovary syndrome and obesity are independently associated with increased expression of 11beta-HSD1. This may lead to increased conversion of cortisone to cortisol in the peripheral adipose tissue and subsequently increased glucocorticoid activity. Decreased peripheral insulin sensitivity and central obesity was associated with increased expression of 11beta-HSD1.

RNA interference targeting α-synuclein attenuates methamphetamine-induced neurotoxicity in SH-SY5Y cells.

PubMed

Chen, Ling; Huang, Enping; Wang, Huijun; Qiu, Pingming; Liu, Chao

2013-07-12

The protein α-synuclein (α-syn) is abundant in neurons and has been claimed to play critical roles in the pathophysiology of Parkinson's disease. Overexpression of α-syn has been shown to be toxicity in methamphetamine (METH)-induced model in vivo and in vitro which has Parkinson's-like pathology. However, the exact mechanisms underlying toxicity of α-syn mediated METH-induced neuron remain unknown. In the present study, human dopaminergic-like neuroblastoma SH-SY5Y cells were used as METH-induced model in vitro. Cell viability was found to be dramatically increased after silencing α-syn expression followed by METH treatment compared with a-syn wild-type cells and the morphological damage to cells after METH treatment was abated through knockdown of α-syn expression in this model. The expression levels of tyrosine hydroxylase (TH), dopamine transporter (DAT) and vesicular monoamine transporter 2(VMAT-2) were significantly decreased and the activity/levels of reactive oxygen species (ROS), nitric oxide synthase (NOS) and nitrogen (NO) were notably increased after METH treatment. However, the changes of these expression levels were reversed in cells transfected with α-syn-shRNA. These results suggested that TH, DAT, VMAT-2, ROS and NOS maybe involved in α-syn mediated METH-induced neuronal toxicity. Copyright © 2013 Elsevier B.V. All rights reserved.

Prenatal administration of retinoic acid upregulates insulin-like growth factor receptors in the nitrofen-induced hypoplastic lung.

PubMed

Ruttenstock, Elke; Doi, Takashi; Dingemann, Jens; Puri, Prem

2011-04-01

Pulmonary hypoplasia (PH) is the main cause of mortality in newborns with congenital diaphragmatic hernia (CDH). Prenatal administration of retinoic acid (RA) stimulates alveologenesis in the nitrofen-induced pulmonary hypoplasia. Insulin-like growth factor receptors (IGFRs) play a crucial role in alveologenesis during lung development. We recently demonstrated that IGFRs were downregulated in later stages of lung development in the nitrofen CDH model. Several studies suggest the ability of RA to regulate insulin-like growth factor signaling. We hypothesized that IGFRs pulmonary gene expression is upregulated after the administration of RA in the nitrofen-induced CDH model. Pregnant rats were exposed to either olive oil or nitrofen on day 9 (D9) of gestation. RA was given intraperitoneally on days D18, D19, and D20. Fetal lungs were dissected on D21 and divided into control, control + RA, CDH, and CDH + RA group. IGFRs gene and protein expression were determined using RT-PCR and immunohistochemistry. mRNA expression levels of IGFRs were significantly increased in control + RA and CDH + RA compared with CDH group. Immunoreactivity of IGFRs was markedly increased in control + RA and CDH + RA compared with CDH lungs. Upregulation of pulmonary gene and protein expression of IGFRs after prenatal RA treatment in the nitrofen model suggests that RA may promote lung growth by stimulating IGFRs mediated alveologenesis. © 2011 Wiley-Liss, Inc.

Lycopene Enhances Docetaxel's Effect in Castration-Resistant Prostate Cancer Associated with Insulin-like Growth Factor I Receptor Levels1

PubMed Central

Tang, Yaxiong; Parmakhtiar, Basmina; Simoneau, Anne R; Xie, Jun; Fruehauf, John; Lilly, Michael; Zi, Xiaolin

2011-01-01

Docetaxel is currently the most effective drug for the treatment of castration-resistant prostate cancer (CRPC), but it only extends life by an average of 2 months. Lycopene, an antioxidant phytochemical, has antitumor activity against prostate cancer (PCa) in several models and is generally safe. We present data on the interaction between docetaxel and lycopene in CRPC models. The growth-inhibitory effect of lycopene on PCa cell lines was positively associated with insulin-like growth factor I receptor (IGF-IR) levels. In addition, lycopene treatment enhanced the growth-inhibitory effect of docetaxel more effectively on DU145 cells with IGF-IR high expression than on those PCa cell lines with IGF-IR low expression. In a DU145 xenograft tumor model, docetaxel plus lycopene caused tumor regression, with a 38% increase in antitumor efficacy (P = .047) when compared with docetaxel alone. Lycopene inhibited IGF-IR activation through inhibiting IGF-I stimulation and by increasing the expression and secretion of IGF-BP3. Downstream effects include inhibition of AKT kinase activity and survivin expression, followed by apoptosis. Together, the enhancement of docetaxel's antitumor efficacy by lycopene supplementation justifies further clinical investigation of lycopene and docetaxel combination for CRPC patients. CRPC patients with IGF-IR-overexpressing tumors may be most likely to benefit from this combination. PMID:21403837

Changes in matrix metalloproteinase network in a spontaneous autoimmune uveitis model.

PubMed

Hofmaier, Florian; Hauck, Stefanie M; Amann, Barbara; Degroote, Roxane L; Deeg, Cornelia A

2011-04-08

Autoimmune uveitis is a sight-threatening disease in which autoreactive T cells cross the blood-retinal barrier. Molecular mechanisms contributing to the loss of eye immune privilege in this autoimmune disease are not well understood. In this study, the authors investigated the changes in the matrix metalloproteinase network in spontaneous uveitis. Matrix metalloproteinase (MMP) MMP2, MMP9, and MMP14 expression and tissue inhibitor of metalloproteinase (TIMP)-2 and lipocalin 2 (LCN2) expression were analyzed using Western blot quantification. Enzyme activities were examined with zymography. Expression patterns of network candidates were revealed with immunohistochemistry, comparing physiological appearance and changes in a spontaneous recurrent uveitis model. TIMP2 protein expression was found to be decreased in both the vitreous and the retina of a spontaneous model for autoimmune uveitis (equine recurrent uveitis [ERU]), and TIMP2 activity was significantly reduced in ERU vitreous. Functionally associated MMPs such as MMP2, MMP14, and MMP9 were found to show altered or shifted expression and activity. Although MMP2 decreased in ERU vitreous, MMP9 expression and activity were found to be increased. These changes were reflected by profound changes within uveitic target tissue, where TIMP2, MMP9, and MMP14 decreased in expression, whereas MMP2 displayed a shifted expression pattern. LCN2, a potential stabilizer of MMP9, was found prominently expressed in equine healthy retina and displayed notable changes in expression patterns accompanied by significant upregulation in autoimmune conditions. Invading cells expressed MMP9 and LCN2. This study implicates a dysregulation or a change in functional protein-protein interactions in this TIMP2-associated protein network, together with altered expression of functionally related MMPs.

