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Sample records for mice express enhanced

  1. Treadmill exercise enhances NMDA receptor expression in schizophrenia mice

    PubMed Central

    Park, Joon-Ki; Lee, Sam-Jun; Kim, Tae-Won

    2014-01-01

    Schizophrenia is a serious psychiatric disorder with several symptoms including cognitive dysfunction. Although the causes of schizophrenia are still unclear, there is a strong suspicion that the abnormality in N-methyl-D-aspartate (NMDA) receptor may contribute to schizophrenia symptoms. In the present study, the effect of treadmill exercise on the NMDA receptor expression was evaluated using MK-801-induced schizophrenia mice. Immunohistochemistry for expressions of NMDA receptor tyrosine hydroxylase (TH) was conducted. Western blot for brain-derived neurotrophic factor (BDNF) was also performed. In the present results, the mice in the MK-801-treated group displayed reduced NMDA receptor expression. Enhanced TH expression and suppressed BDNF expression were also observed in the MK-801-treated mice. Treadmill exercise improved NMDA receptor expression in the MK-801-induced schizophrenia mice. Treadmill exercise also suppressed TH expression and enhanced BDNF expression in the MK-801-induced schizophrenia mice. The present study showed that down-regulation of NMDA receptor demonstrated schizophrenia-like parameters, meanwhile treadmill running improved schizophrenia-related parameters through enhancing NMDA receptor expression. PMID:24678500

  2. Enhanced neurofibrillary degeneration in transgenic mice expressing mutant tau and APP.

    PubMed

    Lewis, J; Dickson, D W; Lin, W L; Chisholm, L; Corral, A; Jones, G; Yen, S H; Sahara, N; Skipper, L; Yager, D; Eckman, C; Hardy, J; Hutton, M; McGowan, E

    2001-08-24

    JNPL3 transgenic mice expressing a mutant tau protein, which develop neurofibrillary tangles and progressive motor disturbance, were crossed with Tg2576 transgenic mice expressing mutant beta-amyloid precursor protein (APP), thus modulating the APP-Abeta (beta-amyloid peptide) environment. The resulting double mutant (tau/APP) progeny and the Tg2576 parental strain developed Abeta deposits at the same age; however, relative to JNPL3 mice, the double mutants exhibited neurofibrillary tangle pathology that was substantially enhanced in the limbic system and olfactory cortex. These results indicate that either APP or Abeta influences the formation of neurofibrillary tangles. The interaction between Abeta and tau pathologies in these mice supports the hypothesis that a similar interaction occurs in Alzheimer's disease. PMID:11520987

  3. Lymphocytes from wasted mice express enhanced spontaneous and {gamma}-ray-induced apoptosis

    SciTech Connect

    Woloschak, G.E. |; Chang-Liu, Chin-Mei; Chung, Jen; Libertin, C.R.

    1993-09-01

    Mice bearing the autosomal recessive mutation wasted (wst/wst) display a disease pattern including faulty repair of DNA damage in lymphocytes after radiation exposure, neurologic abnormalities, and immunodeficiency. Many of the features of this mouse model have suggested a premature or increased spontaneous frequency of apoptosis in thymocytes; past work has shown an inability to establish cultured T cell lines, an abnormally high death rate of stimulated T cells in culture, and an increased sensitivity of T cells to the killing effects of ionizing radiations in wst/wst mice relative to controls. The experiments reported here were designed to examine splenic and thymic lymphocytes from wasted and control mice for signs of early apoptosis. Our results revealed enhanced expression of Rp-8 mRNA (associated with apoptosis) in thymic lymphocytes and reduced expression in splenic lymphocytes of wst/wst mice relative to controls; expression of Rp-2 and Td-30 mRNA (induced during apoptosis) were not detectable in spleen or thymus. Higher spontaneous DNA fragmentation was observed in wasted mice than in controls; however, {gamma}-ray-induced DNA fragmentation peaked at a lower dose and occurred to a greater extent in wasted mice relative to controls. These results provide evidence for high spontaneous and {gamma}-ray-induced apoptosis in T cells of wasted mice as a mechanism underlying the observed lymphocyte and DNA repair abnormalities.

  4. Enhanced human papillomavirus type 8 oncogene expression levels are crucial for skin tumorigenesis in transgenic mice

    SciTech Connect

    Hufbauer, M.; Lazic, D.; Akguel, B.; Brandsma, J.L.; Pfister, H.; Weissenborn, S.J.

    2010-08-01

    Human papillomavirus 8 (HPV8) is involved in skin cancer development in epidermodysplasia verruciformis patients. Transgenic mice expressing HPV8 early genes (HPV8-CER) developed papillomas, dysplasias and squamous cell carcinomas. UVA/B-irradiation and mechanical wounding of HPV8-CER mouse skin led to prompt papilloma induction in about 3 weeks. The aim of this study was to analyze the kinetics and level of transgene expression in response to skin irritations. Transgene expression was already enhanced 1 to 2 days after UVA/B-irradiation or tape-stripping and maintained during papilloma development. The enhanced transgene expression could be assigned to UVB and not to UVA. Papilloma development was thus always paralleled by an increased transgene expression irrespective of the type of skin irritation. A knock-down of E6 mRNA by tattooing HPV8-E6-specific siRNA led to a delay and a lower incidence of papilloma development. This indicates that the early increase of viral oncogene expression is crucial for induction of papillomatosis.

  5. Muscle-specific transgenic expression of porcine myostatin propeptide enhances muscle growth in mice.

    PubMed

    Wang, Kaiyun; Li, Zicong; Li, Yang; Zeng, Jinyong; He, Chang; Yang, Jinzeng; Liu, Dewu; Wu, Zhenfang

    2013-10-01

    Myostatin is a well-known negative regulator of skeletal muscle growth. Inhibition of myostatin activity results in increased muscle mass. Myostatin propeptide, as a myostatin antagonist, could be applied to promote meat production in livestock such as pigs. In this study, we generated a transgenic mouse model expressing porcine myostatin propeptide under the control of muscle-specific regulatory elements. The mean body weight of transgenic mice from a line expressing the highest level of porcine myostatin propeptide was increased by 5.4 % (P = 0.023) and 3.2 % (P = 0.031) in males and females, respectively, at 8 weeks of age. Weight of carcass, fore limb and hind limb was respectively increased by 6.0 % (P = 0.038), 9.0 % (P = 0.014), 8.7 % (P = 0.036) in transgenic male mice, compared to wild-type male controls at the age of 9 weeks. Similarly, carcass, fore limb and hind limb of transgenic female mice was 11.4 % (P = 0.002), 14.5 % (P = 0.006) and 14.5 % (P = 0.03) respectively heavier than that of wild-type female mice. The mean cross-section area of muscle fiber was increased by 17 % (P = 0.002) in transgenic mice, in comparison with wild-type controls. These results demonstrated that porcine myostatin propeptide is effective in enhancement of muscle growth. The present study provided useful information for future study on generation of transgenic pigs overexpressing porcine myostatin propeptide for improvement of muscle mass. PMID:23543410

  6. Hematopoietic Stem Cell Regeneration Enhanced by Ectopic Expression of ROS-detoxifying Enzymes in Transplant Mice

    PubMed Central

    Miao, Weimin; XuFeng, Richard; Park, Moo-Rim; Gu, Haihui; Hu, Linping; Kang, Jin Wook; Ma, Shihui; Liang, Paulina H; Li, Yanxin; Cheng, Haizi; Yu, Hui; Epperly, Michael; Greenberger, Joel; Cheng, Tao

    2013-01-01

    High levels of reactive oxygen species (ROS) can exhaust hematopoietic stem cells (HSCs). Thus, maintaining a low state of redox in HSCs by modulating ROS-detoxifying enzymes may augment the regeneration potential of HSCs. Our results show that basal expression of manganese superoxide dismutase (MnSOD) and catalase were at low levels in long-term and short-term repopulating HSCs, and administration of a MnSOD plasmid and lipofectin complex (MnSOD-PL) conferred radiation protection on irradiated recipient mice. To assess the intrinsic role of elevated MnSOD or catalase in HSCs and hematopoietic progenitor cells, the MnSOD or catalase gene was overexpressed in mouse hematopoietic cells via retroviral transduction. The impact of MnSOD and catalase on hematopoietic progenitor cells was mild, as measured by colony-forming units (CFUs). However, overexpressed catalase had a significant beneficial effect on long-term engraftment of transplanted HSCs, and this effect was further enhanced after an insult of low-dose γ-irradiation in the transplant mice. In contrast, overexpressed MnSOD exhibited an insignificant effect on long-term engraftment of transplanted HSCs, but had a significant beneficial effect after an insult of sublethal irradiation. Taken together, these results demonstrate that HSC function can be enhanced by ectopic expression of ROS-detoxifying enzymes, especially after radiation exposure in vivo. PMID:23295952

  7. Hematopoietic stem cell regeneration enhanced by ectopic expression of ROS-detoxifying enzymes in transplant mice.

    PubMed

    Miao, Weimin; Xufeng, Richard; Park, Moo-Rim; Gu, Haihui; Hu, Linping; Kang, Jin Wook; Ma, Shihui; Liang, Paulina H; Li, Yanxin; Cheng, Haizi; Yu, Hui; Epperly, Michael; Greenberger, Joel; Cheng, Tao

    2013-02-01

    High levels of reactive oxygen species (ROS) can exhaust hematopoietic stem cells (HSCs). Thus, maintaining a low state of redox in HSCs by modulating ROS-detoxifying enzymes may augment the regeneration potential of HSCs. Our results show that basal expression of manganese superoxide dismutase (MnSOD) and catalase were at low levels in long-term and short-term repopulating HSCs, and administration of a MnSOD plasmid and lipofectin complex (MnSOD-PL) conferred radiation protection on irradiated recipient mice. To assess the intrinsic role of elevated MnSOD or catalase in HSCs and hematopoietic progenitor cells, the MnSOD or catalase gene was overexpressed in mouse hematopoietic cells via retroviral transduction. The impact of MnSOD and catalase on hematopoietic progenitor cells was mild, as measured by colony-forming units (CFUs). However, overexpressed catalase had a significant beneficial effect on long-term engraftment of transplanted HSCs, and this effect was further enhanced after an insult of low-dose γ-irradiation in the transplant mice. In contrast, overexpressed MnSOD exhibited an insignificant effect on long-term engraftment of transplanted HSCs, but had a significant beneficial effect after an insult of sublethal irradiation. Taken together, these results demonstrate that HSC function can be enhanced by ectopic expression of ROS-detoxifying enzymes, especially after radiation exposure in vivo. PMID:23295952

  8. Monocyte/macrophage androgen receptor suppresses cutaneous wound healing in mice by enhancing local TNF-alpha expression.

    PubMed

    Lai, Jiann-Jyh; Lai, Kuo-Pao; Chuang, Kuang-Hsiang; Chang, Philip; Yu, I-Chen; Lin, Wen-Jye; Chang, Chawnshang

    2009-12-01

    Cutaneous wounds heal more slowly in elderly males than in elderly females, suggesting a role for sex hormones in the healing process. Indeed, androgen/androgen receptor (AR) signaling has been shown to inhibit cutaneous wound healing. AR is expressed in several cell types in healing skin, including keratinocytes, dermal fibroblasts, and infiltrating macrophages, but the exact role of androgen/AR signaling in these different cell types remains unclear. To address this question, we generated and studied cutaneous wound healing in cell-specific AR knockout (ARKO) mice. General and myeloid-specific ARKO mice exhibited accelerated wound healing compared with WT mice, whereas keratinocyte- and fibroblast-specific ARKO mice did not. Importantly, the rate of wound healing in the general ARKO mice was dependent on AR and not serum androgen levels. Interestingly, although dispensable for wound closure, keratinocyte AR promoted re-epithelialization, while fibroblast AR suppressed it. Further analysis indicated that AR suppressed wound healing by enhancing the inflammatory response through a localized increase in TNF-alpha expression. Furthermore, AR enhanced local TNF-alpha expression via multiple mechanisms, including increasing the inflammatory monocyte population, enhancing monocyte chemotaxis by upregulating CCR2 expression, and enhancing TNF-alpha expression in macrophages. Finally, targeting AR by topical application of a compound (ASC-J9) that degrades AR protein resulted in accelerated healing, suggesting a potential new therapeutic approach that may lead to better treatment of wound healing. PMID:19907077

  9. Enhanced expression of Nrf2 in mice attenuates the fatty liver produced by a methionine- and choline-deficient diet☆

    PubMed Central

    Zhang, Yu-Kun Jennifer; Yeager, Ronnie L.; Tanaka, Yuji; Klaassen, Curtis D.

    2011-01-01

    Oxidative stress has been proposed as an important promoter of the progression of fatty liver diseases. The current study investigates the potential functions of the Nrf2–Keap1 signaling pathway, an important hepatic oxidative stress sensor, in a rodent fatty liver model. Mice with no (Nrf2-null), normal (wild type, WT), and enhanced (Keap1 knockdown, K1-kd) expression of Nrf2 were fed a methionine- and choline-deficient (MCD) diet or a control diet for 5 days. Compared to WT mice, the MCD diet-caused hepatosteatosis was more severe in the Nrf2-null mice and less in the K1-kd mice. The Nrf2-null mice had lower hepatic glutathione and exhibited more lipid peroxidation, whereas the K1-kd mice had the highest amount of glutathione in the liver and developed the least lipid peroxidation among the three genotypes fed the MCD diet. The Nrf2 signaling pathway was activated by the MCD diet, and the Nrf2-targeted cytoprotective genes Nqo1 and Gstα1/2 were induced in WT and even more in K1-kd mice. In addition, Nrf2-null mice on both control and MCD diets exhibited altered expression profiles of fatty acid metabolism genes, indicating Nrf2 may influence lipid metabolism in liver. For example, mRNA levels of long chain fatty acid translocase CD36 and the endocrine hormone Fgf21 were higher in livers of Nrf2-null mice and lower in the K1-kd mice than WT mice fed the MCD diet. Taken together, these observations indicate that Nrf2 could decelerate the onset of fatty livers caused by the MCD diet by increasing hepatic antioxidant and detoxification capabilities. PMID:20350562

  10. Enhanced expression of Nrf2 in mice attenuates the fatty liver produced by a methionine- and choline-deficient diet

    SciTech Connect

    Zhang, Yu-Kun Jennifer; Yeager, Ronnie L.; Tanaka, Yuji; Klaassen, Curtis D.

    2010-06-15

    Oxidative stress has been proposed as an important promoter of the progression of fatty liver diseases. The current study investigates the potential functions of the Nrf2-Keap1 signaling pathway, an important hepatic oxidative stress sensor, in a rodent fatty liver model. Mice with no (Nrf2-null), normal (wild type, WT), and enhanced (Keap1 knockdown, K1-kd) expression of Nrf2 were fed a methionine- and choline-deficient (MCD) diet or a control diet for 5 days. Compared to WT mice, the MCD diet-caused hepatosteatosis was more severe in the Nrf2-null mice and less in the K1-kd mice. The Nrf2-null mice had lower hepatic glutathione and exhibited more lipid peroxidation, whereas the K1-kd mice had the highest amount of glutathione in the liver and developed the least lipid peroxidation among the three genotypes fed the MCD diet. The Nrf2 signaling pathway was activated by the MCD diet, and the Nrf2-targeted cytoprotective genes Nqo1 and Gst{alpha}1/2 were induced in WT and even more in K1-kd mice. In addition, Nrf2-null mice on both control and MCD diets exhibited altered expression profiles of fatty acid metabolism genes, indicating Nrf2 may influence lipid metabolism in liver. For example, mRNA levels of long chain fatty acid translocase CD36 and the endocrine hormone Fgf21 were higher in livers of Nrf2-null mice and lower in the K1-kd mice than WT mice fed the MCD diet. Taken together, these observations indicate that Nrf2 could decelerate the onset of fatty livers caused by the MCD diet by increasing hepatic antioxidant and detoxification capabilities.

  11. Clavulanic acid enhances glutamate transporter subtype I (GLT-1) expression and decreases reinforcing efficacy of cocaine in mice.

    PubMed

    Kim, Jae; John, Joel; Langford, Dianne; Walker, Ellen; Ward, Sara; Rawls, Scott M

    2016-03-01

    The β-lactam antibiotic ceftriaxone (CTX) reduces cocaine reinforcement and relapse in preclinical assays through a mechanism involving activation of glutamate transporter subtype 1 (GLT-1). However, its poor brain penetrability and intravenous administration route may limit its therapeutic utility for indications related to CNS diseases. An alternative is clavulanic acid (CA), a structural analog of CTX that retains the β-lactam core required for GLT-1 activity but displays enhanced brain penetrability and oral activity relative to CTX. Here, we tested the hypothesis that CA (1, 10 mg/kg ip) would enhance GLT-1 expression and decrease cocaine self-administration (SA) in mice, but at lower doses than CTX. Experiments revealed that GLT-1 transporter expression in the nucleus accumbens of mice treated with repeated CA (1, 10 mg/kg) was enhanced relative to saline-treated mice. Repeated CA treatment (1 mg/kg) reduced the reinforcing efficacy of cocaine (0.56 mg/kg/inf) in mice maintained on a progressive-ratio (PR) schedule of reinforcement but did not affect acquisition of cocaine SA under fixed-ratio responding or acquisition or retention of learning. These findings suggest that the β-lactamase inhibitor CA can activate the cellular glutamate reuptake system in the brain reward circuit and reduce cocaine's reinforcing efficacy at 100-fold lower doses than CTX. PMID:26543027

  12. Enhanced expression of transient receptor potential channel 3 in uterine smooth muscle tissues of lipopolysaccharide-induced preterm delivery mice

    PubMed Central

    Zheng, Dongming; Zhang, Lijuan; Na, Quan; Liu, Sishi; Zhuang, Yanyan; Lv, Yuan; Liu, Caixia

    2016-01-01

    Objective(s): We aimed to investigate the influence of transient receptor potential channel 3 (TRPC3) on lipopolysaccharide-induced (LPS) preterm delivery mice. Materials and Methods: Mice were randomly assigned to the four groups: an unpregnant group, a mid-pregnancy group (E15), a term delivery group, and an LPS-induced preterm delivery group (intraperitoneal injection LPS at 15 days). Uterine smooth muscles were obtained through caesarean section; TRPC3 expression was measured by real-time PCR, western blotting, and immunohistochemistry. A specific inhibitor of TRPC3 (SKF96365) was injected into the LPS-induced preterm delivery group to determine whether the delivery interval was prolonged. Results: TRPC3 was primarily expressed in the uterine smooth muscle layer. In addition, the LPS-induced preterm delivery group had an obviously higher expression level of TRPC3 mRNA and protein compared with the unpregnant and E15 groups, which were close to term delivery. More importantly, SKF96365 prolongs the delivery interval of LPS-induced preterm delivery mice. Conclusion: Enhanced expression of TRPC3 may be associated with LPS-induced preterm delivery in mice. The specific inhibitor of TRPC3 (SKF96365) may be helpful for clinical treatment of preterm delivery. PMID:27403264

  13. Expression of the c-myc oncogene under control of an immunoglobulin enhancer in E mu-myc transgenic mice.

    PubMed

    Alexander, W S; Schrader, J W; Adams, J M

    1987-04-01

    Transgenic mice bearing a cellular myc oncogene coupled to the immunoglobulin heavy-chain enhancer (E mu) exhibit perturbed B-lymphocyte development and succumb to B lymphoid tumors. To investigate how the enhancer has affected myc expression, we analyzed the structure and abundance of myc transcripts in tissues of prelymphomatous mice and in the lymphomas. Expression of the E mu-myc transgene appeared to be confined largely to B lymphoid cells, being dominant in bone marrow, spleen, and lymph nodes, with no detectable expression in T cells or other hematopoietic lineages examined. The myc transcripts initiated very predominantly at the normal myc promoters, although use of the more upstream myc promoter was accentuated and an enhancer-associated promoter may be used infrequently. The level of E mu-myc transcripts in the preneoplastic lymphoid tissues and in the E mu-myc tumors was not markedly higher than myc RNA levels in proliferating normal lymphocytes. Thus, enforced expression of structurally normal myc transcripts at only a modestly elevated level has profound biological consequences. The absence of detectable endogenous c-myc RNA in any tumor, or in preneoplastic bone marrow, supports a negative feedback model for normal c-myc regulation. PMID:3037318

  14. Exenatide enhances cognitive performance and upregulates neurotrophic factor gene expression levels in diabetic mice.

    PubMed

    Gumuslu, Esen; Mutlu, Oguz; Celikyurt, Ipek K; Ulak, Guner; Akar, Furuzan; Erden, Faruk; Ertan, Merve

    2016-08-01

    Exenatide is a potent and selective agonist for the GLP-1 (glucagon-like peptide-1) receptor. Recent studies are focused on the effects of GLP-1 analogues on hippocampal neurogenesis, cognition, learning and memory functions. The aim of this study was to assess the effects of chronic exenatide treatment (0.1 μg/kg, s.c, twice daily for 2 weeks) on spatial memory functions by using the modified elevated plus maze (mEPM) test and emotional memory functions by using the passive avoidance (PA) test in streptozotocin/nicotinamide (STZ-NA)-induced diabetic mice. As the genes involved in neurite remodelling are among the primary targets of regulation, the effects of diabetes and chronic administration of exenatide on brain-derived neurotrophic factor (BDNF) and cyclic adenosine monophosphate (cAMP) response element binding protein (CREB) messenger ribonucleic acid (mRNA) levels in the hippocampus of mice were also determined using quantitative real-time polymerase chain reaction (RT-PCR). This study revealed that in the mEPM and PA tests, type-2 diabetes-induced mice exhibited significant impairment of learning and memory which were ameliorated by GLP-1 receptor agonist exenatide. Quantitative RT-PCR revealed that CREB and BDNF gene expression levels were downregulated in diabetic mice, and these alterations were increased by exenatide treatment. Since, exenatide improves cognitive ability in STZ/NA-induced diabetic mice and activates molecular mechanisms of memory storage in response to a learning experience, it may be a candidate for alleviation of mood and cognitive disorder. PMID:26935863

  15. Suppression of Parkin enhances nigrostriatal and motor neuron lesion in mice over-expressing human-mutated tau protein.

    PubMed

    Menéndez, J; Rodríguez-Navarro, J A; Solano, R M; Casarejos, M J; Rodal, I; Guerrero, R; Sánchez, M P; Avila, J; Mena, M A; de Yébenes, J G

    2006-07-01

    Abnormal deposition of protein tau takes place in the brain of patients with several neurodegenerative diseases. Few of these patients present frontotemporal dementia with parkinsonism and amyotrophy (FTDPA-17), an autosomal dominant tauopathy related to mutations of the gene that codes for protein tau, localized in chromosome 17. The great majority of patients with tauopathies such as Alzheimer's disease, sporadic frontotemporal dementia or progressive supranuclear palsy do not show a Mendelian pattern of inheritance. We have occasionally seen tauopathies in patients with parkin mutations and, therefore, hypothesized that the protein tau interacts with parkin. We have tested that hypothesis in mice with combined genetic modifications of tau (over-expression of human tau with three mutations known to produce FTDPA-17) and parkin (deleted) proteins. Homozygote parkin null or over-expressing mutated-human tau mice have subtle behavioral and molecular abnormalities but do not express a clinical phenotype of neurodegenerative disease. Mice with combined homozygous mutations of these two genes show progressively abnormal walking already noticeable at 3 months of age, loss of dopamine and dopamine markers in striatum, nuclear tau immunoreactive deposits in motor neurons of the spinal cord, abnormal expression of glial markers and enhanced levels of pro-apoptotic proteins; findings that were absent or less pronounced in homozygote animals with deletions of parkin or over-expression of tau. The double transgenic mice do not express normal mechanisms of adaptation to stress such as increased levels of GSH and Hsp-70. In addition, they have reduced levels of CHIP-Hsc70, a complex known to attenuate aggregation of tau and to enhance ubiquitination of phosphorylated tau. We have found high levels of phosphorylated tau in parkin-/-+tau(VLW) mice and a relative decrease of the inactivated pSer9 to total GSK-3 levels. Our data reveal that there are interactions between tau and

  16. A fast-evolving human NPAS3 enhancer gained reporter expression in the developing forebrain of transgenic mice

    PubMed Central

    Kamm, Gretel B.; López-Leal, Rodrigo; Lorenzo, Juan R.; Franchini, Lucía F.

    2013-01-01

    The developmental brain gene NPAS3 stands out as a hot spot in human evolution because it contains the largest number of human-specific, fast-evolving, conserved, non-coding elements. In this paper we studied 2xHAR142, one of these elements that is located in the fifth intron of NPAS3. Using transgenic mice, we show that the mouse and chimp 2xHAR142 orthologues behave as transcriptional enhancers driving expression of the reporter gene lacZ to a similar NPAS3 expression subdomain in the mouse central nervous system. Interestingly, the human 2xHAR142 orthologue drives lacZ expression to an extended expression pattern in the nervous system. Thus, molecular evolution of 2xHAR142 provides the first documented example of human-specific heterotopy in the forebrain promoted by a transcriptional enhancer and suggests that it may have contributed to assemble the unique properties of the human brain. PMID:24218632

  17. Enhanced Gastrointestinal Expression of Cytosolic Malic Enzyme (ME1) Induces Intestinal and Liver Lipogenic Gene Expression and Intestinal Cell Proliferation in Mice

    PubMed Central

    Al-Dwairi, Ahmed; Brown, Adam R.; Pabona, John Mark P.; Van, Trang H.; Hamdan, Hamdan; Mercado, Charles P.; Quick, Charles M.; Wight, Patricia A.; Simmen, Rosalia C. M.; Simmen, Frank A.

    2014-01-01

    The small intestine participates in lipid digestion, metabolism and transport. Cytosolic malic enzyme 1 (ME1) is an enzyme that generates NADPH used in fatty acid and cholesterol biosynthesis. Previous work has correlated liver and adipose ME1 expression with susceptibility to obesity and diabetes; however, the contributions of intestine-expressed ME1 to these conditions are unknown. We generated transgenic (Tg) mice expressing rat ME1 in the gastrointestinal epithelium under the control of the murine villin1 promoter/enhancer. Levels of intestinal ME1 protein (endogenous plus transgene) were greater in Tg than wildtype (WT) littermates. Effects of elevated intestinal ME1 on body weight, circulating insulin, select adipocytokines, blood glucose, and metabolism-related genes were examined. Male Tg mice fed a high-fat (HF) diet gained significantly more body weight than WT male littermates and had heavier livers. ME1-Tg mice had deeper intestinal and colon crypts, a greater intestinal 5-bromodeoxyuridine labeling index, and increased expression of intestinal lipogenic (Fasn, Srebf1) and cholesterol biosynthetic (Hmgcsr, Hmgcs1), genes. The livers from HF diet-fed Tg mice also exhibited an induction of cholesterol and lipogenic pathway genes and altered measures (Irs1, Irs2, Prkce) of insulin sensitivity. Results indicate that gastrointestinal ME1 via its influence on intestinal epithelial proliferation, and lipogenic and cholesterologenic genes may concomitantly impact signaling in liver to modify this tissue’s metabolic state. Our work highlights a new mouse model to address the role of intestine-expressed ME1 in whole body metabolism, hepatomegaly, and crypt cell proliferation. Intestinal ME1 may thus constitute a therapeutic target to reduce obesity-associated pathologies. PMID:25402228

  18. Treadmill exercise inhibits hippocampal apoptosis through enhancing N-methyl-D-aspartate receptor expression in the MK-801-induced schizophrenic mice

    PubMed Central

    Chung, Jin Woo; Seo, Jin-Hee; Baek, Sang-Bin; Kim, Chang-Ju; Kim, Tae-Woon

    2014-01-01

    Schizophrenia is a severe mental disorder characterized by abnormal mental functioning and disruptive behaviors. Abnormal expression of N-methyl-D-aspartate (NMDA) receptor, one of the glutamate receptor subtypes, has also been suggested to contribute to the symptoms of schizophrenia. The effect of treadmill exercise on schizophrenia-induced apoptosis in relation with NMDA receptor has not been evaluated. In the present study, we investigated the effect of treadmill exercise on neuronal apoptosis in the hippocampus using MK-801-induced schizophrenic mice. MK-801 was intraperitoneally injected once a day for 2 weeks. The mice in the exercise groups were forced to run on a treadmill exercise for 60 min, once a day for 2 weeks. In the present results, repeated injection of the NMDA receptor antagonist MK-801 reduced expression of NMDA receptor in hippocampal CA2-3 regions. MK-801 injection increased casapse-3 expression and enhanced cytochrome c release in the hippocampus. The ratio of Bax to Bcl-2 was higher in the MK-801-induced schizophrenia mice than the normal mice. In contrast, treadmill exercise enhanced NMDA receptor expression, suppressed caspae-3 activation and cytochrome c release, and inhibited the ratio of Bax to Bcl-2. Based on present finding, we concluded that NMDA receptor hypofunctioning induced neuronal apoptosis in MK-801-induced schizophrenic mice. Treadmill exercise suppressed neuronal apoptosis through enhancing NMDA receptor expression in schizophrenic mice. PMID:25210696

  19. Enhanced Human Immunodeficiency Virus Type 1 Expression and Neuropathogenesis in Knockout Mice Lacking Type I Interferon Responses

    PubMed Central

    He, Hongxia; Sharer, Leroy R.; Chao, Wei; Gu, Chao-Jiang; Borjabad, Alejandra; Hadas, Eran; Kelschenbach, Jennifer; Ichiyama, Koji; Do, Meilan; Potash, Mary Jane

    2014-01-01

    Abstract The roles of Type I interferon (IFN) in human immunodeficiency virus Type 1 (HIV-1) neuropathogenesis are poorly understood; both protective and deleterious effects of IFN signaling have been described. We used genetically modified mice deficient in the Type I IFN receptor (IFNRKO) to analyze the progress of HIV-1 brain infection and neuropathogenesis in the absence of IFN signaling. IFNRKO and wild-type (WT) mice on the 129xSv/Ev or C57BL/6 strain backgrounds were infected systemically with EcoHIV, a chimeric HIV-1 that productively infects mice. IFNRKO mice showed higher HIV-1 expression in spleen and peritoneal macrophages and greater virus infiltration into the brain compared to WT mice. Neuropathogenesis was studied by histopathological, immunohistochemical, immunofluorescence, and polymerase chain reaction analyses of brain tissues after the virus was inoculated into the brain by stereotaxic intracerebral injection. Both IFNRKO and WT mice showed readily detectable HIV-1 and brain lesions, including microglial activation, astrocytosis, and increased expression of genes coding for inflammatory cytokines and chemokines typical of human HIV-1 brain disease. Parameters of HIV-1 neuropathogenesis, including HIV-1 expression in microglia/macrophages, were significantly greater in IFNRKO than in WT mice. Our results show unequivocally that Type I IFN signaling and responses limit HIV-1 infection and pathogenesis in the brains of mice. PMID:24335529

  20. Estradiol replacement enhances fear memory formation, impairs extinction and reduces COMT expression levels in the hippocampus of ovariectomized female mice.

    PubMed

    McDermott, Carmel M; Liu, Dan; Ade, Catherine; Schrader, Laura A

    2015-02-01

    Females experience depression, posttraumatic stress disorder (PTSD), and anxiety disorders at approximately twice the rate of males, but the mechanisms underlying this difference remain undefined. The effect of sex hormones on neural substrates presents a possible mechanism. We investigated the effect of ovariectomy at two ages, before puberty and in adulthood, and 17β-estradiol (E2) replacement administered chronically in drinking water on anxiety level, fear memory formation, and extinction. Based on previous studies, we hypothesized that estradiol replacement would impair fear memory formation and enhance extinction rate. Females, age 4 weeks and 10 weeks, were divided randomly into 4 groups; sham surgery, OVX, OVX+low E2 (200nM), and OVX+high E2 (1000nM). Chronic treatment with high levels of E2 significantly increased anxiety levels measured in the elevated plus maze. In both age groups, high levels of E2 significantly increased contextual fear memory but had no effect on cued fear memory. In addition, high E2 decreased the rate of extinction in both ages. Finally, catechol-O-methyltransferase (COMT) is important for regulation of catecholamine levels, which play a role in fear memory formation and extinction. COMT expression in the hippocampus was significantly reduced by high E2 replacement, implying increased catecholamine levels in the hippocampus of high E2 mice. These results suggest that estradiol enhanced fear memory formation, and inhibited fear memory extinction, possibly stabilizing the fear memory in female mice. This study has implications for a neurobiological mechanism for PTSD and anxiety disorders. PMID:25555360

  1. Expression of human hormone-sensitive lipase in white adipose tissue of transgenic mice increases lipase activity but does not enhance in vitro lipolysis.

    PubMed

    Lucas, Stéphanie; Tavernier, Geneviève; Tiraby, Claire; Mairal, Aline; Langin, Dominique

    2003-01-01

    Hormone-sensitive lipase (HSL) catalyzes the hydrolysis of acylglycerols and cholesteryl esters (CEs). The enzyme is highly expressed in adipose tissues (ATs), where it is thought to play an important role in fat mobilization. The purpose of the present work was to study the effect of a physiological increase of HSL expression in vivo. Transgenic mice were produced with a 21 kb human genomic fragment encompassing the exons encoding the adipocyte form of HSL. hHSL mRNA was expressed at 3-fold higher levels than murine HSL mRNA in white adipocytes. Transgene expression was also observed in brown adipose tissue (BAT) and skeletal muscle. The human protein was detected in ATs of transgenic (Tg) mice. The hydrolytic activities against triacylglycerol (TG), diacylglycerol (DG) analog, and CE were increased in transgenic mouse AT. However, cAMP-inducible adipocyte lipolysis was lower in transgenic animals. In the B6CBA genetic background, transgenic mice up to 14 weeks of age showed lower body weight and fat mass. The phenotype was not observed in older animals and in mice fed a high-fat diet (HFD). In the OF1 genetic background, there was no difference in fat mass of mice fed ad libitum. However, transgenic mice became leaner than their wild-type (WT) littermates after a 4 day calorie restriction. The data show that overexpression of HSL, despite increased lipase activity, does not lead to enhanced lipolysis. PMID:12518034

  2. Enhanced Hypertrophy In ob/ob Mice Due To An Impairment in Expression Of Atrial Natriuretic Peptide

    PubMed Central

    Mascareno, Eduardo; Beckles, Daniel; Dhar-Mascareno, Manya; Siddiqui, M.A.Q.

    2009-01-01

    Rationale We investigated the molecular mechanism(s) that play a role in leptin signaling during the development of left ventricular hypertrophy (LVH) due to pressure overload. To this end, ob/ob leptin deficient and C57BL/6J control mice were subjected transverse aortic constriction (TAC). Methods Control sham C57BL/6J and ob/ob mice, along with C57BL/6J and ob/ob leptin deficient mice were subjected transverse aortic constriction (TAC) for 15 days and then evaluated for morphological, physiological, and molecular changes associated with pressure overload hypertrophy. Results Evaluation by echocardiography revealed a significant increase in left ventricular mass (LVmass) and wall thickness in ob/ob mice subjected to transverse aortic constriction (TAC) as compared to C57BL/6J. Analysis of the expression of molecular markers of LVH, such as atrial natriuretic peptide (ANP), revealed a blunted increase in the level of ANP in ob/ob mice as compared to C57BL/6J mice. We observed that leptin plays a role in modulating the transcriptional activity of the promoter of the ANP gene. Leptin acts by regulating NFATc4, a member of the nuclear factor activated T cell (NFAT) family of transcription factors in cardiomyocytes. Our in vivo studies revealed that ob/ob mice subjected to TAC failed to activate the NFATc4 in the heart, however, intraperitoneal injection of leptin in ob/ob mice restored the NFATc4 DNA-binding activity and induced expression of the ANP gene. Conclusion This study establishes the role of leptin as an anti-hypertrophic agent during pressure overload hypertrophy, and suggests that a key molecular event is the leptin mediated activation of NFATc4 that regulates the transcriptional activation of the ANP gene promoter. PMID:19560554

  3. Diosgenin, a steroidal sapogenin, enhances antigen-specific IgG2a and interferon-gamma expression in ovalbumin-sensitized BALB/c mice.

    PubMed

    Jan, Tong-Rong; Wey, Shiaw-Pyng; Kuan, Chio-Chin; Liao, Mei-Hsiu; Wu, Hsin-Yin

    2007-05-01

    The effect of diosgenin, the most abundant sapogenin in Chinese yam, on humoral immunity was investigated. Ovalbumin (OVA)-sensitized and challenged BALB/c mice were administered daily with diosgenin for 34 days. The production of OVA-specific serum IgG2a was significantly enhanced by diosgenin treatment, whereas total IgE and OVA-specific IgG1, IgG2a and IgM were unaffected. In parallel with the enhancement of IgG2a, OVA-induced IFN-gamma secretion and mRNA expression were markedly elevated in splenocytes of diosgenin-treated mice, whereas IL-4 expression was unaltered. Furthermore, the expression of T-bet, but not of GATA-3, in splenocytes was up-regulated by diosgenin administration. However, diosgenin treatment did not modulate IL-4 mRNA expression and inflammatory cell infiltration in the lung of OVA-sensitized and challenged mice. Collectively, these data suggest that diosgenin regulates the systemic immune response towards the Th1 direction in response to OVA sensitization. The present study provides evidence to show that intake of diosgenin modulates certain aspects of acquired immunity, including the enhancement of antigen-specific IgG2a and IFN-gamma expression, which may be mediated through the up-regulation of Th1 differentiation. PMID:17566144

  4. Folic acid deficiency enhances abeta accumulation in APP/PS1 mice brain and decreases amyloid-associated miRNAs expression.

    PubMed

    Liu, Huan; Tian, Tian; Qin, Shanchun; Li, Wen; Zhang, Xumei; Wang, Xuan; Gao, Yuxia; Huang, Guowei

    2015-12-01

    Recent efforts have revealed the microRNA (miRNA) pathways in the pathogenesis of Alzheimer's disease (AD). Epidemiological studies have revealed an association between folic acid deficiency and AD risk. However, the effects of folic acid deficiency on miRNA expression in AD animals have not been observed. We aimed to find if folic acid deficiency may enhance amyloid-β (Aβ) peptide deposition and regulate amyloid-associated miRNAs and their target genes expression in APP/PS1 mice. APP/PS1 mice and N2a cells were treated with folic acid-deficient diet or medium. Cognitive function of mice was assessed using the Morris water maze. miRNA profile was tested by polymerase chain reaction (PCR) array. Different expressional miRNAs were validated by real-time PCR. The deposition of Aβ plaques was evaluated by immunohistochemistry and enzyme-linked immunosorbent assay. APP and BACE1 proteins in mice brain and N2a cells were determined by Western blot. Folic acid deficiency aggravated amyloid pathology in AD mice. The AD+FD group showed shorter time spent in the target zone during the probe test. Analysis of miRNAs predicted to target these genes revealed several miRNA candidates that were differentially modulated by folic acid deficiency. In APP/PS1 mice brains and N2a cells with folic acid-deficient treatment, miR-106a-5p, miR-200b-3p and miR-339-5p were down-regulated, and their target genes APP and BACE1 were up-regulated. In conclusion, folic acid deficiency can enhance Aβ accumulation in APP/PS1 mice brain and decrease amyloid-associated miRNAs expression. PMID:26345540

  5. Mycobacterium abscessus phospholipase C expression is induced during coculture within amoebae and enhances M. abscessus virulence in mice.

    PubMed

    Bakala N'Goma, Jean Claude; Le Moigne, Vincent; Soismier, Nathalie; Laencina, Laura; Le Chevalier, Fabien; Roux, Anne-Laure; Poncin, Isabelle; Serveau-Avesque, Carole; Rottman, Martin; Gaillard, Jean-Louis; Etienne, Gilles; Brosch, Roland; Herrmann, Jean-Louis; Canaan, Stéphane; Girard-Misguich, Fabienne

    2015-02-01

    Mycobacterium abscessus is a pathogenic, rapidly growing mycobacterium involved in pulmonary and cutaneo-mucous infections worldwide, to which cystic fibrosis patients are exquisitely susceptible. The analysis of the genome sequence of M. abscessus showed that this bacterium is endowed with the metabolic pathways typically found in environmental microorganisms that come into contact with soil, plants, and aquatic environments, where free-living amoebae are frequently present. M. abscessus also contains several genes that are characteristically found only in pathogenic bacteria. One of them is MAB_0555, encoding a putative phospholipase C (PLC) that is absent from most other rapidly growing mycobacteria, including Mycobacterium chelonae and Mycobacterium smegmatis. Here, we report that purified recombinant M. abscessus PLC is highly cytotoxic to mouse macrophages, presumably due to hydrolysis of membrane phospholipids. We further showed by constructing and using an M. abscessus PLC knockout mutant that loss of PLC activity is deleterious to M. abscessus intracellular survival in amoebae. The importance of PLC is further supported by the fact that M. abscessus PLC was found to be expressed only in amoebae. Aerosol challenge of mice with M. abscessus strains that were precultured in amoebae enhanced M. abscessus lung infectivity relative to M. abscessus grown in broth culture. Our study underlines the importance of PLC for the virulence of M. abscessus. Despite the difficulties of isolating M. abscessus from environmental sources, our findings suggest that M. abscessus has evolved in close contact with environmental protozoa, which supports the argument that amoebae may contribute to the virulence of opportunistic mycobacteria. PMID:25486995

  6. Mycobacterium abscessus Phospholipase C Expression Is Induced during Coculture within Amoebae and Enhances M. abscessus Virulence in Mice

    PubMed Central

    Bakala N'Goma, Jean Claude; Le Moigne, Vincent; Soismier, Nathalie; Laencina, Laura; Le Chevalier, Fabien; Roux, Anne-Laure; Poncin, Isabelle; Serveau-Avesque, Carole; Rottman, Martin; Gaillard, Jean-Louis; Etienne, Gilles; Brosch, Roland; Canaan, Stéphane

    2014-01-01

    Mycobacterium abscessus is a pathogenic, rapidly growing mycobacterium involved in pulmonary and cutaneo-mucous infections worldwide, to which cystic fibrosis patients are exquisitely susceptible. The analysis of the genome sequence of M. abscessus showed that this bacterium is endowed with the metabolic pathways typically found in environmental microorganisms that come into contact with soil, plants, and aquatic environments, where free-living amoebae are frequently present. M. abscessus also contains several genes that are characteristically found only in pathogenic bacteria. One of them is MAB_0555, encoding a putative phospholipase C (PLC) that is absent from most other rapidly growing mycobacteria, including Mycobacterium chelonae and Mycobacterium smegmatis. Here, we report that purified recombinant M. abscessus PLC is highly cytotoxic to mouse macrophages, presumably due to hydrolysis of membrane phospholipids. We further showed by constructing and using an M. abscessus PLC knockout mutant that loss of PLC activity is deleterious to M. abscessus intracellular survival in amoebae. The importance of PLC is further supported by the fact that M. abscessus PLC was found to be expressed only in amoebae. Aerosol challenge of mice with M. abscessus strains that were precultured in amoebae enhanced M. abscessus lung infectivity relative to M. abscessus grown in broth culture. Our study underlines the importance of PLC for the virulence of M. abscessus. Despite the difficulties of isolating M. abscessus from environmental sources, our findings suggest that M. abscessus has evolved in close contact with environmental protozoa, which supports the argument that amoebae may contribute to the virulence of opportunistic mycobacteria. PMID:25486995

  7. A highly conserved NF-κB-responsive enhancer is critical for thymic expression of Aire in mice.

    PubMed

    Haljasorg, Uku; Bichele, Rudolf; Saare, Mario; Guha, Mithu; Maslovskaja, Julia; Kõnd, Karin; Remm, Anu; Pihlap, Maire; Tomson, Laura; Kisand, Kai; Laan, Martti; Peterson, Pärt

    2015-12-01

    Autoimmune regulator (Aire) has a unique expression pattern in thymic medullary epithelial cells (mTECs), in which it plays a critical role in the activation of tissue-specific antigens. The expression of Aire in mTECs is activated by receptor activator of nuclear factor κB (RANK) signaling; however, the molecular mechanism behind this activation is unknown. Here, we characterize a conserved noncoding sequence 1 (CNS1) containing two NF-κB binding sites upstream of the Aire coding region. We show that CNS1-deficient mice lack thymic expression of Aire and share several features of Aire-knockout mice, including downregulation of Aire-dependent genes, impaired terminal differentiation of the mTEC population, and reduced production of thymic Treg cells. In addition, we show that CNS1 is indispensable for RANK-induced Aire expression and that CNS1 is activated by NF-κB pathway complexes containing RelA. Together, our results indicate that CNS1 is a critical link between RANK signaling, NF-κB activation, and thymic expression of Aire. PMID:26364592

  8. Transgenic cyclooxygenase-2 expression and high salt enhanced susceptibility to chemical-induced gastric cancer development in mice.

    PubMed

    Leung, Wai K; Wu, Kai-chun; Wong, Christine Y P; Cheng, Alfred S L; Ching, Arthur K K; Chan, Anthony W H; Chong, Wilson W S; Go, Minnie Y Y; Yu, Jun; To, Ka-Fai; Wang, Xin; Chui, Y L; Fan, D M; Sung, Joseph J Y

    2008-08-01

    Cyclooxoygenase (COX)-2 overexpression is involved in gastric carcinogenesis. While high-salt intake is a known risk factor for gastric cancer development, we determined the effects of high salt on gastric chemical carcinogenesis in COX-2 transgenic (TG) mice. COX-2 TG mice were developed in C57/BL6 strain using the full-length human cox-2 complementary DNA construct. Six-week-old COX-2 TG and wild-type (WT) littermates were randomly allocated to receive alternate week of N-methyl-N-nitrosourea (MNU, 240 p.p.m.) in drinking water or control for 10 weeks. Two groups of mice were further treated with 10% NaCl during the initial 10 weeks. All mice were killed at the end of week 50. Both forced COX-2 overexpression and high-salt intake significantly increased the frequency of gastric cancer development in mice as compared with WT littermates treated with MNU alone. However, no additive effect was observed on the combination of high salt and COX-2 expression. We further showed that MNU and high-salt treatment increased chronic inflammatory infiltrates and induced prostaglandin E(2) (PGE(2)) production in the non-cancerous stomach. Whereas high-salt treatment markedly increased the expression of inflammatory cytokines (tumor necrosis factor-alpha, interferon-gamma, interleukin (IL)-1 beta and IL-6) in the gastric mucosa, COX-2 overexpression significantly altered the cell kinetics in the MNU-induced gastric cancer model. In conclusion, both high salt and COX-2 overexpression promote chemical-induced gastric carcinogenesis, possibly related to chronic inflammation, induction of PGE(2), disruption of cell kinetics and induction of inflammatory cytokines. PMID:18611916

  9. B-lymphocyte targeting of gene expression in transgenic mice with the immunoglobulin heavy-chain enhancer.

    PubMed Central

    Gerlinger, P; LeMeur, M; Irrmann, C; Renard, P; Wasylyk, C; Wasylyk, B

    1986-01-01

    A hybrid gene containing rabbit beta-globin structural sequences (-9 to +1650), and a chicken conalbumin gene promoter (+62 to -102) in the place of the beta-globin promoter (upstream from -9), was inactive in 5 different transgenic mouse line. Adding the mouse immunoglobulin heavy-chain (IgH) enhancer to this construction specifically stimulated expression in B-cells. These results show that IgH enhancer is specifically active in B-cells. Expression of the hybrid gene was low compared to the endogenous immunoglobulin heavy and light-chain genes. Substituting the mouse immunoglobulin kappa light-chain gene (Ig kappa) promoter (+4 to -800) for the heterologous conalbumin promoter was not sufficient to restore gene expression to level of the endogenous genes. In addition to the reproducible B cell expression, we also found inheritable unexpected expression in certain tissues, which varied from line to line. Images PMID:3092186

  10. A cell-specific enhancer far upstream of the mouse tyrosinase gene confers high level and copy number-related expression in transgenic mice.

    PubMed Central

    Ganss, R; Montoliu, L; Monaghan, A P; Schütz, G

    1994-01-01

    The tyrosinase gene encodes the key enzyme of melanin production and is tightly regulated during development. A yeast artificial chromosome covering the mouse tyrosinase gene has been shown to rescue completely the albino phenotype of recipient mouse strains, conferring copy number-dependent, position-independent expression. To investigate the presence of cis-acting regulatory elements responsible for the appropriate expression of the tyrosinase gene, DNase I hypersensitive site mapping was performed. A melanoma cell-specific DNase I hypersensitive site was identified at -12 kb upstream of the tyrosinase gene. Functional analysis of the corresponding cis-acting element in transgenic mice and transient transfection assays revealed properties of a strong cell-specific enhancer. RNA expression levels of the transgene correlate with copy number, which is reflected in coat colour and eye pigmentation of transgenic mice. Full enhancer activity in transient transfections is obtained with a minimal sequence of 200 bp. Protein binding analysis reveals the presence of a melanoma cell-specific complex which might contribute to the faithful expression of the tyrosinase gene. Images PMID:8039502

  11. Vermectomy enhances parvalbumin expression and improves motor performance in weaver mutant mice: an animal model for cerebellar ataxia.

    PubMed

    Grüsser-Cornehls, U; Grüsser, C; Bäurle, J

    1999-01-01

    In the Weaver mutant mouse (wv/wv), an animal model for hereditary cerebellar ataxia, electrophysiological experiments have revealed a disorganized output of cerebellar Purkinje cells (the latter using GABA as an inhibitory transmitter) which, by a cascade of mechanisms, was thought to be the cause of the poor motor abilities. In Purkinje cell degeneration mice (pcd/pcd) lacking nearly all Purkinje cells and displaying milder motor deficiencies than wv, in comparison to wild-type mice, a strong increase in parvalbumin- and (co-localized with parvalbumin) glycine-immunopositive somata in the deep cerebellar and vestibular nuclei has recently been found. It was therefore intriguing to investigate whether motor performance in weaver mutants could be ameliorated by applying cerebellar lesions to eliminate the faulty output and to look for a change in transmitter weighting, indicated by a strong increase in parvalbumin-positive somata in areas (the respective target areas) which were formerly devoid of it. Ten Weaver mutants were subjected to cerebellar lesions. After removal of the vermis a total abolition of tremor, a definite improvement in the balance of affected body parts, an increase in locomotor activity when tested in an open-field matrix, and a strong increase in parvalbumin expression in Weaver mutant deep cerebellar and vestibular nuclei in comparison to wild-types have indeed been found. Increase in motor activity (or explorative behaviour) has been placed in relation to learning mechanisms. The increase in parvalbumin expression and the observed improvement in motor abilities and mechanisms probably related to learning underline the hypothesis that any change in the physiological equilibrium of the brain function by removal of input or output related to an assembly of nerve cells leads to a cascade of changes at the transmitter and neuronal level in near or distant connected brain structures. PMID:10336081

  12. Spherical nucleic acid targeting microRNA-99b enhances intestinal MFG-E8 gene expression and restores enterocyte migration in lipopolysaccharide-induced septic mice

    PubMed Central

    Wang, Xiao; Hao, Liangliang; Bu, Heng-Fu; Scott, Alexander W.; Tian, Ke; Liu, Fangyi; De Plaen, Isabelle G.; Liu, Yulan; Mirkin, Chad A.; Tan, Xiao-Di

    2016-01-01

    Milk fat globule-EGF factor 8 (MFG-E8) maintains the intestinal homeostasis by enhancing enterocyte migration and attenuating inflammation. We previously reported that sepsis is associated with down-regulation of intestinal MFG-E8 and impairment of enterocyte migration. Here, we showed that impairment of intestinal epithelial cell migration occurred in lipopolysaccharide (LPS)-induced septic mice. Treatment of RAW264.7 cells (a murine macrophage-like cell line) with LPS increased expression of miR-99b, a microRNA that is predicted to target mouse MFG-E8 3′UTR. Using a luciferase assay, we showed that miR-99b mimic suppressed the activity of a reporter containing MFG-E8 3′UTR. This suggests the role of miR-99b in inhibition of MFG-E8 gene expression. In addition, we developed an anti-miR99b spherical nucleic acid nanoparticle conjugate (SNA-NCanti-miR99b). Treatment of both naïve and LPS-challenged cells with SNA-NCanti-miR99b enhanced MFG-E8 expression in the cells. Administration of SNA-NCanti-miR99b rescued intestinal MFG-E8 expression in LPS-induced septic mice and attenuated LPS inhibitory effects on intestinal epithelial cell migration along the crypt-villus axis. Collectively, our study suggests that LPS represses MFG-E8 expression and disrupts enterocyte migration via a miR-99b dependent mechanism. Furthermore, this work shows that SNA-NCanti-miR99b is a novel nanoparticle-conjugate capable of rescuing MFG-E8 gene expression and maintaining intestinal epithelial homeostasis in sepsis. PMID:27538453

  13. Spherical nucleic acid targeting microRNA-99b enhances intestinal MFG-E8 gene expression and restores enterocyte migration in lipopolysaccharide-induced septic mice.

    PubMed

    Wang, Xiao; Hao, Liangliang; Bu, Heng-Fu; Scott, Alexander W; Tian, Ke; Liu, Fangyi; De Plaen, Isabelle G; Liu, Yulan; Mirkin, Chad A; Tan, Xiao-Di

    2016-01-01

    Milk fat globule-EGF factor 8 (MFG-E8) maintains the intestinal homeostasis by enhancing enterocyte migration and attenuating inflammation. We previously reported that sepsis is associated with down-regulation of intestinal MFG-E8 and impairment of enterocyte migration. Here, we showed that impairment of intestinal epithelial cell migration occurred in lipopolysaccharide (LPS)-induced septic mice. Treatment of RAW264.7 cells (a murine macrophage-like cell line) with LPS increased expression of miR-99b, a microRNA that is predicted to target mouse MFG-E8 3'UTR. Using a luciferase assay, we showed that miR-99b mimic suppressed the activity of a reporter containing MFG-E8 3'UTR. This suggests the role of miR-99b in inhibition of MFG-E8 gene expression. In addition, we developed an anti-miR99b spherical nucleic acid nanoparticle conjugate (SNA-NC(anti-miR99b)). Treatment of both naïve and LPS-challenged cells with SNA-NC(anti-miR99b) enhanced MFG-E8 expression in the cells. Administration of SNA-NC(anti-miR99b) rescued intestinal MFG-E8 expression in LPS-induced septic mice and attenuated LPS inhibitory effects on intestinal epithelial cell migration along the crypt-villus axis. Collectively, our study suggests that LPS represses MFG-E8 expression and disrupts enterocyte migration via a miR-99b dependent mechanism. Furthermore, this work shows that SNA-NC(anti-miR99b) is a novel nanoparticle-conjugate capable of rescuing MFG-E8 gene expression and maintaining intestinal epithelial homeostasis in sepsis. PMID:27538453

  14. Enhanced FGF23 production in mice expressing PI3K-insensitive GSK3 is normalized by β-blocker treatment.

    PubMed

    Fajol, Abul; Chen, Hong; Umbach, Anja T; Quarles, L Darryl; Lang, Florian; Föller, Michael

    2016-02-01

    Glycogen synthase kinase (GSK)-3 is a ubiquitously expressed kinase inhibited by insulin-dependent Akt/PKB/SGK. Mice expressing Akt/PKB/SGK-resistant GSK3α/GSK3β (gsk3(KI)) exhibit enhanced sympathetic nervous activity and phosphaturia with decreased bone density. Hormones participating in phosphate homeostasis include fibroblast growth factor (FGF)-23, a bone-derived hormone that inhibits 1,25-dihydroxyvitamin D3 (1,25(OH)2D3; calcitriol) formation and phosphate reabsorption in the kidney and counteracts vascular calcification and aging. FGF23 secretion is stimulated by the sympathetic nervous system. We studied the role of GSK3-controlled sympathetic activity in FGF23 production and phosphate metabolism. Serum FGF23, 1,25(OH)2D3, and urinary vanillylmandelic acid (VMA) were measured by ELISA, and serum and urinary phosphate and calcium were measured by photometry in gsk3(KI) and gsk3(WT) mice, before and after 1 wk of oral treatment with the β-blocker propranolol. Urinary VMA excretion, serum FGF23, and renal phosphate and calcium excretion were significantly higher, and serum 1,25(OH)2D3 and phosphate concentrations were lower in gsk3(KI) mice than in gsk3(WT) mice. Propranolol treatment decreased serum FGF23 and loss of renal calcium and phosphate and increased serum phosphate concentration in gsk3(KI) mice. We conclude that Akt/PKB/SGK-sensitive GSK3 inhibition participates in the regulation of FGF23 release, 1,25(OH)2D3 formation, and thus mineral metabolism, by controlling the activity of the sympathetic nervous system. PMID:26527066

  15. Enhanced neuronal nitric oxide synthase expression is central to cardiac vagal phenotype in exercise-trained mice

    PubMed Central

    Danson, E J F; Paterson, D J

    2003-01-01

    We investigated whether enhanced cardiac vagal responsiveness elicited by exercise training is dependent on neuronal nitric oxide synthase (NOS-1), since the NO-cGMP pathway facilitates acetylcholine release. Isolated atria with intact right vagal innervation were taken from male mice (18-22 weeks old) after a period of 10 weeks voluntary wheel-running (+EX, n = 27; peaked 9.8 ± 0.6 km day−1 at 5 weeks), and from mice housed in cages without wheels (-EX, n = 27). Immunostaining of whole atria for NOS-1 identified intrinsic neurones, all of which co-localized with choline acetyltransferase-positive ganglia. Western blot analysis confirmed that NOS-1 protein level was significantly greater in +EX compared to -EX atria (P < 0.05, unpaired t test). Basal heart rates (HR) were slower in +EX than in -EX atria (322 ± 6 versus 360 ± 7 beats min−1; P < 0.05, unpaired t test) However, in +EX atria, HR responses to vagal stimulation (VNS, 3 and 5 Hz) were significantly enhanced compared to -EX atria (3 Hz, +EX: −76 ± 8 beats min−1versus -EX: −62 ± 7 beats min−1; 5 Hz, +EX: −106 ± 4 beats min−1versus -EX: −93 ± 3 beats min−1; P < 0.01, unpaired t test). Inhibition of NOS-1 with vinyl-l-N-5-(1-imino-3-butenyl)-l-ornithine (l-VNIO, 100 μm) or soluble guanylyl cyclase with 1H-[1, 2, 4]oxadiazolo[4, 3-a]quinoxalin-1-one (ODQ, 10 μm) abolished the difference in HR responses to VNS between +EX and -EX atria, and effects of l-VNIO were reversed by excess l-arginine (1 mm; P < 0.01, ANOVA). There were no differences between the HR responses to the bath-applied acetylcholine analogue carbamylcholine chloride in +EX and -EX atria (IC50 concentrations were 5.9 ± 0.4 μm (-EX) and 5.7 ± 0.4 μm (+EX)), suggesting that the changes in vagal responsiveness resulted from presynaptic facilitation of neurotransmission. In conclusion, NOS-1 appears to be a key protein in generating the cardiac vagal gain of function elicited by exercise training. PMID:12509490

  16. In vivo expression of the Arf6 Guanine-nucleotide exchange factor cytohesin-1 in mice exhibits enhanced myelin thickness in nerves.

    PubMed

    Torii, Tomohiro; Miyamoto, Yuki; Onami, Naoko; Tsumura, Hideki; Nemoto, Noriko; Kawahara, Katsumasa; Kato, Minoru; Kotera, Jun; Nakamura, Kazuaki; Tanoue, Akito; Yamauchi, Junji

    2013-10-01

    The myelin sheath consists of a unique multiple layer structure that acts as an insulator between neuronal axons to enhance the propagation of the action potential. In neuropathies such as demyelinating or dismyelinating diseases, chronic demyelination and defective remyelination occur repeatedly, leading to more severe neuropathy. As yet, little is known about the possibility of drug target-specific medicine for such diseases. In the developing peripheral nervous system (PNS), myelin sheaths form as Schwann cells wrap individual axons. It is thought that the development of a drug promoting myelination by Schwann cells would provide effective therapy against peripheral nerve disorders: to test such treatment, genetically modified mice overexpressing the drug target molecules are needed. We previously identified an Arf6 activator, the guanine-nucleotide exchange factor cytohesin-1, as the signaling molecule controlling myelination of peripheral axons by Schwann cells; yet, the important issue of whether cytohesin-1 itself promotes myelin thickness in vivo has remained unclear. Herein, we show that, in mouse PNS nerves, Schwann cell-specific expression of wild-type cytohesin-1 exhibits enhanced myelin thickness. Downstream activation of Arf6 is also seen in these transgenic mice, revealing the involvement of the cytohesin-1 and Arf6 signaling unit in promoting myelination. These results suggest that cytohesin-1 may be a candidate for the basis of a therapy for peripheral neuropathies through its enhancement of myelin thickness. PMID:23636892

  17. A novel recombinant BCG-expressing pro-apoptotic protein BAX enhances Th1 protective immune responses in mice.

    PubMed

    Li, Guanghua; Liu, Guoyuan; Song, Na; Kong, Cong; Huang, Qi; Su, Haibo; Bi, Aixiao; Luo, Liulin; Zhu, Lin; Xu, Ying; Wang, Honghai

    2015-08-01

    One-third of the world's population is infected with Mycobacterium tuberculosis (MTB). The protective efficacy of bacille Calmette Guérin (BCG) vaccine against tuberculosis (TB) in adults is highly controversial even though the BCG vaccine has been available for more than 90 years. Because BCG is effective against infantile tuberculosis meningitis and miliary tuberculosis in young children and provides cost-effective prevention from tuberculosis for developing countries, it would be desirable to modify the existing BCG vaccine to provide more comprehensive protection. In our study, we constructed a novel recombinant BCG strain expressing pro-apoptotic BAX (rBCG::BAX) and demonstrated that it significantly induced the apoptosis of macrophages infected with rBCG::BAX both in vitro and in vivo. In addition, it significantly enhanced Ag85B-specific IFN-γ enzyme-linked immunospot responses, IFN-γ secretion, IL-2 secretion and the ratio of Ag85B-specific IgG2b/IgG1, and it significantly decreased Ag85B-specific IL-4. Furthermore, it presumably facilitated antigen presentation by inducing a significant up-regulation in the expression of MHC-II and B7.1 (CD80) co-stimulatory molecules on macrophages. In conclusion, these results suggest that the rBCG::BAX strain elicited predominantly a Th1 protective immune responses and might be a potential tuberculosis vaccine candidate for further study. PMID:25942359

  18. Purple Tea and Its Extract Suppress Diet-induced Fat Accumulation in Mice and Human Subjects by Inhibiting Fat Absorption and Enhancing Hepatic Carnitine Palmitoyltransferase Expression.

    PubMed

    Shimoda, Hiroshi; Hitoe, Shoketsu; Nakamura, Seikou; Matsuda, Hisashi

    2015-06-01

    A number of clinical trials have been completed using green tea and black tea to investigate their effect in controlling weight in overweight adults. The results of these investigations, however, have often been contradictory, with some trials reporting positive effects of tea supplementation and some trials reporting no effect. As a result, the use of these teas for weight loss is controversial. Purple tea is a variety of green tea developed in Kenya (called TRFK306), which in addition to certain tea constituents found in green tea, also contains anthocyanins. The major constituents in the leaves of purple tea are caffeine, theobromine, epigallocatechin (ECG), epigallocatechin gallate (EGCG) and 1,2-di-O-galloyl-4,6-O-(S)-hexahydroxydiphenoyl-β-D-glucose (GHG). We investigated the efficacy of purple tea extract (PTE) on diet-induced fat accumulation in mice. PTE administration (200 mg/kg) significantly suppressed body weight gain, liver weight, abdominal fat and triglycerides in serum and liver. Protein expression of carnitine palmitoyltransferase (CPT) 1A was also enhanced. In olive oil loaded mice, PTE (100 mg/kg) and caffeine (25 mg/kg) suppressed fat absorption. PTE (10 μg/mL) and GHG (10 μg/mL) also enhanced protein expression of CPT1A in HepG2 hepatoma. Moreover, 4-week daily consumption of purple tea drink in humans improved obesity parameters compared to baseline, including body weight (79.9 ± 3.1 kg vs 80.8 ± 3.2, p<0.05), body mass index (BMI) (26.8 ± 0.6 vs 27.0 ± 0.6, p<0.05) and body fat mass (21.0 ± 1.4 kg vs 21.8 ± 1.5, p<0.01). In conclusion, PTE could control diet-induced weight gain by suppression of fat absorption and enhancement of hepatic fat metabolism. PMID:26199579

  19. Purple Tea and Its Extract Suppress Diet-induced Fat Accumulation in Mice and Human Subjects by Inhibiting Fat Absorption and Enhancing Hepatic Carnitine Palmitoyltransferase Expression

    PubMed Central

    Shimoda, Hiroshi; Hitoe, Shoketsu; Nakamura, Seikou; Matsuda, Hisashi

    2015-01-01

    A number of clinical trials have been completed using green tea and black tea to investigate their effect in controlling weight in overweight adults. The results of these investigations, however, have often been contradictory, with some trials reporting positive effects of tea supplementation and some trials reporting no effect. As a result, the use of these teas for weight loss is controversial. Purple tea is a variety of green tea developed in Kenya (called TRFK306), which in addition to certain tea constituents found in green tea, also contains anthocyanins. The major constituents in the leaves of purple tea are caffeine, theobromine, epigallocatechin (ECG), epigallocatechin gallate (EGCG) and 1,2-di-O-galloyl-4,6-O-(S)-hexahydroxydiphenoyl-β-D-glucose (GHG). We investigated the efficacy of purple tea extract (PTE) on diet-induced fat accumulation in mice. PTE administration (200 mg/kg) significantly suppressed body weight gain, liver weight, abdominal fat and triglycerides in serum and liver. Protein expression of carnitine palmitoyltransferase (CPT) 1A was also enhanced. In olive oil loaded mice, PTE (100 mg/kg) and caffeine (25 mg/kg) suppressed fat absorption. PTE (10 μg/mL) and GHG (10 μg/mL) also enhanced protein expression of CPT1A in HepG2 hepatoma. Moreover, 4-week daily consumption of purple tea drink in humans improved obesity parameters compared to baseline, including body weight (79.9 ± 3.1 kg vs 80.8 ± 3.2, p<0.05), body mass index (BMI) (26.8 ± 0.6 vs 27.0 ± 0.6, p<0.05) and body fat mass (21.0 ± 1.4 kg vs 21.8 ± 1.5, p<0.01). In conclusion, PTE could control diet-induced weight gain by suppression of fat absorption and enhancement of hepatic fat metabolism PMID:26199579

  20. Enhanced pan-peroxisome proliferator-activated receptor gene and protein expression in adipose tissue of diet-induced obese mice treated with telmisartan.

    PubMed

    Penna-de-Carvalho, Aline; Graus-Nunes, Francielle; Rabelo-Andrade, Júlia; Mandarim-de-Lacerda, Carlos Alberto; Souza-Mello, Vanessa

    2014-12-01

    Telmisartan has previously been used to target obesity, showing peroxisome proliferator-activated receptor (PPAR) β/δ-related effects in white adipose tissue (WAT). We sought to evaluate whether telmisartan enhances gene and protein expression of all PPAR isoforms in WAT and brown adipose tissue (BAT), as well as their downstream effects upon insulin resistance, adipokine profile and adaptive thermogenesis. Male C57BL/6 mice were fed standard chow (SC; 10% lipids) or high-fat diet (HF; 50% lipids) for 10 weeks. Animals were then randomly allocated into the following four groups: SC, SC-T, HF and HF-T. Telmisartan [10 mg (kg diet)(-1)] was administered for 4 weeks in the diet. Animals in the HF group were overweight and exhibited hypertension, insulin resistance, decreased energy expenditure, a pro-inflammatory adipokine profile and abnormal fat pad mass distribution. Animals in the HF group showed decreased expression of PPARα, β/δ and γ in WAT and BAT, resulting in impaired glucose uptake and insufficient thermogenesis. Due to the improvement in the adipokine profile and enhanced insulin sensitivity with adequate insulin-stimulated glucose uptake after treatment with telmisartan, the activation of all PPAR isoforms in WAT was beneficial. In BAT, telmisartan induced sustained sympathetic activation, because the β3-adrenergic receptor was induced by PPARβ/δ, while uncoupling protein 1 was induced by PPARα to promote thermogenesis. Telmisartan exerted anti-obesity effects through higher pan-PPAR gene and protein expression. Upon PPARα, β/δ and γ (pan-PPAR) agonism in adipose tissue of obese mice, telmisartan ameliorates inflammation and insulin resistance, as well as inducing non-shivering thermogenesis. Our results point to new therapeutic targets for the control of obesity and comorbidities through pan-PPAR-related effects. PMID:25326526

  1. CREB-regulated transcription coactivator 1 enhances CREB-dependent gene expression in spinal cord to maintain the bone cancer pain in mice

    PubMed Central

    Liang, Ying; Liu, Yue; Hou, Bailing; Zhang, Wei; Liu, Ming; Sun, Yu-E; Gu, Xiaoping

    2016-01-01

    Background cAMP response element binding protein (CREB)-dependent gene expression plays an important role in central sensitization. CREB-regulated transcription coactivator 1 (CRTC1) dramatically increases CREB-mediated transcriptional activity. Brain-derived neurotrophic factor, N-methyl-d-aspartate receptor subunit 2B, and miRNA-212/132, which are highly CREB responsive, function downstream from CREB/CRTC1 to mediate activity-dependent synaptic plasticity and in turn loops back to amplify CREB/CRTC1 signaling. This study aimed to investigate the role of spinal CRTC1 in the maintenance of bone cancer pain using an RNA interference method. Results Osteosarcoma cells were implanted into the intramedullary space of the right femurs of C3H/HeNCrlVr mice to induce bone cancer pain. Western blotting was applied to examine the expression of spinal phospho-Ser133 CREB and CRTC1. We further investigated effects of repeated intrathecal administration with Adenoviruses expressing CRTC1-small interfering RNA (siRNA) on nociceptive behaviors and on the upregulation of CREB/CRTC1-target genes associated with bone cancer pain. Inoculation of osteosarcoma cells induced progressive mechanical allodynia and spontaneous pain, and resulted in upregulation of spinal p-CREB and CRTC1. Repeated intrathecal administration with Adenoviruses expressing CRTC1-siRNA attenuated bone cancer–evoked pain behaviors, and reduced CREB/CRTC1-target genes expression in spinal cord, including BDNF, NR2B, and miR-212/132. Conclusions Upregulation of CRTC1 enhancing CREB-dependent gene transcription in spinal cord may play an important role in bone cancer pain. Inhibition of spinal CRTC1 expression reduced bone cancer pain. Interruption to the positive feedback circuit between CREB/CRTC1 and its targets may contribute to the analgesic effects. These findings may provide further insight into the mechanisms and treatment of bone cancer pain. PMID:27060162

  2. Enhancement of fibroblast activation protein α-based vaccines and adenovirus boost immunity by cyclophosphamide through inhibiting IL-10 expression in 4T1 tumor bearing mice.

    PubMed

    Xia, Qiu; Geng, Fei; Zhang, Fang-Fang; Liu, Chen-Lu; Xu, Ping; Lu, Zhen-Zhen; Zhang, Hai-Hong; Kong, Wei; Yu, Xiang-Hui

    2016-08-31

    Fibroblast activation protein α (FAPα) is expressed in cancer-associated fibroblasts (CAFs) of more than 90% of malignant epithelia carcinomas. CAFs are the main type of cells in the tumor microenvironment which offer nutrition and protection to the tumor and regulate immunosuppression. To eliminate CAFs, a vaccine targeting FAPα may be used with a heterologous prime-boost strategy to enhance the FAPα-specific cellular immunity. Here, a FAP vaccine using a recombinant adenovirus (rAd) vector was constructed as well as a DNA vaccine reported in our previous work. Although the DNA prime-rAd boost strategy enhanced FAPα-specific immune responses, improvement of anti-tumor immunity effects was not observed. Examination of immunosuppressive factors revealed that high expression of the IL-10 cytokine was considered the main cause of the failure of the prime-boost strategy. However, heterologous vaccination in combination with a low-dose of cyclophosphamide (CY), which was reported to reduce IL-10 production and promote a shift from immunosuppression to immunopotentiation, resulted in enhanced effects in terms of numbers of effector T cells and tumor growth inhibition rates, compared to the CY alone or DNA alone group. Tumor growth was inhibited markedly when the prime-boost strategy was combined with CY in both the prophylactic and therapeutic settings and the survival time of 4T1 tumor bearing mice was also prolonged significantly. With the reduction of IL-10, enhancement of the anti-tumor effect by the prime-boost strategy was observed. These results suggest that FAPα-targeted rAd boosting in combination with CY is an attractive approach to overcoming immunosuppression in cancer vaccines. PMID:27498213

  3. A pseudotype baculovirus expressing the capsid protein of foot-and-mouth disease virus and a T-Cell immunogen shows enhanced immunogenicity in mice

    PubMed Central

    2011-01-01

    Background Foot-and-mouth disease (FMD) is a highly contagious disease of livestock which causes severe economic loss in cloven-hoofed animals. Vaccination is still a major strategy in developing countries to control FMD. Currently, inactivated vaccine of FMDV has been used in many countries with limited success and safety concerns. Development of a novel effective vaccine is must. Methods In the present study, two recombinant pseudotype baculoviruses, one expressing the capsid of foot-and-mouth disease virus (FMDV) under the control of a cytomegalovirus immediate early enhancer/promoter (CMV-IE), and the other the caspid plus a T-cell immunogen coding region under a CAG promoter were constructed, and their expression was characterized in mammalian cells. In addition, their immunogenicity in a mouse model was investigated. The humoral and cell-mediated immune responses induced by pseudotype baculovirus were compared with those of inactivated vaccine. Results Indirect immunofluorescence assay (IFA) and indirect sandwich-ELISA (IS-ELISA) showed both recombinant baculoviruses (with or without T-cell epitopes) were transduced efficiently and expressed target proteins in BHK-21 cells. In mice, intramuscular inoculation of recombinants with 1 × 109 or 1 × 1010 PFU/mouse induced the production of FMDV-specific neutralizing antibodies and gamma interferon (IFN-γ). Furthermore, recombinant baculovirus with T-cell epitopes had better immunogenicity than the recombinant without T-cell epitopes as demonstrated by significantly enhanced IFN-γ production (P < 0.01) and higher neutralizing antibody titer (P < 0.05). Although the inactivated vaccine produced the highest titer of neutralizing antibodies, a lower IFN-γ expression was observed compared to the two recombinant pseudotype baculoviruses. Conclusions These results indicate that pseudotype baculovirus-mediated gene delivery could be a alternative strategy to develop a new generation of vaccines against FMDV infection

  4. Infantile Postnatal Exposure to Lead (Pb) Enhances Tau Expression in the Cerebral Cortex of Aged Mice: Relevance to AD

    PubMed Central

    Bihaqi, Syed Waseem; Bahmani, Azadeh; Adem, Abdu; Zawia, Nasser H.

    2014-01-01

    The sporadic nature in over 90% of Alzheimer’s disease (AD) cases, the differential susceptibility and course of illness, and latent onset of the disease suggest involvement of an environmental component in the etiology of late onset AD (LOAD). Recent reports from our lab have demonstrated that molecular alterations favor abundant tau phosphorylation and immunoreactivity in the frontal cortex of aged primates with infantile lead (Pb) exposure (Bihaqi and Zawia, 2013). Here we report that developmental Pb exposure results in elevation of protein and mRNA levels of tau in aged mice. Western blot analysis revealed aberrant site-specific tau hyperphosphorylation accompanied by elevated cyclin dependent kinase 5 (CDK5) levels in aged mice with prior Pb exposure. Mice with developmental Pb exposure also displayed altered protein ratio of p35/p25 with more Serine/Threonine phosphatase activity at old age. These changes favored increase in tau phosphorylation, thus providing evidence that neurodegenerative diseases may be in part due to environmental influences that occur during development. PMID:24954411

  5. LONG-TERM INSULIN-LIKE GROWTH FACTOR-I EXPRESSION IN SKELETAL MUSCLES ATTENUATES THE ENHANCED IN VITRO PROLIFERATION ABILITY OF THE RESIDENT SATELLITE CELLS IN TRANSGENIC MICE

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Insulin-like growth factor-I (IGF-I) overexpression for 1-month in mouse skeletal muscle increases satellite cell proliferation potential. However, it is unknown whether this beneficial enhancement by IGF-I expression would persist over a longer-term duration in aged mice. This is an important issue...

  6. Exaggerated IL-15 and Altered Expression of foxp3+ Cell-Derived Cytokines Contribute to Enhanced Colitis in Nlrp3-/- Mice.

    PubMed

    Hirota, Simon A; Ueno, Aito; Tulk, Sarah E; Becker, Helen M; Schenck, L Patrick; Potentier, Mireille S; Li, Yan; Ghosh, Subrata; Muruve, Daniel A; MacDonald, Justin A; Beck, Paul L

    2016-01-01

    The pathogenesis of Crohn's disease (CD) involves defects in the innate immune system, impairing responses to microbes. Studies have revealed that mutations NLRP3 are associated with CD. We reported previously that Nlrp3-/- mice were more susceptible to colitis and exhibited reduced colonic IL-10 expression. In the current study, we sought to determine how the loss of NLRP3 might be altering the function of regulatory T cells, a major source of IL-10. Colitis was induced in wild-type (WT) and Nlrp3-/- mice by treatment with dextran sulphate sodium (DSS). Lamina propria (LP) cells were assessed by flow cytometry and cytokine expression was assessed. DSS-treated Nlrp3-/- mice exhibited increased numbers of colonic foxp3+ T cells that expressed significantly lower levels of IL-10 but increased IL-17. This was associated with increased expression of colonic IL-15 and increased surface expression of IL-15 on LP dendritic cells. Neutralizing IL-15 in Nlrp3-/- mice attenuated the severity of colitis, decreased the number of colonic foxp3+ cells, and reduced the colonic expression of IL-12p40 and IL-17. These data suggest that the NLRP3 inflammasome can regulate intestinal inflammation through noncanonical mechanisms, providing additional insight as to how NLRP3 variants may contribute to the pathogenesis of CD. PMID:27610005

  7. Exaggerated IL-15 and Altered Expression of foxp3+ Cell-Derived Cytokines Contribute to Enhanced Colitis in Nlrp3−/− Mice

    PubMed Central

    Tulk, Sarah E.; Becker, Helen M.; Potentier, Mireille S.; Li, Yan; Ghosh, Subrata; MacDonald, Justin A.; Beck, Paul L.

    2016-01-01

    The pathogenesis of Crohn's disease (CD) involves defects in the innate immune system, impairing responses to microbes. Studies have revealed that mutations NLRP3 are associated with CD. We reported previously that Nlrp3−/− mice were more susceptible to colitis and exhibited reduced colonic IL-10 expression. In the current study, we sought to determine how the loss of NLRP3 might be altering the function of regulatory T cells, a major source of IL-10. Colitis was induced in wild-type (WT) and Nlrp3−/− mice by treatment with dextran sulphate sodium (DSS). Lamina propria (LP) cells were assessed by flow cytometry and cytokine expression was assessed. DSS-treated Nlrp3−/− mice exhibited increased numbers of colonic foxp3+ T cells that expressed significantly lower levels of IL-10 but increased IL-17. This was associated with increased expression of colonic IL-15 and increased surface expression of IL-15 on LP dendritic cells. Neutralizing IL-15 in Nlrp3−/− mice attenuated the severity of colitis, decreased the number of colonic foxp3+ cells, and reduced the colonic expression of IL-12p40 and IL-17. These data suggest that the NLRP3 inflammasome can regulate intestinal inflammation through noncanonical mechanisms, providing additional insight as to how NLRP3 variants may contribute to the pathogenesis of CD. PMID:27610005

  8. Arctigenin Efficiently Enhanced Sedentary Mice Treadmill Endurance

    PubMed Central

    Chen, Jing; Yu, Liang; Hu, Lihong; Jiang, Hualiang; Shen, Xu

    2011-01-01

    Physical inactivity is considered as one of the potential risk factors for the development of type 2 diabetes and other metabolic diseases, while endurance exercise training could enhance fat oxidation that is associated with insulin sensitivity improvement in obesity. AMP-activated protein kinase (AMPK) as an energy sensor plays pivotal roles in the regulation of energy homeostasis, and its activation could improve glucose uptake, promote mitochondrial biogenesis and increase glycolysis. Recent research has even suggested that AMPK activation contributed to endurance enhancement without exercise. Here we report that the natural product arctigenin from the traditional herb Arctium lappa L. (Compositae) strongly increased AMPK phosphorylation and subsequently up-regulated its downstream pathway in both H9C2 and C2C12 cells. It was discovered that arctigenin phosphorylated AMPK via calmodulin-dependent protein kinase kinase (CaMKK) and serine/threonine kinase 11(LKB1)-dependent pathways. Mice treadmill based in vivo assay further indicated that administration of arctigenin improved efficiently mice endurance as reflected by the increased fatigue time and distance, and potently enhanced mitochondrial biogenesis and fatty acid oxidation (FAO) related genes expression in muscle tissues. Our results thus suggested that arctigenin might be used as a potential lead compound for the discovery of the agents with mimic exercise training effects to treat metabolic diseases. PMID:21887385

  9. Expression of Growth Hormone Genes in Transgenic Mice

    PubMed Central

    Palmiter, Richard D.; Hammer, Robert E.; Brinster, Ralph L.

    2016-01-01

    OVERVIEW Human or rat growth hormone (GH) genes have been introduced into all cells of a mouse by microinjection of fertilized eggs but they were not expressed under their own promoters. However, substitution of a mouse metallothionein (MT) promoter allowed expression and regulation comparable to that of the endogenous MT genes. These fusion genes have been used to stimulate the growth of both normal mice and dwarf mice that lack sufficient GH. Substitution of a rat elastase-I promoter directed expression of GH exclusively to the acinar cells of the pancreas. Progress has been made towards developing the hGH gene into a vector that is not expressed in vivo unless an enhancer element is inserted. Recombination between overlapping DNA fragments derived from a MThGH gene, each of which is nonfunctional, has been observed when they are coinjected into mouse eggs. In some cases, functional hGH was produced as evidenced by enhanced growth of the mice.

  10. Long-term insulin-like growth factor-I expression in skeletal muscles attenuates the enhanced in vitro proliferation ability of the resident satellite cells in transgenic mice

    NASA Technical Reports Server (NTRS)

    Chakravarthy, M. V.; Fiorotto, M. L.; Schwartz, R. J.; Booth, F. W.

    2001-01-01

    Insulin-like growth factor-I (IGF-I) overexpression for 1-month in mouse skeletal muscle increases satellite cell proliferation potential. However, it is unknown whether this beneficial enhancement by IGF-I expression would persist over a longer-term duration in aged mice. This is an important issue to address if a prolonged course of IGF-I is to be used clinically in muscle-wasting conditions where satellite cells may become limiting. Using the IGF-I transgenic (IGF-I Tg) mouse that selectively expresses the IGF-I transgene in striated muscles, we found that 18-months of continuous IGF-I overexpression led to a loss in the enhanced in vitro proliferative capacity of satellite cells from Tg skeletal muscles. Also 18-month-old IGF-I Tg satellite cells lost the enhanced BrdU incorporation, greater pRb and Akt phosphorylations, and decreased p27(Kip1) levels initially observed in cells from 1-month-old IGF-I Tg mice. The levels of those biochemical markers reverted to similar values seen in the 18-months WT littermates. These findings, therefore, suggest that there is no further beneficial effect on enhancing satellite cell proliferation ability with persistent long-term expression of IGF-I in skeletal muscles of these transgenic mice.

  11. Introduction of the human AVPR1A gene substantially alters brain receptor expression patterns and enhances aspects of social behavior in transgenic mice

    PubMed Central

    Charles, Rhonda; Sakurai, Takeshi; Takahashi, Nagahide; Elder, Gregory A.; Gama Sosa, Miguel A.; Young, Larry J.; Buxbaum, Joseph D.

    2014-01-01

    Central arginine vasopressin receptor 1A (AVPR1A) modulates a wide range of behaviors, including stress management and territorial aggression, as well as social bonding and recognition. Inter- and intra-species variations in the expression pattern of AVPR1A in the brain and downstream differential behavioral phenotypes have been attributed to differences in the non-coding regions of the AVPR1A gene, including polymorphic elements within upstream regulatory areas. Gene association studies have suggested a link between AVPR1A polymorphisms and autism, and AVPR1A has emerged as a potential pharmacological target for treatment of social cognitive impairments and mood and anxiety disorders. To further investigate the genetic mechanism giving rise to species differences in AVPR1A expression patterns and associated social behaviors, and to create a preclinical mouse model useful for screening drugs targeting AVPR1A, we engineered and extensively characterized bacterial artificial chromosome (BAC) transgenic mice harboring the entire human AVPR1A locus with the surrounding regulatory elements. Compared with wild-type animals, the humanized mice displayed a more widely distributed ligand-AVPR1A binding pattern, which overlapped with that of primates. Furthermore, humanized AVPR1A mice displayed increased reciprocal social interactions compared with wild-type animals, but no differences in social approach and preference for social novelty were observed. Aspects of learning and memory, specifically novel object recognition and spatial relocation recognition, were unaffected. The biological alterations in humanized AVPR1A mice resulted in the rescue of the prepulse inhibition impairments that were observed in knockout mice, indicating conserved functionality. Although further behavioral paradigms and additional cohorts need to be examined in humanized AVPR1A mice, the results demonstrate that species-specific variations in the genomic content of regulatory regions surrounding

  12. Enhanced Expression of Contractile-Associated Proteins and Ion Channels in Preterm Delivery Model Mice With Chronic Odontogenic Porphyromonas Gingivalis Infection.

    PubMed

    Miyoshi, Hiroshi; Konishi, Haruhisa; Teraoka, Yuko; Urabe, Satoshi; Furusho, Hisako; Miyauchi, Mutsumi; Takata, Takashi; Kudo, Yoshiki

    2016-07-01

    Inflammation and infection have been reported to induce preterm delivery. We have studied the relationship between inflammation and various ion channels, including the L-type Ca(2+) channel and P2X7 receptor, during acute inflammation of the pregnant rat uterus induced by lipopolysaccharides. Recently, we found that mice with odontogenic Porphyromonas gingivalis (P.g, an important odontogenic pathogen) infection delivered at day 18.3 of gestation (vs. day 20.5 in normal mice). The purpose of this study was to investigate the expression of myometrial contractile-associated proteins inducing contractions and confirm that these mice are useful as a model for preterm delivery induced by chronic inflammation. We examined the expression of the oxytocin receptor, connexin 43, prostaglandin F receptors, L-type Ca(2+) channel, and P2X7 receptor in the myometrium at day 18 of gestation by real-time PCR and western blot analyses. We also measured TNF-α and IL-1β levels in the blood serum, placenta, fetal membrane and myometrium on the same day. mRNA expression of the oxytocin receptor, connexin 43, prostaglandin F receptors, L-type Ca(2+) channel, and P2X7 receptor was elevated by 5.4, 3.2, 2.4, 2.5, and 1.7 fold, respectively, in the P.g-infected mice. Protein levels of the oxytocin receptor and connexin 43 also increased. Serum levels of TNF-α and IL-1β were elevated, showing that systemic inflammation continued during pregnancy. IL-1β levels in the placenta and fetal membrane also increased, suggesting inflammatory reactions were induced. Thus, mice with odontogenic infection may be useful as a model of chronic inflammation-induced preterm delivery. PMID:26692542

  13. Expression of key regulators of mitochondrial biogenesis in growth hormone receptor knockout (GHRKO) mice is enhanced but is not further improved by other potential life-extending interventions.

    PubMed

    Gesing, Adam; Masternak, Michal M; Wang, Feiya; Joseph, Anna-Maria; Leeuwenburgh, Christiaan; Westbrook, Reyhan; Lewinski, Andrzej; Karbownik-Lewinska, Malgorzata; Bartke, Andrzej

    2011-10-01

    Mitochondrial biogenesis is essential for cell viability. Growth hormone receptor knockout (GHRKO), calorie restriction, and surgical visceral fat removal constitute experimental interventions to delay aging and increase life span. We examined the expression of known regulators of mitochondriogenesis: peroxisome proliferator-activated receptor γ co-activator 1α (PGC-1α), adenosine monophosphate (AMP)-activated protein kinase (AMPK), sirtuin-1 (SIRT-1) and sirtuin-3 (SIRT-3), endothelial nitric oxide synthase (eNOS), nuclear respiratory factor-1, mitochondrial transcription factor A (TFAM), and mitofusin-2 (MFN-2) in the skeletal muscles and hearts of control and calorie-restricted female GHRKO mice and in the kidneys of male GHRKOs after visceral fat removal or sham surgery. Expression of PGC-1α in skeletal muscles, AMPK, SIRT-1, SIRT-3, eNOS, and MFN-2 in the heart and PGC-1α, AMPK, SIRT-3, eNOS, and MFN-2 in kidneys was increased in GHRKO mice but was not affected by calorie restriction or visceral fat removal. GHRKO mice have increased expression of key regulators of mitochondriogenesis, which is not improved further by calorie restriction or visceral fat removal. PMID:21788651

  14. Co-expression of Ubiquitin gene and capsid protein gene enhances the potency of DNA immunization of PCV2 in mice

    PubMed Central

    2011-01-01

    A recombinant plasmid that co-expressed ubiquitin and porcine circovirus type 2 (PCV2) virus capsid protein (Cap), denoted as pc-Ub-Cap, and a plasmid encoding PCV2 virus Cap alone, denoted as pc-Cap, were transfected into 293T cells. Indirect immunofluorescence (IIF) and confocal microscopy were performed to measure the cellular expression of Cap. Three groups of mice were then vaccinated once every three weeks for a total of three doses with pc-Ub-Cap, pc-Cap or the empty vector pCAGGS, followed by challenging all mice intraperitoneally with 0.5 mL 106.5 TCID50/mL PCV2. To characterize the protective immune response against PCV2 infection in mice, assays of antibody titer (including different IgG isotypes), flow cytometric analysis (FCM), lymphocyte proliferation, cytokine production and viremia were evaluated. The results showed that pc-Ub-Cap and pc-Cap were efficiently expressed in 293T cells. However, pc-Ub-Cap-vaccinated animals had a significantly higher level of Cap-specific antibody and induced a stronger Th1 type cellular immune response than did pc-Cap-vaccinated animals, suggesting that ubiquitin conjugation improved both the cellular and humoral immune responses. Additionally, viral replication in blood was lower in the pc-Ub-Cap-vaccinated group than in the pc-Cap and empty vector groups, suggesting that the protective immunity induced by pc-Ub-Cap is superior to that induced by pc-Cap. PMID:21624113

  15. The Sirt1 activator SRT3025 provides atheroprotection in Apoe−/− mice by reducing hepatic Pcsk9 secretion and enhancing Ldlr expression

    PubMed Central

    Miranda, Melroy X.; van Tits, Lambertus J.; Lohmann, Christine; Arsiwala, Tasneem; Winnik, Stephan; Tailleux, Anne; Stein, Sokrates; Gomes, Ana P.; Suri, Vipin; Ellis, James L.; Lutz, Thomas A.; Hottiger, Michael O.; Sinclair, David A.; Auwerx, Johan; Schoonjans, Kristina; Staels, Bart; Lüscher, Thomas F.; Matter, Christian M.

    2015-01-01

    Aims The deacetylase sirtuin 1 (Sirt1) exerts beneficial effects on lipid metabolism, but its roles in plasma LDL-cholesterol regulation and atherosclerosis are controversial. Thus, we applied the pharmacological Sirt1 activator SRT3025 in a mouse model of atherosclerosis and in hepatocyte culture. Methods and results Apolipoprotein E-deficient (Apoe−/−) mice were fed a high-cholesterol diet (1.25% w/w) supplemented with SRT3025 (3.18 g kg−1 diet) for 12 weeks. In vitro, the drug activated wild-type Sirt1 protein, but not the activation-resistant Sirt1 mutant; in vivo, it increased deacetylation of hepatic p65 and skeletal muscle Foxo1. SRT3025 treatment decreased plasma levels of LDL-cholesterol and total cholesterol and reduced atherosclerosis. Drug treatment did not change mRNA expression of hepatic LDL receptor (Ldlr) and proprotein convertase subtilisin/kexin type 9 (Pcsk9), but increased their protein expression indicating post-translational effects. Consistent with hepatocyte Ldlr and Pcsk9 accumulation, we found reduced plasma levels of Pcsk9 after pharmacological Sirt1 activation. In vitro administration of SRT3025 to cultured AML12 hepatocytes attenuated Pcsk9 secretion and its binding to Ldlr, thereby reducing Pcsk9-mediated Ldlr degradation and increasing Ldlr expression and LDL uptake. Co-administration of exogenous Pcsk9 with SRT3025 blunted these effects. Sirt1 activation with SRT3025 in Ldlr−/− mice reduced neither plasma Pcsk9, nor LDL-cholesterol levels, nor atherosclerosis. Conclusion We identify reduction in Pcsk9 secretion as a novel effect of Sirt1 activity and uncover Ldlr as a prerequisite for Sirt1-mediated atheroprotection in mice. Pharmacological activation of Sirt1 appears promising to be tested in patients for its effects on plasma Pcsk9, LDL-cholesterol, and atherosclerosis. PMID:24603306

  16. Angiopoietin-like 4 Stimulates STAT3-mediated iNOS Expression and Enhances Angiogenesis to Accelerate Wound Healing in Diabetic Mice

    PubMed Central

    Chong, Han Chung; Chan, Jeremy Soon Kiat; Goh, Chi Qin; Gounko, Natalia V; Luo, Baiwen; Wang, Xiaoling; Foo, Selin; Wong, Marcus Thien Chong; Choong, Cleo; Kersten, Sander; Tan, Nguan Soon

    2014-01-01

    Impaired wound healing is a major source of morbidity in diabetic patients. Poor outcome has, in part, been related to increased inflammation, poor angiogenesis, and deficiencies in extracellular matrix components. Despite the enormous impact of these chronic wounds, effective therapies are lacking. Here, we showed that the topical application of recombinant matricellular protein angiopoietin-like 4 (ANGPTL4) accelerated wound reepithelialization in diabetic mice, in part, by improving angiogenesis. ANGPTL4 expression is markedly elevated upon normal wound injury. In contrast, ANGPTL4 expression remains low throughout the healing period in diabetic wounds. Exogenous ANGPTL4 modulated several regulatory networks involved in cell migration, angiogenesis, and inflammation, as evidenced by an altered gene expression signature. ANGPTL4 influenced the expression profile of endothelial-specific CD31 in diabetic wounds, returning its profile to that observed in wild-type wounds. We showed ANGPTL4-induced nitric oxide production through an integrin/JAK/STAT3-mediated upregulation of inducible nitric oxide synthase (iNOS) expression in wound epithelia, thus revealing a hitherto unknown mechanism by which ANGPTL4 regulated angiogenesis via keratinocyte-to-endothelial-cell communication. These data show that the replacement of ANGPTL4 may be an effective adjunctive or new therapeutic avenue for treating poor healing wounds. The present finding also confirms that therapeutic angiogenesis remains an attractive treatment modality for diabetic wound healing. PMID:24903577

  17. Interleukin-17 enhances the removal of respiratory syncytial virus in mice by promoting neutrophil migration and reducing interferon-gamma expression.

    PubMed

    Zhang, G; Zhou, K F; Lu, Z H

    2016-01-01

    The aim of this study was to observe the effect of interleukin (IL)-17 on early immune response and inflammation in the lungs of respiratory syncytial virus (RSV)-infected mice. Specific pathogen-free BALB/c mice were randomly assigned to control, RSV-infected, RSV-infected with phosphate-buffered saline, and RSV-infected + IL-17 intervention groups. The RSV infection model was set up by nasal mucosa immunization. The intervention group was provided with restructured IL-17 (intranasal). The viral load and cytokine concentrations in the lung tissues and broncho-alveolar lavage fluid (BALF) were determined by real-time-polymerase chain reaction and enzyme-linked immunosorbent assay. RSV caused acute lung inflammation in mice with a significantly higher number of neutrophils and cytokines such as interferon-gamma (IFN-γ), IL-1β, IL-6, and G-CSF in the BALF than that in the control group. IL-17 intervention led to a significant increase in the number of neutrophilic granulocytes in the BALF. Alternately, IL-17 intervention led to a significant decrease in the IFN-γ concentration and a significant increase in the IL-1β, IL-6 and G-CSF levels in the BALF. IL-17 induced a reduction in the viral load and an increase in the survival rate of mice on the third day of infection. IL- 17 mucosal immunity enhances the removal of RSV and strengthens the immune defense by promoting neutrophil migration and reducing the IFN-γ levels in mouse lungs. PMID:27050987

  18. Adjuvant-enhanced antibody and cellular responses to inclusion bodies expressing FhSAP2 correlates with protection of mice to Fasciola hepatica.