In vitro effects of nanosized diamond particles on macrophages.

PubMed

Shkurupy, V A; Arkhipov, S A; Neshchadim, D V; Akhramenko, E S; Troitskii, A V

2015-02-01

The effects of synthetic diamond nanoparticles (4-6 nm) on mouse macrophage biotropism and biocompatibility and the modulation of the macrophage functions (expression of IL-1α, TNF-α, GM-CSF, bFGF, and TGF-β) by nanoparticles in different concentrations were studied in vitro during exposure of different duration. Macrophage endocytosis of nanodiamonds increased with increasing the concentration of nanoparticles in culture and incubation time. Nanodiamonds exhibited high biotropism and biocompatibility towards macrophages; in doses of 10-20 μg/ml, they induced expression of GM-CSF and TGF-β, inhibited expression of bFGF, and did not stimulate IL-1α and TNF-α. These data indicate that nanodiamond capture by macrophages in the studied experimental model led to modulation of the functional status of macrophages that determine their capacity to stimulate reparative processes without increasing proinflammatory and profibrogenic status.

Age-associated increase in aneuploidy and changes in gene expression in mouse eggs

PubMed Central

Pan, Hua; Ma, Pengpeng; Zhu, Wenting; Schultz, Richard M.

2008-01-01

An increase in the incidence of aneuploidy is well documented with increasing maternal age, in particular in human females. Remarkably, little is known regarding the underlying molecular basis for the age-associated increase in aneuploidy, which is a major source of decreased fertility in humans. Using mouse as a model system we find that eggs obtained from old mice (60–70 weeks of age) display a six-fold increase in the incidence of hyperploidy as assessed by chromosome spreads. Expression profiling of transcripts in oocytes and eggs obtained from young and old mice reveals that ~5% of the transcripts are differentially expressed in oocytes obtained from old females when compared to oocytes obtained from young females (6–12 weeks of age) and that this fraction increases to ~33% in eggs. The latter finding indicates that the normal pattern of degradation of maternal mRNAs that occurs during oocyte maturation is dramatically altered in eggs obtained from old mice and could therefore be a contributing source to the decline in fertility. Analysis of the differentially expressed transcripts also indicated that the strength of the spindle assembly checkpoint is weakened and that higher errors of microtubule-kinetochore interactions constitute part of molecular basis for the ageassociated increase in aneuploidy in females. Last, BRCA1 expression is reduced in oocytes obtained from old females and RNAi-mediated reduction of BRCA1 in oocytes obtained from young females results in perturbing spindle formation and chromosome congression following maturation. PMID:18342300

Secoisolariciresinol Diglucoside Abrogates Oxidative Stress-Induced Damage in Cardiac Iron Overload Condition

PubMed Central

Puukila, Stephanie; Bryan, Sean; Laakso, Anna; Abdel-Malak, Jessica; Gurney, Carli; Agostino, Adrian; Belló-Klein, Adriane; Prasad, Kailash; Khaper, Neelam

2015-01-01

Cardiac iron overload is directly associated with cardiac dysfunction and can ultimately lead to heart failure. This study examined the effect of secoisolariciresinol diglucoside (SDG), a component of flaxseed, on iron overload induced cardiac damage by evaluating oxidative stress, inflammation and apoptosis in H9c2 cardiomyocytes. Cells were incubated with 50 μ5M iron for 24 hours and/or a 24 hour pre-treatment of 500 μ M SDG. Cardiac iron overload resulted in increased oxidative stress and gene expression of the inflammatory mediators tumor necrosis factor-α, interleukin-10 and interferon γ, as well as matrix metalloproteinases-2 and -9. Increased apoptosis was evident by increased active caspase 3/7 activity and increased protein expression of Forkhead box O3a, caspase 3 and Bax. Cardiac iron overload also resulted in increased protein expression of p70S6 Kinase 1 and decreased expression of AMP-activated protein kinase. Pre-treatment with SDG abrogated the iron-induced increases in oxidative stress, inflammation and apoptosis, as well as the increased p70S6 Kinase 1 and decreased AMP-activated protein kinase expression. The decrease in superoxide dismutase activity by iron treatment was prevented by pre-treatment with SDG in the presence of iron. Based on these findings we conclude that SDG was cytoprotective in an in vitro model of iron overload induced redox-inflammatory damage, suggesting a novel potential role for SDG in cardiac iron overload. PMID:25822525

Secoisolariciresinol diglucoside abrogates oxidative stress-induced damage in cardiac iron overload condition.

PubMed

Puukila, Stephanie; Bryan, Sean; Laakso, Anna; Abdel-Malak, Jessica; Gurney, Carli; Agostino, Adrian; Belló-Klein, Adriane; Prasad, Kailash; Khaper, Neelam

2015-01-01

Cardiac iron overload is directly associated with cardiac dysfunction and can ultimately lead to heart failure. This study examined the effect of secoisolariciresinol diglucoside (SDG), a component of flaxseed, on iron overload induced cardiac damage by evaluating oxidative stress, inflammation and apoptosis in H9c2 cardiomyocytes. Cells were incubated with 50 μ5M iron for 24 hours and/or a 24 hour pre-treatment of 500 μ M SDG. Cardiac iron overload resulted in increased oxidative stress and gene expression of the inflammatory mediators tumor necrosis factor-α, interleukin-10 and interferon γ, as well as matrix metalloproteinases-2 and -9. Increased apoptosis was evident by increased active caspase 3/7 activity and increased protein expression of Forkhead box O3a, caspase 3 and Bax. Cardiac iron overload also resulted in increased protein expression of p70S6 Kinase 1 and decreased expression of AMP-activated protein kinase. Pre-treatment with SDG abrogated the iron-induced increases in oxidative stress, inflammation and apoptosis, as well as the increased p70S6 Kinase 1 and decreased AMP-activated protein kinase expression. The decrease in superoxide dismutase activity by iron treatment was prevented by pre-treatment with SDG in the presence of iron. Based on these findings we conclude that SDG was cytoprotective in an in vitro model of iron overload induced redox-inflammatory damage, suggesting a novel potential role for SDG in cardiac iron overload.