    PubMed

    Rivera, Francheska; Espino, Ana M

    2016-01-01

    Fasciola hepatica saposin-like protein-2 (FhSAP2) is a protein differentially expressed in various developmental stages of F. hepatica. Recombinant FhSAP2 has demonstrated the induction of partial protection in mice and rabbits when it is administered subcutaneously (SC) in Freund's adjuvant. Because FhSAP2 is overexpressed in bacteria in the form of inclusion bodies (IBs), we isolated IBs expressing FhSAP2 and tested their immunogenicity when administered SC in mice emulsified in two different adjuvants: QS-21 and Montanide TM ISA720. Animals received three injections containing 20 μg of protein two weeks apart and 4 weeks after the third injection, mice were infected with 10 F. hepatica metacercariae by oral route. The percentages of protection induced by FhSAP2-IBs were estimated to be between 60.0 and 62.5% when compared with adjuvant-vaccinated, infected controls. By determining the levels of IgG1 and IgG2a antibodies and IL-4 and IFNγ cytokines in the serum of experimental animals, it was found that both Th1 and Th2 immune responses were significantly increased in the FhSAP2-IBs vaccinated groups compared with the adjuvant-vaccinated, infected control groups. The adjuvant-vaccinated groups had significantly lower IgG1 to IgG2a ratios and lower IL-4 to IFNγ ratios than the FhSAP2-IBs vaccinated animals, which is indicative of higher levels of Th2 immune responses. Irrespective to the adjuvant used, animals vaccinated with FhSAP2-IBs exhibited significantly higher survival percentage and less liver damage than the adjuvant-control groups. This study suggests that FhSAP2 has potential as vaccine against F. hepatica and that the protection elicited by this molecule could be linked to a mechanism driven by the CD4-Th1 cells. PMID:26632503

  19. Deletion of the Distal Tnfsf11 RL-D2 Enhancer That Contributes to PTH-Mediated RANKL Expression in Osteoblast Lineage Cells Results in a High Bone Mass Phenotype in Mice.

    PubMed

    Onal, Melda; St John, Hillary C; Danielson, Allison L; Pike, J Wesley

    2016-02-01

    Receptor activator of nuclear factor-κB ligand (RANKL) is a tumor necrosis factor (TNF)-like cytokine that is necessary for osteoclast formation and survival. Elevated RANKL synthesis is associated with both increased osteoclast number and bone resorption. Earlier studies identified an enhancer 76 kb upstream of the Tnfsf11 transcriptional start site (TSS) termed RL-D5 or the distal control region (DCR) that modulates RANKL expression in response to PTH, 1,25(OH)2D3,, and an array of cytokines. Mice lacking RL-D5 exhibit high bone mass associated with decreased RANKL expression in bone, spleen, and thymus. In addition to RL-D5, genome-wide studies have identified 9 additional Tnfsf11 enhancers residing upstream of the gene's TSS, which provide RANKL cell type-specificity and responsiveness to local and systemic factors. ChIP-chip analyses has revealed inducible vitamin D receptor (VDR) and cAMP response element-binding protein (CREB) binding at an enhancer termed RL-D2 23 kb upstream of the Tnfsf11 TSS in osteoblastic ST2 cells. Herein, we use ChIP-seq analyses to confirm this finding and then delete this enhancer from the mouse genome to determine its physiological role in vivo. RL-D2(-/-) primary stromal cells showed decreased RANKL-induction by both forskolin and 1,25(OH)2D3 ex vivo. Consistent with this, the parathyroid hormone (PTH) induction of RANKL expression was significantly blunted in RL-D2(-/-) mice in vivo. In contrast, lack of RL-D2 had no effect on 1,25(OH)2D3 induction of RANKL in vivo. Similar to the results found in RL-D5(-/-) mice, lack of RL-D2 led to decreased skeletal RANKL expression, resulting in decreased osteoclast numbers and a progressive increase in bone mineral density. Lack of RL-D2 increased cancellous bone mass in femur and spine but did not alter femoral cortical bone thickness. These results highlight the role of distal enhancers in the regulation of RANKL expression by PTH and perhaps 1,25(OH)2D3 and suggest that the RL-D2 and

  20. Enhanced colonic tumorigenesis in alkaline sphingomyelinase (NPP7) knockout mice.

    PubMed

    Chen, Ying; Zhang, Ping; Xu, Shu-Chang; Yang, Liping; Voss, Ulrikke; Ekblad, Eva; Wu, Yunjin; Min, Yalan; Hertervig, Erik; Nilsson, Åke; Duan, Rui-Dong

    2015-01-01

    Intestinal alkaline sphingomyelinase (alk-SMase) generates ceramide and inactivates platelet-activating factor (PAF) and was previously suggested to have anticancer properties. The direct evidence is still lacking. We studied colonic tumorigenesis in alk-SMase knockout (KO) mice. Formation of aberrant crypt foci (ACF) was examined after azoxymethane (AOM) injection. Tumor was induced by AOM alone, a conventional AOM/dextran sulfate sodium (DSS) treatment, and an enhanced AOM/DSS method. β-Catenin was determined by immunohistochemistry, PAF levels by ELISA, and sphingomyelin metabolites by mass spectrometry. Without treatment, spontaneous tumorigenesis was not identified but the intestinal mucosa appeared thicker in KO than in wild-type (WT) littermates. AOM alone induced more ACF in KO mice but no tumors 28 weeks after injection. However, combination of AOM/DSS treatments induced colonic tumors and the incidence was significantly higher in KO than in WT mice. By the enhanced AOM/DSS method, tumor number per mouse increased 4.5 times and tumor size 1.8 times in KO compared with WT mice. Although all tumors were adenomas in WT mice, 32% were adenocarcinomas in KO mice. Compared with WT mice, cytosol expression of β-catenin was significantly increased and nuclear translocation in tumors was more pronounced in KO mice. Lipid analysis showed decreased ceramide in small intestine and increased sphingosine-1-phosphate (S1P) in both small intestine and colon in nontreated KO mice. PAF levels in feces were significantly higher in the KO mice after AOM/DSS treatment. In conclusion, lack of alk-SMase markedly increases AOM/DSS-induced colonic tumorigenesis associated with decreased ceramide and increased S1P and PAF levels. PMID:25381265

  1. Myocardial Fibrosis Induced by Exposure to Subclinical Lipopolysaccharide Is Associated with Decreased miR-29c and Enhanced NOX2 Expression in Mice

    PubMed Central

    Lew, Wilbur Y. W.; Bayna, Evelyn; Dalle Molle, Erminia; Contu, Riccardo; Condorelli, Gianluigi; Tang, Tong

    2014-01-01

    Background Exposure to subclinical levels of lipopolysaccharide (LPS) occurs commonly and is seemingly well tolerated. However, recurrent LPS exposure induces cardiac fibrosis over 2 to 3 months in a murine model, not mediated by the renin-angiotensin system. Subclinical LPS induces cardiac fibrosis by unique mechanisms. Methods In C57/Bl6 mice, LPS (10 mg/kg) or saline (control) were injected intraperitoneally once a week for 1–4 weeks. Mice showed no signs of distress, change in activity, appetite, or weight loss. Mice were euthanized after 3 days, 1, 2, or 4 weeks to measure cardiac expression of fibrosis-related genes and potential mediators (measured by QRT-PCR), including micro-RNA (miR) and NADPH oxidase (NOX). Collagen fraction area of the left ventricle was measured with picrosirius red staining. Cardiac fibroblasts isolated from adult mouse hearts were incubated with 0, 0.1, 1.0 or 10 ng/ml LPS for 48 hours. Results Cardiac miR expression profiling demonstrated decreased miR-29c after 3 and 7 days following LPS, which were confirmed by QRT-PCR. The earliest changes in fibrosis-related genes and mediators that occurred 3 days after LPS were increased cardiac expression of TIMP-1 and NOX-2 (but not of NOX-4). This persisted at 1 and 2 weeks, with additional increases in collagen Iα1, collagen IIIα1, MMP2, MMP9, TIMP1, TIMP2, and periostin. There was no change in TGF-β or connective tissue growth factor. Collagen fraction area of the left ventricle increased after 2 and 4 weeks of LPS. LPS decreased miR-29c and increased NOX-2 in isolated cardiac fibroblasts. Conclusions Recurrent exposure to subclinical LPS induces cardiac fibrosis after 2–4 weeks. Early changes 3 days after LPS were decreased miR-29c and increased NOX2 and TIMP1, which persisted at 1 and 2 weeks, along with widespread activation of fibrosis-related genes. Decreased miR-29c and increased NOX2, which induce cardiac fibrosis in other conditions, may uniquely mediate LPS-induced cardiac

  2. Allergic Lung Inflammation Reduces Tissue Invasion and Enhances Survival from Pulmonary Pneumococcal Infection in Mice, Which Correlates with Increased Expression of Transforming Growth Factor β1 and SiglecFlow Alveolar Macrophages

    PubMed Central

    Sanfilippo, Alan M.; Furuya, Yoichi; Roberts, Sean; Salmon, Sharon L.

    2015-01-01

    Asthma is generally thought to confer an increased risk for invasive pneumococcal disease (IPD) in humans. However, recent reports suggest that mortality rates from IPD are unaffected in patients with asthma and that chronic obstructive pulmonary disease (COPD), a condition similar to asthma, protects against the development of complicated pneumonia. To clarify the effects of asthma on the subsequent susceptibility to pneumococcal infection, ovalbumin (OVA)-induced allergic lung inflammation (ALI) was induced in mice followed by intranasal infection with A66.1 serotype 3 Streptococcus pneumoniae. Surprisingly, mice with ALI were significantly more resistant to lethal infection than non-ALI mice. The heightened resistance observed following ALI correlated with enhanced early clearance of pneumococci from the lung, decreased bacterial invasion from the airway into the lung tissue, a blunted inflammatory cytokine and neutrophil response to infection, and enhanced expression of transforming growth factor β1 (TGF-β1). Neutrophil depletion prior to infection had no effect on enhanced early bacterial clearance or resistance to IPD in mice with ALI. Although eosinophils recruited into the lung during ALI appeared to be capable of phagocytizing bacteria, neutralization of interleukin-5 (IL-5) to inhibit eosinophil recruitment likewise had no effect on early clearance or survival following infection. However, enhanced resistance was associated with an increase in levels of clodronate-sensitive, phagocytic SiglecFlow alveolar macrophages within the airways following ALI. These findings suggest that, while the risk of developing IPD may actually be decreased in patients with acute asthma, additional clinical data are needed to better understand the risk of IPD in patients with different asthma phenotypes. PMID:25964474

  3. Allergic Lung Inflammation Reduces Tissue Invasion and Enhances Survival from Pulmonary Pneumococcal Infection in Mice, Which Correlates with Increased Expression of Transforming Growth Factor β1 and SiglecF(low) Alveolar Macrophages.

    PubMed

    Sanfilippo, Alan M; Furuya, Yoichi; Roberts, Sean; Salmon, Sharon L; Metzger, Dennis W

    2015-07-01

    Asthma is generally thought to confer an increased risk for invasive pneumococcal disease (IPD) in humans. However, recent reports suggest that mortality rates from IPD are unaffected in patients with asthma and that chronic obstructive pulmonary disease (COPD), a condition similar to asthma, protects against the development of complicated pneumonia. To clarify the effects of asthma on the subsequent susceptibility to pneumococcal infection, ovalbumin (OVA)-induced allergic lung inflammation (ALI) was induced in mice followed by intranasal infection with A66.1 serotype 3 Streptococcus pneumoniae. Surprisingly, mice with ALI were significantly more resistant to lethal infection than non-ALI mice. The heightened resistance observed following ALI correlated with enhanced early clearance of pneumococci from the lung, decreased bacterial invasion from the airway into the lung tissue, a blunted inflammatory cytokine and neutrophil response to infection, and enhanced expression of transforming growth factor β1 (TGF-β1). Neutrophil depletion prior to infection had no effect on enhanced early bacterial clearance or resistance to IPD in mice with ALI. Although eosinophils recruited into the lung during ALI appeared to be capable of phagocytizing bacteria, neutralization of interleukin-5 (IL-5) to inhibit eosinophil recruitment likewise had no effect on early clearance or survival following infection. However, enhanced resistance was associated with an increase in levels of clodronate-sensitive, phagocytic SiglecF(low) alveolar macrophages within the airways following ALI. These findings suggest that, while the risk of developing IPD may actually be decreased in patients with acute asthma, additional clinical data are needed to better understand the risk of IPD in patients with different asthma phenotypes. PMID:25964474

  4. Enhancing immune responses of EV71 VP1 DNA vaccine by co-inoculating plasmid IL-12 or GM-CSF expressing vector in mice.

    PubMed

    Peng, X; Fang, X; Li, J; Kong, L; Li, B; Ding, X

    2016-01-01

    Enterovirus 71 (EV71) is a major causative viral agent for large outbreaks of hand, foot, and mouth disease in children and infants, yet there is no vaccine or effective antiviral treatment for severe EV71 infection. The immunogenicity of EV71 VP1 DNA vaccine and the immunoregulatory activity of interleukin-12 (IL-12) or granulocyte-monocyte colony stimulating factor (GM-CSF) were investigated. DNA vaccine plasmids, pcDNA-VP1, pcDNA-IL-12 and pcDNA-GM-CSF were constructed and inoculated into BALB/c mice with or without pcDNA-IL-12 or pcDNA-GM-CSF by intramuscular injection. Cellular and humoral immune responses were assessed by indirect ELISA, lymphocyte proliferation assays, cytokine release assay and FACS. The VP1 DNA vaccine had good immunogenicity and can induce specific humoral and cellular immunity in BALB/c mice, while IL-2 or GM-CSF plays an immunoadjuvant role and enhances specific immune responses. This study provides a frame of reference for the design of DNA vaccines against EV71. PMID:27188732

  5. Ablation of Chop Transiently Enhances Photoreceptor Survival but Does Not Prevent Retinal Degeneration in Transgenic Mice Expressing Human P23H Rhodopsin

    PubMed Central

    Chiang, Wei-Chieh; Joseph, Victory; Matthes, Michael T.; Lewin, Alfred S.; Gorbatyuk, Marina S.; Ahern, Kelly; LaVail, Matthew M.

    2016-01-01

    RHO (Rod opsin) encodes a G-protein coupled receptor that is expressed exclusively by rod photoreceptors of the retina and forms the essential photopigment, rhodopsin, when coupled with 11-cis-retinal. Many rod opsin disease mutations cause rod opsin protein misfolding and trigger endoplasmic reticulum (ER) stress, leading to activation of the Unfolded Protein Response (UPR) signal transduction network. Chop is a transcriptional activator that is induced by ER stress and promotes cell death in response to chronic ER stress. Here, we examined the role of Chop in transgenic mice expressing human P23H rhodopsin (hP23H Rho Tg) that undergo retinal degeneration. With the exception of one time point, we found no significant induction of Chop in these animals and no significant change in retinal degeneration by histology and electrophysiology when hP23H Rho Tg animals were bred into a Chop−/− background. Our results indicate that Chop does not play a significant causal role during retinal degeneration in these animals. We suggest that other modules of the ER stress-induced UPR signaling network may be involved photoreceptor disease induced by P23H rhodopsin. PMID:26427410

  6. Adenoviral vector expressing murine β-defensin 2 enhances immunogenicity of an adenoviral vector based H5N1 influenza vaccine in aged mice.

    PubMed

    Vemula, Sai V; Pandey, Aseem; Singh, Neetu; Katz, Jacqueline M; Donis, Ruben; Sambhara, Suryaprakash; Mittal, Suresh K

    2013-10-01

    The ability to resist infections and respond to vaccinations is greatly reduced in the older adult population owing to a general decline in innate and adaptive immune functions with aging. Over the years several strategies such as increasing the vaccine dose, number of immunizations and using adjuvants have been evaluated to improve the immunogenicity and efficacy of vaccines in the older adult population. Murine β-defensin 2 (Mbd2) has been shown to function as a molecular adjuvant by recruiting and activating immature dendritic cells (DCs), professional antigen-presenting cells (APC), to the site of the immunization. In this study, we evaluated the potential utility of Mbd2 to enhance the efficacy of an adenoviral vector-based H5N1 influenza vaccine expressing hemagglutinin (HA) and nucleoprotein (NP) (HAd-HA-NP) in an aged mouse model. Our results indicated that immunostimulation with an adenoviral vector expressing Mbd2 (HAd-Mbd2) activated DCs and significantly enhanced the humoral and cellular immune responses induced by HAd-HA-NP. Furthermore, immunostimulation with HAd-Mbd2 followed by immunization with HAd-HA-NP resulted in significantly lower virus titers in the lungs following challenge with a H5N1 influenza virus compared to the group immunized with HAd-HA-NP without immunostimulation. Overall, our results highlight the potential utility of Mbd2 as a molecular adjuvant to enhance the immunogenicity and protective efficacy of vaccines for the elderly. PMID:23892144

  7. Adenoviral Vector Expressing Murine β-Defensin 2 Enhances Immunogenicity of an Adenoviral Vector based H5N1 Influenza Vaccine in Aged Mice

    PubMed Central

    Vemula, Sai V.; Pandey, Aseem; Singh, Neetu; Katz, Jacqueline M; Donis, Ruben; Sambhara, Suryaprakash; Mittal, Suresh K.

    2013-01-01

    The ability to resist infections and respond to vaccinations is greatly reduced in the older adult population owing to a general decline in innate and adaptive immune functions with aging. Over the years several strategies such as increasing the vaccine dose, number of immunizations and using adjuvants have been evaluated to improve the immunogenicity and efficacy of vaccines in the older adult population. Murine ß-defensin 2 (Mbd2) has been shown to function as a molecular adjuvant by recruiting and activating immature dendritic cells (DCs), professional antigen-presenting cells (APC), to the site of the immunization. In this study, we evaluated the potential utility of Mbd2 to enhance the efficacy of an adenoviral vector-based H5N1 influenza vaccine expressing hemagglutinin (HA) and nucleoprotein (NP) (HAd-HA-NP) in an aged mouse model. Our results indicated that immunostimulation with an adenoviral vector expressing Mbd2 (HAd-Mbd2) activated DCs and significantly enhanced the humoral and cellular immune responses induced by HAd-HA-NP. Furthermore, immunostimulation with HAd-Mbd2 followed by immunization with HAd-HA-NP resulted in significantly lower virus titers in the lungs following challenge with a H5N1 influenza virus compared to the group immunized with HAd-HA-NP without immunostimulation. Overall, our results highlight the potential utility of Mbd2 as a molecular adjuvant to enhance the immunogenicity and protective efficacy of vaccines for the elderly. PMID:23892144

  8. Attenuated Escherichia coli strains expressing the colonization factor antigen I (CFA/I) and a detoxified heat-labile enterotoxin (LThK63) enhance clearance of ETEC from the lungs of mice and protect mice from intestinal ETEC colonization and LT-induced fluid accumulation.

    PubMed

    Byrd, Wyatt; Boedeker, Edgar C

    2013-03-15

    Although enterotoxigenic Escherichia coli (ETEC) infections are important causes of infantile and traveler's diarrhea there is no licensed vaccine available for those at-risk. Our goal is to develop a safe, live attenuated ETEC vaccine. We used an attenuated E. coli strain (O157:H7, Δ-intimin, Stx1-neg, Stx2-neg) as a vector (ZCR533) to prepare two vaccine strains, one strain expressing colonization factor antigen I (ZCR533-CFA/I) and one strain expressing CFA/I and a detoxified heat-labile enterotoxin (ZCR533-CFA/I+LThK63) to deliver ETEC antigens to mucosal sites in BALB/c mice. Following intranasal and intragastric immunization with the vaccine strains, serum IgG and IgA antibodies were measured to the CFA/I antigen, however, only serum IgG antibodies were detected to the heat-labile enterotoxin. Intranasal administration of the vaccine strains induced respiratory and intestinal antibody responses to the CFA/I and LT antigens, while intragastric administration induced only intestinal antibody responses with no respiratory antibodies detected to the CFA/I and LT antigens. Mice immunized intranasally with the vaccine strains showed enhanced clearance of wild-type (wt) ETEC bacteria from the lungs. Mice immunized intranasally and intragastrically with the vaccine strains were protected from intestinal colonization following oral challenge with ETEC wt bacteria. Mice immunized intragastrically with the ZCR533-CFA/I+LThK63 vaccine strain had less fluid accumulate in their intestine following challenge with ETEC wt bacteria or with purified LT as compared to the sham mice indicating that the immunized mice were protected from LT-induced intestinal fluid accumulation. Thus, mice intragastrically immunized with the ZCR533-CFA/I+LThK63 vaccine strain were able to effectively neutralize the activity of the LT enterotoxin. However, no difference in intestinal fluid accumulation was detected in the mice immunized intranasally with the vaccine strain as compared to the sham

  9. MondoA deficiency enhances sprint performance in mice

    PubMed Central

    Imamura, Minako; Chang, Benny Hung-Junn; Kohjima, Motoyuki; Li, Ming; Hwang, Byounghoon; Taegtmeyer, Heinrich; Harris, Robert A.; Chan, Lawrence

    2015-01-01

    MondoA is a basic helix–loop–helix (bHLH)/leucine zipper (ZIP) transcription factor that is expressed predominantly in skeletal muscle. Studies in vitro suggest that the Max-like protein X (MondoA:Mlx) heterodimer senses the intracellular energy status and directly targets the promoter region of thioredoxin interacting protein (Txnip) and possibly glycolytic enzymes. We generated MondoA-inactivated (MondoA−/−) mice by gene targeting. MondoA−/− mice had normal body weight at birth, exhibited normal growth and appeared to be healthy. However, they exhibited unique metabolic characteristics. MondoA−/− mice built up serum lactate and alanine levels and utilized fatty acids for fuel during exercise. Gene expression and promoter analysis suggested that MondoA functionally represses peroxisome-proliferator-activated receptor γ co-activator-1α (PGC-1α)–mediated activation of pyruvate dehydrogenase kinase 4 (PDK4) transcription. PDK4 normally down-regulates the activity of pyruvate dehydrogenase, an enzyme complex that catalyses the decarboxylation of pyruvate to acetyl-CoA for entry into the Krebs cycle; in the absence of MondoA, pyruvate is diverted towards lactate and alanine, both products of glycolysis. Dynamic testing revealed that MondoA−/− mice excel in sprinting as their skeletal muscles display an enhanced glycolytic capacity. Our studies uncover a hitherto unappreciated function of MondoA in fuel selection in vivo. Lack of MondoA results in enhanced exercise capacity with sprinting. PMID:25145386

  10. Lifespan of mice and primates correlates with immunoproteasome expression

    PubMed Central

    Pickering, Andrew M.; Lehr, Marcus; Miller, Richard A.

    2015-01-01

    There is large variation in lifespan among different species, and there is evidence that modulation of proteasome function may contribute to longevity determination. Comparative biology provides a powerful tool for identifying genes and pathways that control the rate of aging. Here, we evaluated skin-derived fibroblasts and demonstrate that among primate species, longevity correlated with an elevation in proteasomal activity as well as immunoproteasome expression at both the mRNA and protein levels. Immunoproteasome enhancement occurred with a concurrent increase in other elements involved in MHC class I antigen presentation, including β-2 microglobulin, (TAP1), and TAP2. Fibroblasts from long-lived primates also appeared more responsive to IFN-γ than cells from short-lived primate species, and this increase in IFN-γ responsiveness correlated with elevated expression of the IFN-γ receptor protein IFNGR2. Elevation of immunoproteasome and proteasome activity was also observed in the livers of long-lived Snell dwarf mice and in mice exposed to drugs that have been shown to extend lifespan, including rapamycin, 17-α-estradiol, and nordihydroguaiaretic acid. This work suggests that augmented immunoproteasome function may contribute to lifespan differences in mice and among primate species. PMID:25866968

  11. Lymphopoiesis in transgenic mice over-expressing Artemis.

    PubMed

    Rivera-Munoz, P; Abramowski, V; Jacquot, S; André, P; Charrier, S; Lipson-Ruffert, K; Fischer, A; Galy, A; Cavazzana, M; de Villartay, J-P

    2016-02-01

    Artemis is a factor of the non-homologous end joining pathway involved in DNA double-strand break repair that has a critical role in V(D)J recombination. Mutations in DCLRE1C/ARTEMIS gene result in radiosensitive severe combined immunodeficiency in humans owing to a lack of mature T and B cells. Given the known drawbacks of allogeneic hematopoietic stem cell transplantation (HSCT), gene therapy appears as a promising alternative for these patients. However, the safety of an unregulated expression of Artemis has to be established. We developed a transgenic mouse model expressing human Artemis under the control of the strong CMV early enhancer/chicken beta actin promoter through knock-in at the ROSA26 locus to analyze this issue. Transgenic mice present a normal development, maturation and function of T and B cells with no signs of lymphopoietic malignancies for up to 15 months. These results suggest that the over-expression of Artemis in mice (up to 40 times) has no deleterious effects in early and mature lymphoid cells and support the safety of gene therapy as a possible curative treatment for Artemis-deficient patients. PMID:26361272

  12. An enhancer from the 8q24 prostate cancer risk region is sufficient to direct reporter gene expression to a subset of prostate stem-like epithelial cells in transgenic mice

    PubMed Central

    Ting, Man-Chun; Liao, Chun-Peng; Yan, Chunli; Jia, Li; Groshen, Susan; Frenkel, Baruch; Roy-Burman, Pradip; Coetzee, Gerhard A.; Maxson, Robert

    2012-01-01

    SUMMARY Regions in the 8q24 gene desert contribute significantly to the risk of prostate cancer and other adult cancers. This region contains several DNA regions with enhancer activity in cultured cells. One such segment, histone acetylation peak 10 (AcP10), contains a risk single nucleotide polymorphism (SNP) that is significantly associated with the pathogenesis of colorectal, prostate and other cancers. The mechanism by which AcP10 influences cancer risk remains unknown. Here we show that AcP10 contains a sequence that is highly conserved across terrestrial vertebrates and is capable in transgenic mice of directing reporter gene expression to a subset of prostate lumenal epithelial cells. These cells include a small population of Nkx3.1-positive cells that persist even after androgen ablation. Castration-resistant Nkx3.1-positive (CARN) cells were shown by others to function both as stem cells and cells of origin of prostate cancer. Our results thus provide a mechanism by which AcP10 could influence prostate cancer risk. PMID:22279083

  13. Both core and F proteins of hepatitis C virus could enhance cell proliferation in transgenic mice

    SciTech Connect

    Hu, Wen-Ta; Li, Hui-Chun; Lee, Shen-Kao; Ma, Hsin-Chieh; Yang, Chee-Hing; Chen, Hung-Ling; Lo, Shih-Yen

    2013-05-24

    Highlights: •HCV core and F proteins could induce hepatocyte proliferation in the transgenic mice. •β-Catenin signaling pathway was activated by core protein in the transgenic mice. •β-Catenin signaling pathway was activated by myc-F protein in the transgenic mice. •Expression of SMA protein was enhanced by core but not myc-F protein. -- Abstract: The role of the protein encoded by the alternative open reading frame (ARF/F/core+1) of the Hepatitis C virus (HCV) genome in viral pathogenesis remains unknown. The different forms of ARF/F/core+1 protein were labile in cultured cells, a myc-tag fused at the N-terminus of the F protein made it more stable. To determine the role of core and F proteins in HCV pathogenesis, transgenic mice with either protein expression under the control of Albumin promoter were generated. Expression of core protein and F protein with myc tag (myc-F) could be detected by Western blotting analysis in the livers of these mice. The ratio of liver to body weight is increased for both core and myc-F transgenic mice compared to that of wild type mice. Indeed, the proliferating cell nuclear antigen protein, a proliferation marker, was up-regulated in the transgenic mice with core or myc-F protein. Further analyses by microarray and Western blotting suggested that β-catenin signaling pathway was activated by either core or myc-F protein in the transgenic mice. These transgenic mice were further treated with either Diethynitrosamine (a tumor initiator) or Phenobarbital (a tumor promoter). Phenobarbital but not Diethynitrosamine treatment could increase the liver/body weight ratio of these mice. However, no tumor formation was observed in these mice. In conclusion, HCV core and myc-F proteins could induce hepatocyte proliferation in the transgenic mice possibly through β-catenin signaling pathway.

  14. Endothelial Expression of Scavenger Receptor Class B, Type I Protects against Development of Atherosclerosis in Mice

    PubMed Central

    Vaisman, Boris L.; Vishnyakova, Tatyana G.; Freeman, Lita A.; Amar, Marcelo J.; Demosky, Stephen J.; Liu, Chengyu; Stonik, John A.; Sampson, Maureen L.; Pryor, Milton; Bocharov, Alexander V.; Eggerman, Thomas L.; Patterson, Amy P.; Remaley, Alan T.

    2015-01-01

    The role of scavenger receptor class B, type I (SR-BI) in endothelial cells (EC) was examined in several novel transgenic mouse models expressing SR-BI in endothelium of mice with normal C57Bl6/N, apoE-KO, or Scarb1-KO backgrounds. Mice were also created expressing SR-BI exclusively in endothelium and liver. Endothelial expression of the Tie2-Scarb1 transgene had no significant effect on plasma lipoprotein levels in mice on a normal chow diet but on an atherogenic diet, significantly decreased plasma cholesterol levels, increased plasma HDL cholesterol (HDL-C) levels, and protected mice against atherosclerosis. In 8-month-old apoE-KO mice fed a normal chow diet, the Tie2-Scarb1 transgene decreased aortic lesions by 24%. Mice expressing SR-BI only in EC and liver had a 1.5 ± 0.1-fold increase in plasma cholesterol compared to mice synthesizing SR-BI only in liver. This elevation was due mostly to increased HDL-C. In EC culture studies, SR-BI was found to be present in both basolateral and apical membranes but greater cellular uptake of cholesterol from HDL was found in the basolateral compartment. In summary, enhanced expression of SR-BI in EC resulted in a less atherogenic lipoprotein profile and decreased atherosclerosis, suggesting a possible role for endothelial SR-BI in the flux of cholesterol across EC. PMID:26504816

  15. In vivo modulation of epidermal growth factor receptor phosphorylation in mice expressing different gangliosides.

    PubMed

    Daniotti, Jose L; Crespo, Pilar M; Yamashita, Tadashi

    2006-12-01

    We studied in this work the in vivo phosphorylation of the epidermal growth factor receptor (EGFr) in skin from knockout mice lacking different ganglioside glycosyltransferases. Results show an enhancement of EGFr phosphorylation, after EGF stimulation, in skin from Sial-T2 knockout and Sial-T2/GalNAc-T double knockout mice as compared with wild-type and Sial-T1 knockout mice. Qualitative analysis of ganglioside composition in mice skin suggest that the increase of EGFr phosphorylation observed in skin from Sial-T2 knockout and Sial-T2/GalNAc-T double knockout mice in response to EGF might not be primary attributed to the expression of GD3 or a-series gangliosides in mice skin. These studies provide, for the first time, an approach for studying the molecular mechanisms involved in the in vivo regulation of EGFr function by gangliosides. PMID:16817235

  16. Idiotypic manipulation in mice: BALB/c mice can express the crossreactive idiotype of A/J mice.

    PubMed Central

    Moser, M; Leo, O; Hiernaux, J; Urbain, J

    1983-01-01

    The response of A/J mice to arsonate-coupled keyhole limpet hemocyanin is characterized by a crossreactive idiotype (CRIA). CRIA+ antibodies are restricted to the Igh-Ic haplotype and are never expressed in BALB/c mice after immunization with antigen. Studies at the DNA level suggest that the gene encoding the CRIA heavy chain in A/J mice is probably absent in the genome of BALB/c mice. Despite this, using the immunization cascade tool, we have been able to induce the expression of CRIA+ antibodies in BALB/c mice. These studies led to an apparent paradox, whose understanding will provide new insights into the regulatory mechanisms of the immune system. We suggest that clones secreting CRIA-like Igs in BALB/c mice are "somatic variants" that could arise from gene conversion events. PMID:6576348

  17. Enhancing Methotrexate Tolerance with Folate Tagged Liposomes in Arthritic Mice.

    PubMed

    Nogueira, Eugénia; Lager, Franck; Le Roux, Delphine; Nogueira, Patrícia; Freitas, Jaime; Charvet, Celine; Renault, Gilles; Loureiro, Ana; Almeida, Catarina R; Ohradanova-Repic, Anna; Machacek, Christian; Bernardes, Gonçalo J L; Moreira, Alexandra; Stockinger, Hannes; Burnet, Michael; Carmo, Alexandre M; Gomes, Andreia C; Preto, Ana; Bismuth, Georges; Cavaco-Paulo, Artur

    2015-12-01

    Methotrexate is the first line of treatment of rheumatoid arthritis. Since many patients become unresponsive to methotrexate treatment, only very expensive biological therapies are effective and increased methotrexate tolerance strategies need to be identified. Here we propose the encapsulation of methotrexate in a new liposomal formulation using a hydrophobic fragment of surfactant protein conjugated to a linker and folate to enhance their tolerance and efficacy. In this study we aim to evaluate the efficiency of this system to treat rheumatoid arthritis, by targeting folate receptor β present at the surface of activated macrophages, key effector cells in this pathology. The specificity of our liposomal formulation to target folate receptor β was investigated both in vitro as in vivo using a mouse model of arthritis (collagen-induced arthritis in DBA/1J mice strain). In both systems, the liposomal constructs were shown to be highly specific and efficient in targeting folate receptor β. These liposomal formulations also significantly increase the clinical benefit of the encapsulated methotrexate in vivo in arthritic mice, together with reduced expression of CD39 and CD73 ectonucleotidases by joint-infiltrating macrophages. Thus, our formulation might be a promising cost effective way to treat rheumatoid arthritis and delay or reduce methotrexate intolerance. PMID:26510317

  18. The transcription factor ATF4 regulates glucose metabolism in mice through its expression in osteoblasts

    PubMed Central

    Yoshizawa, Tatsuya; Hinoi, Eiichi; Jung, Dae Young; Kajimura, Daisuke; Ferron, Mathieu; Seo, Jin; Graff, Jonathan M.; Kim, Jason K.; Karsenty, Gerard

    2009-01-01

    The recent demonstration that osteoblasts have a role in controlling energy metabolism suggests that they express cell-specific regulatory genes involved in this process. Activating transcription factor 4 (ATF4) is a transcription factor that accumulates predominantly in osteoblasts, where it regulates virtually all functions linked to the maintenance of bone mass. Since Atf4–/– mice have smaller fat pads than littermate controls, we investigated whether ATF4 also influences energy metabolism. Here, we have shown, through analysis of Atf4–/–mice, that ATF4 inhibits insulin secretion and decreases insulin sensitivity in liver, fat, and muscle. Several lines of evidence indicated that this function of ATF4 occurred through its osteoblastic expression. First, insulin sensitivity is enhanced in the liver of Atf4–/– mice, but not in cultured hepatocytes from these mice. Second, mice overexpressing ATF4 in osteoblasts only [termed here α1(I)Collagen-Atf4 mice] displayed a decrease in insulin secretion and were insulin insensitive. Third, the α1(I)Collagen-Atf4 transgene corrected the energy metabolism phenotype of Atf4–/– mice. Fourth, and more definitely, mice lacking ATF4 only in osteoblasts presented the same metabolic abnormalities as Atf4–/– mice. Molecularly, ATF4 favored expression in osteoblasts of Esp, which encodes a product that decreases the bioactivity of osteocalcin, an osteoblast-specific secreted molecule that enhances secretion of and sensitivity to insulin. These results provide a transcriptional basis to the observation that osteoblasts fulfill endocrine functions and identify ATF4 as a regulator of most functions of osteoblasts. PMID:19726872

  19. Caloric restriction enhances fear extinction learning in mice.

    PubMed

    Riddle, Megan C; McKenna, Morgan C; Yoon, Yone J; Pattwell, Siobhan S; Santos, Patricia Mae G; Casey, B J; Glatt, Charles E

    2013-05-01

    Fear extinction learning, the ability to reassess a learned cue of danger as safe when it no longer predicts aversive events, is often dysregulated in anxiety disorders. Selective serotonin reuptake inhibitors (SSRI's) enhance neural plasticity and their ability to enhance fear extinction learning may explain their anxiolytic properties. Caloric restriction (CR) has SSRI-like effects on neural plasticity and anxiety-related behavior. We implemented CR in mice to determine its effects on conditioned-fear responses. Wild type and serotonin transporter (SERT) knockout mice underwent CR for 7 days leading to significant weight loss. Mice were then tested for cued fear learning and anxiety-related behavior. CR markedly enhanced fear extinction learning and its retention in adolescent female mice, and adults of both sexes. These effects of CR were absent in SERT knockout mice. Moreover, CR phenocopied behavioral and molecular effects of chronic fluoxetine, but there was no additive effect of CR in fluoxetine-treated mice. These results demonstrate that CR enhances fear extinction learning through a SERT-dependent mechanism. These results may have implications for eating disorders such as anorexia nervosa (AN), in which there is a high prevalence of anxiety before the onset of dietary restriction and support proposals that in AN, CR is a motivated effort to control dysregulated fear responses and elevated anxiety. PMID:23303073

  20. Caloric Restriction Enhances Fear Extinction Learning in Mice

    PubMed Central

    Riddle, Megan C; McKenna, Morgan C; Yoon, Yone J; Pattwell, Siobhan S; Santos, Patricia Mae G; Casey, B J; Glatt, Charles E

    2013-01-01

    Fear extinction learning, the ability to reassess a learned cue of danger as safe when it no longer predicts aversive events, is often dysregulated in anxiety disorders. Selective serotonin reuptake inhibitors (SSRI's) enhance neural plasticity and their ability to enhance fear extinction learning may explain their anxiolytic properties. Caloric restriction (CR) has SSRI-like effects on neural plasticity and anxiety-related behavior. We implemented CR in mice to determine its effects on conditioned-fear responses. Wild type and serotonin transporter (SERT) knockout mice underwent CR for 7 days leading to significant weight loss. Mice were then tested for cued fear learning and anxiety-related behavior. CR markedly enhanced fear extinction learning and its retention in adolescent female mice, and adults of both sexes. These effects of CR were absent in SERT knockout mice. Moreover, CR phenocopied behavioral and molecular effects of chronic fluoxetine, but there was no additive effect of CR in fluoxetine-treated mice. These results demonstrate that CR enhances fear extinction learning through a SERT-dependent mechanism. These results may have implications for eating disorders such as anorexia nervosa (AN), in which there is a high prevalence of anxiety before the onset of dietary restriction and support proposals that in AN, CR is a motivated effort to control dysregulated fear responses and elevated anxiety. PMID:23303073

  1. Cytokine expression profile over time in burned mice

    PubMed Central

    Finnerty, Celeste C; Przkora, Rene; Herndon, David N; Jeschke, Marc G

    2009-01-01

    The persistent inflammatory response induced by a severe burn increases patient susceptibility to infections and sepsis, potentially leading to multi-organ failure and death. In order to use murine models to develop interventions that modulate the post-burn inflammatory response, the response in mice and the similarities to the human response must first be determined. Here we present the temporal serum cytokine expression profiles in burned in comparison to sham mice and human burn patients. Male C57BL/6 mice were randomized to control (n=47) or subjected to a 35% TBSA scald burn (n=89). Mice were sacrificed 3, 6, 9, 12, 24, 48 hours and 7, 10, and 14 days post-burn; cytokines were measured by multi-plex array. Following the burn injury, IL-6, IL-1β, KC, G-CSF, TNF, IL-17, MIP-1α, RANTES, and GM-CSF were increased, p<0.05. IL-2, IL-3, and IL-5 were decreased, p<0.05. IL-10, IFN-γ, and IL-12p70 were expressed in a biphasic manner, p<0.05. This temporal cytokine expression pattern elucidates the pathogenesis of the inflammatory response in burned mice. Expression of 11 cytokines were similar in mice and children, returning to lowest levels by post-burn day 14, confirming the utility of the burned mouse model for development of therapeutic interventions to attenuate the post-burn inflammatory response. PMID:19019696

  2. Murine Cytomegalovirus Influences Foxj1 Expression, Ciliogenesis, and Mucus Plugging in Mice with Allergic Airway Disease

    PubMed Central

    Wu, Carol A.; Peluso, John J.; Shanley, John D.; Puddington, Lynn; Thrall, Roger S.

    2008-01-01

    We have followed throughout time the development of allergic airway disease (AAD) in both uninfected mice and mice infected intranasally with murine cytomegalovirus (MCMV). Histological evaluation of lung tissue from uninfected mice with AAD demonstrated mucus plugging after 14 and 21 days of ovalbumin-aerosol challenge, with resolution of mucus plugging occurring by 42 days. In MCMV/AAD mice, mucus plugging was observed after 7 days of ovalbumin-aerosol challenge and remained present at 42 days. The level of interleukin-13 in bronchoalveolar lavage fluid from MCMV/AAD mice was decreased compared with AAD mice and was accompanied by increased levels of interferon-γ. Levels of Muc5A/C, Muc5B, or Muc2 mucin mRNA in the lungs of MCMV/AAD mice were not elevated compared with AAD mice. MCMV was able to infect the airway epithelium, resulting in decreased expression of Foxj1, a transcription factor critical for ciliogenesis, and a loss of ciliated epithelial cells. In addition, an increase in the number of epithelial cells staining positive for periodic acid-Schiff was observed in MCMV/AAD airways. Together, these findings suggest that MCMV infection of the airway epithelium enhances goblet cell metaplasia and diminishes efficient mucociliary clearance in mice with AAD, resulting in increased mucus plugging. PMID:18258850

  3. Cognitive-enhancing effects of hydrolysate of polygalasaponin in SAMP8 mice*

    PubMed Central

    Xu, Pan; Xu, Shu-ping; Wang, Ke-zhu; Lu, Cong; Zhang, Hong-xia; Pan, Rui-le; Qi, Chang; Yang, Yan-yan; Li, Ying-hui; Liu, Xin-min

    2016-01-01

    Objectives: The aim of the study is to evaluate the cognitive-enhancing effects of hydrolysate of polygalasaponin (HPS) on senescence accelerate mouse P8 (SAMP8) mice, an effective Alzheimer’s disease (AD) model, and to research the relevant mechanisms. Methods: The cognitive-enhancing effects of HPS on SAMP8 mice were assessed using Morris water maze (MWM) and step-through passive avoidance tests. Then N-methyl-D-aspartate (NMDA) receptor subunit expression for both the cortex and hippocampus of mice was observed using Western blotting. Results: HPS (25 and 50 mg/kg) improved the escape rate and decreased the escape latency and time spent in the target quadrant for the SAMP8 mice in the MWM after oral administration of HPS for 10 d. Moreover, it decreased error times in the passive avoidance tests. Western blotting showed that HPS was able to reverse the levels of NMDAR1 and NMDAR2B expression in the cortex or hippocampus of model mice. Conclusions: The present study suggested that HPS can improve cognitive deficits in SAMP8 mice, and this mechanism might be associated with NMDA receptor (NMDAR)-related pathways. PMID:27381727

  4. Angiopoietin-1 enhances skeletal muscle regeneration in mice

    PubMed Central

    Mofarrahi, Mahroo; McClung, Joseph M.; Kontos, Christopher D.; Davis, Elaine C.; Tappuni, Bassman; Moroz, Nicolay; Pickett, Amy E.; Huck, Laurent; Harel, Sharon; Danialou, Gawiyou

    2015-01-01

    Activation of muscle progenitor cell myogenesis and endothelial cell angiogenesis is critical for the recovery of skeletal muscle from injury. Angiopoietin-1 (Ang-1), a ligand of Tie-2 receptors, enhances angiogenesis and skeletal muscle satellite cell survival; however, its role in skeletal muscle regeneration after injury is unknown. We assessed the effects of Ang-1 on fiber regeneration, myogenesis, and angiogenesis in injured skeletal muscle (tibialis anterior, TA) in mice. We also assessed endogenous Ang-1 levels and localization in intact and injured TA muscles. TA fiber injury was triggered by cardiotoxin injection. Endogenous Ang-1 mRNA levels immediately decreased in response to cardiotoxin then increased during the 2 wk. Ang-1 protein was expressed in satellite cells, both in noninjured and recovering TA muscles. Positive Ang-1 staining was present in blood vessels but not in nerve fibers. Four days after the initiation of injury, injection of adenoviral Ang-1 into injured muscles resulted in significant increases in in situ TA muscle contractility, muscle fiber regeneration, and capillary density. In cultured human skeletal myoblasts, recombinant Ang-1 protein increased survival, proliferation, migration, and differentiation into myotubes. The latter effect was associated with significant upregulation of the expression of the myogenic regulatory factors MyoD and Myogenin and certain genes involved in cell cycle regulation. We conclude that Ang-1 strongly enhances skeletal muscle regeneration in response to fiber injury and that this effect is mediated through induction of the myogenesis program in muscle progenitor cells and the angiogenesis program in endothelial cells. PMID:25608750

  5. Enhanced Functional Recovery in MRL/MpJ Mice after Spinal Cord Dorsal Hemisection

    PubMed Central

    Thuret, Sandrine; Thallmair, Michaela; Horky, Laura L.; Gage, Fred H.

    2012-01-01

    Adult MRL/MpJ mice have been shown to possess unique regeneration capabilities. They are able to heal an ear-punched hole or an injured heart with normal tissue architecture and without scar formation. Here we present functional and histological evidence for enhanced recovery following spinal cord injury (SCI) in MRL/MpJ mice. A control group (C57BL/6 mice) and MRL/MpJ mice underwent a dorsal hemisection at T9 (thoracic vertebra 9). Our data show that MRL/MpJ mice recovered motor function significantly faster and more completely. We observed enhanced regeneration of the corticospinal tract (CST). Furthermore, we observed a reduced astrocytic response and fewer micro-cavities at the injury site, which appear to create a more growth-permissive environment for the injured axons. Our data suggest that the reduced astrocytic response is in part due to a lower lesion-induced increase of cell proliferation post-SCI, and a reduced astrocytic differentiation of the proliferating cells. Interestingly, we also found an increased number of proliferating microglia, which could be involved in the MRL/MpJ spinal cord repair mechanisms. Finally, to evaluate the molecular basis of faster spinal cord repair, we examined the difference in gene expression changes in MRL/MpJ and C57BL/6 mice after SCI. Our microarray data support our histological findings and reveal a transcriptional profile associated with a more efficient spinal cord repair in MRL/MpJ mice. PMID:22348029

  6. Aberrant Calreticulin Expression in Articular Cartilage of Dio2 Deficient Mice

    PubMed Central

    Bomer, Nils; Cornelis, Frederique M. F.; Ramos, Yolande F. M.; den Hollander, Wouter; Lakenberg, Nico; van der Breggen, Ruud; Storms, Lies; Slagboom, P. Eline; Lories, Rik J. U.; Meulenbelt, Ingrid

    2016-01-01

    Objective To identify intrinsic differences in cartilage gene expression profiles between wild-type- and Dio2-/--mice, as a mechanism to investigate factors that contribute to prolonged healthy tissue homeostasis. Methods Previously generated microarray-data (Illumina MouseWG-6 v2) of knee cartilage of wild-type and Dio2 -/- -mice were re-analyzed to identify differential expressed genes independent of mechanical loading conditions by forced treadmill-running. RT-qPCR and western blot analyses of overexpression and knockdown of Calr in mouse chondro-progenitor cells (ATDC5) were applied to assess the direct effect of differential Calr expression on cartilage deposition. Results Differential expression analyses of articular cartilage of Dio2-/- (N = 9) and wild-type-mice (N = 11) while applying a cutoff threshold (P < 0.05 (FDR) and FC > |1,5|) resulted in 1 probe located in Calreticulin (Calr) that was found significantly downregulated in Dio2-/- mice (FC = -1.731; P = 0.044). Furthermore, overexpression of Calr during early chondrogenesis in ATDC5 cells leads to decreased proteoglycan deposition and corresponding lower Aggrecan expression, whereas knocking down Calr expression does not lead to histological differences of matrix composition. Conclusion We here demonstrate that the beneficial homeostatic state of articular cartilage in Dio2-/- mice is accompanied with significant lower expression of Calr. Functional analyses further showed that upregulation of Calr expression could act as an initiator of cartilage destruction. The consistent association between Calr and Dio2 expression suggests that enhanced expression of these genes facilitate detrimental effects on cartilage integrity. PMID:27163789

  7. Immune enhancement during chronic ethanol feeding in mice - Autoimmune phenomena

    SciTech Connect

    Honchel, R.; Rhoads, C.A.; Fitzpatrick, E.A.; McClain, C.J.; Kaplan, A.M.; Cohen, D.A. )

    1991-03-11

    Chronic alcohol abuse in humans is often associated with diminished immune reactivity and enhanced susceptibility to infections. However, many alcohol-dependent individuals display signs of autoimmunity, which has been implicated in alcohol-associated liver damage. This study demonstrates that C57Bl/6 mice placed on the Lieber-DeCarli liquid ethanol diet for up to 9 weeks displayed augmented immune reactivity as compared to mice placed on an isocaloric control diet. Spleen cells were significantly more responsive to the mitogens, LPS and ConA, as early as 3 weeks after initiation of EtOH feeding and this hyperresponsiveness persisted throughout the 9 week feeding period. Similar enhancement of the mixed lymphocyte response was also seen in EtOH fed mice. The enhancement of immune responsiveness was not related to a change in the numbers or percentages of B cells, T cells, or in the CD4/CD8 T cell ratios as determined by flow cytometry. These studies indicate that under certain conditions of ethanol feeding in mice, enhancement rather than suppression of the immune system may occur. This system may be a model to evaluate possible induction of autoimmune responses during chronic ethanol abuse. Studies are underway to measure the presence of auto-antibodies in the sera of these ethanol fed mice.

  8. Expression of a Chimeric Antigen Receptor in Multiple Leukocyte Lineages in Transgenic Mice.

    PubMed

    Yong, Carmen S M; Westwood, Jennifer A; Schröder, Jan; Papenfuss, Anthony T; von Scheidt, Bianca; Moeller, Maria; Devaud, Christel; Darcy, Phillip K; Kershaw, Michael H

    2015-01-01

    Genetically modified CD8+ T lymphocytes have shown significant anti-tumor effects in the adoptive immunotherapy of cancer, with recent studies highlighting a potential role for a combination of other immune subsets to enhance these results. However, limitations in present genetic modification techniques impose difficulties in our ability to fully explore the potential of various T cell subsets and assess the potential of other leukocytes armed with chimeric antigen receptors (CARs). To address this issue, we generated a transgenic mouse model using a pan-hematopoietic promoter (vav) to drive the expression of a CAR specific for a tumor antigen. Here we present a characterization of the immune cell compartment in two unique vav-CAR transgenic mice models, Founder 9 (F9) and Founder 38 (F38). We demonstrate the vav promoter is indeed capable of driving the expression of a CAR in cells from both myeloid and lymphoid lineage, however the highest level of expression was observed in T lymphocytes from F38 mice. Lymphoid organs in vav-CAR mice were smaller and had reduced cell numbers compared to the wild type (WT) controls. Furthermore, the immune composition of F9 mice differed greatly with a significant reduction in lymphocytes found in the thymus, lymph node and spleen of these mice. To gain insight into the altered immune phenotype of F9 mice, we determined the chromosomal integration site of the transgene in both mouse strains using whole genome sequencing (WGS). We demonstrated that compared to the 7 copies found in F38 mice, F9 mice harbored almost 270 copies. These novel vav-CAR models provide a ready source of CAR expressing myeloid and lymphoid cells and will aid in facilitating future experiments to delineate the role for other leukocytes for adoptive immunotherapy against cancer. PMID:26505904

  9. Expression of a Chimeric Antigen Receptor in Multiple Leukocyte Lineages in Transgenic Mice

    PubMed Central

    Yong, Carmen S. M.; Westwood, Jennifer A.; Schröder, Jan; Papenfuss, Anthony T.; von Scheidt, Bianca; Moeller, Maria; Devaud, Christel

    2015-01-01

    Genetically modified CD8+ T lymphocytes have shown significant anti-tumor effects in the adoptive immunotherapy of cancer, with recent studies highlighting a potential role for a combination of other immune subsets to enhance these results. However, limitations in present genetic modification techniques impose difficulties in our ability to fully explore the potential of various T cell subsets and assess the potential of other leukocytes armed with chimeric antigen receptors (CARs). To address this issue, we generated a transgenic mouse model using a pan-hematopoietic promoter (vav) to drive the expression of a CAR specific for a tumor antigen. Here we present a characterization of the immune cell compartment in two unique vav-CAR transgenic mice models, Founder 9 (F9) and Founder 38 (F38). We demonstrate the vav promoter is indeed capable of driving the expression of a CAR in cells from both myeloid and lymphoid lineage, however the highest level of expression was observed in T lymphocytes from F38 mice. Lymphoid organs in vav-CAR mice were smaller and had reduced cell numbers compared to the wild type (WT) controls. Furthermore, the immune composition of F9 mice differed greatly with a significant reduction in lymphocytes found in the thymus, lymph node and spleen of these mice. To gain insight into the altered immune phenotype of F9 mice, we determined the chromosomal integration site of the transgene in both mouse strains using whole genome sequencing (WGS). We demonstrated that compared to the 7 copies found in F38 mice, F9 mice harbored almost 270 copies. These novel vav-CAR models provide a ready source of CAR expressing myeloid and lymphoid cells and will aid in facilitating future experiments to delineate the role for other leukocytes for adoptive immunotherapy against cancer. PMID:26505904

  10. Microarray expression profiling identifies genes with altered expression in HDL-deficient mice

    SciTech Connect

    Callow, Matthew J.; Dudoit, Sandrine; Gong, Elaine L.; Speed, Terence P.; Rubin, Edward M.

    2000-05-05

    Based on the assumption that severe alterations in the expression of genes known to be involved in HDL metabolism may affect the expression of other genes we screened an array of over 5000 mouse expressed sequence tags (ESTs) for altered gene expression in the livers of two lines of mice with dramatic decreases in HDL plasma concentrations. Labeled cDNA from livers of apolipoprotein AI (apo AI) knockout mice, Scavenger Receptor BI (SR-BI) transgenic mice and control mice were co-hybridized to microarrays. Two-sample t-statistics were used to identify genes with altered expression levels in the knockout or transgenic mice compared with the control mice. In the SR-BI group we found 9 array elements representing at least 5 genes to be significantly altered on the basis of an adjusted p value of less than 0.05. In the apo AI knockout group 8 array elements representing 4 genes were altered compared with the control group (p < 0.05). Several of the genes identified in the SR-BI transgenic suggest altered sterol metabolism and oxidative processes. These studies illustrate the use of multiple-testing methods for the identification of genes with altered expression in replicated microarray experiments of apo AI knockout and SR-BI transgenic mice.

  11. Electroacupuncture Upregulates SIRT1-Dependent PGC-1α Expression in SAMP8 Mice

    PubMed Central

    Dong, Weiguo; Guo, Wanqing; Wang, Feng; Li, Changzheng; Xie, Yongcai; Zheng, Xueha; Shi, Hong

    2015-01-01

    Background Abnormalities of brain energy metabolism are involved in Alzheimer disease (AD). Sirtuin 1 (SIRT1) is a class III histone deacetylase and activates peroxisome proliferator-activated receptor-γ co-activator-1α (PGC-1α), which enhances mitochondrial biogenesis and energy homeostasis. Electroacupuncture (EA) has been reported to improve brain energy metabolism in AD. However, the effect of EA on SIRT1 and PGC-1α in AD remains unclear. Material/Methods ATP levels were measured using assay kits in the hippocampus and frontal cortex of senescence-accelerated mouse prone 8 (SAMP8) mice. Western blotting analysis and quantitative real-time RT-PCR were performed to measure the expression of SIRT1 and PGC-1α in the hippocampus of SAMP8 mice. PGC-1α acetylation was analyzed using immunoprecipitation. Results Compared with senescence-accelerated resistant mice 1 (SAMR1) mice, SAMP8 mice had a decline in ATP levels and the expression of SIRT1 and PGC-1α. EA treatment improved ATP levels, upregulated the expression of SIRT1 and PGC-1α, and decreased PGC-1α acetylation. Conclusions These data suggest that EA improved brain energy metabolism, potentially associated with the upregulation of SIRT1-dependent PGC-1α expression. PMID:26530101

  12. Mice Expressing RHAG and RHD Human Blood Group Genes

    PubMed Central

    Goossens, Dominique; da Silva, Nelly; Metral, Sylvain; Cortes, Ulrich; Callebaut, Isabelle; Picot, Julien; Mouro-Chanteloup, Isabelle; Cartron, Jean-Pierre

    2013-01-01

    Anti-RhD prophylaxis of haemolytic disease of the fetus and newborn (HDFN) is highly effective, but as the suppressive mechanism remains uncertain, a mouse model would be of interest. Here we have generated transgenic mice expressing human RhAG and RhD erythrocyte membrane proteins in the presence and, for human RhAG, in the absence, of mouse Rhag. Human RhAG associates with mouse Rh but not mouse Rhag on red blood cells. In Rhag knockout mice transgenic for human RHAG, the mouse Rh protein is “rescued” (re-expressed), and co-immunoprecipitates with human RhAG, indicating the presence of hetero-complexes which associate mouse and human proteins. RhD antigen was expressed from a human RHD gene on a BAC or from RHD cDNA under control of β-globin regulatory elements. RhD was never observed alone, strongly indicative that its expression absolutely depends on the presence of transgenic human RhAG. This first expression of RhD in mice is an important step in the creation of a mouse model of RhD allo-immunisation and HDFN, in conjunction with the Rh-Rhag knockout mice we have developed previously. PMID:24260394

  13. Mice expressing RHAG and RHD human blood group genes.

    PubMed

    Goossens, Dominique; da Silva, Nelly; Metral, Sylvain; Cortes, Ulrich; Callebaut, Isabelle; Picot, Julien; Mouro-Chanteloup, Isabelle; Cartron, Jean-Pierre

    2013-01-01

    Anti-RhD prophylaxis of haemolytic disease of the fetus and newborn (HDFN) is highly effective, but as the suppressive mechanism remains uncertain, a mouse model would be of interest. Here we have generated transgenic mice expressing human RhAG and RhD erythrocyte membrane proteins in the presence and, for human RhAG, in the absence, of mouse Rhag. Human RhAG associates with mouse Rh but not mouse Rhag on red blood cells. In Rhag knockout mice transgenic for human RHAG, the mouse Rh protein is "rescued" (re-expressed), and co-immunoprecipitates with human RhAG, indicating the presence of hetero-complexes which associate mouse and human proteins. RhD antigen was expressed from a human RHD gene on a BAC or from RHD cDNA under control of β-globin regulatory elements. RhD was never observed alone, strongly indicative that its expression absolutely depends on the presence of transgenic human RhAG. This first expression of RhD in mice is an important step in the creation of a mouse model of RhD allo-immunisation and HDFN, in conjunction with the Rh-Rhag knockout mice we have developed previously. PMID:24260394

  14. CD40L expressed from the canarypox vector, ALVAC, can boost immunogenicity of HIV-1 canarypox vaccine in mice and enhance the in-vitro expansion of viral specific CD8+ T cell memory responses from HIV-1-infected and HIV-1-uninfected individuals

    PubMed Central

    Liu, Jun; Yu, Qigui; Stone, Geoffrey W.; Yue, Feng Yun; Ngai, Nicholas; Jones, R. Brad; Kornbluth, Richard S.; Ostrowski, Mario A.

    2011-01-01

    Summary Human immunodeficiency virus-1 (HIV-1) canarypox vaccines are safe but poorly immunogenic. CD40 ligand (CD40L), a member of the tumor necrosis factor superfamily (TNFSF), is a pivotal co-stimulatory molecule for immune responses. To explore whether CD40L can be used as an adjuvant for HIV-1 canarypox vaccine, we constructed recombinant canarypox viruses expressing CD40L. Co-immunization of mice with CD40L expressing canarypox and the canarypox vaccine expressing HIV-1 proteins, vCP1452, augmented HIV-1 specific cytotoxic T lymphocyte (CTL) responses in terms of frequency, polyfunctionality and interleukin (IL)-7 receptor α chain (IL-7Rα, CD127) expression. In addition, CD40L expressed from canarypox virus could significantly augment CD4+ T cell responses against HIV-1 in mice. CD40L expressed from canarypox virus matured human monocyte-derived dendritic cells (MDDCs) in a tumor necrosis factor α (TNF-α) independent manner, which underwent less apoptosis, and could expand ex vivo Epstein-Barr virus (EBV)-specific CTL responses from healthy human individuals and ex vivo HIV-1-specific CTL responses from HIV-1-infected individuals in the presence or absence of CD4+ T cells. Taken together, our results suggest that CD40L incorporation into poxvirus vectors could be used as a strategy to enhance their immunogenicity. PMID:18562053

  15. Changes in gene expression associated with FTO overexpression in mice.

    PubMed

    Merkestein, Myrte; McTaggart, James S; Lee, Sheena; Kramer, Holger B; McMurray, Fiona; Lafond, Mathilde; Boutens, Lily; Cox, Roger; Ashcroft, Frances M

    2014-01-01

    Single nucleotide polymorphisms in the first intron of the fat-mass-and-obesity-related gene FTO are associated with increased body weight and adiposity. Increased expression of FTO is likely underlying this obesity phenotype, as mice with two additional copies of Fto (FTO-4 mice) exhibit increased adiposity and are hyperphagic. FTO is a demethylase of single stranded DNA and RNA, and one of its targets is the m6A modification in RNA, which might play a role in the regulation of gene expression. In this study, we aimed to examine the changes in gene expression that occur in FTO-4 mice in order to gain more insight into the underlying mechanisms by which FTO influences body weight and adiposity. Our results indicate an upregulation of anabolic pathways and a downregulation of catabolic pathways in FTO-4 mice. Interestingly, although genes involved in methylation were differentially regulated in skeletal muscle of FTO-4 mice, no effect of FTO overexpression on m6A methylation of total mRNA was detected. PMID:24842286

  16. Changes in Gene Expression Associated with FTO Overexpression in Mice

    PubMed Central

    Kramer, Holger B.; McMurray, Fiona; Lafond, Mathilde; Boutens, Lily; Cox, Roger; Ashcroft, Frances M.

    2014-01-01

    Single nucleotide polymorphisms in the first intron of the fat-mass-and-obesity-related gene FTO are associated with increased body weight and adiposity. Increased expression of FTO is likely underlying this obesity phenotype, as mice with two additional copies of Fto (FTO-4 mice) exhibit increased adiposity and are hyperphagic. FTO is a demethylase of single stranded DNA and RNA, and one of its targets is the m6A modification in RNA, which might play a role in the regulation of gene expression. In this study, we aimed to examine the changes in gene expression that occur in FTO-4 mice in order to gain more insight into the underlying mechanisms by which FTO influences body weight and adiposity. Our results indicate an upregulation of anabolic pathways and a downregulation of catabolic pathways in FTO-4 mice. Interestingly, although genes involved in methylation were differentially regulated in skeletal muscle of FTO-4 mice, no effect of FTO overexpression on m6A methylation of total mRNA was detected. PMID:24842286

  17. Partially Redundant Enhancers Cooperatively Maintain Mammalian Pomc Expression Above a Critical Functional Threshold

    PubMed Central

    Lam, Daniel D.; de Souza, Flavio S. J.; Nasif, Sofia; Yamashita, Miho; López-Leal, Rodrigo; Meece, Kana; Sampath, Harini; Mercer, Aaron J.; Wardlaw, Sharon L.

    2015-01-01

    Cell-specific expression of many genes is conveyed by multiple enhancers, with each individual enhancer controlling a particular expression domain. In contrast, multiple enhancers drive similar expression patterns of some genes involved in embryonic development, suggesting regulatory redundancy. Work in Drosophila has indicated that functionally overlapping enhancers canalize development by buffering gene expression against environmental and genetic disturbances. However, little is known about regulatory redundancy in vertebrates and in genes mainly expressed during adulthood. Here we study nPE1 and nPE2, two phylogenetically conserved mammalian enhancers that drive expression of the proopiomelanocortin gene (Pomc) to the same set of hypothalamic neurons. The simultaneous deletion of both enhancers abolished Pomc expression at all ages and induced a profound metabolic dysfunction including early-onset extreme obesity. Targeted inactivation of either nPE1 or nPE2 led to very low levels of Pomc expression during early embryonic development indicating that both enhancers function synergistically. In adult mice, however, Pomc expression is controlled additively by both enhancers, with nPE1 being responsible for ∼80% and nPE2 for ∼20% of Pomc transcription. Consequently, nPE1 knockout mice exhibit mild obesity whereas nPE2-deficient mice maintain a normal body weight. These results suggest that nPE2-driven Pomc expression is compensated by nPE1 at later stages of development, essentially rescuing the earlier phenotype of nPE2 deficiency. Together, these results reveal that cooperative interactions between the enhancers confer robustness of Pomc expression against gene regulatory disturbances and preclude deleterious metabolic phenotypes caused by Pomc deficiency in adulthood. Thus, our study demonstrates that enhancer redundancy can be used by genes that control adult physiology in mammals and underlines the potential significance of regulatory sequence mutations in

  18. Enhanced resistance against Listeria monocytogenes at an early phase of primary infection in pregnant mice: activation of macrophages during pregnancy.

    PubMed Central

    Watanabe, Y; Mitsuyama, M; Sano, M; Nakano, H; Nomoto, K

    1986-01-01

    We investigated the pregnancy-induced changes in macrophage activity which are important in the expression of host defense against infections. Several macrophage functions were examined by using Listeria monocytogenes. In pregnant mice, prolonged survival and enhanced in vivo elimination of bacteria were observed in the early phase of primary infection. Functions of peritoneal macrophages, including in vitro phagocytosis intracellular killing, glucose consumption, generation of superoxide anion, and intracellular beta-glucuronidase activity were shown to be enhanced in pregnant mice. These findings indicate that pregnancy enhances macrophage functions qualitatively. Possible mechanisms for this enhancement and the significance of macrophage activation for pregnant hosts are discussed. PMID:3011673

  19. Self-Exposure to the Male Pheromone ESP1 Enhances Male Aggressiveness in Mice.

    PubMed

    Hattori, Tatsuya; Osakada, Takuya; Matsumoto, Ayaka; Matsuo, Naoki; Haga-Yamanaka, Sachiko; Nishida, Takaya; Mori, Yuji; Mogi, Kazutaka; Touhara, Kazushige; Kikusui, Takefumi

    2016-05-01

    Exocrine gland-secreting peptide 1 (ESP1) released into male tear fluids is a male pheromone that stimulates sexually receptive behavior in female mice via the vomeronasal sensory system. ESP1 also induces c-Fos expression in male brain regions distinct from those in females. However, behavior in males following ESP1 exposure has not been examined. In the present study, we show that ESP1, in conjunction with unfamiliar male urine, enhances male aggression via the specific vomeronasal receptor V2Rp5. In addition, male mice that secrete ESP1 but lack V2Rp5 exhibit a lower level of aggressiveness than do mice that express V2Rp5. These results suggest that ESP1 not only acts as a male pheromone in both sexes but also serves as an auto-stimulatory factor that enhances male aggressiveness by self-exposure. Finally, re-activation of ESP1-induced c-Fos-positive neurons by using the designer receptor exclusively activated by designer drug (DREADD) approach resulted in enhancement of sexual and aggressive behaviors in female and male mice, respectively, indicating that sexually dimorphic activation in the brain is a neural basis for the sex-specific behavioral responses to ESP1. PMID:27151664

  20. Inhibition of estrogen biosynthesis enhances lymphoma growth in mice

    PubMed Central

    Talaber, Gergely; Yakimchuk, Konstantin; Guan, Jiyu; Inzunza, Jose; Okret, Sam

    2016-01-01

    Most lymphomas show higher incidence and poorer prognosis in males compared to females. However, the endocrine contribution to this gender difference is not entirely known. Here we show that castration accelerates lymphoma growth in C57BL6 male mice grafted with murine EG7 T cell lymphoma cells. However, the androgen receptor antagonist Bicalutamide did not affect lymphoma growth, suggesting no impact of androgen receptor signaling on lymphoma progression. In contrast, inhibition of androgen-to-estrogen conversion by the aromatase inhibitor (AI) Letrozole induced faster lymphoma growth in mice, suggesting that androgens impact lymphoma growth through its conversion to estrogens. This was supported by the inability of dihydrotestosterone, which is not converted to estrogens by aromatase, to influence lymphoma growth in castrated male mice. Lymphoma growth was also stimulated in immunocompromised mice grafted with human B cell lymphoma (Granta-519) and treated with either reversible or irreversible AIs, showing that the blockage of estrogen synthesis caused enhanced growth of both murine T and human B cell lymphomas and with different AIs. Additionally, AI-treated EG7 lymphomas showed accelerated growth not only in male but also in intact female mice. Altogether, our results demonstrate that aromatase inhibition accelerates lymphoma growth but not androgens per se, highlighting a protective role of estrogens in lymphoma pathogenesis. These results also raise concern that the use of AIs in women with breast cancer might enhance lymphoma progression. PMID:26943574

  1. Hepatic and renal Bcrp transporter expression in mice treated with perfluorooctanoic acid

    PubMed Central

    Eldasher, Lobna M.; Wen, Xia; Little, Michael S.; Bircsak, Kristin M.; Yacovino, Lindsay L.; Aleksunes, Lauren M.

    2013-01-01

    The breast cancer resistance protein (Bcrp) is an efflux transporter that participates in the biliary and renal excretion of drugs and environmental chemicals. Recent evidence suggests that pharmacological activation of the peroxisome proliferator activated receptor alpha (PPARα) can up-regulate the hepatic expression of Bcrp. The current study investigated the regulation of hepatic and renal Bcrp mRNA and protein in mice treated with the PPARα agonist perfluorooctanoic acid (PFOA) and the ability of PFOA to alter human BCRP function in vitro. Bcrp mRNA and protein expression were quantified in the livers and kidneys of male C57BL/6 mice treated with vehicle or PFOA (1 or 3 mg/kg/day oral gavage) for 7 days. PFOA treatment increased liver weights as well as the hepatic mRNA and protein expression of the PPARα target gene, cytochrome P450 4a14. Compared to vehicle-treated control mice, PFOA increased hepatic Bcrp mRNA and protein between 1.5- and 3-fold. Immunofluorescent staining confirmed enhanced canalicular Bcrp staining in liver sections from PFOA-treated mice. The kidney expression of cytochrome P450 4a14 mRNA, but not Bcrp, was increased in mice treated with PFOA. Micromolar concentrations of PFOA decreased human BCRP ATPase activity and inhibited BCRP-mediated transport in inverted membrane vesicles. Together, these studies demonstrate that PFOA induces hepatic Bcrp expression in mice and may inhibit human BCRP transporter function at concentrations that exceed levels observed in humans. PMID:23435180

  2. Memory-enhancing effects of Cuscuta japonica Choisy via enhancement of adult hippocampal neurogenesis in mice.

    PubMed

    Moon, Minho; Jeong, Hyun Uk; Choi, Jin Gyu; Jeon, Seong Gak; Song, Eun Ji; Hong, Seon-Pyo; Oh, Myung Sook

    2016-09-15

    It is generally accepted that functional and structural changes within the hippocampus are involved in learning and memory and that adult neurogenesis in this region may modulate cognition. The extract of Cuscuta japonica Choisy (CJ) is a well-known traditional Chinese herbal medicine that has been used since ancient times as a rejuvenation remedy. The systemic effects of this herb are widely known and can be applied for the treatment of a number of physiological diseases, but there is a lack of evidence describing its effects on brain function. Thus, the present study investigated whether CJ would enhance memory function and/or increase hippocampal neurogenesis using mice orally administered with CJ water extract or vehicle for 21days. Performance on the novel object recognition and passive avoidance tests revealed that treatment with CJ dose-dependently improved the cognitive function of mice. Additionally, CJ increased the Ki-67-positive proliferating cells and the number of doublecortin-stained neuroblasts in the dentate gyrus (DG) of the hippocampus, and double labeling with 5-bromo-2-deoxyuridine and neuronal specific nuclear protein showed that CJ increased the number of mature neurons in the DG. Finally, CJ resulted in the upregulated expression of neurogenic differentiation factor, which is essential for the maturation and differentiation of granule cells in the hippocampus. Taken together, the present findings indicate that CJ stimulated neuronal cell proliferation, differentiation, and maturation, which are all processes associated with neurogenesis. Additionally, these findings suggest that CJ may improve learning and memory via the enhancement of adult hippocampal neurogenesis. PMID:27185736

  3. The effect of centaurein on interferon-{gamma} expression and Listeria infection in mice

    SciTech Connect

    Chang, S.-L.; Yeh, H.-H.; Lin, Y.-S.; Chiang, Y.-M.; Wu, T.-K. . E-mail: tkwmll@mail.nctu.edu.tw; Yang, W.-C. . E-mail: wcyang@gate.sinica.edu.tw

    2007-02-15

    We previously found that centaurein enhanced IFN-{gamma} transcription in T cells. Here, we demonstrate that centaurein increased the IFN-{gamma} expression in T and NK cells and the serum IFN-{gamma} level in mice. Centaurein elevated the transcription of T-bet but not GATA-3, which is consistent with its effect on that of IFN-{gamma} but not IL-4. Additionally, centaurein effectively protected mice against Listeria infection. Moreover, centaurein per se or in combination with antibiotics could treat Listeria infection. Our mechanistic studies suggest that centaurein augments IFN-{gamma} expression via a transcriptional up-regulation of T-bet and that centaurein protects against or treats Listeria infection via a modulation of IFN-{gamma} expression.

  4. Increase of Calcium Sensing Receptor Expression Is Related to Compensatory Insulin Secretion during Aging in Mice

    PubMed Central

    Oh, Yoon Sin; Seo, Eun-Hui; Lee, Young-Sun; Cho, Sung Chun; Jung, Hye Seung; Park, Sang Chul; Jun, Hee-Sook

    2016-01-01

    Type 2 diabetes is caused by both insulin resistance and relative insulin deficiency. To investigate age-related changes in glucose metabolism and development of type 2 diabetes, we compared glucose homeostasis in different groups of C57BL/6J mice ranging in age from 4 months to 20 months (4, 8, 12, 16 and 20 months). Interestingly, we observed that non-fasting glucose levels were not significantly changed, but glucose tolerance gradually increased by 20 months of age, whereas insulin sensitivity declined with age. We found that the size of islets and glucose-stimulated insulin secretion increased with aging. However, mRNA expression of pancreatic and duodenal homeobox 1 and granuphilin was decreased in islets of older mice compared with that of 4-month-old mice. Serum calcium (Ca2+) levels were significantly decreased at 12, 20 and 28 months of age compared with 4 months and calcium sensing receptor (CaSR) mRNA expression in the islets significantly increased with age. An extracellular calcium depletion agent upregulated CaSR mRNA expression and consequently enhanced insulin secretion in INS-1 cells and mouse islets. In conclusion, we suggest that decreased Ca2+ levels and increased CaSR expression might be involved in increased insulin secretion to compensate for insulin resistance in aged mice. PMID:27441644

  5. Enhanced spontaneous locomotor activity in bovine GH transgenic mice involves peripheral mechanisms.

    PubMed

    Bohlooly-Y, M; Olsson, B; Gritli-Linde, A; Brusehed, O; Isaksson, O G; Ohlsson, C; Söderpalm, B; Törnell, J; Ola, B

    2001-10-01

    Clinical and experimental studies indicate a role for GH in mechanisms related to anhedonia/hedonia, psychic energy, and reward. Recently we showed that transgenic mice with general overexpression of bovine GH display increased spontaneous locomotor activity. In the present study, we investigated whether this behavioral change is owing to a direct action of GH in the central nervous system or to peripheral GH actions. A transgenic construct, containing the glial fibrillary acidic protein promoter directing specific expression of bovine GH to the central nervous system, was designed. The central nervous system-specific expression of bovine GH in the glial fibrillary acidic protein-bovine GH transgenic mice was confirmed, but no effect on spontaneous locomotor activity was observed. Serum bovine GH levels were increased in glial fibrillary acidic protein-bovine GH transgenic mice but clearly lower than in transgenic mice with general overexpression of bovine GH. In contrast to the transgenic mice with general overexpression of bovine GH, glial fibrillary acidic protein-bovine GH mice did not display any difference in serum IGF-I levels. The levels of free T(3) and the conversion of the free T(4) to free T(3) were only increased in transgenic mice with general overexpression of bovine GH, but serum corticosterone levels were similarly increased in both transgenic models. These results suggest that free T(3) and/or IGF-I, affecting dopamine and serotonin systems in the central nervous system, may mediate the enhanced locomotor activity observed in transgenic mice with general overexpression of bovine GH. PMID:11564723

  6. TLR9 expression is required for the development of cigarette smoke-induced emphysema in mice.

    PubMed

    Foronjy, Robert F; Salathe, Matthias A; Dabo, Abdoulaye J; Baumlin, Nathalie; Cummins, Neville; Eden, Edward; Geraghty, Patrick

    2016-07-01

    The expression of Toll-like receptor (TLR)-9, a pathogen recognition receptor that recognizes unmethylated CpG sequences in microbial DNA molecules, is linked to the pathogenesis of several lung diseases. TLR9 expression and signaling was investigated in animal and cell models of chronic obstructive pulmonary disease (COPD). We observed enhanced TLR9 expression in mouse lungs following exposure to cigarette smoke. Tlr9(-/-) mice were resistant to cigarette smoke-induced loss of lung function as determined by mean linear intercept, total lung capacity, lung compliance, and tissue elastance analysis. Tlr9 expression also regulated smoke-mediated immune cell recruitment to the lung; apoptosis; expression of granulocyte-colony stimulating factor (G-CSF), the CXCL5 protein, and matrix metalloproteinase-2 (MMP-2); and protein tyrosine phosphatase 1B (PTP1B) activity in the lung. PTP1B, a phosphatase with anti-inflammatory abilities, was identified as binding to TLR9. In vivo delivery of a TLR9 agonist enhanced TLR9 binding to PTP1B, which inactivated PTP1B. Ptp1b(-/-) mice had elevated lung concentrations of G-CSF, CXCL5, and MMP-2, and tissue expression of type-1 interferon following TLR9 agonist administration, compared with wild-type mice. TLR9 responses were further determined in fully differentiated normal human bronchial epithelial (NHBE) cells isolated from nonsmoker, smoker, and COPD donors, and then cultured at air liquid interface. NHBE cells from smokers and patients with COPD expressed more TLR9 and secreted greater levels of G-CSF, IL-6, CXCL5, IL-1β, and MMP-2 upon TLR9 ligand stimulation compared with cells from nonsmoker donors. Although TLR9 combats infection, our results indicate that TLR9 induction can affect lung function by inactivating PTP1B and upregulating expression of proinflammatory cytokines. PMID:27288485

  7. Severe iron deficiency anemia in transgenic mice expressing liver hepcidin.

    PubMed

    Nicolas, Gaël; Bennoun, Myriam; Porteu, Arlette; Mativet, Sandrine; Beaumont, Carole; Grandchamp, Bernard; Sirito, Mario; Sawadogo, Michèle; Kahn, Axel; Vaulont, Sophie

    2002-04-01

    We recently reported the hemochromatosis-like phenotype observed in our Usf2 knockout mice. In these mice, as in murine models of hemochromatosis and patients with hereditary hemochromatosis, iron accumulates in parenchymal cells (in particular, liver and pancreas), whereas the reticuloendothelial system is spared from this iron loading. We suggested that this phenotypic trait could be attributed to the absence, in the Usf2 knockout mice, of a secreted liver-specific peptide, hepcidin. We conjectured that the reverse situation, namely overexpression of hepcidin, might result in phenotypic traits of iron deficiency. This question was addressed by generating transgenic mice expressing hepcidin under the control of the liver-specific transthyretin promoter. We found that the majority of the transgenic mice were born with a pale skin and died within a few hours after birth. These transgenic animals had decreased body iron levels and presented severe microcytic hypochromic anemia. So far, three mosaic transgenic animals have survived. They were unequivocally identified by physical features, including reduced body size, pallor, hairless and crumpled skin. These pleiotropic effects were found to be associated with erythrocyte abnormalities, with marked anisocytosis, poikylocytosis and hypochromia, which are features characteristic of iron-deficiency anemia. These results strongly support the proposed role of hepcidin as a putative iron-regulatory hormone. The animal models devoid of hepcidin (the Usf2 knockout mice) or overexpressing the peptide (the transgenic mice presented in this paper) represent valuable tools for investigating iron homeostasis in vivo and for deciphering the molecular mechanisms of hepcidin action. PMID:11930010

  8. Severe iron deficiency anemia in transgenic mice expressing liver hepcidin

    PubMed Central

    Nicolas, Gaël; Bennoun, Myriam; Porteu, Arlette; Mativet, Sandrine; Beaumont, Carole; Grandchamp, Bernard; Sirito, Mario; Sawadogo, Michèle; Kahn, Axel; Vaulont, Sophie

    2002-01-01

    We recently reported the hemochromatosis-like phenotype observed in our Usf2 knockout mice. In these mice, as in murine models of hemochromatosis and patients with hereditary hemochromatosis, iron accumulates in parenchymal cells (in particular, liver and pancreas), whereas the reticuloendothelial system is spared from this iron loading. We suggested that this phenotypic trait could be attributed to the absence, in the Usf2 knockout mice, of a secreted liver-specific peptide, hepcidin. We conjectured that the reverse situation, namely overexpression of hepcidin, might result in phenotypic traits of iron deficiency. This question was addressed by generating transgenic mice expressing hepcidin under the control of the liver-specific transthyretin promoter. We found that the majority of the transgenic mice were born with a pale skin and died within a few hours after birth. These transgenic animals had decreased body iron levels and presented severe microcytic hypochromic anemia. So far, three mosaic transgenic animals have survived. They were unequivocally identified by physical features, including reduced body size, pallor, hairless and crumpled skin. These pleiotropic effects were found to be associated with erythrocyte abnormalities, with marked anisocytosis, poikylocytosis and hypochromia, which are features characteristic of iron-deficiency anemia. These results strongly support the proposed role of hepcidin as a putative iron-regulatory hormone. The animal models devoid of hepcidin (the Usf2 knockout mice) or overexpressing the peptide (the transgenic mice presented in this paper) represent valuable tools for investigating iron homeostasis in vivo and for deciphering the molecular mechanisms of hepcidin action. PMID:11930010

  9. Enhanced penetration of exogenous EPCs into brains of APP/PS1 transgenic mice

    PubMed Central

    Yuan, Xiaoyang; Mei, Bin; Zhang, Le; Zhang, Cuntai; Zheng, Miao; Liang, Huifang; Wang, Wei; Zheng, Jie; Ding, Ling; Zheng, Kai

    2016-01-01

    The aim of this study was to investigate the repair function of exogenous Endothelial progenitor cells (EPCs) for brain microvascular damage of the APP/PS1 transgenic mouse model of Alzheimer’s disease (AD). This study used a density-gradient centrifugation method to isolate mononuclear cells (MNCs) from mouse bone marrow, which were subsequently seeded and cultured. Cells were characterized by morphology and detection of the surface markers CD34 and CD133 at different time points by immunofluorescence (IF) and flow cytometry (FCM). Then, EPCs were transfected with GFP adenoviral vectors (GFP-EPCs). Wild-type (WT) and APP/PS1 transgenic mice both received GFP-EPCs injection through the tail vein, and using a PBS buffer injection as the control. Seven days later, the animals’ brain tissue was isolated. Expression of GFP was detected by quantitative polymerase chain reaction (qPCR) and western-blot (WB), while the fluorescence of GFP within the brains of mice was observed under a fluorescence microscope. Higher mRNA and protein expression of GFP, accompanied with increased green fluorescence, were detected in the brain of GFP-EPCs-injected APP/PS1 mice, as compared with GFP-EPCs-injected WT mice. The results show that the APP/PS1 transgenic mouse model of AD exhibited enhanced penetration of exogenous EPCs into brains than the WT mice. PMID:27186272

  10. Enhanced penetration of exogenous EPCs into brains of APP/PS1 transgenic mice.

    PubMed

    Yuan, Xiaoyang; Mei, Bin; Zhang, Le; Zhang, Cuntai; Zheng, Miao; Liang, Huifang; Wang, Wei; Zheng, Jie; Ding, Ling; Zheng, Kai

    2016-01-01

    The aim of this study was to investigate the repair function of exogenous Endothelial progenitor cells (EPCs) for brain microvascular damage of the APP/PS1 transgenic mouse model of Alzheimer's disease (AD). This study used a density-gradient centrifugation method to isolate mononuclear cells (MNCs) from mouse bone marrow, which were subsequently seeded and cultured. Cells were characterized by morphology and detection of the surface markers CD34 and CD133 at different time points by immunofluorescence (IF) and flow cytometry (FCM). Then, EPCs were transfected with GFP adenoviral vectors (GFP-EPCs). Wild-type (WT) and APP/PS1 transgenic mice both received GFP-EPCs injection through the tail vein, and using a PBS buffer injection as the control. Seven days later, the animals' brain tissue was isolated. Expression of GFP was detected by quantitative polymerase chain reaction (qPCR) and western-blot (WB), while the fluorescence of GFP within the brains of mice was observed under a fluorescence microscope. Higher mRNA and protein expression of GFP, accompanied with increased green fluorescence, were detected in the brain of GFP-EPCs-injected APP/PS1 mice, as compared with GFP-EPCs-injected WT mice. The results show that the APP/PS1 transgenic mouse model of AD exhibited enhanced penetration of exogenous EPCs into brains than the WT mice. PMID:27186272

  11. INCREASED LIVER PATHOLOGY IN HEPATITIS C VIRUS TRANSGENIC MICE EXPRESSING THE HEPATITIS B VIRUS X PROTEIN

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Transgenic mice expressing the full-length HCV coding sequence were crossed with mice that express the HBV X gene-encoded regulatory protein HBx (ATX mice) to test the hypothesis that HBx expression accelerates HCV-induced liver pathogenesis. At 16 months (mo) of age, hepatocellular carcinoma was id...

  12. FGF-7 expression enhances the performance of bioengineered skin.

    PubMed

    Erdag, Gulsun; Medalie, Daniel A; Rakhorst, Hinne; Krueger, Gerald G; Morgan, Jeffrey R

    2004-07-01

    To improve the performance of bioengineered skin, we used a recombinant retrovirus encoding FGF-7 to modify diploid human keratinocytes genetically. Control or FGF-7-expressing keratinocytes were seeded onto acellular human dermis to form bioengineered skin. Gene-modified skin secreted significant levels of FGF-7 and formed a thicker and hyperproliferative epidermis with about four times the number of cells per square centimeter. Secretion of an endogenous trophic factor, VEGF, was increased approximately 5-fold. Migration of FGF-7-expressing keratinocytes was stimulated as was the self-healing of bioengineered skin expressing FGF-7. When tested in a bacterial infection model, the antimicrobial properties of FGF-7-expressing skin were increased >500-fold against both gram-negative and gram-positive bacteria. After transplantation to full-thickness wounds on athymic mice, skin expressing FGF-7 was revascularized more rapidly. These results demonstrate that genetic modification can be used to enhance performance and that expression of FGF-7 augments several properties important to the wound-healing properties of bioengineered skin. PMID:15233944

  13. Gomafu lncRNA knockout mice exhibit mild hyperactivity with enhanced responsiveness to the psychostimulant methamphetamine

    PubMed Central

    Ip, Joanna Y.; Sone, Masamitsu; Nashiki, Chieko; Pan, Qun; Kitaichi, Kiyoyuki; Yanaka, Kaori; Abe, Takaya; Takao, Keizo; Miyakawa, Tsuyoshi; Blencowe, Benjamin J.; Nakagawa, Shinichi

    2016-01-01

    The long noncoding RNA Gomafu/MIAT/Rncr2 is thought to function in retinal cell specification, stem cell differentiation and the control of alternative splicing. To further investigate physiological functions of Gomafu, we created mouse knockout (KO) model that completely lacks the Gomafu gene. The KO mice did not exhibit any developmental deficits. However, behavioral tests revealed that the KO mice are hyperactive. This hyperactive behavior was enhanced when the KO mice were treated with the psychostimulant methamphetamine, which was associated with an increase in dopamine release in the nucleus accumbens. RNA sequencing analyses identified a small number of genes affected by the deficiency of Gomafu, a subset of which are known to have important neurobiological functions. These observations suggest that Gomafu modifies mouse behavior thorough a mild modulation of gene expression and/or alternative splicing of target genes. PMID:27251103

  14. Gomafu lncRNA knockout mice exhibit mild hyperactivity with enhanced responsiveness to the psychostimulant methamphetamine.

    PubMed

    Ip, Joanna Y; Sone, Masamitsu; Nashiki, Chieko; Pan, Qun; Kitaichi, Kiyoyuki; Yanaka, Kaori; Abe, Takaya; Takao, Keizo; Miyakawa, Tsuyoshi; Blencowe, Benjamin J; Nakagawa, Shinichi

    2016-01-01

    The long noncoding RNA Gomafu/MIAT/Rncr2 is thought to function in retinal cell specification, stem cell differentiation and the control of alternative splicing. To further investigate physiological functions of Gomafu, we created mouse knockout (KO) model that completely lacks the Gomafu gene. The KO mice did not exhibit any developmental deficits. However, behavioral tests revealed that the KO mice are hyperactive. This hyperactive behavior was enhanced when the KO mice were treated with the psychostimulant methamphetamine, which was associated with an increase in dopamine release in the nucleus accumbens. RNA sequencing analyses identified a small number of genes affected by the deficiency of Gomafu, a subset of which are known to have important neurobiological functions. These observations suggest that Gomafu modifies mouse behavior thorough a mild modulation of gene expression and/or alternative splicing of target genes. PMID:27251103

  15. 2-Deoxy-D-Glucose Enhances Anesthethic Effects in Mice

    PubMed Central

    Wang, Hui; Xu, Zhipeng; Wu, Anshi; Dong, Yuanlin; Zhang, Yiying; Yue, Yun; Xie, Zhongcong

    2014-01-01

    Background The mechanisms of general anesthesia by volatile drugs remain largely unknown. Mitochondrial dysfunction and reduction in energy levels have been suggested to be associated with general anesthesia status. 2-Deoxy-d-glucose (2-DG), an analog of glucose, inhibits hexokinase and reduces cellular levels of adenosine triphosphate (ATP). 3-Nitropropionic acid is another compound which can deplete ATP levels. In contrast, idebenone and L-carnitine could rescue deficits of energy. We therefore sought to determine whether 2-DG and/or 3-nitropropionic acid can enhance the anesthetic effects of isoflurane, and whether idebenone and L-carnitine can reverse the actions of 2-DG. Methods C57BL/6J mice (8 months old) received different concentrations of isoflurane with and without the treatments of 2-DG, 3-nitropropionic acid, idebenone, and L-carnitine. Isoflurane-induced loss of righting reflex (LORR) was determined in the mice. ATP levels in H4 human neuroglioma cells were assessed after these treatments. Finally, 31P-magnetic resonance spectroscopy was used to determine the effects of isoflurane on brain ATP levels in the mice. Results 2-DG enhanced isoflurane-induced LORR (P = 0.002, N = 15). 3-nitropropionic acid also enhanced the anesthetic effects of isoflurane (P = 0.005, N = 15). Idebenone (Idebenone + saline versus Idebenone + 2-DG: P = 0.165, N = 15), but not L-cartinine (L-carnitine + saline versus L-carnitine + 2-DG: P < 0.0001, N = 15), inhibited the effects of 2-DG on enhancing isoflurane-induced LORR in mice, as evidenced by 2-DG not enhancing isoflurane-induced LORR in mice pretreated with idebenone. Idebenone (Idebenone + saline versus Idebenone + 2-DG: P = 0.177, N = 6), but not L-cartinine (L-carnitine + saline versus L-carnitine + 2-DG: P = 0.029, N = 6), also mitigated the effects of 2-DG on reducing ATP levels in cells, as evidenced by 2-DG not decreasing ATP levels in the cell pretreated with idebenone. Finally, isoflurane decreased ATP levels

  16. In vivo photoacoustic imaging of tyrosinase expressing tumours in mice

    NASA Astrophysics Data System (ADS)

    Laufer, Jan; Jathoul, Amit; Johnson, Peter; Zhang, Edward; Lythgoe, Mark; Pedley, R. Barbara; Pule, Martin; Beard, Paul

    2012-02-01

    Two human tumour cell lines (K562, 293T) were stably transfected to achieve the genetic expression of tyrosinase, which is involved in the production of the pigment eumelanin. The cells were injected subcutaneously into nude mice to form tumour xenografts, which were imaged over a period of up to 26 days using an all-optical photoacoustic imaging system. 3D photoacoustic images of the tumours and the surrounding vasculature were acquired at excitation wavelengths ranging from 600nm to 770nm. The images showed tumour growth and continued tyrosinase expression over the full 26 day duration of the study. These findings were confirmed by histological analysis of excised tumour samples.