Expression of TNF-alpha and immunohistochemical distribution of hepatic macrophage surface markers in carbon tetrachloride-induced chronic liver injury in rats.

PubMed

Orfila, C; Lepert, J C; Alric, L; Carrera, G; Beraud, M; Vinel, J P; Pipy, B

1999-10-01

In liver injury induced by carbon tetrachloride, secondary hepatic injury occurs from inflammatory processes originating from products released by activated Kupffer cells, which play a central role in hepatic inflammation. The purpose of our study was to demonstrate, in rats, the relationships between a function of the hepatic macrophages, TNF-alpha production and the state of activation of these cells, characterized by their phenotype, in the different phases of the process and development of fibrosis in a carbon tetrachloride-induced cirrhosis model. The immunohistochemical localization of proinflammatory cytokine TNF-alpha and surface surface makers (ED1 and ED2) was studied in hepatitis and cirrhosis in response to 3 and 9 weeks ingestion of carbon tetrachloride. After carbon tetrachloride ingestion, accompanying the increased necrosis, immunohistochemical analysis of liver tissue sections demonstrated the significantly increased number of cells expressing ED1, ED2 and TNF-alpha, compared to normal. The number of cells expressing the surface phenotypic markers of liver macrophages increased and this change was concomitantly associated with an increased cellular expression of TNF-alpha. Local macrophage proliferation and influx of newly recruited blood monocytes resulted in an increase of the macrophage population. The populational changes involved difference in functional activity and enhanced TNF-alpha expression. This cytokine expressed in the carbon tetrachloride-induced inflammatory process is associated with the development of fibrosis and may contribute to disease severity.

Increased Transendothelial Transport of CCL3 Is Insufficient to Drive Immune Cell Transmigration through the Blood-Brain Barrier under Inflammatory Conditions In Vitro.

PubMed

De Laere, Maxime; Sousa, Carmelita; Meena, Megha; Buckinx, Roeland; Timmermans, Jean-Pierre; Berneman, Zwi; Cools, Nathalie

2017-01-01

Many neuroinflammatory diseases are characterized by massive immune cell infiltration into the central nervous system. Identifying the underlying mechanisms could aid in the development of therapeutic strategies specifically interfering with inflammatory cell trafficking. To achieve this, we implemented and validated a blood-brain barrier (BBB) model to study chemokine secretion, chemokine transport, and leukocyte trafficking in vitro. In a coculture model consisting of a human cerebral microvascular endothelial cell line and human astrocytes, proinflammatory stimulation downregulated the expression of tight junction proteins, while the expression of adhesion molecules and chemokines was upregulated. Moreover, chemokine transport across BBB cocultures was upregulated, as evidenced by a significantly increased concentration of the inflammatory chemokine CCL3 at the luminal side following proinflammatory stimulation. CCL3 transport occurred independently of the chemokine receptors CCR1 and CCR5, albeit that migrated cells displayed increased expression of CCR1 and CCR5. However, overall leukocyte transmigration was reduced in inflammatory conditions, although higher numbers of leukocytes adhered to activated endothelial cells. Altogether, our findings demonstrate that prominent barrier activation following proinflammatory stimulation is insufficient to drive immune cell recruitment, suggesting that additional traffic cues are crucial to mediate the increased immune cell infiltration seen in vivo during neuroinflammation.

Induction of parkinsonism-related proteins in the spinal motor neurons of transgenic mouse carrying a mutant SOD1 gene.

PubMed

Morimoto, Nobutoshi; Nagai, Makiko; Miyazaki, Kazunori; Ohta, Yasuyuki; Kurata, Tomoko; Takehisa, Yasushi; Ikeda, Yoshio; Matsuura, Tohru; Asanuma, Masato; Abe, Koji

2010-06-01

Amyotrophic lateral sclerosis is a progressive and fatal disease caused by selective death of motor neurons, and a number of these patients carry mutations in the superoxide dismutase 1 (SOD1) gene involved in ameliorating oxidative stress. Recent studies indicate that oxidative stress and disruption of mitochondrial homeostasis is a common mechanism for motor neuron degeneration in amyotrophic lateral sclerosis and the loss of midbrain dopamine neurons in Parkinson's disease. Therefore, the present study investigated the presence and alterations of familial Parkinson's disease-related proteins, PINK1 and DJ-1, in spinal motor neurons of G93ASOD1 transgenic mouse model of amyotrophic lateral sclerosis. Following onset of disease, PINK1 and DJ-1 protein expression increased in the spinal motor neurons. The activated form of p53 also increased and translocated to the nuclei of spinal motor neurons, followed by increased expression of p53-activated gene 608 (PAG608). This is the first report demonstrating that increased expression of PAG608 correlates with activation of phosphorylated p53 in spinal motor neurons of an amyotrophic lateral sclerosis model. These results provide further evidence of the profound correlations between spinal motor neurons of amyotrophic lateral sclerosis and parkinsonism-related proteins.

Methionine Deprivation Induces a Targetable Vulnerability in Triple-Negative Breast Cancer Cells by Enhancing TRAIL Receptor-2 Expression.