  17. Afadin requirement for cytokine expressions in keratinocytes during chemically induced inflammation in mice

    PubMed Central

    Yoshida, Toshiyuki; Iwata, Takanori; Takai, Yoshimi; Birchmeier, Walter; Yamato, Masayuki; Okano, Teruo

    2014-01-01

    Afadin is a filamentous actin-binding protein and a mediator of nectin signaling. Nectins are Ig-like cell adhesion molecules, and the nectin family is composed of four members, nectin-1 to nectin-4. Nectins show homophilic and heterophilic interactions with other nectins or proteins on adjacent cells. Nectin signaling induces formation of cell–cell junctions and is required for the development of epithelial tissues, including skin. This study investigated the role of afadin in epithelial tissue development and established epithelium-specific afadin-deficient (CKO) mice. Although showing no obvious abnormality in the skin development and homeostasis, the mice showed the reduced neutrophil infiltration into the epidermis during chemical-induced inflammation with 12-O-tetradecanoylphorbol 13-acetate (TPA). Immunohistochemical and quantitative real-time PCR analyses showed that the expression levels of cytokines including Cxcl2, Il-1β and Tnf-α were reduced in CKO keratinocytes compared with control keratinocytes during TPA-induced inflammation. Primary-cultured skin keratinocytes from CKO mice also showed reduced expression of these cytokines and weak activation of Rap1 compared with those from control mice after the TPA treatment. These results suggested a remarkable function of afadin, which was able to enhance cytokine expression through Rap1 activation in keratinocytes during inflammation. PMID:25297509

  18. Adiponectin exacerbates collagen-induced arthritis via enhancing Th17 response and prompting RANKL expression.

    PubMed

    Sun, Xiaoxuan; Feng, Xiaoke; Tan, Wenfeng; Lin, Na; Hua, Minhui; Wei, Yu; Wang, Fang; Li, Ningli; Zhang, Miaojia

    2015-01-01

    We previously reported adiponectin (AD) is highly expressed in the inflamed synovial joint tissue and correlates closely with progressive bone erosion in Rheumatoid arthritis (RA) patients. Here, we investigate the role of adiponectin in regulating Th17 response and the expression of receptor activator of nuclear factor-κB ligand (RANKL) in mice with CIA mice by intraarticularly injection of adiponectin into knee joints on day 17, day 20 and day 23 post first collagen immunization. The increased adiponectin expression was found in inflamed joint tissue of collagen-induced arthritis (CIA) mice. Adiponectin injection resulted in an earlier onset of arthritis, an aggravated arthritic progression, more severe synovial hyperplasia, bone erosion and osteoporosis in CIA mice. CD4(+)IL-17(+) Th17 cells, IL-17 mRNA and RANKL mRNA expression were markedly increased in the joint tissue of adiponectin treated CIA mice. Moreover, adiponectin treatment markedly enhanced Th17 cell generation from naive CD4(+) T cells in vitro, which accompanied by the high expression of Th17 transcription factor ROR-γt, and Th17 cytokine genes included IL-22 and IL-23. This study reveals a novel effect of adiponectin in exacerbating CIA progression by enhancing Th17 cell response and RANKL expression. PMID:26063682

  19. BAC transgenic mice provide evidence that p53 expression is highly regulated in vivo

    PubMed Central

    Chen, L; Zhang, G X; Zhou, Y; Zhang, C X; Xie, Y Y; Xiang, C; He, X Y; Zhang, Q; Liu, G

    2015-01-01

    p53 is an important tumor suppressor and stress response mediator. Proper control of p53 level and activity is tightly associated with its function. Posttranslational modifications and the interactions with Mdm2 and Mdm4 are major mechanisms controlling p53 activity and stability. As p53 protein is short-lived and hardly detectable in unstressed situations, less is known on its basal level expression and the corresponding controlling mechanisms in vivo. In addition, it also remains obscure how p53 expression might contribute to its functional regulation. In this study, we established bacterial artificial chromosome transgenic E.coli β-galactosidase Z gene reporter mice to monitor p53 expression in mouse tissues and identify important regulatory elements critical for the expression in vivo. We revealed preferentially high level of p53 reporter expressions in the proliferating, but not the differentiated compartments of the majority of tissues during development and tissue homeostasis. In addition, tumors as well as regenerating tissues in the p53 reporter mice also expressed high level of β-gal. Furthermore, both the enhancer box sequence (CANNTG) in the p53 promoter and the 3′ terminal untranslated region element were critical in mediating the high-level expression of the reporter. We also provided evidence that cellular myelocytomatosis oncogene was a critical player regulating p53 mRNA expression in proliferating cells and tissues. Finally, we found robust p53 activation preferentially in the proliferating compartment of mouse tissues upon DNA damage and the proliferating cells exhibited an enhanced p53 response as compared with cells in a quiescent state. Together, these results suggested a highly regulated expression pattern of p53 in the proliferating compartment controlled by both transcriptional and posttranscriptional mechanisms, and such regulated p53 expression may impose functional significance upon stress by setting up a precautionary mode in

  20. Methane limit LPS-induced NF-κB/MAPKs signal in macrophages and suppress immune response in mice by enhancing PI3K/AKT/GSK-3β-mediated IL-10 expression

    PubMed Central

    Zhang, Xu; Li, Na; Shao, Han; Meng, Yan; Wang, Liping; Wu, Qian; Yao, Ying; Li, Jinbao; Bian, Jinjun; Zhang, Yan; Deng, Xiaoming

    2016-01-01

    Inflammatory diseases such as sepsis and autoimmune colitis, characterized by an overwhelming activation of the immune system and the counteracting anti-inflammatory response, remain a major health problem in worldwide. Emerging evidence suggests that methane have a protective effect on many animal models, like ischaemia reperfusion injury and diabetes-associated diseases. Whether methane could modulating inflammatory diseases remains largely unknown. Here we show that methane-rich saline (MS) ip treatment (16 ml/kg) alleviated endotoxin shock, bacteria-induced sepsis and dextran-sulfate-sodium-induced colitis in mice via decreased production of TNF-α and IL-6. In MS-treated macrophages, LPS-induced activation of NF-κb/MAPKs was attenuated. Interestingly, MS treatment significantly elevated the levels of IL-10 both in vitro and in vivo. Neutralization of IL-10 abrogated the therapeutic effect of MS. Moreover, anti-IL10 blockade partially restored the MS-mediated attenuation of NF-κb/MAPKs phosphorylation. We further found that MS resulted in markedly enhanced phosphorylation of GSK-3β and AKT, which both mediate the release of Il-10. Additionally, inhibition of PI3K attenuated MS-mediated p-GSK-3β and IL-10 production and reversed the suppressed activation of NF-κb/ MAPKs in response to LPS. Our results reveal a novel effect and mechanisms of methane and support the potential value of MS as a therapeutic approach in innate inflammatory diseases. PMID:27405597

  1. Methane limit LPS-induced NF-κB/MAPKs signal in macrophages and suppress immune response in mice by enhancing PI3K/AKT/GSK-3β-mediated IL-10 expression.

    PubMed

    Zhang, Xu; Li, Na; Shao, Han; Meng, Yan; Wang, Liping; Wu, Qian; Yao, Ying; Li, Jinbao; Bian, Jinjun; Zhang, Yan; Deng, Xiaoming

    2016-01-01

    Inflammatory diseases such as sepsis and autoimmune colitis, characterized by an overwhelming activation of the immune system and the counteracting anti-inflammatory response, remain a major health problem in worldwide. Emerging evidence suggests that methane have a protective effect on many animal models, like ischaemia reperfusion injury and diabetes-associated diseases. Whether methane could modulating inflammatory diseases remains largely unknown. Here we show that methane-rich saline (MS) ip treatment (16 ml/kg) alleviated endotoxin shock, bacteria-induced sepsis and dextran-sulfate-sodium-induced colitis in mice via decreased production of TNF-α and IL-6. In MS-treated macrophages, LPS-induced activation of NF-κb/MAPKs was attenuated. Interestingly, MS treatment significantly elevated the levels of IL-10 both in vitro and in vivo. Neutralization of IL-10 abrogated the therapeutic effect of MS. Moreover, anti-IL10 blockade partially restored the MS-mediated attenuation of NF-κb/MAPKs phosphorylation. We further found that MS resulted in markedly enhanced phosphorylation of GSK-3β and AKT, which both mediate the release of Il-10. Additionally, inhibition of PI3K attenuated MS-mediated p-GSK-3β and IL-10 production and reversed the suppressed activation of NF-κb/ MAPKs in response to LPS. Our results reveal a novel effect and mechanisms of methane and support the potential value of MS as a therapeutic approach in innate inflammatory diseases. PMID:27405597

  2. Caspase-2 Deficiency Enhances Aging-Related Traits in Mice

    PubMed Central

    Zhang, Yingpei; Padalecki, Susan S; Chaudhuri, Asish R; Waal, Eric De; Goins, Beth A; Grubbs, Barry; Ikeno, Yuji; Richardson, Arlan; Mundy, Gregory R; Herman, Brian

    2007-01-01

    Alteration of apoptotic activity has been observed in a number of tissues in aging mammals, but it remains unclear whether and/or how apoptosis may affect aging. Caspase-2 is a member of the cysteine protease family that plays a critical role in apoptosis. To understand the impact of compromised apoptosis function on mammalian aging, we conducted a comparative study on caspase-2 deficient mice and their wild-type littermates with a specific focus on the aging-related traits at advanced ages. We found that caspase-2 deficiency enhanced a number of traits commonly seen in premature aging animals. Loss of caspase-2 was associated with shortened maximum lifespan, impaired hair growth, increased bone loss, and reduced body fat content. In addition, we found that the livers of caspase-2 deficient mice had higher levels of oxidized proteins than those of age-matched wild-type mice, suggesting that caspase-2 deficiency compromised the animal's ability to clear oxidatively damaged cells. Collectively, these results suggest that caspase-2 deficiency affects aging in the mice. This study thus demonstrates for the first time that disruption of a key apoptotic gene has a significant impact on aging. PMID:17188333

  3. Changes in phagocytosis and expression of microglial cells in craniocerebral injury mice models.

    PubMed

    Guo, F Y; Liu, T; Chen, J J; Gao, W; Yang, F; Zhou, X Y; Liao, Y L

    2016-01-01

    The objective of this study was to investigate the changes in phagocytic function and expression quantities of CD11b and tumor necrosis factor-α (TNF-α) among microglia cells of craniocerebral injury mice. Modified Feeney method was used to establish the craniocerebral injury mice models. Twenty-one male SPF mice were divided into a control group and a trauma group. The scalp was incised and a bone window was opened in the control group without cerebral injury. In the trauma group, the mice were sacrificed after the craniocerebral injury at 1, 3, 6, 12, 24 and 48 h to make frozen sections of cerebral tissues. The phagocytic rate of microglia cells was observed by using fluorescent microsphere. The changes in the expression quantities of CD11b and TNF-α were detected by enzyme-linked immuno sorbent assay (ELISA). The phagocytic ability of the microglia cells after the craniocerebral injury increased at 1 h after injury compared with that of the control group (P less than 0.01). The expression of surface antigen CD11b of the microglia cells and the expression of TNF-α increased at 1, 3, 6, 12, 24 and 48 h after the injury compared with those of the control group (P less than 0.01). The phagocytic ability of the microglia cells increased. The expressions of CD11b and TNF-α were also gradually enhanced in the acute phase after craniocerebral injury, and then gradually decreased to the normal level. The expressions of CD11b and TNF-α indicated a high consistency with the changing trend of the phagocytic ability, suggesting that the microglia cells may participate in the regulation of the inflammatory process of the central nervous system through absorbing apoptotic cells and increasing and secreting inflammatory and anti-inflammatory factors. PMID:27358141

  4. Constitutive adipocyte mTORC1 activation enhances mitochondrial activity and reduces visceral adiposity in mice.

    PubMed

    Magdalon, Juliana; Chimin, Patricia; Belchior, Thiago; Neves, Rodrigo X; Vieira-Lara, Marcel A; Andrade, Maynara L; Farias, Talita S; Bolsoni-Lopes, Andressa; Paschoal, Vivian A; Yamashita, Alex S; Kowaltowski, Alicia J; Festuccia, William T

    2016-05-01

    Mechanistic target of rapamycin complex 1 (mTORC1) loss of function reduces adiposity whereas partial mTORC1 inhibition enhances fat deposition. Herein we evaluated how constitutive mTORC1 activation in adipocytes modulates adiposity in vivo. Mice with constitutive mTORC1 activation in adipocytes induced by tuberous sclerosis complex (Tsc)1 deletion and littermate controls were evaluated for body mass, energy expenditure, glucose and fatty acid metabolism, mitochondrial function, mRNA and protein contents. Adipocyte-specific Tsc1 deletion reduced visceral, but not subcutaneous, fat mass, as well as adipocyte number and diameter, phenotypes that were associated with increased lipolysis, UCP-1 content (browning) and mRNA levels of pro-browning transcriptional factors C/EBPβ and ERRα. Adipocyte Tsc1 deletion enhanced mitochondrial oxidative activity, fatty acid oxidation and the expression of PGC-1α and PPARα in both visceral and subcutaneous fat. In brown adipocytes, however, Tsc1 deletion did not affect UCP-1 content and basal respiration. Adipocyte Tsc1 deletion also reduced visceral adiposity and enhanced glucose tolerance, liver and muscle insulin signaling and adiponectin secretion in mice fed with purified low- or high-fat diet. In conclusion, adipocyte-specific Tsc1 deletion enhances mitochondrial activity, induces browning and reduces visceral adiposity in mice. PMID:26923434

  5. Transient Expression of Proteins by Hydrodynamic Gene Delivery in Mice

    PubMed Central

    Kovacsics, Daniella; Raper, Jayne

    2014-01-01

    Efficient expression of transgenes in vivo is of critical importance in studying gene function and developing treatments for diseases. Over the past years, hydrodynamic gene delivery (HGD) has emerged as a simple, fast, safe and effective method for delivering transgenes into rodents. This technique relies on the force generated by the rapid injection of a large volume of physiological solution to increase the permeability of cell membranes of perfused organs and thus deliver DNA into cells. One of the main advantages of HGD is the ability to introduce transgenes into mammalian cells using naked plasmid DNA (pDNA). Introducing an exogenous gene using a plasmid is minimally laborious, highly efficient and, contrary to viral carriers, remarkably safe. HGD was initially used to deliver genes into mice, it is now used to deliver a wide range of substances, including oligonucleotides, artificial chromosomes, RNA, proteins and small molecules into mice, rats and, to a limited degree, other animals. This protocol describes HGD in mice and focuses on three key aspects of the method that are critical to performing the procedure successfully: correct insertion of the needle into the vein, the volume of injection and the speed of delivery. Examples are given to show the application of this method to the transient expression of two genes that encode secreted, primate-specific proteins, apolipoprotein L-I (APOL-I) and haptoglobin-related protein (HPR). PMID:24837006

  6. Transient expression of proteins by hydrodynamic gene delivery in mice.

    PubMed

    Kovacsics, Daniella; Raper, Jayne

    2014-01-01

    Efficient expression of transgenes in vivo is of critical importance in studying gene function and developing treatments for diseases. Over the past years, hydrodynamic gene delivery (HGD) has emerged as a simple, fast, safe and effective method for delivering transgenes into rodents. This technique relies on the force generated by the rapid injection of a large volume of physiological solution to increase the permeability of cell membranes of perfused organs and thus deliver DNA into cells. One of the main advantages of HGD is the ability to introduce transgenes into mammalian cells using naked plasmid DNA (pDNA). Introducing an exogenous gene using a plasmid is minimally laborious, highly efficient and, contrary to viral carriers, remarkably safe. HGD was initially used to deliver genes into mice, it is now used to deliver a wide range of substances, including oligonucleotides, artificial chromosomes, RNA, proteins and small molecules into mice, rats and, to a limited degree, other animals. This protocol describes HGD in mice and focuses on three key aspects of the method that are critical to performing the procedure successfully: correct insertion of the needle into the vein, the volume of injection and the speed of delivery. Examples are given to show the application of this method to the transient expression of two genes that encode secreted, primate-specific proteins, apolipoprotein L-I (APOL-I) and haptoglobin-related protein (HPR). PMID:24837006

  7. Generation and characterization of transgenic mice expressing cobra venom factor.

    PubMed

    Andrä, Jörg; Halter, Roman; Kock, Michael A; Niemann, Heiner; Vogel, Carl-Wilhelm; Paul, Dieter

    2002-10-01

    Cobra venom factor (CVF), the anticomplementary protein in cobra venom, activates the alternative complement pathway, eventually leading to complement consumption. Here, we describe the development of a transgenic mouse model for CVF. We generated a DNA construct containing the full-length cDNA for single-chain pre-pro-CVF. Expression of CVF was controlled by the alpha(1)-antitrypsin promoter to achieve liver-specific expression. Linearized DNA was microinjected into murine ovary cells (strain CD(2)F(1) (BALB/cxDBA/2J)) and the newborn mice were analyzed for stable integration of CVF DNA. After establishing the transgene, mice were propagated in a BALB/c background. The CVF mRNA was detected in the liver and, in some animals, in the kidney. CVF protein was detected in small amounts in the serum. Serum complement hemolytic activity in CVF-transgenic mice was virtually absent. The concentration of plasma C3 was significantly reduced. The CVF-transgenic animals show no unusual phenotype. They provide an animal model to study the effect of long-term complement depletion by continued activation, as well as the role of complement in host immune response and pathogenesis of disease. PMID:12220893

  8. Frostbite Protection in Mice Expressing an Antifreeze Glycoprotein

    PubMed Central

    Heisig, Martin; Mattessich, Sarah; Rembisz, Alison; Acar, Ali; Shapiro, Martin; Booth, Carmen J.; Neelakanta, Girish; Fikrig, Erol

    2015-01-01

    Ectotherms in northern latitudes are seasonally exposed to cold temperatures. To improve survival under cold stress, they use diverse mechanisms to increase temperature resistance and prevent tissue damage. The accumulation of anti-freeze proteins that improve cold hardiness occurs in diverse species including plants, arthropods, fish, and amphibians. We previously identified an Ixodes scapularis anti-freeze glycoprotein, named IAFGP, and demonstrated its cold protective function in the natural tick host and in a transgenic Drosophila model. Here we show, in a transgenic mouse model expressing an anti-freeze glycoprotein, that IAFGP protects mammalian cells and mice from cold shock and frostbite respectively. Transgenic skin samples showed reduced cell death upon cold storage ex vivo and transgenic mice demonstrated increased resistance to frostbite injury in vivo. IAFGP actively protects mammalian tissue from freezing, suggesting its application for the prevention of frostbite, and other diseases associated with cold exposure. PMID:25714402

  9. Identification of hypothalamic arcuate nucleus-specific enhancer region of Kiss1 gene in mice.

    PubMed

    Goto, Teppei; Tomikawa, Junko; Ikegami, Kana; Minabe, Shiori; Abe, Hitomi; Fukanuma, Tatsuya; Imamura, Takuya; Takase, Kenji; Sanbo, Makoto; Tomita, Koichi; Hirabayashi, Masumi; Maeda, Kei-ichiro; Tsukamura, Hiroko; Uenoyama, Yoshihisa

    2015-01-01

    Pulsatile secretion of GnRH plays a pivotal role in follicular development via stimulating tonic gonadotropin secretion in mammals. Kisspeptin neurons, located in the arcuate nucleus (ARC), are considered to be an intrinsic source of the GnRH pulse generator. The present study aimed to determine ARC-specific enhancer(s) of the Kiss1 gene by an in vivo reporter assay. Three green fluorescent protein (GFP) reporter constructs (long, medium length, and short) were generated by insertion of GFP cDNA at the Kiss1 locus. Transgenic female mice bearing the long and medium-length constructs showed apparent GFP signals in kisspeptin-immunoreactive cells in both the ARC and anteroventral periventricular nucleus, in which another population of kisspeptin neurons are located. On the other hand, transgenic mice bearing 5'-truncated short construct showed few GFP signals in the ARC kisspeptin-immunoreactive cells, whereas they showed colocalization of GFP- and kisspeptin-immunoreactivities in the anteroventral periventricular nucleus. In addition, chromatin immunoprecipitation and chromosome conformation capture assays revealed recruitment of unoccupied estrogen receptor-α in the 5'-upstream region and intricate chromatin loop formation between the 5'-upstream and promoter regions of Kiss1 locus in the ARC. Taken together, the present results indicate that 5'-upstream region of Kiss1 locus plays a critical role in Kiss1 gene expression in an ARC-specific manner and that the recruitment of estrogen receptor-α and formation of a chromatin loop between the Kiss1 promoter and the 5' enhancer region may be required for the induction of ARC-specific Kiss1 gene expression. These results suggest that the 5'-upstream region of Kiss1 locus functions as an enhancer for ARC Kiss1 gene expression in mice. PMID:25486239

  10. Vaccine adjuvant ginsenoside Rg1 enhances immune responses against hepatitis B surface antigen in mice.

    PubMed

    Yuan, Ding; Yuan, Qin; Cui, Qianqian; Liu, Chaoqi; Zhou, Zhiyong; Zhao, Haixia; Dun, Yaoyan; Wang, Ting; Zhang, Changcheng

    2016-06-01

    The adjuvant effect of ginsenoside Rg1 on immune responses against hepatitis B surface antigen (HBsAg) in mice was investigated. Female BALB/c mice were subcutaneously injected with saline or HBsAg antigen with or without Rg1 on days 7 and 21. Samples were collected 2 weeks after the boosting for the detection of anti-HBsAg immunoglobulin G (IgG) isotypes in sera and gamma interferon (IFN-γ) and interleukin-4 (IL-4) produced in splenocytes. The innate and adaptive immune responses were measured in mice immunized as described above. The results showed that ginsenoside Rg1 had adjuvant properties in stimulating IgG, splenocyte proliferation, and mRNA expression of cytokines IFN-γ and IL-4, as well as the expression of cell surface marker TLR4 in the HBsAg-immunized mice. These results indicate that Rg1 enhances both Th1 (IgG2b and IFN-γ) and Th2 (IgG1 and IL-4) responses. In addition, the TLR4 signaling pathway is involved in the adjuvant activities of ginsenoside Rg1. PMID:27095502

  11. Apoptosis inhibitor of macrophage protein enhances intraluminal debris clearance and ameliorates acute kidney injury in mice.

    PubMed

    Arai, Satoko; Kitada, Kento; Yamazaki, Tomoko; Takai, Ryosuke; Zhang, Xizhong; Tsugawa, Yoji; Sugisawa, Ryoichi; Matsumoto, Ayaka; Mori, Mayumi; Yoshihara, Yasunori; Doi, Kent; Maehara, Natsumi; Kusunoki, Shunsuke; Takahata, Akiko; Noiri, Eisei; Suzuki, Yusuke; Yahagi, Naoki; Nishiyama, Akira; Gunaratnam, Lakshman; Takano, Tomoko; Miyazaki, Toru

    2016-02-01

    Acute kidney injury (AKI) is associated with prolonged hospitalization and high mortality, and it predisposes individuals to chronic kidney disease. To date, no effective AKI treatments have been established. Here we show that the apoptosis inhibitor of macrophage (AIM) protein on intraluminal debris interacts with kidney injury molecule (KIM)-1 and promotes recovery from AKI. During AKI, the concentration of AIM increases in the urine, and AIM accumulates on necrotic cell debris within the kidney proximal tubules. The AIM present in this cellular debris binds to KIM-1, which is expressed on injured tubular epithelial cells, and enhances the phagocytic removal of the debris by the epithelial cells, thus contributing to kidney tissue repair. When subjected to ischemia-reperfusion (IR)-induced AKI, AIM-deficient mice exhibited abrogated debris clearance and persistent renal inflammation, resulting in higher mortality than wild-type (WT) mice due to progressive renal dysfunction. Treatment of mice with IR-induced AKI using recombinant AIM resulted in the removal of the debris, thereby ameliorating renal pathology. We observed this effect in both AIM-deficient and WT mice, but not in KIM-1-deficient mice. Our findings provide a basis for the development of potentially novel therapies for AKI. PMID:26726878

  12. Expression of KRAS in the endometrium of early pregnant mice and its effect during embryo implantation.

    PubMed

    Long, Xia; Zhang, Min; Chen, Xuemei; He, Junlin; Ding, Yubin; Zhang, Cuizhen; Liu, Xueqing; Wang, Yingxiong

    2015-07-01

    This study investigated the expression pattern of Ki-ras2 Kirsten rat sarcoma viral oncogene homolog (KRAS) in the endometrium of early-stage pregnant mice and its function during embryo implantation. The expression of KRAS was measured at the mRNA level using real-time polymerase chain reaction (PCR) and at the protein level using immunohistochemistry and western blotting. The expressions of KRAS mRNA and protein were not significantly different in the endometrium of pseudopregnant and early-stage pregnant mice. However, the immunohistochemistry results showed that KRAS was highly expressed in the decidualizing stromal cells on days 5-7 of mouse pregnancy and was enhanced in the epithelial cells as pregnancy progressed. The expression of KRAS protein was higher after the stromal cell was artificially decidualized in vivo and in vitro. Stromal cell proliferation was attenuated after down-regulating KRAS expression. After silencing KRAS in the mouse uterus, the embryo implantation rate was significantly reduced (P < 0.005). We speculate that KRAS may regulate the stromal cell proliferation and differentiation progress and then affect the embryo implantation process. This study reveals that KRAS plays an important role in regulating the embryo implantation process. PMID:25999213

  13. Deinococcus radiodurans pprI expression enhances the radioresistance of eukaryotes.

    PubMed

    Wen, Ling; Yue, Ling; Shi, Yi; Ren, Lili; Chen, Tingting; Li, Na; Zhang, Shuyu; Yang, Wei; Yang, Zhanshan

    2016-03-29

    PprI accelerates radiation-induced DNA damage repair via regulating the expression of DNA repair genes and enhances antioxidative enzyme activity in Deinococcus radiodurans after radiation. The main aim of our study was to determine whether the expression of pprI gene could fulfil its DNA repair function in eukaryotes and enhance the radioresistance of eukaryotic organism or not. In this study, we constructed pEGFP-c1-pprI eukaryotic expression vector and established a human lung epithelial cell line BEAS-2B with stable integration of pprI gene. We found that pprIexpression enhanced radioresistance of BEAS-2B cells, decreased γ-H2AX foci formation and apoptosis in irradiated BEAS-2B cells and alleviated radiation induced G2/M arrest of BEAS-2B cells. Moreover, we transferred pEGFP-c1-pprI vector into muscle of BALB/c mice by in vivo electroporation and studied the protective effect of prokaryotic pprI gene in irradiated mice. We found that pprI expression alleviated acute radiation induced hematopoietic system, lung, small intestine and testis damage and increased survival rate of irradiated mice via regulating Rad51 expression in different organs. These findings suggest that prokaryotic pprI gene expression in mammalian cells could enhance radioresistance in vitro and in vivo. PMID:26992215

  14. Deinococcus radiodurans pprI expression enhances the radioresistance of eukaryotes

    PubMed Central

    Wen, Ling; Yue, Ling; Shi, Yi; Ren, Lili; Chen, Tingting; Li, Na; Zhang, Shuyu; Yang, Wei; Yang, Zhanshan

    2016-01-01

    PprI accelerates radiation-induced DNA damage repair via regulating the expression of DNA repair genes and enhances antioxidative enzyme activity in Deinococcus radiodurans after radiation. The main aim of our study was to determine whether the expression of pprI gene could fulfil its DNA repair function in eukaryotes and enhance the radioresistance of eukaryotic organism or not. In this study, we constructed pEGFP-c1-pprI eukaryotic expression vector and established a human lung epithelial cell line BEAS-2B with stable integration of pprI gene. We found that pprIexpression enhanced radioresistance of BEAS-2B cells, decreased γ-H2AX foci formation and apoptosis in irradiated BEAS-2B cells and alleviated radiation induced G2/M arrest of BEAS-2B cells. Moreover, we transferred pEGFP-c1-pprI vector into muscle of BALB/c mice by in vivo electroporation and studied the protective effect of prokaryotic pprI gene in irradiated mice. We found that pprI expression alleviated acute radiation induced hematopoietic system, lung, small intestine and testis damage and increased survival rate of irradiated mice via regulating Rad51 expression in different organs. These findings suggest that prokaryotic pprI gene expression in mammalian cells could enhance radioresistance in vitro and in vivo. PMID:26992215

  15. Long-term exercise modulates hippocampal gene expression in senescent female mice.

    PubMed

    Alvarez-López, María Jesús; Castro-Freire, Marco; Cosín-Tomás, Marta; Sanchez-Roige, Sandra; Lalanza, Jaume F; Del Valle, Jaume; Párrizas, Marcelina; Camins, Antonio; Pallás, Merce; Escorihuela, Rosa María; Kaliman, Perla

    2013-01-01

    The senescence-accelerated SAMP8 mouse is considered a useful non-transgenic model for studying aspects of progressive cognitive decline and Alzheimer's disease (AD). Using SAMR1 mice as controls, here we explored the effects of 6 months of voluntary wheel running in 10-month-old female SAMP8 mice. Exercise in SAMP8 mice improved phenotypic features associated with premature aging (i.e., skin color and body tremor) and enhanced vascularization and BDNF gene expression in the hippocampus compared with controls. With the aim of identifying genes involved in brain aging responsive to long-term exercise, we performed whole genome microarray studies in hippocampus from sedentary SAMP8 (P8sed), SAMR1 (R1sed), and exercised SAMP8 (P8run) mice. The genes differentially expressed in P8sed versus R1sed were considered as putative aging markers (i) and those differentially expressed in P8run versus P8sed were considered as genes modulated by exercise (ii). Genes differentially expressed in both comparisons (i and ii) were considered as putative aging genes responsive to physical exercise. We identified 34 genes which met both criteria. Gene ontology analysis revealed that they are mainly involved in functions related to extracellular matrix maintenance. Selected genes were validated by real-time quantitative PCR assays, i.e., collagen type 1 alpha 1 (col1a1), collagen type 1 alpha 2 (col1a2), fibromodulin (fmod), prostaglandin D(2) synthase (ptgds), and aldehyde dehydrogenase (Aldh1a2). As a whole, our study suggests that exercise training during adulthood may prevent or delay gene expression alterations and processes associated with hippocampal aging in at-risk subjects. PMID:23168450

  16. Absence of Cx43 selectively from osteocytes enhances responsiveness to mechanical force in mice.

    PubMed

    Bivi, Nicoletta; Pacheco-Costa, Rafael; Brun, Lucas R; Murphy, Thomas R; Farlow, Nathan R; Robling, Alexander G; Bellido, Teresita; Plotkin, Lilian I

    2013-07-01

    The osteocyte network is crucial for the response of bone to mechanical force. Within this network, connexin43 (Cx43) is thought to mediate the communication of osteocytes and osteoblasts among themselves and the exchange of small molecules with the extracellular milieu. Despite recent advances in understanding Cx43 role for the response of bone cells to mechanical stimulation, the contribution of Cx43 specifically in osteocytes to mechanotransduction in vivo is not well-known. We examined the anabolic response to ulnar axial loading of mice lacking Cx43 in osteocytes (Cx43(ΔOt)). Loading induced a greater increase in periosteal bone formation rate in Cx43(ΔOt) mice compared to control littermates, resulting from higher mineralizing surface and enhanced mineral apposition rate. Expression of β-catenin protein, a molecule implicated in mechanotransduction, was higher in bones from Cx43(ΔOt) mice, compared to littermate controls. In addition, MLO-Y4 osteocytic cells knocked-down for Cx43 exhibited higher β-catenin protein expression and enhanced response to mechanical stimulation. These findings suggest that osteocytes lacking Cx43 are "primed" to respond to mechanical stimulation and that absence of Cx43 in osteocytes unleashes bone formation, by a mechanism that might involve accumulation of β-catenin. PMID:23483620

  17. Constitutive telomerase expression promotes mammary carcinomas in aging mice

    PubMed Central

    Artandi, Steven E.; Alson, Scott; Tietze, Maja K.; Sharpless, Norman E.; Ye, Siqin; Greenberg, Roger A.; Castrillon, Diego H.; Horner, James W.; Weiler, Sarah R.; Carrasco, Ruben D.; DePinho, Ronald A.

    2002-01-01

    Telomerase is up-regulated in the vast majority of human cancers and serves to halt the progressive telomere shortening that ultimately blocks would-be cancer cells from achieving a full malignant phenotype. In contrast to humans, the laboratory mouse possesses long telomeres and, even in early generation telomerase-deficient mice, the level of telomere reserve is sufficient to avert telomere-based checkpoint responses and to permit full malignant progression. These features in the mouse provide an opportunity to determine whether enforced high-level telomerase activity can serve functions that extend beyond its ability to sustain telomere length and function. Here, we report the generation and characterization of transgenic mice that express the catalytic subunit of telomerase (mTERT) at high levels in a broad variety of tissues. Expression of mTERT conferred increased telomerase enzymatic activity in several tissues, including mammary gland, splenocytes, and cultured mouse embryonic fibroblasts. In mouse embryonic fibroblasts, mTERT overexpression extended telomere lengths but did not prevent culture-induced replicative arrest, thus reinforcing the view that this phenomenon is not related to occult telomere shortening. Robust telomerase activity, however, was associated with the spontaneous development of mammary intraepithelial neoplasia and invasive mammary carcinomas in a significant proportion of aged females. These data indicate that enforced mTERT expression can promote the development of spontaneous cancers even in the setting of ample telomere reserve. PMID:12034875

  18. Erythropoietin reduces the expression of myostatin in mdx dystrophic mice.

    PubMed

    Feder, D; Rugollini, M; Santomauro, A; Oliveira, L P; Lioi, V P; Santos, R dos; Ferreira, L G; Nunes, M T; Carvalho, M H; Delgado, P O; Carvalho, A A S; Fonseca, F L A

    2014-11-01

    Erythropoietin (EPO) has been well characterized as a renal glycoprotein hormone regulating red blood cell production by inhibiting apoptosis of erythrocyte progenitors in hematopoietic tissues. EPO exerts regulatory effects in cardiac and skeletal muscles. Duchenne muscular dystrophy is a lethal degenerative disorder of skeletal and cardiac muscle. In this study, we tested the possible therapeutic beneficial effect of recombinant EPO (rhEPO) in dystrophic muscles in mdx mice. Total strength was measured using a force transducer coupled to a computer. Gene expression for myostatin, transforming growth factor-β1 (TGF-β1), and tumor necrosis factor-α (TNF-α) was determined by quantitative real time polymerase chain reaction. Myostatin expression was significantly decreased in quadriceps from mdx mice treated with rhEPO (rhEPO = 0.60 ± 0.11, control = 1.07 ± 0.11). On the other hand, rhEPO had no significant effect on the expression of TGF-β1 (rhEPO = 0.95 ± 0.14, control = 1.05 ± 0.16) and TNF-α (rhEPO = 0.73 ± 0.20, control = 1.01 ± 0.09). These results may help to clarify some of the direct actions of EPO on skeletal muscle. PMID:25296358

  19. Efficient Generation of Mice with Consistent Transgene Expression by FEEST.

    PubMed

    Gao, Lei; Jiang, Yonghua; Mu, Libing; Liu, Yanbin; Wang, Fengchao; Wang, Peng; Zhang, Aiqun; Tang, Nan; Chen, Ting; Luo, Minmin; Yu, Lei; Gao, Shaorong; Chen, Liang

    2015-01-01

    Transgenic mouse models are widely used in biomedical research; however, current techniques for producing transgenic mice are limited due to the unpredictable nature of transgene expression. Here, we report a novel, highly efficient technique for the generation of transgenic mice with single-copy integration of the transgene and guaranteed expression of the gene-of-interest (GOI). We refer to this technique as functionally enriched ES cell transgenics, or FEEST. ES cells harboring an inducible Cre gene enabled the efficient selection of transgenic ES cell clones using hygromycin before Cre-mediated recombination. Expression of the GOI was confirmed by assaying for the GFP after Cre recombination. As a proof-of-principle, we produced a transgenic mouse line containing Cre-activatable tTA (cl-tTA6). This tTA mouse model was able to induce tumor formation when crossed with a transgenic mouse line containing a doxycycline-inducible oncogene. We also showed that the cl-tTA6 mouse is a valuable tool for faithfully recapitulating the clinical course of tumor development. We showed that FEEST can be easily adapted for other genes by preparing a transgenic mouse model of conditionally activatable EGFR L858R. Thus, FEEST is a technique with the potential to generate transgenic mouse models at a genome-wide scale. PMID:26573149

  20. Increased hepcidin in transferrin-treated thalassemic mice correlates with increased liver BMP2 expression and decreased hepatocyte ERK activation

    PubMed Central

    Chen, Huiyong; Choesang, Tenzin; Li, Huihui; Sun, Shuming; Pham, Petra; Bao, Weili; Feola, Maria; Westerman, Mark; Li, Guiyuan; Follenzi, Antonia; Blanc, Lionel; Rivella, Stefano; Fleming, Robert E.; Ginzburg, Yelena Z.

    2016-01-01

    Iron overload results in significant morbidity and mortality in β-thalassemic patients. Insufficient hepcidin is implicated in parenchymal iron overload in β-thalassemia and approaches to increase hepcidin have therapeutic potential. We have previously shown that exogenous apo-transferrin markedly ameliorates ineffective erythropoiesis and increases hepcidin expression in Hbbth1/th1 (thalassemic) mice. We utilize in vivo and in vitro systems to investigate effects of exogenous apo-transferrin on Smad and ERK1/2 signaling, pathways that participate in hepcidin regulation. Our results demonstrate that apo-transferrin increases hepcidin expression in vivo despite decreased circulating and parenchymal iron concentrations and unchanged liver Bmp6 mRNA expression in thalassemic mice. Hepatocytes from apo-transferrin-treated mice demonstrate decreased ERK1/2 pathway and increased serum BMP2 concentration and hepatocyte BMP2 expression. Furthermore, hepatocyte ERK1/2 phosphorylation is enhanced by neutralizing anti-BMP2/4 antibodies and suppressed in vitro in a dose-dependent manner by BMP2, resulting in converse effects on hepcidin expression, and hepatocytes treated with MEK/ERK1/2 inhibitor U0126 in combination with BMP2 exhibit an additive increase in hepcidin expression. Lastly, bone marrow erythroferrone expression is normalized in apo-transferrin treated thalassemic mice but increased in apo-transferrin injected wild-type mice. These findings suggest that increased hepcidin expression after exogenous apo-transferrin is in part independent of erythroferrone and support a model in which apo-transferrin treatment in thalassemic mice increases BMP2 expression in the liver and other organs, decreases hepatocellular ERK1/2 activation, and increases nuclear Smad to increase hepcidin expression in hepatocytes. PMID:26635037

  1. Enhanced glucose tolerance in pancreatic-derived factor (PANDER) knockout C57BL/6 mice.

    PubMed

    Moak, Shari L; Dougan, Grace C; MarElia, Catherine B; Danse, Whitney A; Fernandez, Amanda M; Kuehl, Melanie N; Athanason, Mark G; Burkhardt, Brant R

    2014-11-01

    Pancreatic-derived factor (PANDER; also known as FAM3B) is a uniquely structured protein strongly expressed within and secreted from the endocrine pancreas. PANDER has been hypothesized to regulate fasting and fed glucose homeostasis, hepatic lipogenesis and insulin signaling, and to serve a potential role in the onset or progression of type 2 diabetes (T2D). Despite having potentially pivotal pleiotropic roles in glycemic regulation and T2D, there has been limited generation of stable animal models for the investigation of PANDER function, and there are no models on well-established genetic murine backgrounds for T2D. Our aim was to generate an enhanced murine model to further elucidate the biological function of PANDER. Therefore, a pure-bred PANDER knockout C57BL/6 (PANKO-C57) model was created and phenotypically characterized with respect to glycemic regulation and hepatic insulin signaling. The PANKO-C57 model exhibited an enhanced metabolic phenotype, particularly with regard to enhanced glucose tolerance. Male PANKO-C57 mice displayed decreased fasting plasma insulin and C-peptide levels, whereas leptin levels were increased as compared with matched C57BL/6J wild-type mice. Despite similar peripheral insulin sensitivity between both groups, hepatic insulin signaling was significantly increased during fasting conditions, as demonstrated by increased phosphorylation of hepatic PKB/Akt and AMPK, along with mature SREBP-1 expression. Insulin stimulation of PANKO-C57 mice resulted in increased hepatic triglyceride and glycogen content as compared with wild-type C57BL/6 mice. In summary, the PANKO-C57 mouse represents a suitable model for the investigation of PANDER in multiple metabolic states and provides an additional tool to elucidate the biological function and potential role in T2D. PMID:25217499

  2. Enhanced glucose tolerance in pancreatic-derived factor (PANDER) knockout C57BL/6 mice

    PubMed Central

    Moak, Shari L.; Dougan, Grace C.; MarElia, Catherine B.; Danse, Whitney A.; Fernandez, Amanda M.; Kuehl, Melanie N.; Athanason, Mark G.; Burkhardt, Brant R.

    2014-01-01

    Pancreatic-derived factor (PANDER; also known as FAM3B) is a uniquely structured protein strongly expressed within and secreted from the endocrine pancreas. PANDER has been hypothesized to regulate fasting and fed glucose homeostasis, hepatic lipogenesis and insulin signaling, and to serve a potential role in the onset or progression of type 2 diabetes (T2D). Despite having potentially pivotal pleiotropic roles in glycemic regulation and T2D, there has been limited generation of stable animal models for the investigation of PANDER function, and there are no models on well-established genetic murine backgrounds for T2D. Our aim was to generate an enhanced murine model to further elucidate the biological function of PANDER. Therefore, a pure-bred PANDER knockout C57BL/6 (PANKO-C57) model was created and phenotypically characterized with respect to glycemic regulation and hepatic insulin signaling. The PANKO-C57 model exhibited an enhanced metabolic phenotype, particularly with regard to enhanced glucose tolerance. Male PANKO-C57 mice displayed decreased fasting plasma insulin and C-peptide levels, whereas leptin levels were increased as compared with matched C57BL/6J wild-type mice. Despite similar peripheral insulin sensitivity between both groups, hepatic insulin signaling was significantly increased during fasting conditions, as demonstrated by increased phosphorylation of hepatic PKB/Akt and AMPK, along with mature SREBP-1 expression. Insulin stimulation of PANKO-C57 mice resulted in increased hepatic triglyceride and glycogen content as compared with wild-type C57BL/6 mice. In summary, the PANKO-C57 mouse represents a suitable model for the investigation of PANDER in multiple metabolic states and provides an additional tool to elucidate the biological function and potential role in T2D. PMID:25217499

  3. Role of oxidative stress on diesel-enhanced influenza infection in mice

    PubMed Central

    2010-01-01

    Numerous studies have shown that air pollutants, including diesel exhaust (DE), reduce host defenses, resulting in decreased resistance to respiratory infections. This study sought to determine if DE exposure could affect the severity of an ongoing influenza infection in mice, and examine if this could be modulated with antioxidants. BALB/c mice were treated by oropharyngeal aspiration with 50 plaque forming units of influenza A/HongKong/8/68 and immediately exposed to air or 0.5 mg/m3 DE (4 hrs/day, 14 days). Mice were necropsied on days 1, 4, 8 and 14 post-infection and lungs were assessed for virus titers, lung inflammation, immune cytokine expression and pulmonary responsiveness (PR) to inhaled methacholine. Exposure to DE during the course of infection caused an increase in viral titers at days 4 and 8 post-infection, which was associated with increased neutrophils and protein in the BAL, and an early increase in PR. Increased virus load was not caused by decreased interferon levels, since IFN-β levels were enhanced in these mice. Expression and production of IL-4 was significantly increased on day 1 and 4 p.i. while expression of the Th1 cytokines, IFN-γ and IL-12p40 was decreased. Treatment with the antioxidant N-acetylcysteine did not affect diesel-enhanced virus titers but blocked the DE-induced changes in cytokine profiles and lung inflammation. We conclude that exposure to DE during an influenza infection polarizes the local immune responses to an IL-4 dominated profile in association with increased viral disease, and some aspects of this effect can be reversed with antioxidants. PMID:21092162

  4. Genetically Determined Susceptibility to Tuberculosis in Mice Causally Involves Accelerated and Enhanced Recruitment of Granulocytes

    PubMed Central

    Keller, Christine; Hoffmann, Reinhard; Lang, Roland; Brandau, Sven; Hermann, Corinna; Ehlers, Stefan

    2006-01-01

    Classical twin studies and recent linkage analyses of African populations have revealed a potential involvement of host genetic factors in susceptibility or resistance to Mycobacterium tuberculosis infection. In order to identify the candidate genes involved and test their causal implication, we capitalized on the mouse model of tuberculosis, since inbred mouse strains also differ substantially in their susceptibility to infection. Two susceptible and two resistant mouse strains were aerogenically infected with 1,000 CFU of M. tuberculosis, and the regulation of gene expression was examined by Affymetrix GeneChip U74A array with total lung RNA 2 and 4 weeks postinfection. Four weeks after infection, 96 genes, many of which are involved in inflammatory cell recruitment and activation, were regulated in common. One hundred seven genes were differentially regulated in susceptible mouse strains, whereas 43 genes were differentially expressed only in resistant mice. Data mining revealed a bias towards the expression of genes involved in granulocyte pathophysiology in susceptible mice, such as an upregulation of those for the neutrophil chemoattractant LIX (CXCL5), interleukin 17 receptor, phosphoinositide kinase 3 delta, or gamma interferon-inducible protein 10. Following M. tuberculosis challenge in both airways or peritoneum, granulocytes were recruited significantly faster and at higher numbers in susceptible than in resistant mice. When granulocytes were efficiently depleted by either of two regimens at the onset of infection, only susceptible mice survived aerosol challenge with M. tuberculosis significantly longer than control mice. We conclude that initially enhanced recruitment of granulocytes contributes to susceptibility to tuberculosis. PMID:16790804

  5. Muscle-directed gene therapy for phenylketonuria (PKU): Development of transgenic mice with muscle-specific phenylalanine hydroxylase expression

    SciTech Connect

    Harding, C.O.; Messing, A.; Wolff, J.A.

    1994-09-01

    Phenylketonuria (PKU) is an attractive target for gene therapy because of shortcomings in current therapy including lifelong commitment to a difficult and expensive diet, persistent mild cognitive deficits in some children despite adequate dietary therapy, and maternal PKU syndrome. Phenylalanine hydroxylase (PAH) is normally expressed only in liver, but we propose to treat PKU by introducing the gene for PAH into muscle. In order to evaluate both the safety and efficacy of this approach, we have a developed a trangenic mouse which expresses PAH in both cardiac and skeletal muscle. The transgene includes promoter and enhancer sequences from the mouse muscle creatine kinase (MCK) gene fused to the mouse liver PAH cDNA. Mice which have inherited the transgene are healthy, active, and do not exhibit any signs of muscle weakness or wasting. Ectopic PAH expression in muscle is not detrimental to the health, neurologic function, or reproduction of the mice. Pah{sup enu2} hyperphenylalaninemic mice, a model of human PAH deficiency, bred to carry the transgene have substantial PAH expression in cardiac and skeletal muscle but none in liver. Muscle PAH expression alone does not complement the hyperphenylalaninemic phenotype of Pah{sup enu2} mice. However, administration of reduced tetrahydrobiopterin to transgenic Pah{sup enu2} mice is associated with a 25% mean decrease in serum phenylalanine levels. We predict that ectopic expression of PAH in muscle along with adequate muscle supplies of reduced biopterin cofactor will decrease hyperphenylalaninemia in PKU.

  6. Expression levels of neuroimmune biomarkers in hypothalamus of allergic mice after phthalate exposure.

    PubMed

    Win-Shwe, Tin-Tin; Yanagisawa, Rie; Koike, Eiko; Nitta, Hiroshi; Takano, Hirohisa

    2013-10-01

    Previously, we demonstrated that maternal exposure to phthalates enhances atopic dermatitis in male mouse offspring. However, whether phthalate exposure affects neuroimmune biomarkers in allergic mice has not yet been studied. Di-(2-ethylhexyl) phthalate (DEHP) and di-isononyl phthalate (DINP) are environmental chemicals that are commonly used as plasticizers. This study was designed to investigate the expression levels of neuroimmune biomarkers in the hypothalamus of a murine model of allergic asthma after phthalate exposure throughout juvenility until adulthood. Six-week-old C3H/HeJ Jcl male mice were treated with DEHP or DINP (0, 0.02, 0.4 or 8 nmol per body per week) and ovalbumin (OVA; 1 µg per body per 2 weeks) for 7 weeks intratracheally. On the day after the completion of the phthalate and OVA treatment, the hypothalamus from each mouse was collected, and the mRNA expression levels of neuroimmune biomarkers were examined using a real-time RT-PCR analysis. The mRNA expression levels of the proinflammatory cytokines interleukin (IL)-1β and tumor necrosis factor (TNF)-α, the chemokine CCL3, the transcription factor nuclear factor (NF)-κB, the oxidative stress marker heme-oxygenase (HO)1, a nerve growth factor, and the microglia marker Iba1 were remarkably up-regulated in the hypothalami of mice treated with 8 nmol of DEHP in the presence of the allergen. However, no significant changes were observed, except for reductions in the TNF-α and CCL2 mRNA levels, in mice exposed to DINP combined with the allergen. This study is the first report to show that high-dose DEHP exposure throughout juvenility until adulthood may induce neuroinflammation by modulating neuroimmune biomarkers in the hypothalami of allergic mice. PMID:23148021

  7. Human IL-32 expression protects mice against a hypervirulent strain of Mycobacterium tuberculosis

    PubMed Central

    Bai, Xiyuan; Shang, Shaobin; Henao-Tamayo, Marcela; Basaraba, Randall J.; Ovrutsky, Alida R.; Matsuda, Jennifer L.; Takeda, Katsuyuki; Chan, Mallory M.; Dakhama, Azzeddine; Kinney, William H.; Trostel, Jessica; Bai, An; Honda, Jennifer R.; Achcar, Rosane; Hartney, John; Joosten, Leo A. B.; Kim, Soo-Hyun; Orme, Ian; Dinarello, Charles A.; Ordway, Diane J.; Chan, Edward D.

    2015-01-01

    Silencing of interleukin-32 (IL-32) in a differentiated human promonocytic cell line impairs killing of Mycobacterium tuberculosis (MTB) but the role of IL-32 in vivo against MTB remains unknown. To study the effects of IL-32 in vivo, a transgenic mouse was generated in which the human IL-32γ gene is expressed using the surfactant protein C promoter (SPC-IL-32γTg). Wild-type and SPC-IL-32γTg mice were infected with a low-dose aerosol of a hypervirulent strain of MTB (W-Beijing HN878). At 30 and 60 d after infection, the transgenic mice had 66% and 85% fewer MTB in the lungs and 49% and 68% fewer MTB in the spleens, respectively; the transgenic mice also exhibited greater survival. Increased numbers of host-protective innate and adaptive immune cells were present in SPC-IL-32γTg mice, including tumor necrosis factor-alpha (TNFα) positive lung macrophages and dendritic cells, and IFN-gamma (IFNγ) and TNFα positive CD4+ and CD8+ T cells in the lungs and mediastinal lymph nodes. Alveolar macrophages from transgenic mice infected with MTB ex vivo had reduced bacterial burden and increased colocalization of green fluorescent protein-labeled MTB with lysosomes. Furthermore, mouse macrophages made to express IL-32γ but not the splice variant IL-32β were better able to limit MTB growth than macrophages capable of producing both. The lungs of patients with tuberculosis showed increased IL-32 expression, particularly in macrophages of granulomas and airway epithelial cells but also B cells and T cells. We conclude that IL-32γ enhances host immunity to MTB. PMID:25820174

  8. Nck2 Deficiency in Mice Results in Increased Adiposity Associated With Adipocyte Hypertrophy and Enhanced Adipogenesis.

    PubMed

    Dusseault, Julie; Li, Bing; Haider, Nida; Goyette, Marie-Anne; Côté, Jean-François; Larose, Louise

    2016-09-01

    Obesity results from an excessive expansion of white adipose tissue (WAT) from hypertrophy of preexisting adipocytes and enhancement of precursor differentiation into mature adipocytes. We report that Nck2-deficient mice display progressive increased adiposity associated with adipocyte hypertrophy. A negative relationship between the expression of Nck2 and WAT expansion was recapitulated in humans such that reduced Nck2 protein and mRNA levels in human visceral WAT significantly correlate with the degree of obesity. Accordingly, Nck2 deficiency promotes an adipogenic program that not only enhances adipocyte differentiation and lipid droplet formation but also results in dysfunctional elevated lipogenesis and lipolysis activities in mouse WAT as well as in stromal vascular fraction and 3T3-L1 preadipocytes. We provide strong evidence to support that through a mechanism involving primed PERK activation and signaling, Nck2 deficiency in adipocyte precursors is associated with enhanced adipogenesis in vitro and adiposity in vivo. Finally, in agreement with elevated circulating lipids, Nck2-deficient mice develop glucose intolerance, insulin resistance, and hepatic steatosis. Taken together, these findings reveal that Nck2 is a novel regulator of adiposity and suggest that Nck2 is important in limiting WAT expansion and dysfunction in mice and humans. PMID:27325288

  9. Cognitive-Enhancing Effect of Aronia melanocarpa Extract against Memory Impairment Induced by Scopolamine in Mice.

    PubMed

    Lee, Hyeon Yong; Weon, Jin Bae; Jung, Youn Sik; Kim, Nam Young; Kim, Myong Ki; Ma, Choong Je

    2016-01-01

    Aronia melanocarpa (A. melanocarpa) berries are a fruit with a marked antioxidant effect. The objective of this study was to confirm the effect of A. melanocarpa berries extract against scopolamine-induced memory impairment in mice using the Morris water maze and passive avoidance test. Moreover, we determined a possible mechanism of the cognitive-enhancing effect involving AChE activity and BDNF and p-CREB expression in the hippocampus of mice. A. melanocarpa berries extract attenuated the learning and memory impairment induced by scopolamine in the Morris water maze (79.3 ± 0.8 s of 200 mg/kg and 64.4 ± 10.7 s of 400 mg/kg on day 4) and passive avoidance tests (46.0 ± 41.1 s of 200 mg/kg and 25.6 ± 18.7 s of 400 mg/kg). A. melanocarpa berries extract reduced the acetylcholinesterase level in the hippocampus of scopolamine-injected mice and increased BDNF and p-CREB expression in the hippocampus. The major compound, cyanidin-3-O-galactoside, also reversed memory impairment. These results showed that A. melanocarpa berries extract improved memory impairment by inhibiting AChE and increasing BDNF and p-CREB expression, and cyanidin-3-O-galactoside may be responsible for the effect of A. melanocarpa berries extract. PMID:27239211

  10. Cognitive-Enhancing Effect of Aronia melanocarpa Extract against Memory Impairment Induced by Scopolamine in Mice

    PubMed Central

    Lee, Hyeon Yong; Weon, Jin Bae; Jung, Youn Sik; Kim, Nam Young; Kim, Myong Ki; Ma, Choong Je

    2016-01-01

    Aronia melanocarpa (A. melanocarpa) berries are a fruit with a marked antioxidant effect. The objective of this study was to confirm the effect of A. melanocarpa berries extract against scopolamine-induced memory impairment in mice using the Morris water maze and passive avoidance test. Moreover, we determined a possible mechanism of the cognitive-enhancing effect involving AChE activity and BDNF and p-CREB expression in the hippocampus of mice. A. melanocarpa berries extract attenuated the learning and memory impairment induced by scopolamine in the Morris water maze (79.3 ± 0.8 s of 200 mg/kg and 64.4 ± 10.7 s of 400 mg/kg on day 4) and passive avoidance tests (46.0 ± 41.1 s of 200 mg/kg and 25.6 ± 18.7 s of 400 mg/kg). A. melanocarpa berries extract reduced the acetylcholinesterase level in the hippocampus of scopolamine-injected mice and increased BDNF and p-CREB expression in the hippocampus. The major compound, cyanidin-3-O-galactoside, also reversed memory impairment. These results showed that A. melanocarpa berries extract improved memory impairment by inhibiting AChE and increasing BDNF and p-CREB expression, and cyanidin-3-O-galactoside may be responsible for the effect of A. melanocarpa berries extract. PMID:27239211

  11. Conditional Expression of the Androgen Receptor Increases Susceptibility of Bladder Cancer in Mice

    PubMed Central

    Luong, Richard; Yu, Eun-Jeong; He, Yongfeng; Gonzalgo, Mark L.; Sun, Zijie

    2016-01-01

    Bladder cancer represents a significant human tumor burden, accounting for about 7.7% and 2.4% of all cancer cases in males and females, respectively. While men have a higher risk of developing bladder cancer, women tend to present at a later stage of disease and with more aggressive tumors. Previous studies have suggested a promotional role of androgen signaling in enhancing bladder cancer development. To directly assess the role of androgens in bladder tumorigenesis, we have developed a novel transgenic mouse strain, R26hARLoxP/+:Upk3aGCE/+, in which the human AR transgene is conditionally expressed in bladder urothelium. Intriguingly, both male and female R26hARLoxP/+:Upk3aGCE/+ mice display a higher incidence of urothelial cell carcinoma (UCC) than the age and sex matched control littermates in response to the carcinogen, N-butyl-N-(4-hydroxybutyl) nitrosamine (BBN). We detect expression of the human AR transgene in CK5-positive and p63-positive basal cells in bladder urothelium. Further analyses of UCC tissues from R26hARLoxP/+:Upk3aGCE/+ mice showed that the majority of tumor cells are of urothelial basal cell origin. Positive immunostaining of transgenic AR protein was observed in the majority of tumor cells of the transgenic mice, providing a link between transgenic AR expression and oncogenic transformation. We observed an increase in Ki67 positive cells within the UCC lesions of transgenic AR mice. Manipulating endogenous androgen levels by castration and androgen supplementation directly affected bladder tumor development in male and female R26hARLoxP/+:Upk3aGCE/+ mice, respectively. Taken together, our data demonstrate for the first time that conditional activation of transgenic AR expression in bladder urothelium enhances carciongen-induced bladder tumor formation in mice. This new AR transgenic mouse line mimics certain features of human bladder cancer and can be used to study bladder tumorigenesis and for drug development. PMID:26862755

  12. Conditional Expression of the Androgen Receptor Increases Susceptibility of Bladder Cancer in Mice.

    PubMed

    Johnson, Daniel T; Hooker, Erika; Luong, Richard; Yu, Eun-Jeong; He, Yongfeng; Gonzalgo, Mark L; Sun, Zijie

    2016-01-01

    Bladder cancer represents a significant human tumor burden, accounting for about 7.7% and 2.4% of all cancer cases in males and females, respectively. While men have a higher risk of developing bladder cancer, women tend to present at a later stage of disease and with more aggressive tumors. Previous studies have suggested a promotional role of androgen signaling in enhancing bladder cancer development. To directly assess the role of androgens in bladder tumorigenesis, we have developed a novel transgenic mouse strain, R26hARLoxP/+:Upk3aGCE/+, in which the human AR transgene is conditionally expressed in bladder urothelium. Intriguingly, both male and female R26hARLoxP/+:Upk3aGCE/+ mice display a higher incidence of urothelial cell carcinoma (UCC) than the age and sex matched control littermates in response to the carcinogen, N-butyl-N-(4-hydroxybutyl) nitrosamine (BBN). We detect expression of the human AR transgene in CK5-positive and p63-positive basal cells in bladder urothelium. Further analyses of UCC tissues from R26hARLoxP/+:Upk3aGCE/+ mice showed that the majority of tumor cells are of urothelial basal cell origin. Positive immunostaining of transgenic AR protein was observed in the majority of tumor cells of the transgenic mice, providing a link between transgenic AR expression and oncogenic transformation. We observed an increase in Ki67 positive cells within the UCC lesions of transgenic AR mice. Manipulating endogenous androgen levels by castration and androgen supplementation directly affected bladder tumor development in male and female R26hARLoxP/+:Upk3aGCE/+ mice, respectively. Taken together, our data demonstrate for the first time that conditional activation of transgenic AR expression in bladder urothelium enhances carciongen-induced bladder tumor formation in mice. This new AR transgenic mouse line mimics certain features of human bladder cancer and can be used to study bladder tumorigenesis and for drug development. PMID:26862755

  13. Neuroglobin Over Expressing Mice: Expression Pattern and Effect on Brain Ischemic Infarct Size

    PubMed Central

    Raida, Zindy; Hundahl, Christian Ansgar; Nyengaard, Jens R.; Hay-Schmidt, Anders

    2013-01-01

    Background Stroke is a major cause of death and severe disability, but effective treatments are limited. Neuroglobin, a neuronal heme-globin, has been advocated as a novel pharmacological target in combating stroke and neurodegenerative disorders based on cytoprotective properties. Using thoroughly validated antibodies and oligos, we give a detailed brain anatomical characterization of transgenic mice over expressing Neuroglobin. Moreover, using permanent middle artery occlusion the effect of elevated levels of Neuroglobin on ischemic damage was studied. Lastly, the impact of mouse strain genetic background on ischemic damage was investigated. Principal Findings A four to five fold increase in Neuroglobin mRNA and protein expression was seen in the brain of transgenic mice. A β-actin promoter was used to drive Neuroglobin over expression, but immunohistochemistry and in situ hybridization showed over expression to be confined to primarily the cortex, hippocampus, cerebellum, and only in neurons. The level and expression pattern of endogenous Neuroglobin was unaffected by insertion of the over expressing Ngb transgene. Neuroglobin over expression resulted in a significant reduction in infarct volume 24 hours after ischemia. Immunohistochemistry showed no selective sparing of Neuroglobin expressing cells in the ischemic core or penumbra. A significant difference in infarct volume was found between mice of the same strain, but from different colonies. Significance In contrast to some previous reports, Neuroglobin over expression is not global but confined to a few well-defined brain regions, and only in neurons. This study confirms previous reports showing a correlation between reduced infarct volume and elevated Neuroglobin levels, but underlines the need to study the likely contribution from compensatory mechanisms to the phenotype following a genetic perturbation. We also stress, that care should be taken when comparing results where different mouse strains and

  14. Interleukin 6 Deficiency Modulates the Hypothalamic Expression of Energy Balance Regulating Peptides during Pregnancy in Mice

    PubMed Central

    Pazos, Patricia; Lima, Luis; Casanueva, Felipe F.; Diéguez, Carlos; García, María C.

    2013-01-01

    Pregnancy is associated with hyperphagia, increased adiposity and multiple neuroendocrine adaptations. Maternal adipose tissue secretes rising amounts of interleukin 6 (IL6), which acts peripherally modulating metabolic function and centrally increasing energy expenditure and reducing body fat. To explore the role of IL6 in the central mechanisms governing dam's energy homeostasis, early, mid and late pregnant (gestational days 7, 13 and 18) wild-type (WT) and Il6 knockout mice (Il6-KO) were compared with virgin controls at diestrus. Food intake, body weight and composition as well as indirect calorimetry measurements were performed in vivo. Anabolic and orexigenic peptides: neuropeptide Y (Npy) and agouti-related peptide (Agrp); and catabolic and anorectic neuropeptides: proopiomelanocortin (Pomc), corticotrophin and thyrotropin-releasing hormone (Crh and Trh) mRNA levels were determined by in situ hybridization. Real time-PCR and western-blot were used for additional tissue gene expression and protein studies. Non-pregnant Il6-KO mice were leaner than WT mice due to a decrease in fat but not in lean body mass. Pregnant Il6-KO mice had higher fat accretion despite similar body weight gain than WT controls. A decreased fat utilization in absence of Il6 might explain this effect, as shown by increased respiratory exchange ratio (RER) in virgin Il6-KO mice. Il6 mRNA levels were markedly enhanced in adipose tissue but reduced in hypothalamus of mid and late pregnant WT mice. Trh expression was also stimulated at gestational day 13 and lack of Il6 blunted this effect. Conversely, in late pregnant mice lessened hypothalamic Il6 receptor alpha (Il6ra), Pomc and Crh mRNA were observed. Il6 deficiency during this stage up-regulated Npy and Agrp expression, while restoring Pomc mRNA levels to virgin values. Together these results demonstrate that IL6/IL6Ra system modulates Npy/Agrp, Pomc and Trh expression during mouse pregnancy, supporting a role of IL6 in the central

  15. Liver specific expression of Cu/ZnSOD extends the lifespan of Sod1 null mice

    PubMed Central

    Zhang, Yiqiang; Liu, Yuhong; Walsh, Michael; Bokov, Alex; Ikeno, Yuji; Jang, Young C.; Perez, Viviana I.; Van Remmen, Holly; Richardson, Arlan

    2016-01-01

    Genetic ablation of CuZn-superoxide dismutase (Sod1) in mice (Sod1−/− mice) leads to shortened lifespan with a dramatic increase in hepatocellular carcinoma and accelerated aging phenotypes, including early onset sarcopenia. To study the tissue specific effects of oxidative stress in the Sod1−/− mice, we generated mice that only express the human SOD1 gene specifically in the liver of Sod1−/− mice (Sod1−/−/hSOD1alb mice). Expression of hSOD1 in the liver of Sod1−/− mice improved liver function, reduced oxidative damage in liver, and partially restored the expression of several genes involved in tumorigenesis, which are abnormally expressed in the livers of the Sod1−/− mice. However, liver specific expression of hSOD1 did not prevent the loss of body weight and muscle mass and alterations in the structure of neuromuscular junctions. The expression of hSOD1 in the liver of Sod1−/− mice significantly improved the lifespan of Sod1−/− mice; however, the lifespan of the Sod1−/−/hSOD1alb mice was still significantly shorter than wild type mice. PMID:26839948

  16. Increased microvascular density and enhanced leukocyte rolling and adhesion in the skin of VEGF transgenic mice.

    PubMed

    Detmar, M; Brown, L F; Schön, M P; Elicker, B M; Velasco, P; Richard, L; Fukumura, D; Monsky, W; Claffey, K P; Jain, R K

    1998-07-01

    Vascular endothelial growth factor (VEGF) has been implicated in the pathologic angiogenesis observed in psoriasis and other chronic inflammatory skin diseases that are characterized by enhanced expression of VEGF by epidermal keratinocytes and of VEGF receptors by tortuous microvessels in the upper dermis. To investigate the functional importance of chronic VEGF overexpression in vivo, we used a keratin 14 promoter expression cassette containing the gene for murine VEGF164 to selectively target VEGF expression to basal epidermal keratinocytes in transgenic mice. These mice demonstrated an increased density of tortuous cutaneous blood capillaries with elevated expression levels of the high affinity VEGF receptors, VEGFR-1 and VEGFR-2, most prominently during the neonatal period. In contrast, no abnormalities of lymphatic vessels were detected. In addition, the number of mast cells in the upper dermis was significantly increased in transgenic skin. Intravital fluorescence microscopy revealed highly increased leukocyte rolling and adhesion in postcapillary skin venules that were both inhibited after injection of blocking antibodies against E- and P-selectin. Combined blocking antibodies against intercellular adhesion molecule-1 and lymphocyte function-associated antigen-1 were without effect, whereas an anti-vascular cell adhesion molecule-1/VLA-4 antibody combination almost completely normalized the enhanced leukocyte adhesion in transgenic mice. This study reveals VEGF as a growth factor specific for blood vessels, but not lymphatic vessels, and demonstrates that chronic orthotopic overexpression of VEGF in the epidermis is sufficient to induce cardinal features of chronic skin inflammation, providing a molecular link between angiogenesis, mast cell accumulation, and leukocyte recruitment to sites of inflammation. PMID:9665379

  17. The enhancement of immune function and activation of NF-κB by resveratrol-treatment in immunosuppressive mice.

    PubMed

    Lai, Xin; Pei, Qingsheng; Song, Xu; Zhou, Xun; Yin, Zhongqiong; Jia, Renyong; Zou, Yuanfeng; Li, Lixia; Yue, Guizhou; Liang, Xiaoxia; Yin, Lizi; Lv, Cheng; Jing, Bo

    2016-04-01

    Resveratrol, a kind of natural product found in over 70 plants, possesses both immunomodulatory and anticancer effects. Many reports have shown that resveratrol has the bidirectional regulation effects on antigen presenting and cellular immunity. However, few reports have evaluated the effects of resveratrol on reinforcing immunity recovery via activating nuclear factor -κappa B (NF-κB). In the present study, we investigated the effects of resveratrol on recovery and reconstruction of immune function by detecting nonspecific and specific immunity in immunosuppressive mice. We found that, compared to the immunosuppressive mice, the spleen index and spleen lymphocyte proliferation of resveratrol-treated mice (30mg/kg body weight) were enhanced. After resveratrol-treatment (15mg/kg body weight), the function of peritoneal macrophages was enhanced and the CD4+ cells were increased in peripheral blood. The expressions of serum cytokines related to immune function, including interleukin (IL)-1α/β, IL-2, tumor necrosis factor-α and NF-κB were up-regulated in a dose-dependent manner. The expression of the transcription factor NF-κB in spleen was enhanced after resveratrol-treatment. The immuno-enhancement effects of resveratrol were similar to that of levamisole (served as positive control). These results demonstrated that resveratrol had potent immune enhancement activity in immunosuppressive mice, and one possible mechanism of action was to activate the NF-κB. PMID:26854575

  18. Gender-specific reduction of hepatic Mrp2 expression by high-fat diet protects female mice from ANIT toxicity

    SciTech Connect

    Kong, Bo; Csanaky, Iván L.; Aleksunes, Lauren M.; Patni, Meghan; Chen, Qi; Ma, Xiaochao; Jaeschke, Hartmut; Weir, Scott; Broward, Melinda; Klaassen, Curtis D.; Guo, Grace L.

    2012-06-01

    Emerging evidence suggests that feeding a high-fat diet (HFD) to rodents affects the expression of genes involved in drug transport. However, gender-specific effects of HFD on drug transport are not known. The multidrug resistance-associated protein 2 (Mrp2, Abcc2) is a transporter highly expressed in the hepatocyte canalicular membrane and is important for biliary excretion of glutathione-conjugated chemicals. The current study showed that hepatic Mrp2 expression was reduced by HFD feeding only in female, but not male, C57BL/6J mice. In order to determine whether down-regulation of Mrp2 in female mice altered chemical disposition and toxicity, the biliary excretion and hepatotoxicity of the Mrp2 substrate, α-naphthylisothiocyanate (ANIT), were assessed in male and female mice fed control diet or HFD for 4 weeks. ANIT-induced biliary injury is a commonly used model of experimental cholestasis and has been shown to be dependent upon Mrp2-mediated efflux of an ANIT glutathione conjugate that selectively injures biliary epithelial cells. Interestingly, HFD feeding significantly reduced early-phase biliary ANIT excretion in female mice and largely protected against ANIT-induced liver injury. In summary, the current study showed that, at least in mice, HFD feeding can differentially regulate Mrp2 expression and function and depending upon the chemical exposure may enhance or reduce susceptibility to toxicity. Taken together, these data provide a novel interaction between diet and gender in regulating hepatobiliary excretion and susceptibility to injury. -- Highlights: ► High-fat diet decreases hepatic Mrp2 expression only in female but not in male mice. ► HFD significantly reduces early-phase biliary ANIT excretion in female mice. ► HFD protects female mice against ANIT-induced liver injury.

  19. Overexpression of motor protein KIF17 enhances spatial and working memory in transgenic mice.

    PubMed

    Wong, Richard Wing-Chuen; Setou, Mitsutoshi; Teng, Junlin; Takei, Yosuke; Hirokawa, Nobutaka

    2002-10-29

    The kinesin superfamily proteins (KIFs) play essential roles in receptor transportation along the microtubules. KIF17 transports the N-methyl-d-aspartate receptor NR2B subunit in vitro, but its role in vivo is unknown. To clarify this role, we generated transgenic mice overexpressing KIF17 tagged with GFP. The KIF17 transgenic mice exhibited enhanced learning and memory in a series of behavioral tasks, up-regulated NR2B expression with the potential involvement of a transcriptional factor, the cAMP-dependent response element-binding protein, and increased phosphorylation of the cAMP-dependent response element-binding protein. Our results suggest that the motor protein KIF17 contributes to neuronal events required for learning and memory by trafficking fundamental N-methyl-d-aspartate-type glutamate receptors. PMID:12391294

  20. STAT5A expression in Swiss 3T3 cells promotes adipogenesis in vivo in an athymic mice model system.

    PubMed

    Stewart, William C; Pearcy, Lisa A; Floyd, Z Elizabeth; Stephens, Jacqueline M

    2011-09-01

    Many studies from our laboratories and others have shown that STAT5 expression and activity are increased during adipogenesis of murine and human adipocytes. Ectopic expression of STAT5A in fibroblasts or preadipocytes can confer or enhance adipogenesis. To determine whether STAT5A also plays a role in adipogenesis in vivo, we injected athymic mice with Swiss 3T3 cells expressing an empty pBABE retrovirus, Swiss cells expressing a pBABE retrovirus-containing STAT5A, or 3T3-F442A preadipocytes. Athymic mice injected with either 3T3-F442A cells or Swiss 3T3 cells expressing STAT5A resulted in fat pad formation at the site of injection. However, mice injected with Swiss cells containing the parent retroviral vector did not have any observable fat pads. An analysis of the ectopic fat pads obtained from the Swiss 3T3 STAT5A mice revealed abundant expression of peroxisome proliferator-activated receptor-γ (PPAR-γ) and adiponectin. The protein levels of both of these fat cell markers were comparable to expression levels in epididymal fat pads. These results demonstrate that STAT5A can promote adipogenesis in vivo in this model system which supports a role of this transcription factor in adipocyte development in the whole animal. PMID:21494231

  1. The VpreB1 enhancer drives developmental stage-specific gene expression in vivo.

    PubMed

    Licence, Steve; Persson, Christine; Mundt, Cornelia; Mårtensson, Inga-Lill

    2003-04-01

    In adult mice, the VpreB genes are expressed in bone marrow progenitor (pro-) and precursor (pre-) B cells. As part of the pre-B cell receptor, the proteins are crucial for the proliferation of these cells and consequently normal B lymphocyte development. Using cell lines, we identified a lineage- and developmental-stage-specific VpreB1 enhancer. Here, we analyze its specificity in vivo by generating transgenic mice in which expression of a reporter gene (human CD122) is regulated by the VpreB1 enhancer in the context of its own promoter. All transgenic lines expressed the reporter gene in the bone marrow in a copy number-independent manner, whereas expression levels were integration site-dependent. While the enhancer is not tissue specific, within the B cell lineage the expression pattern of human CD122 mimicked that of endogenous VpreB1. Thus, low levels were detected in pro-B cells, high levels in pre-BI and slightly lower levels in pre-BII cells; no expression was detected in immature/mature B cells. Furthermore, when in vitro cultured transgenic pre-B cells differentiated into immature B cells there was concomitant down-regulation of human CD122 and endogenous VpreB1. Thus the VpreB1 enhancer is sufficient to ensure developmental stage-specific expression of a reporter gene in B lymphocytes in vivo. PMID:12672078

  2. Enhancement of alcohol drinking in mice depends on alterations in RNA editing of serotonin 2C receptors

    PubMed Central

    Watanabe, Yoshihisa; Yoshimoto, Kanji; Tatebe, Harutsugu; Kita, Masakazu; Nishikura, Kazuko; Kimura, Minoru; Tanaka, Masaki

    2014-01-01

    Serotonin 2C receptors (5-HT2CR) are G-protein-coupled receptors with various actions, including involvement in drug addiction. 5-HT2CR undergoes mRNA editing, converting genomically encoded adenosine residues to inosines via adenosine deaminases acting on RNA (ADARs). Here we show that enhanced alcohol drinking behaviour in mice is associated with the degree of 5-HT2CR mRNA editing in the nucleus accumbens and dorsal raphe nuceus, brain regions important for reward and addiction. Following chronic alcohol vapour exposure, voluntary alcohol intake increased in C57BL/6J mice, but remained unchanged in C3H/HeJ and DBA/2J mice. 5-HT2CR mRNA editing frequency in both regions increased significantly in C57BL/6J mice, as did expressions of 5-HT2CR, ADAR1 and ADAR2, but not in other strains. Moreover, mice that exclusively express the unedited isoform (INI) of 5-HT2CR mRNA on a C57BL/6J background did not exhibit increased alcohol intake compared with wild-type mice. Our results indicate that alterations in 5-HT2CR mRNA editing underlie alcohol preference in mice. PMID:24345557

  3. RNA Interference of Myocyte Enhancer Factor 2A Accelerates Atherosclerosis in Apolipoprotein E-Deficient Mice

    PubMed Central

    Zhao, Yu-xia; Liu, Gang-qiong; Zhang, Jin-ying

    2015-01-01

    Objective Myocyte enhancer factor-2A (MEF 2A) has been shown to be involved in atherosclerotic lesion development, but its role in preexisting lesions is still unclear. In the present study we aim to assess the role of MEF 2A in the progression of pre-existing atherosclerosis. Methods Eighty apolipoprotein E-deficient mice (APOE KO) were randomly allocated to control, scramble and MEF 2A RNA interference (RNAi) groups, and constrictive collars were used to induce plaque formation. Six weeks after surgery, lentiviral shRNA construct was used to silence the expression of MEF 2A. Carotid plaques were harvested for analysis 4 weeks after viral vector transduction. Inflammatory gene expression in the plasma and carotid plaques was determined by using ELISAs and real-time RT-PCR. Results The expression level of MEF 2A was significantly reduced in plasma and plaque in the RNAi group, compared to the control and NC groups, whereas the expression level of pro-inflammatory cytokines was markedly increased. Silencing MEF 2A using lentiviral shRNA significantly reduced the plaque collagen content and fibrous cap thickness, as well as increased plaque area. However, silencing MEF 2A had no obvious effect on plaque lipid content. Conclusions Lentivirus-mediated MEF 2A shRNA accelerates inflammation and atherosclerosis in APOE KO mice, but has no effect on lipoprotein levels in plasma. PMID:25793529

  4. Lethal cutaneous disease in transgenic mice conditionally expressing type I human T cell leukemia virus Tax.

    PubMed

    Kwon, Hakju; Ogle, Louise; Benitez, Bobby; Bohuslav, Jan; Montano, Mauricio; Felsher, Dean W; Greene, Warner C

    2005-10-21

    Type I human T cell leukemia virus (HTLV-I) is etiologically linked with adult T cell leukemia, an aggressive and usually fatal expansion of activated CD4+ T lymphocytes that frequently traffic to skin. T cell transformation induced by HTLV-I involves the action of the 40-kDa viral Tax transactivator protein. Tax both stimulates the HTLV-I long terminal repeat and deregulates the expression of select cellular genes by altering the activity of specific host transcription factors, including cyclic AMP-responsive element-binding protein (CREB)/activating transcription factor, NF-kappaB/Rel, and serum response factor. To study initiating events involved in HTLV-I Tax-induced T cell transformation, we generated "Tet-off" transgenic mice conditionally expressing in a lymphocyte-restricted manner (EmuSR alpha promoter-enhancer) either wild-type Tax or mutant forms of Tax that selectively compromise the NF-kappaB (M22) or CREB/activating transcription factor (M47) activation pathways. Wild-type Tax and M47 Tax-expressing mice, but not M22-Tax expressing mice, developed progressive alopecia, hyperkeratosis, and skin lesions containing profuse activated CD4 T cell infiltrates with evidence of deregulated inflammatory cytokine production. In addition, these animals displayed systemic lymphadenopathy and splenomegaly. These findings suggest that Tax-mediated activation of NF-kappaB plays a key role in the development of this aggressive skin disease that shares several features in common with the skin disease occurring during the preleukemic stage in HTLV-I-infected patients. Of note, this skin disease completely resolved when Tax transgene expression was suppressed by administration of doxycycline, emphasizing the key role played by this viral oncoprotein in the observed pathology. PMID:16105841

  5. Generation of transgenic mice with liver-specific expression of human nuclear receptor nr5a2.

    PubMed

    Wang, Shui-Liang; Yang, Hua; Xie, You-Hua; Wang, Yuan; Li, Jian-Zhong; Wang, Long; Wang, Zhu-Gang; Fu, Ji-Liang

    2005-12-01

    Human nuclear receptor hb1 f(nr5a2) was cloned and characterized as a novel member of the Ftz-F1 (nr5a) nuclear receptor subfamily,whose its biological function remained largely unidentified. The aim of this study was to establish transgenic mouse model that specifically expressed hB1F in the liver to faciliate the functional study of hB1F. hb1f cDNA was placed downstream of mouse albumin gene enhancer/promoter to construct a liver-specific hb1f expression vector. Transgene fragments were microinjected into fertilized eggs of mice. The manipulated embryos were transferred into the oviducts of pseudopregnant female mice. Four offspring were identified as carrying the transgenes by PCR,from which one was also verified by Southern blotting. RT-PCR and Western blotting results showed that the transgene was expressed in the liver of the transgenic mice. Transgenic founder mice were used to establish transgenic mouse lineages. The F1 mice were identified by PCR analysis. Genetic analysis of the transgenic mice demonstrated that the transgene had been integrated into the chromosome at a single site and could be stably transmitted. PMID:16459652

  6. Dietary wolfberry supplementation enhances the protective effect of flu vaccine against influenza challenge in aged mice.

    PubMed

    Du, Xiaogang; Wang, Junpeng; Niu, Xinli; Smith, Donald; Wu, Dayong; Meydani, Simin Nikbin

    2014-02-01

    Current vaccines for influenza do not fully protect the aged against influenza infection. Although wolfberry (goji berry) has been shown to improve immune response, including enhanced antibody production, after vaccination in the aged, it is not known if this effect would translate to better protection after influenza infection, nor is its underlying mechanism well understood. To address these issues, we conducted a study using a 2 × 2 design in which aged male mice (20-22 mo) were fed a control or a 5% wolfberry diet for 30 d, then immunized with an influenza vaccine or saline (control) on days 31 and 52 of the dietary intervention, and finally challenged with influenza A/Puerto Rico/8/34 virus. Mice fed wolfberry had higher influenza antibody titers and improved symptoms (less postinfection weight loss) compared with the mice treated by vaccine alone. Furthermore, an in vitro mechanistic study showed that wolfberry supplementation enhanced maturation and activity of antigen-presenting dendritic cells (DCs) in aged mice, as indicated by phenotypic change in expression of DC activation markers major histocompatibility complex class II, cluster of differentiation (CD) 40, CD80, and CD86, and functional change in DC production of cytokines interleukin-12 and tumor necrosis factor-α as well as DC endocytosis. Also, adoptive transfer of wolfberry-treated bone marrow DCs (loaded with ovalbumin(323-339)-peptide) promoted antigen-specific T cell proliferation as well as interleukin-4 and interferon-γ production in CD4(+) T cells. In summary, our data indicate that dietary wolfberry enhances the efficacy of influenza vaccination, resulting in better host protection to prevent subsequent influenza infection; this effect may be partly attributed to improved DC function. PMID:24336457

  7. Pleurotus nebrodensis polysaccharide (PN-S) enhances the immunity of immunosuppressed mice.

    PubMed

    Cui, Hai-Yan; Wang, Chang-Lu; Wang, Yu-Rong; Li, Zhen-Jing; Chen, Mian-Hua; Li, Feng-Juan; Sun, Yan-Ping

    2015-10-01

    In the present study, the effects of Pleurotus nebrodensis polysaccharide (PN-S) on the immune functions of immunosuppressed mice were determined. The immunosuppressed mouse model was established by treating the mice with cyclophosphamide (40 mg/kg/2d, CY) through intraperitoneal injection. The results showed that PN-S administration significantly reversed the CY-induced weight loss, increased the thymic and splenic indices, and promoted proliferation of T lymphocyte, B lymphocyte, and macrophages. PN-S also enhanced the activity of natural killer cells and increased the immunoglobulin M (IgM) and immunoglobulin G (IgG) levels in the serum. In addition, PN-S treatment significantly increased the phagocytic activity of mouse peritoneal macrophages. PN-S also increased the levels of interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α), interferon-γ (INF-γ), and nitric oxide (NOS) in splenocytes. qRT-PCR results also indicated that PN-S increased the mRNA expression of IL-6, TNF-α, INF-γ, and nitric oxide synthase (iNOS) in the splenocytes. These results suggest that PN-S treatment enhances the immune function of immunosuppressed mice. This study may provide a basis for the application of this fungus in adjacent immunopotentiating therapy against cancer and in the treatment of chemotherapy-induced immunosuppression. PMID:26481376

  8. C/EBPε mediates nicotinamide-enhanced clearance of Staphylococcus aureus in mice

    PubMed Central

    Kyme, Pierre; Thoennissen, Nils H.; Tseng, Ching Wen; Thoennissen, Gabriela B.; Wolf, Andrea J.; Shimada, Kenichi; Krug, Utz O.; Lee, Kunik; Müller-Tidow, Carsten; Berdel, Wolfgang E.; Hardy, W. David; Gombart, Adrian F.; Koeffler, H. Phillip; Liu, George Y.

    2012-01-01

    The myeloid-specific transcription factor, CCAAT/enhancer-binding protein ε (C/EBPε) is a critical mediator of myelopoiesis. Mutation of this gene is responsible for neutrophil-specific granule deficiency in humans, a condition that confers susceptibility to Staphylococcus aureus infection. We found that C/EBPε-deficient mice are severely affected by infection with S. aureus, and C/EBPε deficiency in neutrophils contributes to the infectious phenotype. Conversely, exposure to the epigenetic modulator nicotinamide (vitamin B3) increased expression of C/EBPε in WT myeloid cells. Further, nicotinamide increased the activity of C/EBPε and select downstream antimicrobial targets, particularly in neutrophils. In a systemic murine infection model as well as in murine and human peripheral blood, nicotinamide enhanced killing of S. aureus by up to 1,000 fold but had no effect when administered to either C/EBPε-deficient mice or mice depleted of neutrophils. Nicotinamide was efficacious in both prophylactic and therapeutic settings. Our findings suggest that C/EBPε is an important target to boost killing of bacteria by the innate immune system. PMID:22922257

  9. Life extension factor klotho prevents mortality and enhances cognition in hAPP transgenic mice.

    PubMed

    Dubal, Dena B; Zhu, Lei; Sanchez, Pascal E; Worden, Kurtresha; Broestl, Lauren; Johnson, Erik; Ho, Kaitlyn; Yu, Gui-Qiu; Kim, Daniel; Betourne, Alexander; Kuro-O, Makoto; Masliah, Eliezer; Abraham, Carmela R; Mucke, Lennart

    2015-02-11

    Aging is the principal demographic risk factor for Alzheimer disease (AD), the most common neurodegenerative disorder. Klotho is a key modulator of the aging process and, when overexpressed, extends mammalian lifespan, increases synaptic plasticity, and enhances cognition. Whether klotho can counteract deficits related to neurodegenerative diseases, such as AD, is unknown. Here we show that elevating klotho expression decreases premature mortality and network dysfunction in human amyloid precursor protein (hAPP) transgenic mice, which simulate key aspects of AD. Increasing klotho levels prevented depletion of NMDA receptor (NMDAR) subunits in the hippocampus and enhanced spatial learning and memory in hAPP mice. Klotho elevation in hAPP mice increased the abundance of the GluN2B subunit of NMDAR in postsynaptic densities and NMDAR-dependent long-term potentiation, which is critical for learning and memory. Thus, increasing wild-type klotho levels or activities improves synaptic and cognitive functions, and may be of therapeutic benefit in AD and other cognitive disorders. PMID:25673831

  10. Life Extension Factor Klotho Prevents Mortality and Enhances Cognition in hAPP Transgenic Mice

    PubMed Central

    Zhu, Lei; Sanchez, Pascal E.; Worden, Kurtresha; Broestl, Lauren; Johnson, Erik; Ho, Kaitlyn; Yu, Gui-Qiu; Kim, Daniel; Betourne, Alexander; Kuro-o, Makoto; Masliah, Eliezer; Abraham, Carmela R.

    2015-01-01

    Aging is the principal demographic risk factor for Alzheimer disease (AD), the most common neurodegenerative disorder. Klotho is a key modulator of the aging process and, when overexpressed, extends mammalian lifespan, increases synaptic plasticity, and enhances cognition. Whether klotho can counteract deficits related to neurodegenerative diseases, such as AD, is unknown. Here we show that elevating klotho expression decreases premature mortality and network dysfunction in human amyloid precursor protein (hAPP) transgenic mice, which simulate key aspects of AD. Increasing klotho levels prevented depletion of NMDA receptor (NMDAR) subunits in the hippocampus and enhanced spatial learning and memory in hAPP mice. Klotho elevation in hAPP mice increased the abundance of the GluN2B subunit of NMDAR in postsynaptic densities and NMDAR-dependent long-term potentiation, which is critical for learning and memory. Thus, increasing wild-type klotho levels or activities improves synaptic and cognitive functions, and may be of therapeutic benefit in AD and other cognitive disorders. PMID:25673831

  11. Lack of LCAT reduces the LPS-neutralizing capacity of HDL and enhances LPS-induced inflammation in mice.

    PubMed

    Petropoulou, Peristera-Ioanna; Berbée, Jimmy F P; Theodoropoulos, Vassilios; Hatziri, Aikaterini; Stamou, Panagiota; Karavia, Eleni A; Spyridonidis, Alexandros; Karagiannides, Iordanes; Kypreos, Kyriakos E

    2015-10-01

    HDL has important immunomodulatory properties, including the attenuation of lipopolysaccharide (LPS)-induced inflammatory response. As lecithin-cholesterol acyltransferase (LCAT) is a critical enzyme in the maturation of HDL we investigated whether LCAT-deficient (Lcat(-/-)) mice present an increased LPS-induced inflammatory response. LPS (100μg/kg body weight)-induced cytokine response in Lcat(-/-) mice was markedly enhanced and prolonged compared to wild-type mice. Importantly, reintroducing LCAT expression using adenovirus-mediated gene transfer reverted their phenotype to that of wild-type mice. Ex vivo stimulation of whole blood with LPS (1-100ng/mL) showed a similar enhanced pro-inflammatory phenotype. Further characterization in RAW 264.7 macrophages in vitro showed that serum and HDL, but not chylomicrons, VLDL or the lipid-free protein fraction of Lcat(-/-) mice, had a reduced capacity to attenuate the LPS-induced TNFα response. Analysis of apolipoprotein composition revealed that LCAT-deficient HDL lacks significant amounts of ApoA-I and ApoA-II and is primarily composed of ApoE, while HDL from Apoa1(-/-) mice is highly enriched in ApoE and ApoA-II. ApoA-I-deficiency did not affect the capacity of HDL to neutralize LPS, though Apoa1(-/-) mice showed a pronounced LPS-induced cytokine response. Additional immunophenotyping showed that Lcat(-/-) , but not Apoa1(-/-) mice, have markedly increased circulating monocyte numbers as a result of increased Cd11b(+)Ly6C(med) monocytes, whereas 'pro-inflammatory' Cd11b(+)Ly6C(hi) monocytes were reduced. In line with this observation, peritoneal macrophages of Lcat(-/-) mice showed a markedly dampened LPS-induced TNFα response. We conclude that LCAT-deficiency increases LPS-induced inflammation in mice due to reduced LPS-neutralizing capacity of immature discoidal HDL and increased monocyte number. PMID:26170061

  12. Enhancing the expressiveness of structured reporting systems.

    PubMed

    Langlotz, C P

    2000-05-01

    The overall goal of this research is to build a structured reporting system that reduces the cost, delays, and inconvenience associated with conventional dictation and speech recognition systems. We have implemented such a structured reporting system for radiology that replaces current dictation and transcription processes by allowing radiologists and other imaging professionals to select imaging findings from a medical lexicon. The system uses an imaging-specific information model, called a "description set,' to organize selected terms in a relational database. Unique features of the knowledge representation that enhance its expressiveness include its ability to codify uncertainty about an imaging observation and to represent explicitly the logical relationships among imaging findings. In addition, the system does not require the user to fill in "blanks' in a static text template. Instead, it allows entry of terms in arbitrary order and uses automated text-generation techniques to create a text report that referring physicians are accustomed to receiving. In parallel, the system also produces a multimedia report that the referring physician can use as a quick reference. Unlike the results of conventional dictation or speech recognition, each finding is coded in a relational database for later information processing. Thus, the structured report database can be used to index images by content, to provide real-time decision support, to enhance radiologists' performance, to conduct exploratory clinical research, and to transmit imaging report data to computer-based patient record systems. PMID:10847362

  13. Enhanced p122RhoGAP/DLC-1 Expression Can Be a Cause of Coronary Spasm

    PubMed Central

    Kinjo, Takahiko; Tanaka, Makoto; Osanai, Tomohiro; Shibutani, Shuji; Narita, Ikuyo; Tanno, Tomohiro; Nishizaki, Kimitaka; Ichikawa, Hiroaki; Kimura, Yoshihiro; Ishida, Yuji; Yokota, Takashi; Shimada, Michiko; Homma, Yoshimi; Tomita, Hirofumi; Okumura, Ken

    2015-01-01

    Background We previously showed that phospholipase C (PLC)-δ1 activity was enhanced by 3-fold in patients with coronary spastic angina (CSA). We also reported that p122Rho GTPase-activating protein/deleted in liver cancer-1 (p122RhoGAP/DLC-1) protein, which was discovered as a PLC-δ1 stimulator, was upregulated in CSA patients. We tested the hypothesis that p122RhoGAP/DLC-1 overexpression causes coronary spasm. Methods and Results We generated transgenic (TG) mice with vascular smooth muscle (VSM)-specific overexpression of p122RhoGAP/DLC-1. The gene and protein expressions of p122RhoGAP/DLC-1 were markedly increased in the aorta of homozygous TG mice. Stronger staining with anti-p122RhoGAP/DLC-1 in the coronary artery was found in TG than in WT mice. PLC activities in the plasma membrane fraction and the whole cell were enhanced by 1.43 and 2.38 times, respectively, in cultured aortic vascular smooth muscle cells from homozygous TG compared with those from WT mice. Immediately after ergometrine injection, ST-segment elevation was observed in 1 of 7 WT (14%), 6 of 7 heterozygous TG (84%), and 7 of 7 homozygous TG mice (100%) (p<0.05, WT versus TGs). In the isolated Langendorff hearts, coronary perfusion pressure was increased after ergometrine in TG, but not in WT mice, despite of the similar response to prostaglandin F2α between TG and WT mice (n = 5). Focal narrowing of the coronary artery after ergometrine was documented only in TG mice. Conclusions VSM-specific overexpression of p122RhoGAP/DLC-1 enhanced coronary vasomotility after ergometrine injection in mice, which is relevant to human CSA. PMID:26624289

  14. Toxicity studies with 5-hydroxymethylfurfural and its metabolite 5-sulphooxymethylfurfural in wild-type mice and transgenic mice expressing human sulphotransferases 1A1 and 1A2.

    PubMed

    Bauer-Marinovic, Morana; Taugner, Felicitas; Florian, Simone; Glatt, Hansruedi

    2012-05-01

    5-Sulphooxymethylfurfural (SMF), an electrophilic metabolite of the abundant Maillard product 5-hydroxymethylfurfural (HMF), was intraperitoneally administered to FVB/N mice. At a dosage of 250 mg/kg, most animals died after 5-11 days due to massive damage to proximal tubules. At lower dosages, administered repeatedly, tubules also were the major target of toxicity, with regeneration and atypical hyperplasia occurring at later periods. Additionally, hepatotoxic effects and serositis of peritoneal tissues were observed. SMF is a minor metabolite of HMF in conventional mice, but HMF is an excellent substrate for a major sulphotransferase (hSULT1A1) in humans. Parental FVB/N mice and FVB/N-hSULT1A1/2 mice, carrying multiple copies of the hSULT1A1/2 gene cluster, were exposed to HMF in drinking water (0, 134 and 536 mg/kg body mass/day) for 12 weeks. Nephrotoxic effects and enhanced proliferation of hepatocytes were only detected at the high dosage. They were mild and, surprisingly, unaffected by hSULT1A1/2 expression. Thus, SMF was a potent nephrotoxicant when administered as a bolus, but did not reach levels sufficient to produce serious toxicity when generated from HMF administered continuously via drinking water. This was even the case in transgenic mice expressing clearly higher HMF sulphation activity in liver and kidney than humans. PMID:22349055

  15. Expression and regulation of Dickkopf2 during periimplantation in mice.

    PubMed

    Zhang, Ying; Peng, Sha; Kuang, Haibin; Chen, Qi; Liu, Shuang; Zhang, Li; Duan, Enkui

    2009-02-01

    Successful implantation depends on active dialogue between the maternal endometrium and the implanting blastocysts that is well controlled by groups of regulators at the molecular level. Dickkopf2 (Dkk2) is a member of Dickkopf family normally acting as an antagonist of canonical Wnt/beta-catenin signaling, which has been proven to participate in tumorigenesis and early embryo development. In order to explore the potential function of Dkk2 in embryo implantation, the present study investigated the uterine expression and regulation profiles of Dkk2 during periimplantation in mice. Using reverse transcription-polymerase chain reaction, immunohistochemistry and Western blotting, we showed that the mRNA and protein levels of Dkk2 began to increase in the glandular epithelium on day 4, continued to increase on day 5 and then decreased from day 6 of pregnancy. Moreover, on days 5-8 of pregnancy, Dkk2 was increasingly expressed in the deciduum of the uterus, especially around the implanting embryos. In addition, upregulation of Dkk2 was also observed in uteri treated with estrogen (estradiol-17beta) as well as in oil-induced artificial decidualization, indicating that the expression of Dkk2 could be induced by both steroid hormone (estrogen) and the process of decidualization. Furthermore, in the postimplantation uterus, the Dkk2 protein showed an inversed expression with active beta-catenin from day 6 onward, supporting the notion that Dkk2 plays an inhibitory role against canonical Wnt signaling in the context of the decidualizing stroma. Collectively, our data suggests that Dkk2 expression is associated with uterine receptivity changes as well as the process of decidualization and that it might play important roles through inhibition of canonical Wnt signaling in the periimplantation uterus. PMID:18689976

  16. Genotype differences in anxiety and fear learning and memory of WT and ApoE4 mice associated with enhanced generation of hippocampal reactive oxygen species.