PubMed

Strekalova, Elena; Malin, Dmitry; Good, David M; Cryns, Vincent L

2015-06-15

Many neoplasms are vulnerable to methionine deficiency by mechanisms that are poorly understood. Because gene profiling studies have revealed that methionine depletion increases TNF-related apoptosis-inducing ligand receptor-2 (TRAIL-R2) mRNA, we postulated that methionine stress sensitizes breast cancer cells to proapoptotic TRAIL-R2 agonists. Human triple (ER/PR/HER2)-negative breast carcinoma cell lines were cultured in control or methionine-free media. The effects of methionine depletion on TRAIL receptor expression and sensitivity to chemotherapy or a humanized agonistic TRAIL-R2 monoclonal antibody (lexatumumab) were determined. The melanoma-associated antigen MAGED2 was silenced to delineate its functional role in sensitizing TNBC cells to methionine stress. An orthotopic TNBC model was utilized to evaluate the effects of dietary methionine deficiency, lexatumumab, or the combination. Methionine depletion sensitized TNBC cells to lexatumumab-induced caspase activation and apoptosis by increasing TRAIL-R2 mRNA and cell surface expression. MCF-10A cells transformed by oncogenic H-Ras, but not untransformed cells, and matrix-detached TNBC cells were highly sensitive to the combination of lexatumumab and methionine depletion. Proteomics analyses revealed that MAGED2, which has been reported to reduce TRAIL-R2 expression, was suppressed by methionine stress. Silencing MAGED2 recapitulated features of methionine deprivation, including enhanced mRNA and cell surface expression of TRAIL receptors and increased sensitivity to TRAIL receptor agonists. Dietary methionine deprivation enhanced the antitumor effects of lexatumumab in an orthotopic metastatic TNBC model. Methionine depletion exposes a targetable defect in TNBC cells by increasing TRAIL-R2 expression. Our findings provide the foundation for a clinical trial combining dietary methionine restriction and TRAIL-R2 agonists. Clin Cancer Res; 21(12); 2780-91. ©2015 AACR. ©2015 American Association for Cancer Research.

Hippocampal synapsin I, growth-associated protein-43, and microtubule-associated protein-2 immunoreactivity in learned helplessness rats and antidepressant-treated rats.

PubMed

Iwata, M; Shirayama, Y; Ishida, H; Kawahara, R

2006-09-01

Learned helplessness rats are thought to be an animal model of depression. To study the role of synapse plasticity in depression, we examined the effects of learned helplessness and antidepressant treatments on synapsin I (a marker of presynaptic terminals), growth-associated protein-43 (GAP-43; a marker of growth cones), and microtubule-associated protein-2 (MAP-2; a marker of dendrites) in the hippocampus by immunolabeling. (1) Learned helplessness rats showed significant increases in the expression of synapsin I two days after the attainment of learned helplessness, and significant decreases in the protein expression eight days after the achievement of learned helplessness. Subchronic treatment of naïve rats with imipramine or fluvoxamine significantly decreased the expression of synapsin I. (2) Learned helplessness increased the expression of GAP-43 two days and eight days after learned helplessness training. Subchronic treatment of naïve rats with fluvoxamine but not imipramine showed a tendency to decrease the expression of synapsin I. (3) Learned helplessness rats showed increased expression of MAP-2 eight days after the attainment of learned helplessness. Naïve rats subchronically treated with imipramine showed a tendency toward increased expression of MAP-2, but those treated with fluvoxamine did not. These results indicate that the neuroplasticity-related proteins synapsin I, GAP-43, and MAP-2 may play a role in the pathophysiology of depression and the mechanisms of antidepressants.

Podocyte-specific RAP1GAP expression contributes to focal segmental glomerulosclerosis–associated glomerular injury

PubMed Central

Potla, Uma; Ni, Jie; Vadaparampil, Justin; Yang, Guozhe; Leventhal, Jeremy S.; Campbell, Kirk N.; Chuang, Peter Y.; Morozov, Alexei; He, John C.; D’Agati, Vivette D.; Klotman, Paul E.; Kaufman, Lewis

2014-01-01

Injury to the specialized epithelial cells of the glomerulus (podocytes) underlies the pathogenesis of all forms of proteinuric kidney disease; however, the specific genetic changes that mediate podocyte dysfunction after injury are not fully understood. Here, we performed a large-scale insertional mutagenic screen of injury-resistant podocytes isolated from mice and found that increased expression of the gene Rap1gap, encoding a RAP1 activation inhibitor, ameliorated podocyte injury resistance. Furthermore, injured podocytes in murine models of disease and kidney biopsies from glomerulosclerosis patients exhibited increased RAP1GAP, resulting in diminished glomerular RAP1 activation. In mouse models, podocyte-specific inactivation of Rap1a and Rap1b induced massive glomerulosclerosis and premature death. Podocyte-specific Rap1a and Rap1b haploinsufficiency also resulted in severe podocyte damage, including features of podocyte detachment. Over-expression of RAP1GAP in cultured podocytes induced loss of activated β1 integrin, which was similarly observed in kidney biopsies from patients. Furthermore, preventing elevation of RAP1GAP levels in injured podocytes maintained β1 integrin–mediated adhesion and prevented cellular detachment. Taken together, our findings suggest that increased podocyte expression of RAP1GAP contributes directly to podocyte dysfunction by a mechanism that involves loss of RAP1-mediated activation of β1 integrin. PMID:24642466

The skin, a novel niche for recirculating B cells1

PubMed Central

Geherin, Skye A.; Fintushel, Sarah R.; Lee, Michael H.; Wilson, R. Paul; Patel, Reema T.; Alt, Carsten; Young, Alan J.; Hay, John B.; Debes, Gudrun F.

2012-01-01

B cells infiltrate the skin in many chronic inflammatory diseases caused by autoimmunity or infection. Despite potential contribution to disease, skin-associated B cells remain poorly characterized. Using an ovine model of granulomatous skin inflammation, we demonstrate that B cells increase in the skin and skin-draining afferent lymph during inflammation. Surprisingly, skin B cells are a heterogeneous population that is distinct from lymph node B cells, with more large lymphocytes as well as B-1-like B cells that co-express high levels IgM and CD11b. Skin B cells have increased MHCII, CD1, and CD80/86 expression compared with lymph node B cells, suggesting that they are well-suited for T cell activation at the site of inflammation. Furthermore, we show that skin accumulation of B cells and antibody-secreting cells during inflammation increases local antibody titers, which could augment host defense and autoimmunity. While skin B cells express typical skin homing receptors such as E-selectin ligand and alpha-4 and beta-1 integrins, they are unresponsive to ligands for chemokine receptors associated with T cell homing into skin. Instead, skin B cells migrate toward the cutaneously expressed CCR6 ligand CCL20. Our data support a model in which B cells use CCR6-CCL20 to recirculate through the skin, fulfilling a novel role in skin immunity and inflammation. PMID:22561151

A framework to understand the variations of PSD-95 expression in brain aging and in Alzheimer's disease.