    PubMed

    Villasana, Laura E; Weber, Sydney; Akinyeke, Tunde; Raber, Jacob

    2016-09-01

    Apolipoprotein E (apoE), involved in cholesterol and lipid metabolism, also influences cognitive function and injury repair. In humans, apoE is expressed in three isoforms. E4 is a risk factor for age-related cognitive decline and Alzheimer's disease, particularly in women. E4 might also be a risk factor for developing behavioral and cognitive changes following (56) Fe irradiation, a component of the space environment astronauts are exposed to during missions. These changes might be related to enhanced generation of reactive oxygen species (ROS). In this study, we compared the behavioral and cognitive performance of sham-irradiated and irradiated wild-type (WT) mice and mice expressing the human E3 or E4 isoforms, and assessed the generation of ROS in hippocampal slices from these mice. E4 mice had greater anxiety-like and conditioned fear behaviors than WT mice, and these genotype differences were associated with greater levels of ROS in E4 than WT mice. The greater generation of ROS in the hippocampus of E4 than WT mice might contribute to their higher anxiety levels and enhanced fear conditioning. In E4, but not WT, mice, phorbol-12-myristate-13-acetate-treated hippocampal slices showed more dihydroxy ethidium oxidation in sham-irradiated than irradiated mice and hippocampal heme oxygenase-1 levels were higher in irradiated than sham-irradiated E4 mice. Mice with apolipoprotein E4 (E4), a risk factor for Alzheimer's disease, have greater anxiety-like and conditioned fear behaviors than wild-type (WT) mice. Generation of reactive oxygen species (ROS, in red) 3 months following (56) Fe irradiation, a component of the space environment astronauts are exposed to, is more pronounced in the hippocampus of E4 than WT mice. In E4, but not WT, mice, hippocampal levels of the oxidative stress-relevant marker heme oxygenase-1 are higher in irradiated than sham-irradiated E4 mice. PMID:27412623

  17. Disruption of Nrf2 enhances susceptibility to severe airway inflammation and asthma in mice.

    PubMed

    Rangasamy, Tirumalai; Guo, Jia; Mitzner, Wayne A; Roman, Jessica; Singh, Anju; Fryer, Allison D; Yamamoto, Masayuki; Kensler, Thomas W; Tuder, Rubin M; Georas, Steve N; Biswal, Shyam

    2005-07-01

    Oxidative stress has been postulated to play an important role in the pathogenesis of asthma; although a defect in antioxidant responses has been speculated to exacerbate asthma severity, this has been difficult to demonstrate with certainty. Nuclear erythroid 2 p45-related factor 2 (Nrf2) is a redox-sensitive basic leucine zipper transcription factor that is involved in the transcriptional regulation of many antioxidant genes. We show that disruption of the Nrf2 gene leads to severe allergen-driven airway inflammation and hyperresponsiveness in mice. Enhanced asthmatic response as a result of ovalbumin sensitization and challenge in Nrf2-disrupted mice was associated with more pronounced mucus cell hyperplasia and infiltration of eosinophils into the lungs than seen in wild-type littermates. Nrf2 disruption resulted in an increased expression of the T helper type 2 cytokines interleukin (IL)-4 and IL-13 in bronchoalveolar lavage fluid and in splenocytes after allergen challenge. The enhanced severity of the asthmatic response from disruption of the Nrf2 pathway was a result of a lowered antioxidant status of the lungs caused by lower basal expression, as well as marked attenuation, of the transcriptional induction of multiple antioxidant genes. Our studies suggest that the responsiveness of Nrf2-directed antioxidant pathways may act as a major determinant of susceptibility to allergen-mediated asthma. PMID:15998787

  18. Conditional deletion of cardiomyocyte peroxisome proliferator-activated receptor γ enhances myocardial ischemia-reperfusion injury in mice.

    PubMed

    Hobson, Michael J; Hake, Paul W; O'Connor, Michael; Schulte, Christine; Moore, Victoria; James, Jeanne M; Piraino, Giovanna; Zingarelli, Basilia

    2014-01-01

    The nuclear transcription factor peroxisome proliferator-activated receptor γ (PPARγ) is a key regulator of the inflammatory response to an array of biologic insults. We have previously demonstrated that PPARγ ligands reduce myocardial ischemia-reperfusion injury in rodents. In the current study, we directly determined the role of cardiomyocyte PPARγ in ischemia-reperfusion injury, using a model of conditional cardiomyocyte-specific deletion of PPARγ in vivo. In mice, α-myosin heavy chain-restricted Cre-mediated PPARγ deficiency was induced by tamoxifen treatment (30 mg/kg intraperitoneally) for 4 days (PPARγ mice), whereas controls included mice treated with the oil diluent vehicle (PPARγ mice). Western blot and histochemical analyses confirmed that expression of PPARγ protein was abolished in cardiomyocytes of mice treated with tamoxifen, but not with vehicle. After tamoxifen or vehicle treatment, animals were subjected to 30-min ligation of the left anterior descending coronary artery followed by 2-h reperfusion. In PPARγ mice, myocardial ischemia and reperfusion induced extensive myocardial damage, which was associated with elevated tissue activity of myeloperoxidase, indicating infiltration of neutrophils, and elevated plasma levels of troponin I when compared with PPARγ mice. Upon echocardiographic analysis, PPARγ mice also demonstrated ventricular dilatation and systolic dysfunction. Plasma levels of the proinflammatory cytokines interleukin 1β and interleukin 6 were higher in PPARγ mice when compared with PPARγ mice. These pathological events in PPARγ mice were associated with enhanced nuclear factor κB DNA binding in the infarcted hearts. Thus, our data suggest that cardiomyocyte PPARγ is a crucial protective receptor and may prevent reperfusion injury by modulating mechanisms of inflammation. PMID:24089001

  19. Conditional deletion of cardiomyocyte peroxisome proliferator-activated receptor-γ enhances myocardial ischemia-reperfusion injury in mice

    PubMed Central

    Hobson, Michael J.; Hake, Paul W.; O’Connor, Michael; Schulte, Christine; Moore, Victoria; James, Jeanne M.; Piraino, Giovanna; Zingarelli, Basilia

    2013-01-01

    The nuclear transcription factor peroxisome proliferator-activated receptor-γ (PPARγ) is a key regulator of the inflammatory response to an array of biologic insults. We have previously demonstrated that PPARγ ligands reduce myocardial ischemia-reperfusion injury in rodents. In the current study, we directly determined the role of cardiomyocyte PPARγ in ischemia-reperfusion injury, employing a model of conditional cardiomyocyte-specific deletion of PPARγ in vivo. In mice, α-myosin heavy chain-restricted Cre-mediated PPARγ deficiency was induced by tamoxifen treatment (30 mg/kg intraperitoneally) for 4 days (PPARγ−/− mice); whereas controls included mice treated with the oil diluent vehicle (PPARγ+/+ mice). Western blot and histochemical analyses confirmed that expression of PPARγ protein was abolished in cardiomyocytes of mice treated with tamoxifen, but not with vehicle. After tamoxifen or vehicle treatment, animals were subjected to 30 min ligation of the left anterior descending coronary artery followed by 2 hrs reperfusion. In PPARγ−/− mice, myocardial ischemia and reperfusion induced extensive myocardial damage, which was associated with elevated tissue activity of myeloperoxidase, indicating infiltration of neutrophils, and elevated plasma levels of troponin-I when compared to PPARγ+/+ mice. PPARγ−/− mice also demonstrated ventricular dilatation and systolic dysfunction upon echocardiographic analysis. Plasma levels of the pro-inflammatory cytokines interleukin-1β and interleukin-6 were higher in PPARγ−/− mice when compared to PPARγ+/+ mice. These pathological events in PPARγ−/− mice were associated with enhanced nuclear factor-κB DNA binding in the infarcted hearts. Thus, our data suggests that cardiomyocyte PPARγ is a crucial protective receptor and may prevent reperfusion injury by modulating mechanisms of inflammation. PMID:24089001

  20. Engineered silica nanoparticles act as adjuvants to enhance allergic airway disease in mice

    PubMed Central

    2013-01-01

    Background With the increase in production and use of engineered nanoparticles (NP; ≤ 100 nm), safety concerns have risen about the potential health effects of occupational or environmental NP exposure. Results of animal toxicology studies suggest that inhalation of NP may cause pulmonary injury with subsequent acute or chronic inflammation. People with chronic respiratory diseases like asthma or allergic rhinitis may be even more susceptible to toxic effects of inhaled NP. Few studies, however, have investigated adverse effects of inhaled NP that may enhance the development of allergic airway disease. Methods We investigated the potential of polyethylene glycol coated amorphous silica NP (SNP; 90 nm diameter) to promote allergic airway disease when co-exposed during sensitization with an allergen. BALB/c mice were sensitized by intranasal instillation with 0.02% ovalbumin (OVA; allergen) or saline (control), and co-exposed to 0, 10, 100, or 400 μg of SNP. OVA-sensitized mice were then challenged intranasally with 0.5% OVA 14 and 15 days after sensitization, and all animals were sacrificed a day after the last OVA challenge. Blood and bronchoalveolar lavage fluid (BALF) were collected, and pulmonary tissue was processed for histopathology and biochemical and molecular analyses. Results Co-exposure to SNP during OVA sensitization caused a dose-dependent enhancement of allergic airway disease upon challenge with OVA alone. This adjuvant-like effect was manifested by significantly greater OVA-specific serum IgE, airway eosinophil infiltration, mucous cell metaplasia, and Th2 and Th17 cytokine gene and protein expression, as compared to mice that were sensitized to OVA without SNP. In saline controls, SNP exposure did cause a moderate increase in airway neutrophils at the highest doses. Conclusions These results suggest that airway exposure to engineered SNP could enhance allergen sensitization and foster greater manifestation of allergic airway disease upon

  1. Radiation Exposure Alters Expression of Metabolic Enzyme Genes In Mice

    NASA Technical Reports Server (NTRS)

    Wotring, Virginia E.; Mangala, L. S.; Zhang, Y.; Wu, H.

    2010-01-01

    Most pharmaceuticals are metabolized by the liver. The health of the liver, especially the rate of its metabolic enzymes, determines the concentration of circulating drugs as well as the duration of their efficacy. Because of the importance of the liver in drug metabolism it is important to understand the effects of spaceflight on the enzymes of the liver. Exposure to cosmic radiation is one aspect of spaceflight that can be modeled in ground experiments. This study is an effort to examine the effects of adaptive mechanisms that may be triggered by early exposure to low radiation doses. Using procedures approved by the JSC Animal Care & Use Committee, C57 male mice were exposed to Cs-137 in groups: controls, low dose (50 mGy), high dose (6Gy) and a fourth group that received both radiation doses separated by 24 hours. Animals were anesthetized and sacrificed 4 hours after their last radiation exposure. Livers were removed immediately and flash-frozen in liquid nitrogen. Tissue was homogenized, RNA extracted and purified (Absolutely RNA, Agilent). Quality of RNA samples was evaluated (Agilent Bioanalyzer 2100). Complementary DNA was prepared from high-quality RNA samples, and used to run RT-qPCR screening arrays for DNA Repair and Drug Metabolism (SuperArray, SABiosciences/Qiagen; BioRad Cfx96 qPCR System). Of 91 drug metabolism genes examined, expression of 7 was altered by at least one treatment condition. Genes that had elevated expression include those that metabolize promethazine and steroids (4-8-fold), many that reduce oxidation products, and one that reduces heavy metal exposure (greater than 200-fold). Of the 91 DNA repair and general metabolism genes examined, expression of 14 was altered by at least one treatment condition. These gene expression changes are likely homeostatic and could lead to development of new radioprotective countermeasures.

  2. Gene expression profiling of gastrocnemius of “minimuscle” mice

    PubMed Central

    Meek, Thomas H.; Pandey, Sachchida Nand; Broitman-Maduro, Gina; Maduro, Morris F.; Bronikowski, Anne M.; Garland, Theodore; Chen, Yi-Wen

    2013-01-01

    Few studies have investigated heterogeneity of selection response in replicate lines subjected to equivalent selection. We developed four replicate lines of mice based on high levels of voluntary wheel running (high runner or HR lines) while also maintaining four nonselected control lines. This led to the unexpected discovery of the HR minimuscle (HRmini) phenotype, recognized by a 50% reduction in hindlimb muscle mass, which became fixed in 1 of the four HR selected lines. Here, we report genome-wide expression profiling describing transcriptome differences between HRnormal and HRmini medial gastrocnemius. Consistent with the known reduction of type IIB fibers in HRmini, Myh4 gene expression was −8.82-fold less (P = 0.0001) in HRmini, which was closely associated with differences in the “calcium signaling” canonical pathway, including structural genes (e.g., Mef2c, twofold greater in HRmini, P = 0.0003) and myogenic factors (e.g., Myog, 3.8-fold greater in HRmini, P = 0.0026) associated with slow-type myofibers. The gene that determines the HRmini phenotype is known to reside in a 2.6335-Mb interval on mouse chromosome 11 and 7 genes (Myh10, Chrnb1, Acadvl, Senp3, Gabarap, Eif5a, and Clec10a) from this region were differentially expressed. Verification by real-time PCR confirmed 1.5-fold greater (P < 0.05) expression of very long chain acyl-CoA dehydrogenase (Acadvl) in HRmini. Ten other genes associated with fatty acid metabolism were also upregulated in HRmini, suggesting differences in the ability to metabolize fatty acids in HRnormal and HRmini muscles. This work provides a resource for understanding differences in muscle phenotypes in populations exhibiting high running capacity. PMID:23362141

  3. Dietary Iron Enhances Colonic Inflammation and IL-6/IL-11-Stat3 Signaling Promoting Colonic Tumor Development in Mice

    PubMed Central

    Ho, Desiree S.; Fu, S. Kristine; Forrest, Cynthia H.; Croft, Kevin D.; Olynyk, John K.; Lawrance, Ian C.; Trinder, Debbie

    2013-01-01

    Chronic intestinal inflammation and high dietary iron are associated with colorectal cancer development. The role of Stat3 activation in iron-induced colonic inflammation and tumorigenesis was investigated in a mouse model of inflammation-associated colorectal cancer. Mice, fed either an iron-supplemented or control diet, were treated with azoxymethane and dextran sodium sulfate (DSS). Intestinal inflammation and tumor development were assessed by endoscopy and histology, gene expression by real-time PCR, Stat3 phosphorylation by immunoblot, cytokines by ELISA and apoptosis by TUNEL assay. Colonic inflammation was more severe in mice fed an iron-supplemented compared with a control diet one week post-DSS treatment, with enhanced colonic IL-6 and IL-11 release and Stat3 phosphorylation. Both IL-6 and ferritin, the iron storage protein, co-localized with macrophages suggesting iron may act directly on IL-6 producing-macrophages. Iron increased DSS-induced colonic epithelial cell proliferation and apoptosis consistent with enhanced mucosal damage. DSS-treated mice developed anemia that was not alleviated by dietary iron supplementation. Six weeks post-DSS treatment, iron-supplemented mice developed more and larger colonic tumors compared with control mice. Intratumoral IL-6 and IL-11 expression increased in DSS-treated mice and IL-6, and possibly IL-11, were enhanced by dietary iron. Gene expression of iron importers, divalent metal transporter 1 and transferrin receptor 1, increased and iron exporter, ferroportin, decreased in colonic tumors suggesting increased iron uptake. Dietary iron and colonic inflammation synergistically activated colonic IL-6/IL-11-Stat3 signaling promoting tumorigenesis. Oral iron therapy may be detrimental in inflammatory bowel disease since it may exacerbate colonic inflammation and increase colorectal cancer risk. PMID:24223168

  4. Increased TNFR1 expression and signaling in injured peripheral nerves of mice with reduced BACE1 activity.

    PubMed

    Liu, Lijuan; Fissel, John A; Tasnim, Aniqa; Borzan, Jasenka; Gocke, Anne; Calabresi, Peter A; Farah, Mohamed H

    2016-09-01

    Hematogenous macrophages remove myelin debris from injured peripheral nerves to provide a micro-environment conducive to axonal regeneration. Previously, we observed that injured peripheral nerves from Beta-site APP Cleaving Enzyme 1 (BACE1) knockout (KO) mice displayed earlier influx of and enhanced phagocytosis by macrophages when compared to wild-type (WT) mice. These observations suggest that BACE1 might regulate macrophage influx into distal stumps of injured nerves. To determine through which pathway BACE1 influences macrophage influx, we used a mouse inflammation antibody array to assay the expression of inflammation-related proteins in injured nerves of BACE1 KO and WT mice. The most significant change was in expression of tumor necrosis factor receptor 1 (TNFR1) in the distal stump of injured BACE1 KO nerves. Western blotting of protein extracts confirmed increased expression of TNFR1 and its downstream transcriptional factor NFκB in the BACE1 KO distal stumps. Additionally, treatment of WT mice with a BACE1 inhibitor resulted in increased TNFR1 expression and signaling in the distal stump of injured nerves. Exogenous TNFα increased nuclear translocation of p65 NFκB in BACE1 KO tissue and cultured fibroblasts compared with control WT. BACE1 regulates TNFR1 expression at the level of gene expression and not through proteolytic processing. The accelerated macrophage influx in injured nerves of BACE1 KO mice correlates with increased expression and signaling via TNFR1, indicating a link between BACE1 activity and TNFR1 expression/signaling that might contribute to repair of the injured nervous system. PMID:27080468

  5. Maitake beta-glucan enhances therapeutic effect and reduces myelosupression and nephrotoxicity of cisplatin in mice.

    PubMed

    Masuda, Yuki; Inoue, Munechika; Miyata, Ayu; Mizuno, Shigeto; Nanba, Hiroaki

    2009-05-01

    Cisplatin is broadly used clinically as an anticancer drug. Despite its significant anticancer activity, cisplatin-induced nephrotoxicity and myelosuppression limit its use. MD-Fraction is glucan purified from maitake (Grifola frondosa), which has beta-1, 6-main chain with beta-1, 3-branches, has been reported to exhibit antitumor and antimetastatic activities by enhancing the immune system. In this study, we demonstrate that MD-Fraction in combination with cisplatin significantly enhanced antitumor and antimetastatic activity compared to cisplatin alone. MD-Fraction reduced decreases in body weight, spleen weight and the number of immunocompetent cells such as macrophages, DCs and NK cells in cisplatin-treated mice. MD-Fraction also induced IL-12p70 production by splenocytes, resulting in increased NK cell activity in cisplatin-treated mice. MD-Fraction significantly increased the mRNA expression of GM-CSF, G-CSF, M-CSF, IFN-gamma, IL-12 p40 in splenocytes and reduced the decrease in the number of CFU-GM colonies in cisplatin-treated bone marrow. These facts suggest that MD-Fraction can reduce cisplatin-induced myelosuppression. Moreover, treatment with MD-Fraction significantly reduced cisplatin-induced nephrotoxicity accompanied by increases in serum creatinine level, necrosis and apoptosis of renal tubular cells. These results suggest that MD-Fraction in combination with cisplatin cannot only enhance antitumor and antimentastatic acitivity, but also reduce cisplatin-induced myelotoxicity and nephrotoxicity. PMID:19249389

  6. Berberine enhances antidiabetic effects and attenuates untoward effects of canagliflozin in streptozotocin-induced diabetic mice.

    PubMed

    Tian, Cai-Ming; Jiang, Xin; Ouyang, Xiao-Xi; Zhang, Ya-Ou; Xie, Wei-Dong

    2016-07-01

    The present study aimed at determining whether berberine can enhance the antidiabetic effects and alleviate the adverse effects of canagliflozin in diabetes mellitus. Streptozotocin-induced diabetic mice were introduced, and the combined effects of berberine and canagliflozin on glucose metabolism and kidney functions were investigated. Our results showed that berberine combined with canagliflozin (BC) increased reduction of fasting and postprandial blood glucose, diet, and water intake compared with berberine or canagliflozin alone. Interestingly, BC showed greater decrease in blood urea nitrogen and creatinine levels and lower total urine glucose excretion than canagliflozin alone. In addition, BC showed increased phosphorylated 5' AMP-activated protein kinase (pAMPK) expression and decreased tumor necrosis factor alpha (TNFα) levels in kidneys, compared with berberine or canagliflozin alone. These results indicated that BC was a stronger antidiabetic than berberine or canagliflozin alone with less negative side effects on the kidneys in the diabetic mice. The antidiabetic effect was likely to be mediated by synergically promoting the expression of pAMPK and reducing the expression of TNFα in kidneys. The present study represented the first report that canagliflozin combined with berberine was a promising treatment for diabetes mellitus. The exact underlying mechanisms of action should be investigated in future studies. PMID:27507202

  7. Traumatic brain injury enhances neuroinflammation and lesion volume in caveolin deficient mice

    PubMed Central

    2014-01-01

    Background Traumatic brain injury (TBI) enhances pro-inflammatory responses, neuronal loss and long-term behavioral deficits. Caveolins (Cavs) are regulators of neuronal and glial survival signaling. Previously we showed that astrocyte and microglial activation is increased in Cav-1 knock-out (KO) mice and that Cav-1 and Cav-3 modulate microglial morphology. We hypothesized that Cavs may regulate cytokine production after TBI. Methods Controlled cortical impact (CCI) model of TBI (3 m/second; 1.0 mm depth; parietal cortex) was performed on wild-type (WT; C57Bl/6), Cav-1 KO, and Cav-3 KO mice. Histology and immunofluorescence microscopy (lesion volume, glia activation), behavioral tests (open field, balance beam, wire grip, T-maze), electrophysiology, electron paramagnetic resonance, membrane fractionation, and multiplex assays were performed. Data were analyzed by unpaired t tests or analysis of variance (ANOVA) with post-hoc Bonferroni’s multiple comparison. Results CCI increased cortical and hippocampal injury and decreased expression of MLR-localized synaptic proteins (24 hours), enhanced NADPH oxidase (Nox) activity (24 hours and 1 week), enhanced polysynaptic responses (1 week), and caused hippocampal-dependent learning deficits (3 months). CCI increased brain lesion volume in both Cav-3 and Cav-1 KO mice after 24 hours (P < 0.0001, n = 4; one-way ANOVA). Multiplex array revealed a significant increase in expression of IL-1β, IL-9, IL-10, KC (keratinocyte chemoattractant), and monocyte chemoattractant protein 1 (MCP-1) in ipsilateral hemisphere and IL-9, IL-10, IL-17, and macrophage inflammatory protein 1 alpha (MIP-1α) in contralateral hemisphere of WT mice after 4 hours. CCI increased IL-2, IL-6, KC and MCP-1 in ipsilateral and IL-6, IL-9, IL-17 and KC in contralateral hemispheres in Cav-1 KO and increased all 10 cytokines/chemokines in both hemispheres except for IL-17 (ipsilateral) and MIP-1α (contralateral) in Cav-3 KO (versus WT CCI). Cav-3 KO CCI

  8. Cell-type specific expression of a dominant negative PKA mutation in mice.

    PubMed

    Willis, Brandon S; Niswender, Colleen M; Su, Thomas; Amieux, Paul S; McKnight, G Stanley

    2011-01-01

    We employed the Cre recombinase/loxP system to create a mouse line in which PKA activity can be inhibited in any cell-type that expresses Cre recombinase. The mouse line carries a mutant Prkar1a allele encoding a glycine to aspartate substitution at position 324 in the carboxy-terminal cAMP-binding domain (site B). This mutation produces a dominant negative RIα regulatory subunit (RIαB) and leads to inhibition of PKA activity. Insertion of a loxP-flanked neomycin cassette in the intron preceding the site B mutation prevents expression of the mutant RIαB allele until Cre-mediated excision of the cassette occurs. Embryonic stem cells expressing RIαB demonstrated a reduction in PKA activity and inhibition of cAMP-responsive gene expression. Mice expressing RIαB in hepatocytes exhibited reduced PKA activity, normal fasting induced gene expression, and enhanced glucose disposal. Activation of the RIαB allele in vivo provides a novel system for the analysis of PKA function in physiology. PMID:21533282

  9. Expression of A152T human tau causes age-dependent neuronal dysfunction and loss in transgenic mice.

    PubMed

    Maeda, Sumihiro; Djukic, Biljana; Taneja, Praveen; Yu, Gui-Qiu; Lo, Iris; Davis, Allyson; Craft, Ryan; Guo, Weikun; Wang, Xin; Kim, Daniel; Ponnusamy, Ravikumar; Gill, T Michael; Masliah, Eliezer; Mucke, Lennart

    2016-04-01

    A152T-variant human tau (hTau-A152T) increases risk for tauopathies, including Alzheimer's disease. Comparing mice with regulatable expression of hTau-A152T or wild-type hTau (hTau-WT), we find age-dependent neuronal loss, cognitive impairments, and spontaneous nonconvulsive epileptiform activity primarily in hTau-A152T mice. However, overexpression of either hTau species enhances neuronal responses to electrical stimulation of synaptic inputs and to an epileptogenic chemical. hTau-A152T mice have higher hTau protein/mRNA ratios in brain, suggesting that A152T increases production or decreases clearance of hTau protein. Despite their functional abnormalities, aging hTau-A152T mice show no evidence for accumulation of insoluble tau aggregates, suggesting that their dysfunctions are caused by soluble tau. In human amyloid precursor protein (hAPP) transgenic mice, co-expression of hTau-A152T enhances risk of early death and epileptic activity, suggesting copathogenic interactions between hTau-A152T and amyloid-β peptides or other hAPP metabolites. Thus, the A152T substitution may augment risk for neurodegenerative diseases by increasing hTau protein levels, promoting network hyperexcitability, and synergizing with the adverse effects of other pathogenic factors. PMID:26931567

  10. Erythropoietin attenuates the sequels of ischaemic spinal cord injury with enhanced recruitment of CD34+ cells in mice

    PubMed Central

    Hirano, Koji; Wagner, Klaus; Mark, Peter; Pittermann, Erik; Gäbel, Ralf; Furlani, Dario; Li, Wenzhong; Vollmar, Brigitte; Yamada, Tomomi; Steinhoff, Gustav; Ma, Nan

    2012-01-01

    Abstract Erythropoietin has been shown to promote tissue regeneration after ischaemic injury in various organs. Here, we investigated whether Erythropoietin could ameliorate ischaemic spinal cord injury in the mouse and sought an underlying mechanism. Spinal cord ischaemia was developed by cross-clamping the descending thoracic aorta for 7 or 9 min. in mice. Erythropoietin (5000 IU/kg) or saline was administrated 30 min. before aortic cross-clamping. Neurological function was assessed using the paralysis score for 7 days after the operation. Spinal cords were histologically evaluated 2 and 7 days after the operation. Immunohistochemistry was used to detect CD34+ cells and the expression of brain-derived neurotrophic factor and vascular endothelial growth factor. Each mouse exhibited either mildly impaired function or complete paralysis at day 2. Erythropoietin-treated mice with complete paralysis demonstrated significant improvement of neurological function between day 2 and 7, compared to saline-treated mice with complete paralysis. Motor neurons in erythropoietin-treated mice were more preserved at day 7 than those in saline-treated mice with complete paralysis. CD34+ cells in the lumbar spinal cord of erythropoietin-treated mice were more abundant at day 2 than those of saline-treated mice. Brain-derived neurotrophic factor and vascular endothelial growth factor were markedly expressed in lumbar spinal cords in erythropoietin-treated mice at day 7. Erythropoietin demonstrated neuroprotective effects in the ischaemic spinal cord, improving neurological function and attenuating motor neuron loss. These effects may have been mediated by recruited CD34+ cells, and enhanced expression of brain-derived neurotrophic factor and vascular endothelial growth factor. PMID:22145921

  11. Improved muscle healing through enhanced regeneration and reduced fibrosis in myostatin-null mice.

    PubMed

    McCroskery, Seumas; Thomas, Mark; Platt, Leanne; Hennebry, Alex; Nishimura, Takanori; McLeay, Lance; Sharma, Mridula; Kambadur, Ravi

    2005-08-01

    Numerous stimulatory growth factors that can influence muscle regeneration are known. Recently, it has been demonstrated that neutralization of muscle growth inhibitory factors, such as myostatin (Mstn; also known as growth differentiation factor 8, Gdf8), also leads to increased muscle regeneration in mdx mice that are known to have cycles of degeneration. However, the precise mechanism by which Mstn regulates muscle regeneration has not yet been fully determined. To investigate the role of Mstn in adult skeletal muscle regeneration, wild-type and myostatin-null (Mstn-/-) mice were injured with notexin. Forty-eight hours after injury, accelerated migration and enhanced accretion of myogenic cells (MyoD1+) and macrophages (Mac-1+) was observed at the site of regeneration in Mstn-/- muscle as compared with wild-type muscle. Inflammatory cell numbers decreased more rapidly in the Mstn-/- muscle, indicating that the whole process of inflammatory cell response is accelerated in Mstn-/- mice. Consistent with this result, the addition of recombinant Mstn reduced the activation of satellite cells (SCs) and chemotactic movements of both myoblasts and macrophages ex vivo. Examination of regenerated muscle (28 days after injury) also revealed that Mstn-/- mice showed increased expression of decorin mRNA, reduced fibrosis and improved healing as compared with wild-type mice. On the basis of these results, we propose that Mstn negatively regulates muscle regeneration not only by controlling SC activation but also by regulating the migration of myoblasts and macrophages to the site of injury. Thus, antagonists of Mstn could potentially be useful as pharmacological agents for the treatment of disorders of overt degeneration and regeneration. PMID:16079293

  12. Enhanced expression of rabies virus surface G-protein in Escherichia coli using SUMO fusion.

    PubMed

    Singh, Ankit; Yadav, Dinesh; Rai, Krishan Mohan; Srivastava, Meenal; Verma, Praveen C; Singh, Pradhyumna K; Tuli, Rakesh

    2012-01-01

    Fusion systems are known to increase the expression of difficult to express recombinant proteins in soluble form to facilitate their purification. Rabies glycoprotein was also tough to express at sufficient level in soluble form in both E. coli and plant. The present work was aimed to over-express and purify this membrane protein from soluble extract of E. coli. Fusion of Small Ubiqutin like Modifier (SUMO) with rabies glycoprotein increased ~1.5 fold higher expression and ~3.0 fold solubility in comparison to non-fused in E. coli. The SUMO fusion also simplified the purification process. Previously engineered rabies glycoprotein gene in tobacco plants provides complete protection to mice, but the expression was very low for purification. Our finding demonstrated that the SUMO-fusion was useful for enhancing expression and solubility of the membrane protein and again proves to be a good alternative technology for applications in biomedical and pharmaceutical research. PMID:22134654

  13. Variants in an Hdac9 intronic enhancer plasmid impact Twist1 expression in vitro.

    PubMed

    Siekmann, Tyler E; Gerber, Madelyn M; Toland, Amanda Ewart

    2016-04-01

    Skin tumor susceptibility 5 (Skts5) was previously mapped to mouse chromosome 12 through linkage analysis of skin tumor susceptible Mus musculus (NIH/Ola-S) and skin tumor resistant outbred Mus spretus (SPRET/Out-R) mice. Hdac9 was identified as a potential candidate for Skts5 based on conserved non-synonymous sequence variants and expression analyses. Studies by others identified an enhancer in human HDAC9 that correlated with TWIST1 expression. We identified 45 sequence variants between NIH/Ola-S and SPRET/Out-R mice from the orthologous region of the human HDAC9 enhancer. Variants mapping to intron 18 differentially affected luciferase expression in vitro. NIH/Ola-S clones showed an approximate 1.7-fold increased luciferase expression relative to vector alone or the equivalent clones from SPRET/Out-R-R. Furthermore, cells transfected with a portion of the NIH/Ola-S intron induced 2.2-fold increases in Twist1 expression, but the same region from SPRET/Out-R mice resulted in no up-regulation of Twist1. In silico transcription factor analyses identified multiple transcription factors predicted to differentially bind NIH/Ola-S and SPRET/Out-R polymorphic sites. Chromatin immunoprecipitation studies of two transcription factors, Gata3 and Oct1, demonstrated differential binding between NIH/Ola-S and SPRET/Out-R plasmids that corroborated the in silico predictions. Together these studies provide evidence that the murine orthologous region to a human HDAC9 enhancer also acts as a transcriptional enhancer for mouse Twist1. As ectopic sequence variants between NIH/Ola-S and SPRET/Out-R differentially impacted luciferase expression, correlated with Twist1 expression in vitro, and affected Gata3 and Oct1 binding, these variants may explain part of the observed differences in skin tumor susceptibility at Skts5 between NIH/Ola-S and SPRET/Out-R. PMID:26721262

  14. MicroRNA loss enhances learning and memory in mice.

    PubMed

    Konopka, Witold; Kiryk, Anna; Novak, Martin; Herwerth, Marina; Parkitna, Jan Rodriguez; Wawrzyniak, Marcin; Kowarsch, Andreas; Michaluk, Piotr; Dzwonek, Joanna; Arnsperger, Tabea; Wilczynski, Grzegorz; Merkenschlager, Matthias; Theis, Fabian J; Köhr, Georg; Kaczmarek, Leszek; Schütz, Günther

    2010-11-01

    Dicer-dependent noncoding RNAs, including microRNAs (miRNAs), play an important role in a modulation of translation of mRNA transcripts necessary for differentiation in many cell types. In vivo experiments using cell type-specific Dicer1 gene inactivation in neurons showed its essential role for neuronal development and survival. However, little is known about the consequences of a loss of miRNAs in adult, fully differentiated neurons. To address this question, we used an inducible variant of the Cre recombinase (tamoxifen-inducible CreERT2) under control of Camk2a gene regulatory elements. After induction of Dicer1 gene deletion in adult mouse forebrain, we observed a progressive loss of a whole set of brain-specific miRNAs. Animals were tested in a battery of both aversively and appetitively motivated cognitive tasks, such as Morris water maze, IntelliCage system, or trace fear conditioning. Compatible with rather long half-life of miRNAs in hippocampal neurons, we observed an enhancement of memory strength of mutant mice 12 weeks after the Dicer1 gene mutation, before the onset of neurodegenerative process. In acute brain slices, immediately after high-frequency stimulation of the Schaffer collaterals, the efficacy at CA3-to-CA1 synapses was higher in mutant than in control mice, whereas long-term potentiation was comparable between genotypes. This phenotype was reflected at the subcellular and molecular level by the elongated filopodia-like shaped dendritic spines and an increased translation of synaptic plasticity-related proteins, such as BDNF and MMP-9 in mutant animals. The presented work shows miRNAs as key players in the learning and memory process of mammals. PMID:21048142

  15. Immunisation with proteins expressed during chronic murine melioidosis provides enhanced protection against disease.

    PubMed

    Champion, Olivia L; Gourlay, Louise J; Scott, Andrew E; Lassaux, Patricia; Conejero, Laura; Perletti, Lucia; Hemsley, Claudia; Prior, Joann; Bancroft, Gregory; Bolognesi, Martino; Titball, Richard W

    2016-03-29

    There is an urgent need for an effective vaccine against human disease caused by Burkholderia pseudomallei, and although a wide range of candidates have been tested in mice none provide high level protection. We considered this might reflect the inability of these vaccine candidates to protect against chronic disease. Using Q-RT PCR we have identified 6 genes which are expressed in bacteria colonising spleens and lungs of chronically infected mice. Three of the genes (BPSL1897, BPSL3369 and BPSL2287) have been expressed in Escherichia coli and the encoded proteins purified. We have also included BPSL2765, a protein known to induce immune responses associated with a reduced incidence of chronic/recurrent disease in humans. Immunisation of mice with a combination of these antigens resulted in the induction of antibody responses against all of the proteins. Compared with mice immunised with capsular polysaccharide or LolC protein, mice immunised with the combination of chronic stage antigens showed enhanced protection against experimental disease in mice. PMID:26917010

  16. Comparative gene expression and phenotype analyses of skeletal muscle from aged wild-type and PAPP-A-deficient mice.

    PubMed

    Conover, Cheryl A; Bale, Laurie K; Nair, K Sreekumaran

    2016-07-01

    Mice deficient in pregnancy-associated plasma protein-A (PAPP-A) have extended lifespan associated with decreased incidence and severity of degenerative diseases of age, such as cardiomyopathy and nephropathy. In this study, the effect of PAPP-A deficiency on aging skeletal muscle was investigated. Whole-genome expression profiling was performed on soleus muscles from 18-month-old wild-type (WT) and PAPP-A knock-out (KO) mice of the same sex and from the same litter ('womb-mates') to identify potential mechanisms of skeletal muscle aging and its retardation in PAPP-A deficiency. Top genes regulated in PAPP-A KO compared to WT muscle were associated with increased muscle function, increased metabolism, in particular lipid metabolism, and decreased stress. Fiber cross-sectional area was significantly increased in solei from PAPP-A KO mice. In vitro contractility experiments indicated increased specific force and decreased fatigue in solei from PAPP-A KO mice. Intrinsic mitochondrial oxidative capacity was significantly increased in skeletal muscle of aged PAPP-A KO compared to WT mice. Moreover, 18-month-old PAPP-A KO mice exhibited significantly enhanced endurance running on a treadmill. Thus, PAPP-A deficiency in mice is associated with indices of healthy skeletal muscle function with age. PMID:27086066

  17. Exposure to diesel exhaust upregulates COX-2 expression in ApoE knockout mice

    PubMed Central

    Bai, Ni; Tranfield, Erin M.; Kavanagh, Terrance J.; Kaufman, Joel D.; Rosenfeld, Michael E.; van Eeden, Stephan F.

    2015-01-01

    Introduction We have shown that diesel exhaust (DE) inhalation caused progression of atherosclerosis; however, the mechanisms are not fully understood. We hypothesize that exposure to DE upregulates cyclooxygenase (COX) expression and activity, which could play a role in DE-induced atherosclerosis. Methods ApoE knockout mice (30-week old) fed with regular chow were exposed to DE (at 200 μg/m3 of particulate matter) or filtered air (control) for 7 weeks (6 h/day, 5 days/week). The protein and mRNA expression of COX-1 and COX-2 were evaluated by immunohistochemistry analysis and quantitative real-time PCR, respectively. To examine COX activity, thoracic aortae were mounted in a wire myograph, and phenylephrine (PE)-stimulated vasoconstriction was measured with and without the presence of COX antagonists (indomethacin). COX-2 activity was further assessed by urine 2,3-dinor-6-keto PGF1α level, a major metabolite of prostacyclin I2 (PGI2). Results Immunohistochemistry analysis demonstrates that DE exposure enhanced COX-2 expression in both thoracic aorta (p < 0.01) and aortic root (p < 0.03), with no modification of COX-1 expression. The increased COX-2 expression was positively correlated with smooth muscle cell content in aortic lesions (R2 = 0.4081, p < 0.008). The fractional changes of maximal vasoconstriction in the presence of indomethacin was attenuated by 3-fold after DE exposure (p < 0.02). Urine 2,3-dinor-6-keto PGF1α level was 15-fold higher in DE group than the control (p < 0.007). The mRNA expression of COX-2 (p < 0.006) and PGI synthase (p < 0.02), but not COX-1, was significantly augmented after DE exposure. Conclusion We show that DE inhalation enhanced COX-2 expression, which is also associated with phenotypic changes of aortic lesion. PMID:22746401

  18. Thromboxane prostanoid receptors enhance contractions, endothelin-1 and oxidative stress in microvessels from mice with CKD

    PubMed Central

    Wang, Cheng; Luo, Zaiming; Kohan, Donald; Wellstein, Anton; Jose, Pedro A.; Welch, William J.; Wilcox, Christopher S.; Wang, Dan

    2015-01-01

    Cardiovascular disease (CVD) is frequent in chronic kidney disease (CKD) and has been related to angiotensin II (ANG II), endothelin-1 (ET-1), thromboxane A2 (TxA2) and reactive oxygen species (ROS). Since activation of thromboxane prostanoid receptors (TP-Rs) can generate ROS which can generate ET-1, we tested the hypothesis that CKD induces cyclooxygenase (COX)-2 whose products activate TP-Rs to enhance ET-1 and ROS generation and contractions. Mesenteric resistance arterioles were isolated from C57/BL6, or TP-R +/+ and TP-R −/− mice 3 months after SHAM-operation (SHAM) or surgical reduced renal mass (RRM, n=6/group). Microvascular contractions were studied on a wire myograph. Cellular (ethidium: dihydroethidium) and mitochondrial (mitoSOX) ROS were measured by fluorescence microscopy. Mice with RRM had increased excretion of markers of oxidative stress, thromboxane, and microalbumin, increased plasma ET-1 and increased microvascular expression of p22phox, COX-2, TP-Rs, preproendothelin and endothelin-A receptors and increased arteriolar remodeling. They had increased contractions to U-46,619 (118±3 vs. 87±6, P<0.05) and ET-1 (108±5 vs. 89±4, P<0.05), which were dependent on cellular and mitochondrial ROS, COX-2, and TP-Rs. RRM doubled the ET-1-induced cellular and mitochondrial ROS generation (P<0.05). TP-R −/− mice with RRM lacked these abnormal structural and functional microvascular responses and lacked the increased systemic and the increased microvascular oxidative stress and circulating ET-1. In conclusion, RRM leads to microvascular remodeling and enhanced ET-1-induced cellular and mitochondrial ROS and contractions that are mediated by COX-2 products activating TP-Rs. Thus, TP-Rs can be upstream from enhanced ROS, ET-1, microvascular remodeling and contractility and may thereby coordinate vascular dysfunction in CKD. PMID:25733239

  19. Citalopram Enhances Neurovascular Regeneration and Sensorimotor Functional Recovery after Ischemic Stroke in Mice

    PubMed Central

    Espinera, Alyssa R.; Ogle, Molly E.; Gu, Xiaohuan; Wei, Ling

    2013-01-01

    Recent clinical trials have demonstrated that treatment with selective serotonin reuptake inhibitors (SSRIs) after stroke enhances motor functional recovery; however, the underlying mechanisms remain to be further elucidated. We hypothesized that daily administration of the clinical drug citalopram would produce these functional benefits via enhancing neurovascular repair in the ischemic peri-infarct region. To test this hypothesis, focal ischemic stroke was induced in male C57/B6 mice by permanent ligation of distal branches of the middle cerebral artery to the barrel cortex and 7-min occlusion of the bilateral common carotid arteries. Citalopram (10 mg/kg, i.p.) was injected 24 hrs after stroke and daily thereafter. To label proliferating cells, bromo-deoxyuridine was injected daily beginning 3 days after stroke. Immunohistochemical and functional assays were performed to elucidate citalopram-mediated cellular and sensorimotor changes after stroke. Citalopram treatment had no significant effect on infarct formation or edema 3 days after stroke; however, citalopram-treated mice had better functional recovery than saline-treated controls 3 and 14 days after stroke in the adhesive removal test. Increased expression of brain derived neurotrophic factor was detected in the peri-infarct region 7 days after stroke in citalopram-treated animals. The number of proliferating neural progenitor cells and the distance of neuroblast migration from the sub-ventricular zone towards the ischemic cortex were significantly greater in citalopram-treated mice at 7 days after stroke. Immunohistochemical staining and co-localization analysis showed that citalopram-treated animals generated more new neurons and microvessels in the peri-infarct region 21 and 28 days after stroke. Taken together, these results suggest that citalopram promotes post-stroke sensorimotor recovery likely via enhancing neurogenesis, neural cell migration and the microvessel support in the peri-infarct region of

  20. Genetic Background Modulates Gene Expression Profile Induced by Skin Irradiation in Ptch1 Mice

    SciTech Connect

    Galvan, Antonella; Noci, Sara; Mancuso, Mariateresa; Pazzaglia, Simonetta; Saran, Anna; Dragani, Tommaso A.

    2008-12-01

    Purpose: Ptch1 germ-line mutations in mice predispose to radiation-induced basal cell carcinoma of the skin, with tumor incidence modulated by the genetic background. Here, we examined the possible mechanisms underlying skin response to radiation in F1 progeny of Ptch1{sup neo67/+} mice crossed with either skin tumor-susceptible (Car-S) or -resistant (Car-R) mice and X-irradiated (3 Gy) at 2 days of age or left untreated. Methods and Materials: We conducted a gene expression profile analysis in mRNA samples extracted from the skin of irradiated or control mice, using Affymetrix whole mouse genome expression array. Confirmation of the results was done using real-time reverse-transcriptase polymerase chain reaction. Results: Analysis of the gene expression profile of normal skin of F1 mice at 4 weeks of age revealed a similar basal profile in the nonirradiated mice, but alterations in levels of 71 transcripts in irradiated Ptch1{sup neo67/+} mice of the Car-R cross and modulation of only eight genes in irradiated Ptch1{sup neo67/+} mice of the Car-S cross. Conclusions: These results indicate that neonatal irradiation causes a persistent change in the gene expression profile of the skin. The tendency of mice genetically resistant to skin tumorigenesis to show a more complex pattern of transcriptional response to radiation than do genetically susceptible mice suggests a role for this response in genetic resistance to basal cell tumorigenesis.

  1. Regulatory region in choline acetyltransferase gene directs developmental and tissue-specific expression in transgenic mice.

    PubMed Central

    Lönnerberg, P; Lendahl, U; Funakoshi, H; Arhlund-Richter, L; Persson, H; Ibáñez, C F

    1995-01-01

    Acetylcholine, one of the main neurotransmitters in the nervous system, is synthesized by the enzyme choline acetyltransferase (ChAT; acetyl-CoA:choline O-acetyltransferase, EC 2.3.1.6). The molecular mechanisms controlling the establishment, maintenance, and plasticity of the cholinergic phenotype in vivo are largely unknown. A previous report showed that a 3800-bp, but not a 1450-bp, 5' flanking segment from the rat ChAT gene promoter directed cell type-specific expression of a reporter gene in cholinergic cells in vitro. Now we have characterized a distal regulatory region of the ChAT gene that confers cholinergic specificity on a heterologous downstream promoter in a cholinergic cell line and in transgenic mice. A 2342-bp segment from the 5' flanking region of the ChAT gene behaved as an enhancer in cholinergic cells but as a repressor in noncholinergic cells in an orientation-independent manner. Combined with a heterologous basal promoter, this fragment targeted transgene expression to several cholinergic regions of the central nervous system of transgenic mice, including basal forebrain, cortex, pons, and spinal cord. In eight independent transgenic lines, the pattern of transgene expression paralleled qualitatively and quantitatively that displayed by endogenous ChAT mRNA in various regions of the rat central nervous system. In the lumbar enlargement of the spinal cord, 85-90% of the transgene expression was targeted to the ventral part of the cord, where cholinergic alpha-motor neurons are located. Transgene expression in the spinal cord was developmentally regulated and responded to nerve injury in a similar way as the endogenous ChAT gene, indicating that the 2342-bp regulatory sequence contains elements controlling the plasticity of the cholinergic phenotype in developing and injured neurons. Images Fig. 1 Fig. 2 PMID:7732028

  2. Distinct Effects of Chronic Dopaminergic Stimulation on Hippocampal Neurogenesis and Striatal Doublecortin Expression in Adult Mice

    PubMed Central

    Salvi, Rachele; Steigleder, Tobias; Schlachetzki, Johannes C. M.; Waldmann, Elisabeth; Schwab, Stefan; Winner, Beate; Winkler, Jürgen; Kohl, Zacharias

    2016-01-01

    While adult neurogenesis is considered to be restricted to the hippocampal dentate gyrus (DG) and the subventricular zone (SVZ), recent studies in humans and rodents provide evidence for newly generated neurons in regions generally considered as non-neurogenic, e.g., the striatum. Stimulating dopaminergic neurotransmission has the potential to enhance adult neurogenesis in the SVZ and the DG most likely via D2/D3 dopamine (DA) receptors. Here, we investigated the effect of two distinct preferential D2/D3 DA agonists, Pramipexole (PPX), and Ropinirole (ROP), on adult neurogenesis in the hippocampus and striatum of adult naïve mice. To determine newly generated cells in the DG incorporating 5-bromo-2′-deoxyuridine (BrdU) a proliferation paradigm was performed in which two BrdU injections (100 mg/kg) were applied intraperitoneally within 12 h after a 14-days-DA agonist treatment. Interestingly, PPX, but not ROP significantly enhanced the proliferation in the DG by 42% compared to phosphate buffered saline (PBS)-injected control mice. To analyze the proportion of newly generated cells differentiating into mature neurons, we quantified cells co-expressing BrdU and Neuronal Nuclei (NeuN) 32 days after the last of five BrdU injections (50 mg/kg) applied at the beginning of 14-days DA agonist or PBS administration. Again, PPX only enhanced neurogenesis in the DG significantly compared to ROP- and PBS-injected mice. Moreover, we explored the pro-neurogenic effect of both DA agonists in the striatum by quantifying neuroblasts expressing doublecortin (DCX) in the entire striatum, as well as in the dorsal and ventral sub-regions separately. We observed a significantly higher number of DCX+ neuroblasts in the dorsal compared to the ventral sub-region of the striatum in PPX-injected mice. These results suggest that the stimulation of hippocampal and dorsal striatal neurogenesis may be up-regulated by PPX. The increased generation of neural cells, both in constitutively active

  3. Distinct Effects of Chronic Dopaminergic Stimulation on Hippocampal Neurogenesis and Striatal Doublecortin Expression in Adult Mice.