PubMed

Savioz, Armand; Leuba, Geneviève; Vallet, Philippe G

2014-11-01

The postsynaptic density protein PSD-95 is a major element of synapses. PSD-95 is involved in aging, Alzheimer's disease (AD) and numerous psychiatric disorders. However, contradictory data about PSD-95 expression in aging and AD have been reported. Indeed in AD versus control brains PSD-95 varies according to regions, increasing in the frontal cortex, at least in a primary stage, and decreasing in the temporal cortex. In contrast, in transgenic mouse models of aging and AD PSD-95 expression is decreased, in behaviorally aged impaired versus unimpaired rodents it can decrease or increase and finally, it is increased in rodents grown in enriched environments. Different factors explain these contradictory results in both animals and humans, among others concomitant psychiatric endophenotypes, such as depression. The possible involvement of PSD-95 in reactive and/or compensatory mechanisms during AD progression is underscored, at least before the occurrence of important synaptic elimination. Thus, in AD but not in AD transgenic mice, enhanced expression might precede the diminution commonly observed in advanced aging. A two-compartments cell model, separating events taking place in cell bodies and synapses, is presented. Overall these data suggest that AD research will progress by untangling pathological from protective events, a prerequisite for effective therapeutic strategies. Copyright © 2014 Elsevier B.V. All rights reserved.

The Effects of Myo-Inositol and B and D Vitamin Supplementation in the db/+ Mouse Model of Gestational Diabetes Mellitus

PubMed Central

Plows, Jasmine F.; Budin, Florence; Andersson, Rebecka A. M.; Mills, Valerie J.; Mace, Katherine; Davidge, Sandra T.; Vickers, Mark H.; Baker, Philip N.; Silva-Zolezzi, Irma; Stanley, Joanna L.

2017-01-01

Gestational diabetes mellitus (GDM) is a growing concern, affecting an increasing number of pregnant women worldwide. By predisposing both the affected mothers and children to future disease, GDM contributes to an intergenerational cycle of obesity and diabetes. In order to stop this cycle, safe and effective treatments for GDM are required. This study sought to determine the treatment effects of dietary supplementation with myo-inositol (MI) and vitamins B2, B6, B12, and D in a mouse model of GDM (pregnant db/+ dams). In addition, the individual effects of vitamin B2 were examined. Suboptimal B2 increased body weight and fat deposition, decreased GLUT4 adipose tissue expression, and increased expression of inflammatory markers. MI supplementation reduced weight and fat deposition, and reduced expression of inflammatory markers in adipose tissue of mice on suboptimal B2. MI also significantly reduced the hyperleptinemia observed in db/+ mice, when combined with supplemented B2. MI was generally associated with adipose tissue markers of improved insulin sensitivity and glucose uptake, while the combination of vitamins B2, B6, B12, and D was associated with a reduction in adipose inflammatory marker expression. These results suggest that supplementation with MI and vitamin B2 could be beneficial for the treatment/prevention of GDM. PMID:28212289

The YAP1/SIX2 axis is required for DDX3-mediated tumor aggressiveness and cetuximab resistance in KRAS-wild-type colorectal cancer

PubMed Central

Wu, De-Wei; Lin, Po-Lin; Wang, Lee; Huang, Chi-Chou; Lee, Huei

2017-01-01

The mechanism underlying tumor aggressiveness and cetuximab (CTX) resistance in KRAS-wild-type (KRAS -WT) colorectal cancer remains obscure. We here provide evidence that DDX3 promoted soft agar growth and invasiveness of KRAS-WT cells, as already confirmed in KRAS-mutated cells. Mechanistically, increased KRAS expression induced ROS production, which elevated HIF-1α and YAP1 expression. Increased HIF-1α persistently promoted DDX3 expression via a KRAS/ROS/HIF-1α feedback loop. DDX3-mediated aggressiveness and CTX resistance were regulated by the YAP1/SIX2 axis in KRAS-WT cells and further confirmed in animal models. Kaplan-Meier and Cox regression analysis indicated that DDX3, KRAS, and YAP1 expression had prognostic value for OS and RFS in KRAS-WT and KRAS-mutated tumors, but SIX2 and YAP1/SIX2 were prognostic value only in KRAS-WT patients. The observation from patients seemed to support the mechanistic action of cell and animal models. We therefore suggest that combining YAP1 inhibitors with CTX may therefore suppress DDX3-mediated tumor aggressiveness and enhance CTX sensitivity in KRAS-WT colorectal cancer. PMID:28435452

Mena invasive (MenaINV) promotes multicellular streaming motility and transendothelial migration in a mouse model of breast cancer

PubMed Central

Roussos, Evanthia T.; Balsamo, Michele; Alford, Shannon K.; Wyckoff, Jeffrey B.; Gligorijevic, Bojana; Wang, Yarong; Pozzuto, Maria; Stobezki, Robert; Goswami, Sumanta; Segall, Jeffrey E.; Lauffenburger, Douglas A.; Bresnick, Anne R.; Gertler, Frank B.; Condeelis, John S.

2011-01-01

We have shown previously that distinct Mena isoforms are expressed in invasive and migratory tumor cells in vivo and that the invasion isoform (MenaINV) potentiates carcinoma cell metastasis in murine models of breast cancer. However, the specific step of metastatic progression affected by this isoform and the effects on metastasis of the Mena11a isoform, expressed in primary tumor cells, are largely unknown. Here, we provide evidence that elevated MenaINV increases coordinated streaming motility, and enhances transendothelial migration and intravasation of tumor cells. We demonstrate that promotion of these early stages of metastasis by MenaINV is dependent on a macrophage–tumor cell paracrine loop. Our studies also show that increased Mena11a expression correlates with decreased expression of colony-stimulating factor 1 and a dramatically decreased ability to participate in paracrine-mediated invasion and intravasation. Our results illustrate the importance of paracrine-mediated cell streaming and intravasation on tumor cell dissemination, and demonstrate that the relative abundance of MenaINV and Mena11a helps to regulate these key stages of metastatic progression in breast cancer cells. PMID:21670198

Hypoperfusion induces overexpression of beta-amyloid precursor protein mRNA in a focal ischemic rodent model.

PubMed

Shi, J; Yang, S H; Stubley, L; Day, A L; Simpkins, J W

2000-01-17

Silent stroke is one of the risk factors of dementia. In the present study, we used a novel focal ischemic animal model to investigate the effects of comparatively small changes of cerebral blood flow (CBF) on the expression of beta-amyloid precursor protein (APP) mRNA. Focal ischemia was achieved by introducing a 4-0 monofilament to the bifurcation of anterior and middle cerebral arteries. Brain samples were harvested from ischemic core and penumbra of cortices at 1, 4 and 7 days following ischemia. The expression of APP mRNA was assessed by RT-PCR. The CBF was decreased to 50% for 1 day after stroke and recovered to 90% at the fourth day after stroke. The changes of CBF were accompanied by an increase in the expression of APP mRNA. APP mRNA increased to 208% and 152% in the penumbra and core ischemic regions, respectively, on the fourth day after MCAO and remained high through the seventh day of ischemia. This study suggests brain hypoperfusion enhances APP mRNA expression and may contribute to the progression of cognitive impairment after silent stroke.

Mena invasive (MenaINV) promotes multicellular streaming motility and transendothelial migration in a mouse model of breast cancer.

PubMed

Roussos, Evanthia T; Balsamo, Michele; Alford, Shannon K; Wyckoff, Jeffrey B; Gligorijevic, Bojana; Wang, Yarong; Pozzuto, Maria; Stobezki, Robert; Goswami, Sumanta; Segall, Jeffrey E; Lauffenburger, Douglas A; Bresnick, Anne R; Gertler, Frank B; Condeelis, John S

2011-07-01

We have shown previously that distinct Mena isoforms are expressed in invasive and migratory tumor cells in vivo and that the invasion isoform (Mena(INV)) potentiates carcinoma cell metastasis in murine models of breast cancer. However, the specific step of metastatic progression affected by this isoform and the effects on metastasis of the Mena11a isoform, expressed in primary tumor cells, are largely unknown. Here, we provide evidence that elevated Mena(INV) increases coordinated streaming motility, and enhances transendothelial migration and intravasation of tumor cells. We demonstrate that promotion of these early stages of metastasis by Mena(INV) is dependent on a macrophage-tumor cell paracrine loop. Our studies also show that increased Mena11a expression correlates with decreased expression of colony-stimulating factor 1 and a dramatically decreased ability to participate in paracrine-mediated invasion and intravasation. Our results illustrate the importance of paracrine-mediated cell streaming and intravasation on tumor cell dissemination, and demonstrate that the relative abundance of Mena(INV) and Mena11a helps to regulate these key stages of metastatic progression in breast cancer cells.

Cancer and systemic sclerosis: novel insights into pathogenesis and clinical implications

PubMed Central

Shah, Ami A.; Rosen, Antony

2012-01-01

Purpose of review Most epidemiologic studies have demonstrated an increased risk of cancer in scleroderma patients. Reasons for this risk increase have been poorly understood and often attributed to cytotoxic therapies or damage from scleroderma. Recognition that some patients have a close temporal relationship between cancer diagnosis and scleroderma clinical onset has focused attention on the possibility that scleroderma may be a paraneoplastic syndrome in a subset of patients. This review will discuss the latest epidemiologic data linking cancer and scleroderma and explore a model for the development of paraneoplastic scleroderma. Recent findings New investigations have demonstrated an association between RNA polymerase III autoantibodies and a close temporal relationship between cancer diagnosis and the development of clinical scleroderma. A unique nucleolar RNA polymerase III expression pattern has been identified in malignant tissue from these scleroderma patients suggesting that autoantigen expression in the cancer and the autoantibody response are associated. Similar data in inflammatory myositis have illustrated that disease-specific autoantigens may be expressed in cancers and damaged target tissues (muscle) undergoing regeneration. Summary These data suggest a model of paraneoplastic autoimmunity in which cross-reactive immune responses may target autoantigens that are expressed in both cancers and diseased autoimmune target tissues. PMID:21825998

Expression of cancer stem markers could be influenced by silencing of p16 gene in HeLa cervical carcinoma cells.

PubMed

Wu, H; Zhang, J; Shi, H

2016-01-01

Effect of the tumor suppression gene p16 on the biological characteristics of HeLa cervical carcinoma cells was explored. The expression of p16 protein was increased in HeLa tumor sphere cells, and no significant difference in tumor spheres from the first to the fourth passages. Compared with those of parental HeLa cells, the proportion of CD44+/CD24- and ABCG2+ cells increased significantly in tumor spheres. However after the cells were silenced by the p16-sh289 vector, expression of P16 protein and the cell number of CD44+/CD24- and ABCG2+ decreased. Moreover, HeLa cells with p16 gene silencing showed decreased abilities of sphere formation and matrigel invasion. More HeLa cells with p16 gene silence were needed for tumor formation in nude mice. Tumor size and weight in mouse model established with p16 gene silenced HeLa cells were less than those with HeLa parental cell model. The present results indicate that silencing of the p16 gene inhibits expression of cancer stem cell markers and tumorigenic ability of HeLa cells.

[Effect of Electroacupuncture at "Neiguan" (PC 6) and "Tianshu" (ST 25) for Colonic Motility and D 2 Receptor in Irritable Bowel Syndrome Rats].

PubMed

Wang, Shan; Guo, Meng-Wei; Gao, Yu-Shan; Ren, Xiao-Xuan; Lan, Ying; Ji, Mao-Xian; Wu, Yan-Ying; Li, Kai-Ge; Tan, Li-Hua; Sui, Ming-He

2018-01-25

To observe and compare the effects of electroacupuncture (EA) at "Tianshu" (ST 25) and "Neiguan" (PC 6) for colonic motility and the expression of colon dopamine D 2 in irritable bowel syndrome (IBS) rats, and to explore the specificity of different meridians and different acupoints. Forty Wistar newborn rats were randomly divided into blank, model, Tianshu and Neiguan groups. Separation of mother and child and acetic acid coloclyster combined with colorectal distension were used to establish IBS model in the model, Tianshu and Neiguan groups. At the age of 9 weeks, EA at bilateral ST 25 and PC 6 were applied in the corresponding groups 5 times, once every other day. After the intervention, the Bristol fecal score, the latent period of abdominal retraction reflex and the number of contraction waves were recorded. The expression of dopamine D 2 receptor was detected by immunohistochemistry. Compared with the blank group, the Bristol fecal score of the model group was higher ( P <0.01), the 1 st contraction wave latent period was shorter ( P <0.01), the number of contraction waves in 90 s increased ( P <0.01), the immunoreactive expression of D 2 receptor in colon decreased ( P <0.01). Compared with the model group, the Bristol fecal scores of the Tianshu and Neiguan groups decreased ( P <0.01), the 1 st contraction wave latent periods were longer ( P <0.01), the numbers of contraction waves in 90 s decreased ( P <0.01), the positive expressions of D 2 receptor in colon increased ( P <0.01, P <0.05). Compared with the Tianshu group, the immunoreactive expression of D 2 receptor in the Neiguan group decreased ( P <0.01). EA at ST 25 and PC 6 can improve the symptoms of colonic motility in IBS rats. The effect of EA at ST 25 is better, which indicates that different meridians and different acupoints play specific effects.

ANALYSIS OF TMEFF2 ALLOGRAFTS AND TRANSGENIC MOUSE MODELS REVEALS ROLES IN PROSTATE REGENERATION AND CANCER

PubMed Central

Corbin, JM.; Overcash, RF.; Wren, JD.; Coburn, A.; Tipton, GJ.; Ezzell, JA.; McNaughton, KK.; Fung, KM; Kosanke, SD.; Ruiz-Echevarria, MJ

2015-01-01

BACKGROUND Previous results from our lab indicate a tumor suppressor role for the transmembrane protein with epidermal growth factor and two follistatin motifs 2 (TMEFF2) in prostate cancer (PCa). Here, we further characterize this role and uncover new functions for TMEFF2 in cancer and adult prostate regeneration. METHODS The role of TMEFF2 was examined in PCa cells using Matrigel™ cultures and allograft models of PCa cells. In addition, we developed a transgenic mouse model that expresses TMEFF2 from a prostate specific promoter. Anatomical, histological and metabolic characterizations of the transgenic mouse prostate were conducted. The effect of TMEFF2 in prostate regeneration was studied by analyzing branching morphogenesis in the TMEFF2-expressing mouse lobes and alterations in branching morphogenesis were correlated with the metabolomic profiles of the mouse lobes. The role of TMEFF2 in prostate tumorigenesis in whole animals was investigated by crossing the TMEFF2 transgenic mice with the TRAMP mouse model of PCa and analyzing the histopathological changes in the progeny. RESULTS Ectopic expression of TMEFF2 impairs growth of PCa cells in Matrigel or allograft models. Surprisingly, while TMEFF2 expression in the TRAMP mouse did not have a significant effect on the glandular prostate epithelial lesions, the double TRAMP/TMEFF2 transgenic mice displayed an increased incidence of neuroendocrine type tumors. In addition, TMEFF2 promoted increased branching specifically in the dorsal lobe of the prostate suggesting a potential role in developmental processes. These results correlated with data indicating an alteration in the metabolic profile of the dorsal lobe of the transgenic TMEFF2 mice. CONCLUSIONS Collectively, our results confirm the tumor suppressor role of TMEFF2 and suggest that ectopic expression of TMEFF2 in mouse prostate leads to additional lobe-specific effects in prostate regeneration and tumorigenesis. This points to a complex and multifunctional role for TMEFF2 during PCa progression. PMID:26417683

Analysis of TMEFF2 allografts and transgenic mouse models reveals roles in prostate regeneration and cancer.

PubMed

Corbin, Joshua M; Overcash, Ryan F; Wren, Jonathan D; Coburn, Anita; Tipton, Greg J; Ezzell, Jennifer A; McNaughton, Kirk K; Fung, Kar-Ming; Kosanke, Stanley D; Ruiz-Echevarria, Maria J

2016-01-01

Previous results from our lab indicate a tumor suppressor role for the transmembrane protein with epidermal growth factor and two follistatin motifs 2 (TMEFF2) in prostate cancer (PCa). Here, we further characterize this role and uncover new functions for TMEFF2 in cancer and adult prostate regeneration. The role of TMEFF2 was examined in PCa cells using Matrigel(TM) cultures and allograft models of PCa cells. In addition, we developed a transgenic mouse model that expresses TMEFF2 from a prostate specific promoter. Anatomical, histological, and metabolic characterizations of the transgenic mouse prostate were conducted. The effect of TMEFF2 in prostate regeneration was studied by analyzing branching morphogenesis in the TMEFF2-expressing mouse lobes and alterations in branching morphogenesis were correlated with the metabolomic profiles of the mouse lobes. The role of TMEFF2 in prostate tumorigenesis in whole animals was investigated by crossing the TMEFF2 transgenic mice with the TRAMP mouse model of PCa and analyzing the histopathological changes in the progeny. Ectopic expression of TMEFF2 impairs growth of PCa cells in Matrigel or allograft models. Surprisingly, while TMEFF2 expression in the TRAMP mouse did not have a significant effect on the glandular prostate epithelial lesions, the double TRAMP/TMEFF2 transgenic mice displayed an increased incidence of neuroendocrine type tumors. In addition, TMEFF2 promoted increased branching specifically in the dorsal lobe of the prostate suggesting a potential role in developmental processes. These results correlated with data indicating an alteration in the metabolic profile of the dorsal lobe of the transgenic TMEFF2 mice. Collectively, our results confirm the tumor suppressor role of TMEFF2 and suggest that ectopic expression of TMEFF2 in mouse prostate leads to additional lobe-specific effects in prostate regeneration and tumorigenesis. This points to a complex and multifunctional role for TMEFF2 during PCa progression. © 2015 Wiley Periodicals, Inc.

Precision-cut rat, mouse, and human intestinal slices as novel models for the early-onset of intestinal fibrosis.

PubMed

Pham, Bao Tung; van Haaften, Wouter Tobias; Oosterhuis, Dorenda; Nieken, Judith; de Graaf, Inge Anne Maria; Olinga, Peter

2015-04-01

Intestinal fibrosis (IF) is a major complication of inflammatory bowel disease. IF research is limited by the lack of relevant in vitro and in vivo models. We evaluated precision-cut intestinal slices (PCIS) prepared from human, rat, and mouse intestine as ex vivo models mimicking the early-onset of (human) IF. Precision-cut intestinal slices prepared from human (h), rat (r), and mouse (m) jejunum, were incubated up to 72 h, the viability of PCIS was assessed by ATP content and morphology, and the gene expression of several fibrosis markers was determined. The viability of rPCIS decreased after 24 h of incubation, whereas mPCIS and hPCIS were viable up to 72 h of culturing. Furthermore, during this period, gene expression of heat shock protein 47 and plasminogen activator inhibitor 1 increased in all PCIS in addition to augmented expression of synaptophysin in hPCIS, fibronectin (Fn2) and TGF-β1 in rPCIS, and Fn2 and connective tissue growth factor (Ctgf) in mPCIS. Addition of TGF-β1 to rPCIS or mPCIS induced the gene expression of the fibrosis markers Pro-collagen1a1, Fn2, and Ctgf in both species. However, none of the fibrosis markers was further elevated in hPCIS. We successfully developed a novel ex vivo model that can mimic the early-onset of fibrosis in the intestine using human, rat, and mouse PCIS. Furthermore, in rat and mouse PCIS, TGF-β1 was able to even further increase the gene expression of fibrosis markers. This indicates that PCIS can be used as a model for the early-onset of IF. © 2015 The Authors. Physiological Reports published by Wiley Periodicals, Inc. on behalf of the American Physiological Society and The Physiological Society.

[Effect of qihuang decoction on mRNA expressions of Bcl-2, Bax, and Caspase-3, 9 in intestinal mucosa epithelium of ischemia/reperfusion injured rats].

PubMed

Yu, Qing-Sheng; Yu, Hong-Liang; Pan, Jin-Fang

2011-02-01

To observe the effect of Qihuang Decoction (QHD) on mRNA expression of apoptosis genes Bcl-2, Bax, and signal transduction molecules Caspase-3, 9 in intestinal mucosa epithelium of ischemia/ reperfusion (I/R) injured rats. Forty Wistar rats were randomized equally into 4 groups, the control group, the model group, the glutamine group, and the QHD group. Rats in the latter two groups were gastric infused with glutamine and QHD respectively for 3 days, but saline was infused instead to rats in the control group and model group. After then, except those in the control group intervened only by sham operation, rats were made into I/R injured model by 45 min occlusion of superior mesenteric artery followed by 1 h reperfusion. Immediately after modeling, mRNA expressions of Bcl-2, Bax, Caspase-3, and Caspase-9 in intestinal mucosa epithelium of rats were detected by reverse transcription-polymerase chain reaction (RT-PCR). Compared with the control group, mRNA expressions of Bcl-2, Bax, Caspase-3 and Caspase-9 were higher in the other three groups (P < 0.05). Compared with the model group, Bcl-2 mRNA expression was higher, while the expressions of the other three indices were lower in both the glutamine group and the QHD group (P < 0.05); and comparisons between the glutamine group and the QHD group showed a more depressed Bax mRNA expression (0.281 +/- 0.087 vs 0.350 +/- 0.053) and higher Bcl-2/Bax ratio (1.648 vs 1. 374) in the QHD group. QHD can reduce the I/R injury in the intestinal mucosa epithelium by inhibiting the cell apoptosis. The mechanism may be correlated with increased Bcl-2 mRNA expressions and decreased mRNA expressions of Bax, Caspase-3 and Caspase-9.

Bisphenol A Exposure May Induce Hepatic Lipid Accumulation via Reprogramming the DNA Methylation Patterns of Genes Involved in Lipid Metabolism

NASA Astrophysics Data System (ADS)

Ke, Zhang-Hong; Pan, Jie-Xue; Jin, Lu-Yang; Xu, Hai-Yan; Yu, Tian-Tian; Ullah, Kamran; Rahman, Tanzil Ur; Ren, Jun; Cheng, Yi; Dong, Xin-Yan; Sheng, Jian-Zhong; Huang, He-Feng

2016-08-01

Accumulating evidence suggests a role of bisphenol A (BPA) in metabolic disorders. However, the underlying mechanism is still unclear. Using a mouse BPA exposure model, we investigated the effects of long-term BPA exposure on lipid metabolism and the underlying mechanisms. The male mice exposed to BPA (0.5 μg BPA /kg/day, a human relevant dose) for 10 months exhibited significant hepatic accumulation of triglycerides and cholesterol. The liver cells from the BPA-exposed mice showed significantly increased expression levels of the genes related to lipid synthesis. These liver cells showed decreased DNA methylation levels of Srebf1 and Srebf2, and increased expression levels of Srebf1 and Srebf2 that may upregulate the genes related to lipid synthesis. The expression levels of DNA methyltransferases were decreased in BPA-exposed mouse liver. Hepa1-6 cell line treated with BPA showed decreased expression levels of DNA methyltransferases and increased expression levels of genes involved in lipid synthesis. DNA methyltransferase knockdown in Hepa1-6 led to hypo-methylation and increased expression levels of genes involved in lipid synthesis. Our results suggest that long-term BPA exposure could induce hepatic lipid accumulation, which may be due to the epigenetic reprogramming of the genes involved in lipid metabolism, such as the alterations of DNA methylation patterns.

Bisphenol A Exposure May Induce Hepatic Lipid Accumulation via Reprogramming the DNA Methylation Patterns of Genes Involved in Lipid Metabolism

PubMed Central

Ke, Zhang-Hong; Pan, Jie-Xue; Jin, Lu-Yang; Xu, Hai-Yan; Yu, Tian-Tian; Ullah, Kamran; Rahman, Tanzil Ur; Ren, Jun; Cheng, Yi; Dong, Xin-Yan; Sheng, Jian-Zhong; Huang, He-Feng

2016-01-01

Accumulating evidence suggests a role of bisphenol A (BPA) in metabolic disorders. However, the underlying mechanism is still unclear. Using a mouse BPA exposure model, we investigated the effects of long-term BPA exposure on lipid metabolism and the underlying mechanisms. The male mice exposed to BPA (0.5 μg BPA /kg/day, a human relevant dose) for 10 months exhibited significant hepatic accumulation of triglycerides and cholesterol. The liver cells from the BPA-exposed mice showed significantly increased expression levels of the genes related to lipid synthesis. These liver cells showed decreased DNA methylation levels of Srebf1 and Srebf2, and increased expression levels of Srebf1 and Srebf2 that may upregulate the genes related to lipid synthesis. The expression levels of DNA methyltransferases were decreased in BPA-exposed mouse liver. Hepa1-6 cell line treated with BPA showed decreased expression levels of DNA methyltransferases and increased expression levels of genes involved in lipid synthesis. DNA methyltransferase knockdown in Hepa1-6 led to hypo-methylation and increased expression levels of genes involved in lipid synthesis. Our results suggest that long-term BPA exposure could induce hepatic lipid accumulation, which may be due to the epigenetic reprogramming of the genes involved in lipid metabolism, such as the alterations of DNA methylation patterns. PMID:27502578