    PubMed

    Salvi, Rachele; Steigleder, Tobias; Schlachetzki, Johannes C M; Waldmann, Elisabeth; Schwab, Stefan; Winner, Beate; Winkler, Jürgen; Kohl, Zacharias

    2016-01-01

    While adult neurogenesis is considered to be restricted to the hippocampal dentate gyrus (DG) and the subventricular zone (SVZ), recent studies in humans and rodents provide evidence for newly generated neurons in regions generally considered as non-neurogenic, e.g., the striatum. Stimulating dopaminergic neurotransmission has the potential to enhance adult neurogenesis in the SVZ and the DG most likely via D2/D3 dopamine (DA) receptors. Here, we investigated the effect of two distinct preferential D2/D3 DA agonists, Pramipexole (PPX), and Ropinirole (ROP), on adult neurogenesis in the hippocampus and striatum of adult naïve mice. To determine newly generated cells in the DG incorporating 5-bromo-2'-deoxyuridine (BrdU) a proliferation paradigm was performed in which two BrdU injections (100 mg/kg) were applied intraperitoneally within 12 h after a 14-days-DA agonist treatment. Interestingly, PPX, but not ROP significantly enhanced the proliferation in the DG by 42% compared to phosphate buffered saline (PBS)-injected control mice. To analyze the proportion of newly generated cells differentiating into mature neurons, we quantified cells co-expressing BrdU and Neuronal Nuclei (NeuN) 32 days after the last of five BrdU injections (50 mg/kg) applied at the beginning of 14-days DA agonist or PBS administration. Again, PPX only enhanced neurogenesis in the DG significantly compared to ROP- and PBS-injected mice. Moreover, we explored the pro-neurogenic effect of both DA agonists in the striatum by quantifying neuroblasts expressing doublecortin (DCX) in the entire striatum, as well as in the dorsal and ventral sub-regions separately. We observed a significantly higher number of DCX(+) neuroblasts in the dorsal compared to the ventral sub-region of the striatum in PPX-injected mice. These results suggest that the stimulation of hippocampal and dorsal striatal neurogenesis may be up-regulated by PPX. The increased generation of neural cells, both in constitutively active

  4. Hepatocyte-targeted HFE and TFR2 control hepcidin expression in mice

    PubMed Central

    Gao, Junwei; Chen, Juxing; De Domenico, Ivana; Koeller, David M.; Harding, Cary O.; Fleming, Robert E.; Koeberl, Dwight D.

    2010-01-01

    Hereditary hemochromatosis is caused by mutations in the hereditary hemochromatosis protein (HFE), transferrin-receptor 2 (TfR2), hemojuvelin, hepcidin, or ferroportin genes. Hepcidin is a key iron regulator, which is secreted by the liver, and decreases serum iron levels by causing the down-regulation of the iron transporter, ferroportin. Mutations in either HFE or TfR2 lower hepcidin levels, implying that both HFE and TfR2 are necessary for regulation of hepcidin expression. In this study, we used a recombinant adeno-associated virus, AAV2/8, for hepatocyte-specific expression of either Hfe or Tfr2 in mice. Expression of Hfe in Hfe-null mice both increased Hfe and hepcidin mRNA and lowered hepatic iron and Tf saturation. Expression of Tfr2 in Tfr2-deficient mice had a similar effect, whereas expression of Hfe in Tfr2-deficient mice or of Tfr2 in Hfe-null mice had no effect on liver or serum iron levels. Expression of Hfe in wild-type mice increased hepcidin mRNA and lowered iron levels. In contrast, expression of Tfr2 had no effect on wild-type mice. These findings suggest that Hfe is limiting in formation of the Hfe/Tfr2 complex that regulates hepcidin expression. In addition, these studies show that the use of recombinant AAV vector to deliver genes is a promising approach for studying physiologic consequences of protein complexes. PMID:20177050

  5. Hepatocyte-targeted HFE and TFR2 control hepcidin expression in mice.

    PubMed

    Gao, Junwei; Chen, Juxing; De Domenico, Ivana; Koeller, David M; Harding, Cary O; Fleming, Robert E; Koeberl, Dwight D; Enns, Caroline A

    2010-04-22

    Hereditary hemochromatosis is caused by mutations in the hereditary hemochromatosis protein (HFE), transferrin-receptor 2 (TfR2), hemojuvelin, hepcidin, or ferroportin genes. Hepcidin is a key iron regulator, which is secreted by the liver, and decreases serum iron levels by causing the down-regulation of the iron transporter, ferroportin. Mutations in either HFE or TfR2 lower hepcidin levels, implying that both HFE and TfR2 are necessary for regulation of hepcidin expression. In this study, we used a recombinant adeno-associated virus, AAV2/8, for hepatocyte-specific expression of either Hfe or Tfr2 in mice. Expression of Hfe in Hfe-null mice both increased Hfe and hepcidin mRNA and lowered hepatic iron and Tf saturation. Expression of Tfr2 in Tfr2-deficient mice had a similar effect, whereas expression of Hfe in Tfr2-deficient mice or of Tfr2 in Hfe-null mice had no effect on liver or serum iron levels. Expression of Hfe in wild-type mice increased hepcidin mRNA and lowered iron levels. In contrast, expression of Tfr2 had no effect on wild-type mice. These findings suggest that Hfe is limiting in formation of the Hfe/Tfr2 complex that regulates hepcidin expression. In addition, these studies show that the use of recombinant AAV vector to deliver genes is a promising approach for studying physiologic consequences of protein complexes. PMID:20177050

  6. Short-duration intermittent hypoxia enhances endurance capacity by improving muscle fatty acid metabolism in mice.

    PubMed

    Suzuki, Junichi

    2016-04-01

    This study was designed to (1) investigate the effects of acute short-duration intermittent hypoxia on musclemRNAand microRNAexpression levels; and (2) clarify the mechanisms by which short-duration intermittent hypoxia improves endurance capacity. Experiment-1: Male mice were subjected to either acute 1-h hypoxia (12% O2), acute short-duration intermittent hypoxia (12% O2for 15 min, room air for 10 min, 4 times, Int-Hypo), or acute endurance exercise (Ex). The expression of vascular endothelial growth factor-AmRNAwas significantly greater than the control at 0 h post Ex and 6 h post Int-Hypo in the deep red region of the gastrocnemius muscle. miR-16 expression levels were significantly lower at 6 and 10 h post Int-Hypo. Peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α)mRNAlevels were significantly greater than the control at 3 h post Ex and 6 h post Int-Hypo. miR-23a expression levels were lower than the control at 6-24 h post Int-Hypo. Experiment-2: Mice were subjected to normoxic exercise training with or without intermittent hypoxia for 3 weeks. Increases in maximal exercise capacity were significantly greater by training with short-duration intermittent hypoxia (IntTr) than without hypoxia. Both 3-Hydroxyacyl-CoA-dehydrogenase and total carnitine palmitoyl transferase activities were significantly enhanced in IntTr. Peroxisome proliferator-activated receptor delta andPGC-1α mRNAlevels were both significantly greater in IntTr than in the sedentary controls. These results suggest that exercise training under normoxic conditions with exposure to short-duration intermittent hypoxia represents a beneficial strategy for increasing endurance performance by enhancing fatty acid metabolism in skeletal muscle. PMID:27044851

  7. Oligodendroglial and Pan-neural Crest Expression of Cre Recombinase Directed by Sox10 Enhancer

    PubMed Central

    Stine, Zachary E.; Huynh, Jimmy L.; Loftus, Stacie K.; Gorkin, David U.; Salmasi, Amirali H.; Novak, Thomas; Purves, Todd; Miller, Ronald A.; Antonellis, Anthony; Gearhart, John P.; Pavan, William J.; McCallion, Andrew S.

    2010-01-01

    Utilizing a recently identified Sox10 distal enhancer directing Cre expression, we report S4F:Cre, a transgenic mouse line capable of inducing recombination in oligodendroglia and all examined neural crest derived tissues. Assayed using R26R:LacZ reporter mice expression was detected in neural crest derived tissues including the forming facial skeleton, dorsal root ganglia, sympathetic ganglia, enteric nervous system, aortae and melanoblasts, consistent with Sox10 expression. LacZ reporter expression was also detected in non-neural crest derived tissues including the oligodendrocytes and the ventral neural tube. This line provides appreciable differences in Cre expression pattern from other transgenic mouse lines that mark neural crest populations, including additional populations defined by the expression of other SoxE proteins. The S4F:Cre transgenic line will thus serve as a powerful tool for lineage tracing, gene function characterization and genome manipulation in these populations. PMID:19830815

  8. Dietary trans fats enhance doxorubicin-induced cardiotoxicity in mice.

    PubMed

    Mong, Mei-chin; Hsia, Te-chun; Yin, Mei-chin

    2013-10-01

    This study investigated the combined effects of trans fat diet (TFD) and doxorubicin upon cardiac oxidative, inflammatory, and coagulatory stress. TFD increased trans fatty acid deposit in heart (P < 0.05), and decreased protein C and antithrombin-III activities in circulation (P < 0.05). TFD plus doxorubicin treatment elevated activities of plasminogen activator inhibitor-1, lactate dehydrogenase, and creatine phosphokinase (P < 0.05). This combination also raised xanthine oxidase activity, and enhanced cardiac levels of reactive oxygen species, interleukin (IL)-6, IL-10, tumor necrosis factor-alpha, and monocyte chemoattractant protein-1 than TFD or doxorubicin treatment alone (P < 0.05). TFD alone increased cardiac nuclear factor kappa B (NF-κB) activity (P < 0.05), but failed to affect expression of NF-κB and mitogen-activated protein kinase (MAPK) (P > 0.05). Doxorubicin treatment alone augmented cardiac activity, mRNA expression, and protein production of NF-κB and MAPK (P < 0.05). TFD plus doxorubicin treatment further upregulated cardiac expression of NF-κB p65, p-p38, and p-ERK1/2 (P < 0.05). These findings suggest that TFD exacerbates doxorubicin-induced cardiotoxicity. PMID:24024564

  9. In vitro vitamin K(2) and 1α,25-dihydroxyvitamin D(3) combination enhances osteoblasts anabolism of diabetic mice.

    PubMed

    Poon, Christina C W; Li, Rachel W S; Seto, Sai Wang; Kong, Siu Kai; Ho, Ho Pui; Hoi, Maggie P M; Lee, Simon M Y; Ngai, Sai Ming; Chan, Shun Wan; Leung, George P H; Kwan, Yiu Wa

    2015-11-15

    In this study, we evaluated the anabolic effect and the underlying cellular mechanisms involved of vitamin K2 (10 nM) and 1α,25-dihydroxyvitamin D3 (1,25(OH)2D3) (10 nM), alone and in combination, on primary osteoblasts harvested from the iliac crests of C57BL/KsJ lean (+/+) and obese/diabetic (db/db) mice. A lower alkaline phosphatase (ALP) activity plus a reduced expression of bone anabolic markers and bone formation transcription factors (osteocalcin, Runx2, Dlx5, ATF4 and OSX) were consistently detected in osteoblasts of db/db mice compared to lean mice. A significantly higher calcium deposits formation in osteoblasts was observed in lean mice when compared to db/db mice. Co-administration of vitamin K2 (10 nM) and 1,25(OH)2D3 (10 nM) caused an enhancement of calcium deposits in osteoblasts in both strains of mice. Vitamins K2 and 1,25(OH)2D3 co-administration time-dependently (7, 14 and 21 days) increased the levels of bone anabolic markers and bone formation transcription factors, with a greater magnitude of increase observed in osteoblasts of db/db mice. Combined vitamins K2 plus 1,25(OH)2D3 treatment significantly enhanced migration and the re-appearance of surface microvilli and ruffles of osteoblasts of db/db mice. Thus, our results illustrate that vitamins K2 plus D3 combination could be a novel therapeutic strategy in treating diabetes-associated osteoporosis. PMID:26452518

  10. Absence of nonhematopoietic MHC class II expression protects mice from experimental autoimmune myocarditis.

    PubMed

    Thelemann, Christoph; Haller, Sergio; Blyszczuk, Przemyslaw; Kania, Gabriela; Rosa, Muriel; Eriksson, Urs; Rotman, Samuel; Reith, Walter; Acha-Orbea, Hans

    2016-03-01

    Experimental autoimmune myocarditis (EAM) is a CD4(+) T-cell-mediated model of human inflammatory dilated cardiomyopathies. Heart-specific CD4(+) T-cell activation is dependent on autoantigens presented by MHC class II (MHCII) molecules expressed on professional APCs. In this study, we addressed the role of inflammation-induced MHCII expression by cardiac nonhematopoietic cells on EAM development. EAM was induced in susceptible mice lacking inducible expression of MHCII molecules on all nonhematopoietic cells (pIV-/- K14 class II transactivator (CIITA) transgenic (Tg) mice) by immunization with α-myosin heavy chain peptide in CFA. Lack of inducible nonhematopoietic MHCII expression in pIV-/- K14 CIITA Tg mice conferred EAM resistance. In contrast, cardiac pathology was induced in WT and heterozygous mice, and correlated with elevated cardiac endothelial MHCII expression. Control mice with myocarditis displayed an increase in infiltrating CD4(+) T cells and in expression of IFN-γ, which is the major driver of nonhematopoietic MHCII expression. Mechanistically, IFN-γ neutralization in WT mice shortly before disease onset resulted in reduced cardiac MHCII expression and pathology. These findings reveal a previously overlooked contribution of IFN-γ to induce endothelial MHCII expression in the heart and to progress cardiac pathology during myocarditis. PMID:26621778

  11. Enhancement of leptin receptor signaling by SOCS3 deficiency induces development of gastric tumors in mice.

    PubMed

    Inagaki-Ohara, K; Mayuzumi, H; Kato, S; Minokoshi, Y; Otsubo, T; Kawamura, Y I; Dohi, T; Matsuzaki, G; Yoshimura, A

    2014-01-01

    Leptin acts on its receptor (ObR) in the hypothalamus to inhibit food intake and energy expenditure. Leptin and ObR are also expressed in the gastrointestinal tract; however, the physiological significance of leptin signaling in the gut remains uncertain. Suppressor of cytokine signaling 3 (SOCS3) is a key negative feedback regulator of ObR-mediated signaling in the hypothalamus. We now show that gastrointestinal epithelial cell-specific SOCS3 conditional knockout (T3b-SOCS3 cKO) mice developed gastric tumors by enhancing leptin production and the ObRb/signal transducer and activator of transcription 3 (STAT3) signaling pathway. All T3b-SOCS3 cKO mice developed tumors in the stomach but not in the bowels by 2 months of age, even though the SOCS3 deletion occurred in both the epithelium of stomach and bowels. The tumors developed in the absence of the inflammatory response and all cKO mice died within 6 months. These tumors displayed pathology and molecular alterations, such as an increase in MUC2 (Mucin 2, oligomeric mucus/gel-forming) and TFF3 (trefoil factor 3), resembling human intestinal-type gastric tumors. Administration of antileptin antibody to T3b-SOCS3 cKO mice reduced hyperplasia of gastric mucosa, which is the step of the initiation of gastric tumor. These data suggest that SOCS3 is an antigastric tumor gene that suppresses leptin overexpression and ObRb/STAT3 hyperactivation, supporting the hypothesis that the leptin/ObRb/STAT3 axis accelerates tumorigenesis and that it may represent a new therapeutic target for the treatment of gastric cancer. PMID:23178499

  12. Enhanced Energy Expenditure, Glucose Utilization, and Insulin Sensitivity in VAMP8 Null Mice

    PubMed Central

    Zong, Haihong; Wang, Cheng-Chun; Vaitheesvaran, Bhavapriya; Kurland, Irwin J.; Hong, Wanjin; Pessin, Jeffrey E.

    2011-01-01

    OBJECTIVE Previous studies have demonstrated that the VAMP8 protein plays a complex role in the control of granule secretion, transport vesicle trafficking, phagocytosis, and endocytosis. The present study was aimed to investigate the role of VAMP8 in mediating GLUT4 trafficking and therefore insulin action in mice. RESEARCH DESIGN AND METHODS Physiological parameters were measured using Oxymax indirect calorimetry system in 12-week-old VAMP8 null mice. Dynamic analysis of glucose homeostasis was assessed using euglycemic–hyperinsulinemic clamp coupled with tracer radioactively labeled 2-deoxyglucose. Insulin stimulated GLUT4 protein expressions on muscle cell surface were examined by immunofluorescence microscopy. RESULTS VAMP8 null mice display reduced adiposity with increased energy expenditure despite normal food intake and reduced spontaneous locomotor activity. In parallel, the VAMP8 null mice also had fasting hypoglycemia (84 ± 11 vs. 115 ± 4) and enhanced glucose tolerance with increased insulin sensitivity due to increases in both basal and insulin-stimulated glucose uptake in skeletal muscle (0.19 ± 0.04 vs. 0.09 ± 0.01 mmol/kg/min during basal, 0.6 ± 0.04 vs. 0.31 ± 0.06 mmol/kg/min during clamp in red-gastrocnemius muscle, P < 0.05). Consistent with a role for VAMP8 in the endocytosis of the insulin-responsive GLUT4, sarcolemma GLUT4 protein levels were increased in both the basal and insulin-stimulated states without any significant change in the total amount of GLUT4 protein or related facilitative glucose transporters present in skeletal muscle, GLUT1, GLUT3, and GLUT11. CONCLUSIONS These data demonstrate that, in the absence of VAMP8, the relative subcellular distribution of GLUT4 is altered, resulting in increased sarcolemma levels that can account for increased glucose clearance and insulin sensitivity. PMID:20876717

  13. Opposing regulation of cytochrome P450 expression by CAR and PXR in hypothyroid mice

    SciTech Connect

    Park, Young Joo; Lee, Eun Kyung; Lee, Yoon Kwang; Park, Do Joon; Jang, Hak Chul; Moore, David D.

    2012-09-01

    Clinical hypothyroidism affects various metabolic processes including drug metabolism. CYP2B and CYP3A are important cytochrome P450 drug metabolizing enzymes that are regulated by the xenobiotic receptors constitutive androstane receptor (CAR, NR1I3) and pregnane X receptor (PXR, NR1I2). We evaluated the regulation of the hepatic expression of CYPs by CAR and PXR in the hypothyroid state induced by a low-iodine diet containing 0.15% propylthiouracil. Expression of Cyp3a11 was suppressed in hypothyroid C57BL/6 wild type (WT) mice and a further decrement was observed in hypothyroid CAR{sup −/−} mice, but not in hypothyroid PXR{sup −/−} mice. In contrast, expression of Cyp2b10 was induced in both WT and PXR{sup −/−} hypothyroid mice, and this induction was abolished in CAR{sup −/−} mice and in and CAR{sup −/−} PXR{sup −/−} double knockouts. CAR mRNA expression was increased by hypothyroidism, while PXR expression remained unchanged. Carbamazepine (CBZ) is a commonly used antiepileptic that is metabolized by CYP3A isoforms. After CBZ treatment of normal chow fed mice, serum CBZ levels were highest in CAR{sup −/−} mice and lowest in WT and PXR{sup −/−} mice. Hypothyroid WT or PXR{sup −/−} mice survived chronic CBZ treatment, but all hypothyroid CAR{sup −/−} and CAR{sup −/−} PXR{sup −/−} mice died, with CAR{sup −/−}PXR{sup −/−} mice surviving longer than CAR{sup −/−} mice (12.3 ± 3.3 days vs. 6.3 ± 2.1 days, p = 0.04). All these findings suggest that hypothyroid status affects xenobiotic metabolism, with opposing responses of CAR and PXR and their CYP targets that can cancel each other out, decreasing serious metabolic derangement in response to a xenobiotic challenge. -- Highlights: ► Hypothyroid status activates CAR in mice and induces Cyp2b10 expression. ► Hypothyroid status suppresses PXR activity in mice and represses Cyp3a11 expression. ► These responses balance each other out in normal mice.

  14. TNF-α upregulates sclerostin expression in obese mice fed a high-fat diet.

    PubMed

    Baek, Kyunghwa; Hwang, Hyo Rin; Park, Hyun-Jung; Kwon, Arang; Qadir, Abdul S; Ko, Seong-Hee; Woo, Kyung Mi; Ryoo, Hyun-Mo; Kim, Gwan-Shik; Baek, Jeong-Hwa

    2014-05-01

    Sclerostin decreases bone mass by antagonizing the Wnt signaling pathway. We examined whether obesity-induced bone loss is associated with the expression of sclerostin. Five-week-old male mice were assigned to one of two groups (n = 10 each) and fed either a control diet (10% kcal from fat; CON) or a high-fat diet (60% kcal from fat; HF) for 12 weeks. Thex final body weight and whole body fat mass of the HF mice were higher than those of the CON mice. The distal femur cancellous bone mineral density and bone formation rate was lower in HF mice than in CON mice. The percent erosion surface was higher in the HF mice than the CON mice. The serum levels and femoral osteocytic protein expression levels of tumor necrosis factor-α (TNF-α) were significantly higher in HF mice than in CON mice. Sclerostin mRNA levels and osteocytic sclerostin protein levels in femoral cortex were also higher in HF mice than in CON mice. Sclerostin expression in MLO-Y4 osteocytes increased with TNF-α treatment, and TNF-α-induced sclerostin expression was blocked by the inhibition of NF-κB activation. Chromatin immunoprecipitation and a luciferase reporter assay demonstrated that NF-κB directly binds to the NF-κB binding elements on the mouse sost promoter and stimulates sclerostin expression. These results support a model in which, in the context of obesity or other inflammatory diseases that increase the production of TNF-α, TNF-α upregulates the expression of sclerostin through NF-κB signaling pathway, thus contributing to bone loss. PMID:24446199

  15. A single oral dose of flavan-3-ols enhances energy expenditure by sympathetic nerve stimulation in mice.

    PubMed

    Kamio, Naoya; Suzuki, Takuma; Watanabe, Yuto; Suhara, Yoshitomo; Osakabe, Naomi

    2016-02-01

    Numerous clinical studies have found that ingestion of chocolate reduces the risk of metabolic syndrome, however, the mechanisms were remain unclear. We have reported that a single dose of a flavan-3-ol fraction derived from cocoa (FL) enhanced energy expenditure (EE) and increased the mRNA expression levels of uncoupling proteins (UCPs) and peroxisome proliferator-activated receptor gamma coactivator-1 alpha (PGC-1α), and the protein level of phosphorylated AMP-activated protein kinase (AMPK)α in tissues, along with plasma adrenaline level. In the present study, we examined whether the EE enhancing activity of FL is mediated by adrenergic effect using several adrenalin receptor (AR) blockers. In the first study, mice were butoxamine, as β2AR blocker, with vehicle or 10mg/kg FL orally. We found that pretreatment with butoxamine prevented the increases of EE, the mRNA expression of UCP-3, and phosphorylated AMPKα that were induced in the gastrocnemius muscle of mice by 10mg/kg FL. Secondly, mice were given SR52930, as β3AR blocker. Pretreatment with SR52930 prevented the increases of EE, the mRNA expression of UCP-3, and phosphorylated AMPKα that were induced in the gastrocnemius muscle of mice by 10mg/kg FL. Pretreatment with a combination of both blockers also reduced the increments in mRNA expression levels of UCPs and PGC-1α, however, phosphorylated AMPKα in skeletal muscle was rather increased. These results suggest that the ability of a single oral dose of FL to enhance metabolic activity is mediated by sympathetic nerve system (SNS). PMID:26738802

  16. Altered connexin 43 expression underlies age-dependent decrease of regulatory T cell suppressor function in nonobese diabetic mice.

    PubMed

    Kuczma, Michal; Wang, Cong-Yi; Ignatowicz, Leszek; Gourdie, Robert; Kraj, Piotr

    2015-06-01

    Type 1 diabetes is one of the most extensively studied autoimmune diseases, but the cellular and molecular mechanisms leading to T cell-mediated destruction of insulin-producing β cells are still not well understood. In this study, we show that regulatory T cells (T(regs)) in NOD mice undergo age-dependent loss of suppressor functions exacerbated by the decreased ability of activated effector T cells to upregulate Foxp3 and generate T(regs) in the peripheral organs. This age-dependent loss is associated with reduced intercellular communication mediated by gap junctions, which is caused by impaired upregulation and decreased expression of connexin 43. Regulatory functions can be corrected, even in T cells isolated from aged, diabetic mice, by a synergistic activity of retinoic acid, TGF-β, and IL-2, which enhance connexin 43 and Foxp3 expression in T(regs) and restore the ability of conventional CD4(+) T cells to upregulate Foxp3 and generate peripherally derived T(regs). Moreover, we demonstrate that suppression mediated by T(regs) from diabetic mice is enhanced by a novel reagent, which facilitates gap junction aggregation. In summary, our report identifies gap junction-mediated intercellular communication as an important component of the T(reg) suppression mechanism compromised in NOD mice and suggests how T(reg) mediated immune regulation can be improved. PMID:25911751

  17. Altered CD45 isoform expression affects lymphocyte function in CD45 Tg mice.

    PubMed

    Tchilian, Elma Z; Dawes, Ritu; Hyland, Lisa; Montoya, Maria; Le Bon, Agnes; Borrow, Persephone; Hou, Sam; Tough, David; Beverley, Peter C L

    2004-09-01

    Transgenic mice have been constructed expressing high (CD45RABC) and low (CD45R0) molecular weight CD45 isoforms on a CD45-/- background. Phenotypic analysis and in vivo challenge of these mice with influenza and lymphocytic choriomeningitis viruses shows that T cell differentiation and peripheral T cell function are related to the level of CD45 expression but not to which CD45 isoform is expressed. In contrast, B cell differentiation is not restored, irrespective of the level of expression of a single isoform. All CD45 trangenic mice have T cells with an activated phenotype and increased T cell turnover. These effects are more prominent in CD8 than CD4 cells. The transgenic mice share several properties with humans expressing variant CD45 alleles and provide a model to understand immune function in variant individuals. PMID:15302847

  18. Senescence-associated phenotypes in Akita diabetic mice are enhanced by absence of bradykinin B2 receptors

    PubMed Central

    Kakoki, Masao; Kizer, Catherine M.; Yi, Xianwen; Takahashi, Nobuyuki; Kim, Hyung-Suk; Bagnell, C. Robert; Edgell, Cora-Jean S.; Maeda, Nobuyo; Jennette, J. Charles; Smithies, Oliver

    2006-01-01

    We have previously reported that genetically increased angiotensin-converting enzyme levels, or absence of the bradykinin B2 receptor, increase kidney damage in diabetic mice. We demonstrate here that this is part of a more general phenomenon — diabetes and, to a lesser degree, absence of the B2 receptor, independently but also largely additively when combined, enhance senescence-associated phenotypes in multiple tissues. Thus, at 12 months of age, indicators of senescence (alopecia, skin atrophy, kyphosis, osteoporosis, testicular atrophy, lipofuscin accumulation in renal proximal tubule and testicular Leydig cells, and apoptosis in the testis and intestine) are virtually absent in WT mice, detectable in B2 receptor–null mice, clearly apparent in mice diabetic because of a dominant mutation (Akita) in the Ins2 gene, and most obvious in Akita diabetic plus B2 receptor–null mice. Renal expression of several genes that encode proteins associated with senescence and/or apoptosis (TGF-β1, connective tissue growth factor, p53, α-synuclein, and forkhead box O1) increases in the same progression. Concomitant increases occur in 8-hydroxy-2′-deoxyguanosine, point mutations and deletions in kidney mitochondrial DNA, and thiobarbituric acid–reactive substances in plasma, together with decreases in the reduced form of glutathione in erythrocytes. Thus, absence of the bradykinin B2 receptor increases the oxidative stress, mitochondrial DNA damage, and many senescence-associated phenotypes already present in untreated Akita diabetic mice. PMID:16604193

  19. Spatiotemporal Expression and Functional Implication of CXCL14 in the Developing Mice Cerebellum

    PubMed Central

    Park, Cho Rong; Kim, Dong-Kyu; Cho, Eun Bee; You, Dong-Joo; do Rego, Jean Luc; Vaudry, David; Sun, Woong; Kim, Hyun; Seong, Jae Young; Hwang, Jong-Ik

    2012-01-01

    Cerebellar granule neurons migrate from the external granule cell layer (EGL) to the internal granule cell layer (IGL) during postnatal morphogenesis. This migration process through 4 different layers is a complex mechanism which is highly regulated by many secreted proteins. Although chemokines are well-known peptides that trigger cell migration, but with the exception of CXCL12, which is responsible for prenatal EGL formation, their functions have not been thoroughly studied in granule cell migration. In the present study, we examined cerebellar CXCL14 expression in neonatal and adult mice. CXCL14 mRNA was expressed at high levels in adult mouse cerebellum, but the protein was not detected. Nevertheless, Western blotting analysis revealed transient expression of CXCL14 in the cerebellum in early postnatal days (P1, P8), prior to the completion of granule cell migration. Looking at the distribution of CXCL14 by immunohistochemistry revealed a strong immune reactivity at the level of the Purkinje cell layer and molecular layer which was absent in the adult cerebellum. In functional assays, CXCL14 stimulated transwell migration of cultured granule cells and enhanced the spreading rate of neurons from EGL microexplants. Taken together, these results revealed the transient expression of CXCL14 by Purkinje cells in the developing cerebellum and demonstrate the ability of the chemokine to stimulate granule cell migration, suggesting that it must be involved in the postnatal maturation of the cerebellum. PMID:22843118

  20. Phycocyanin ameliorates alloxan-induced diabetes mellitus in mice: Involved in insulin signaling pathway and GK expression.

    PubMed

    Ou, Yu; Ren, Zhiheng; Wang, Jianhui; Yang, Xuegan

    2016-03-01

    The therapeutic potential and molecular mechanism of phycocyanin from Spirulina on alloxan-induced diabetes mice was investigated. In the experiment, 4-week treatment of phycocyanin at the dose of 100 and 200 mg/kg body weight in alloxan-induced diabetes mice resulted in improved metrics in comparison with alloxan-induced diabetes group. These metrics include blood glucose levels, glycosylated serum protein (GSP), glycosylated hemoglobin (GHb) and fasting serum insulin (FINS) levels. As its molecular mode of action, phycocyanin leads to the increase of IRS-1 tyrosine phosphorylation and the decrease of IRS-1 serine phosphorylation, also accompany with increased level of Akt phosphorylation on Ser473 in the liver and pancreas in diabetic mice. In addition, phycocyanin treatment enhanced the glucokinase (GK) level in the liver and pancreas, and the glucokinase regulatory protein (GKRP) level in the liver in diabetic mice. The results suggest that phycocyanin ameliorates alloxan-induced diabetes mellitus in mice by activating insulin signaling pathway and GK expression in pancreas and liver in diabetic mice. PMID:26827782

  1. Rescue of NGF-deficient mice I: transgenic expression of NGF in skin rescues mice lacking endogenous NGF.

    PubMed

    Harrison, Susan M W; Davis, Brian M; Nishimura, Merry; Albers, Kathryn M; Jones, Marc E; Phillips, Heidi S

    2004-03-30

    Mice lacking a functional NGF gene (ngf-/- mice) have less than one third of the normal complement of sensory neurons, few sympathetic postganglionic neurons and die shortly after birth. We report here that transgenic expression of NGF under control of the K14 keratin promoter can rescue some elements of the peripheral nervous system and restore normal growth and viability to ngf-/- mice. While hybrid transgenic-ngf-/- mice (ngfTKOs) displayed marginal rescue of trigeminal ganglion neurons, the percentage of CGRP-positive neurons was restored to normal. Restoration of CGRP-positive terminals in skin and spinal cord was also found and accompanied by recovery of behavioral responses to noxious stimuli. ngfTKO mice displayed a normal number of superior cervical ganglion neurons and recovery of sympathetic innervation of skin. These results demonstrate that substitution of a functional NGF locus by a transgene directing expression largely to skin can result in normal growth and viability. Thus, the most vital functions of NGF are not dependent on faithful recapitulation of the normal spatiotemporal pattern of gene expression. PMID:15010204

  2. Tolerance, fermentation, and cytokine expression in healthy aged male C57BL/6J mice fed resistant starch

    PubMed Central

    Zhou, June; Keenan, Michael J.; Keller, Jeffrey; Fernandez-Kim, Sun Ok; Pistell, Paul J.; Tulley, Richard T.; Raggio, Anne M.; Shen, Li; Zhang, Hanjie; Martin, Roy J.; Blackman, Marc R.

    2013-01-01

    Health benefits of resistant starch (RS), a dietary fermentable fiber, have been well documented in young, but not in old populations. As the essential step of more comprehensive evaluations of RS on healthy aging, we examined the effects of dietary RS on tolerance, colonic fermentation, and cytokine expression in aged mice. Healthy older (18–20 months) C57BL/6J male mice were fed control, 18% RS, or 36% RS diets for 10 weeks. Body weight gain, body composition, and fat pad weights did not differ among the three groups after 10 weeks, indicating good tolerance of the RS diet. Fermentation indicators (cecum weights, and cecal proglucagon and PYY mRNA expression) were enhanced in a RS dose dependent manner (P<0.01). Serum concentrations of soluble cytokine receptors (sTNF-Rb; sIL-4R; sIL-2Rα sVEGFR1; and sRAGE) and TNFα expression (gene and protein) in visceral fat did not differ significantly among groups. Adiponectin protein concentrations, but not gene expression, were greater in epididymal fat of the 36% RS versus control groups (P<0.05). Conclusion: in aged mice, dietary RS is well tolerated, fermented in the colon, and stimulates colonic expression of proglucagon and PYY mRNA, and adiponectin protein in visceral fat. PMID:22174009

  3. Transgenic mice expressing yellow fluorescent protein under control of the human tyrosine hydroxylase promoter.

    PubMed

    Choi, Eun Yang; Yang, Jae Won; Park, Myung Sun; Sun, Woong; Kim, Hyun; Kim, Seung U; Lee, Myung Ae

    2012-10-01

    Pathogenesis of Parkinson's disease and related catecholaminergic neurological disorders is closely associated with changes in the levels of tyrosine hydroxylase (TH). Therefore, investigation of the regulation of the TH gene system should assist in understanding the pathomechanisms involved in these neurological disorders. To identify regulatory domains that direct human TH expression in the central nervous system (CNS), we generated two transgenic mouse lines in which enhanced yellow fluorescent protein (EYFP) is expressed under the control of either 3.2-kb (hTHP-EYFP construct) human TH promoter or 3.2-kb promoter with 2-kb 3'-flanking regions (hTHP-ex3-EYFP construct) of the TH gene. In the adult transgenic mouse brain, the hTHP-EYFP construct directs neuron-specific EYFP expression in various CNS areas, such as olfactory bulb, striatum, interpeduncular nucleus, cerebral cortex, hippocampus, and particularly dentate gyrus. Although these EYFP-positive cells were identified as mature neurons, few EYFP-positive cells were TH-positive neurons. On the other hand, we could detect the EYFP mRNA expression in a subset of neurons in the olfactory bulb, midbrain, and cerebellum, in which expression of endogenous TH is enriched, with hTHP-ex3-EYFP transgenic mice. These results indicate that the 3.2-kb sequence upstream of the TH gene is not sufficient for proper expression and that the 2-kb sequence from the translation start site to exon 3 is necessary for expression of EYFP in a subset of catecholaminergic neurons. PMID:22714400

  4. HCELL Expression on Murine MSC Licenses Pancreatotropism and Confers Durable Reversal of Autoimmune Diabetes in NOD Mice

    PubMed Central

    Abdi, Reza; Moore, Robert; Sakai, Shinobu; Donnelly, Conor B.; Mounayar, Marwan; Sackstein, Robert

    2015-01-01

    Type 1 diabetes (T1D) is an immune-mediated disease resulting in destruction of insulin-producing pancreatic beta cells. Mesenchymal stem cells (MSCs) possess potent immunomodulatory properties, garnering increasing attention as cellular therapy for T1D and other immunologic diseases. However, MSCs generally lack homing molecules, hindering their colonization at inflammatory sites following intravenous (IV) administration. Here we analyzed whether enforced E-selectin ligand expression on murine MSCs could impact their effect in reversing hyperglycemia in non-obese diabetic (NOD) mice. Though murine MSCs natively do not express the E-selectin binding determinant sialyl Lewisx (sLex), we found that fucosyltransferase-mediated α(1,3)-exofucosylation of murine MSCs resulted in sLex display uniquely on cell surface CD44 thereby creating HCELL, the E-selectin-binding glycoform of CD44. Following IV infusion into diabetic NOD mice, allogeneic HCELL+ MSCs showed 3-fold greater peri-islet infiltrates compared to buffer-treated (i.e., HCELL−) MSCs, with distribution in proximity to E-selectin-expressing microvessels. Exofucosylation had no effect on MSC immunosuppressive capacity in in vitro assays, however, though engraftment was temporary for both HCELL+ and HCELL− MSCs, administration of HCELL+ MSCs resulted in durable reversal of hyperglycemia, whereas only transient reversal was observed following administration of HCELL− MSCs. Notably, exofucosylation of MSCs generated from CD44−/− mice induced prominent membrane expression of sLex, but IV administration of these MSCs into hyperglycemic NOD mice showed no enhanced pancreatotropism or reversal of hyperglycemia. These findings provide evidence that glycan engineering to enforce HCELL expression boosts trafficking of infused MSCs to pancreatic islets of NOD mice and substantially improves their efficacy in reversing autoimmune diabetes. PMID:25641589

  5. Taurine-Induced Long-Lasting Enhancement of Synaptic Transmission in Mice: Role of Transporters

    PubMed Central

    Sergeeva, O A; Chepkova, A N; Doreulee, N; Eriksson, K S; Poelchen, W; Mönnighoff, I; Heller-Stilb, B; Warskulat, U; Häussinger, D; Haas, H L

    2003-01-01

    Taurine, a major osmolyte in the brain evokes a long-lasting enhancement (LLETAU) of synaptic transmission in hippocampal and cortico-striatal slices. Hippocampal LLETAU was abolished by the GABA uptake blocker nipecotic acid (NPA) but not by the taurine-uptake inhibitor guanidinoethyl sulphonate (GES). Striatal LLETAU was sensitive to GES but not to NPA. Semiquantitative PCR analysis and immunohistochemistry revealed that taurine transporter expression is significantly higher in the striatum than in the hippocampus. Taurine transporter-deficient mice displayed very low taurine levels in both structures and a low ability to develop LLETAU in the striatum, but not in the hippocampus. The different mechanisms of taurine-induced synaptic plasticity may reflect the different vulnerabilities of these brain regions under pathological conditions that are accompanied by osmotic changes such as hepatic encephalopathy. PMID:12824447

  6. Generation of an ABCG2{sup GFPn-puro} transgenic line - A tool to study ABCG2 expression in mice

    SciTech Connect

    Orford, Michael; Mean, Richard; Lapathitis, George; Genethliou, Nicholas; Panayiotou, Elena; Panayi, Helen; Malas, Stavros

    2009-06-26

    The ATP-binding cassette (ABC) transporter 2 (ABCG2) is expressed by stem cells in many organs and in stem cells of solid tumors. These cells are isolated based on the side population (SP) phenotype, a Hoechst 3342 dye efflux property believed to be conferred by ABCG2. Because of the limitations of this approach we generated transgenic mice that express Nuclear GFP (GFPn) coupled to the Puromycin-resistance gene, under the control of ABCG2 promoter/enhancer sequences. We show that ABCG2 is expressed in neural progenitors of the developing forebrain and spinal cord and in embryonic and adult endothelial cells of the brain. Using the neurosphere assay, we isolated tripotent ABCG2-expressing neural stem cells from embryonic mouse brain. This transgenic line is a powerful tool for studying the expression of ABCG2 in many tissues and for performing functional studies in different experimental settings.

  7. Increased Bacterial Load and Expression of Antimicrobial Peptides in Skin of Barrier-Deficient Mice with Reduced Cancer Susceptibility.

    PubMed

    Natsuga, Ken; Cipolat, Sara; Watt, Fiona M

    2016-01-01

    Mice lacking three epidermal barrier proteins-envoplakin, periplakin, and involucrin (EPI-/- mice)-have a defective cornified layer, reduced epidermal γδ T cells, and increased dermal CD4(+) T cells. They are also resistant to developing skin tumors. The tumor-protective mechanism involves signaling between Rae-1 expressing keratinocytes and the natural killer group 2D receptor on immune cells, which also plays a role in host defenses against infection. Given the emerging link between bacteria and cancer, we investigated whether EPI-/- mice have an altered skin microbiota. The bacterial phyla were similar in wild-type and EPI-/- skin. However, bacteria were threefold more abundant in EPI-/- skin and penetrated deeper into the epidermis. The major epithelial defense mechanism against bacteria is production of antimicrobial proteins (AMPs). EPI-/- skin exhibited enhanced expression of antimicrobial peptides. However, reducing the bacterial load by antibiotic treatment or breeding mice under specific pathogen-free conditions did not reduce AMP expression or alleviate the abnormalities in T-cell populations. We conclude that the atopic characteristics of EPI-/- skin are a consequence of the defective barrier rather than a response to the increased bacterial load. It is therefore unlikely that the increase in skin microbiota contributes directly to the observed cancer resistance. PMID:26763429

  8. Increased Bacterial Load and Expression of Antimicrobial Peptides in Skin of Barrier-Deficient Mice with Reduced Cancer Susceptibility

    PubMed Central

    Natsuga, Ken; Cipolat, Sara; Watt, Fiona M.

    2016-01-01

    Mice lacking three epidermal barrier proteins—envoplakin, periplakin, and involucrin (EPI-/- mice)—have a defective cornified layer, reduced epidermal γδ T cells, and increased dermal CD4+ T cells. They are also resistant to developing skin tumors. The tumor-protective mechanism involves signaling between Rae-1 expressing keratinocytes and the natural killer group 2D receptor on immune cells, which also plays a role in host defenses against infection. Given the emerging link between bacteria and cancer, we investigated whether EPI-/- mice have an altered skin microbiota. The bacterial phyla were similar in wild-type and EPI-/- skin. However, bacteria were threefold more abundant in EPI-/- skin and penetrated deeper into the epidermis. The major epithelial defense mechanism against bacteria is production of antimicrobial proteins (AMPs). EPI-/- skin exhibited enhanced expression of antimicrobial peptides. However, reducing the bacterial load by antibiotic treatment or breeding mice under specific pathogen-free conditions did not reduce AMP expression or alleviate the abnormalities in T-cell populations. We conclude that the atopic characteristics of EPI-/- skin are a consequence of the defective barrier rather than a response to the increased bacterial load. It is therefore unlikely that the increase in skin microbiota contributes directly to the observed cancer resistance. PMID:26763429

  9. Minocycline enhances hippocampal memory, neuroplasticity and synapse-associated proteins in aged C57 BL/6 mice.

    PubMed

    Jiang, Ying; Liu, Yingying; Zhu, Cansheng; Ma, Xiaomeng; Ma, Lili; Zhou, Linli; Huang, Qiling; Cen, Lei; Pi, Rongbiao; Chen, Xiaohong

    2015-05-01

    Previous studies have suggested that minocycline can attenuate cognitive deficits in animal models of conditions such as Alzheimer's disease and cerebral ischemia through inhibiting microglia associated anti-inflammatory actions. However the pathway that minocycline targets to enhance cognitive performance is not fully defined. Here we examined the effects of minocycline on learning and memory in aged (22-month-old) C57 BL/6 mice. We treated one group of mice with minocycline (30 mg/kg/day), and another group of mice with donepezil (2 mg/kg/day) as a positive control. The Morris water maze and passive avoidance tests were used to evaluate the effects of minocycline on learning and memory deficits. We also used high-frequency stimulation-induced long-term potentiation and Golgi-Cox staining to assess the effect of minocycline on synaptic plasticity and synaptogenesis. The effects of minocycline on synapse-associated signaling proteins were determined by western blot. We found that minocycline ameliorates cognitive deficits, enhances neuroplasticity, activates brain-derived neurotrophic factor- extracellular signal-regulated kinases signaling and increases expression of Arc, EGR1 and PSD-95 in the CA1 and dentate gyrus regions of the hippocampus in aged mice. The effects of minocycline in aged mice were similar to those of donepezil. Our results suggest that minocycline could improve learning and memory through enhancing synaptic plasticity and synaptogenesis, modulating the expression of synapse-associated signaling proteins, which provide a rationale for exploring the viability of using minocycline treatment in cognitive deficits. PMID:25838119

  10. Olfactory discrimination varies in mice with different levels of α7-nicotinic acetylcholine receptor expression

    PubMed Central

    Hellier, Jennifer L.; Arevalo, Nicole L.; Blatner, Megan J.; Dang, An K.; Clevenger, Amy C.; Adams, Catherine E.; Restrepo, Diego

    2010-01-01

    Previous studies have shown that schizophrenics have decreased expression of α7-nicotinic acetylcholine (α7) receptors in the hippocampus and other brain regions, paranoid delusions, disorganized speech, deficits in auditory gating (i.e., inability to inhibit neuronal responses to repetitive auditory stimuli), and difficulties in odor discrimination and detection. Here we use mice with decreased α7 expression that also show a deficit in auditory gating to determine if these mice have similar deficits in olfaction. In the adult mouse olfactory bulb (OB), α7 expression localizes in the glomerular layer; however, the functional role of α7 is unknown. We show that inbred mouse strains (i.e., C3H and C57) with varying α7 expression (e.g., α7 wild-type [α7+/+], α7 heterozygous knock-out [α7+/−] and α7 homozygous knockout mice [α7−/−]) significantly differ in odor discrimination and detection of chemically related odorant pairs. Using [125I] α-bungarotoxin (α-BGT) autoradiography, α7 expression was measured in the OB. As previously demonstrated, α-BGT binding was localized to the glomerular layer. Significantly more expression of α7 was observed in C57 α7+/+ mice compared to C3H α7+/+ mice. Furthermore, C57 α7+/+ mice were able to detect a significantly lower concentration of an odor in a mixture compared to C3H α7+/+ mice. Both C57 and C3H α7+/+ mice discriminated between chemically related odorants sooner than α7+/− or α7−/− mice. These data suggest that α7-nicotinic-receptors contribute strongly to olfactory discrimination and detection in mice and may be one of the mechanisms producing olfactory dysfunction in schizophrenics. PMID:20713028

  11. The progesterone and estrogen modify the uterine prolactin and prolactin receptor expression of hyperprolactinemic mice.

    PubMed

    do Amaral, Vinícius Cestari; Carvalho, Kátia Candido; Maciel, Gustavo Arantes Rosa; Simoncini, Tommaso; da Silva, Priscilla Ludovico; Marcondes, Rodrigo Rodrigues; Soares, José Maria; Baracat, Edmund Chada

    2015-02-01

    The aim of this study was to evaluate the effects of metoclopramide-induced hyperprolactinemia on the prolactin (PRL) and PRL receptor's expression in the uterus of mice. For this purpose, 49 Swiss mice were divided into the following groups: GrSS (non-ovariectomized mice given vehicle); GrMET (non-ovariectomized mice treated with metoclopramide); OvSS (ovariectomized mice given vehicle); OvMET (ovariectomized mice treated with metoclopramide); OvMET+17βE (ovariectomized mice treated with metoclopramide and 17β estradiol); OvMET+MP (ovariectomized mice treated with metoclopramide and micronized progesterone); OvMET+17βE+MP (ovariectomized mice treated with metoclopramide and a solution of 17β estradiol and micronized progesterone). Immunohistochemical analyzes were evaluated semi-quantitatively. Our results showed that GrMET, OvMET+MP, and OvMET+17βE+MP presented strong PRL expression. OvMET and OvMET+17βE presented mild reaction, while GrSS and OvSS presented weak reaction. Concerning PRL receptor, OvMET+MP and OvMET+17βE+MP showed strong reaction; GrMET, OvSS, and OvMET+17βE showed mild reaction; and GrSS and OvMET showed weak reaction. These findings suggest that progesterone alone or in combination with estrogen may increase the expression of uterine PRL and PRL receptor. PMID:25299230

  12. Female Mice Lacking Estrogen Receptor-α in Hypothalamic Proopiomelanocortin (POMC) Neurons Display Enhanced Estrogenic Response on Cortical Bone Mass

    PubMed Central

    Farman, H. H.; Windahl, S. H.; Westberg, L.; Isaksson, H.; Egecioglu, E.; Schele, E.; Ryberg, H.; Jansson, J. O.; Tuukkanen, J.; Koskela, A.; Xie, S. K.; Hahner, L.; Zehr, J.; Clegg, D. J.; Lagerquist, M. K.

    2016-01-01

    Estrogens are important regulators of bone mass and their effects are mainly mediated via estrogen receptor (ER)α. Central ERα exerts an inhibitory role on bone mass. ERα is highly expressed in the arcuate (ARC) and the ventromedial (VMN) nuclei in the hypothalamus. To test whether ERα in proopiomelanocortin (POMC) neurons, located in ARC, is involved in the regulation of bone mass, we used mice lacking ERα expression specifically in POMC neurons (POMC-ERα−/−). Female POMC-ERα−/− and control mice were ovariectomized (OVX) and treated with vehicle or estradiol (0.5 μg/d) for 6 weeks. As expected, estradiol treatment increased the cortical bone thickness in femur, the cortical bone mechanical strength in tibia and the trabecular bone volume fraction in both femur and vertebrae in OVX control mice. Importantly, the estrogenic responses were substantially increased in OVX POMC-ERα−/− mice compared with the estrogenic responses in OVX control mice for cortical bone thickness (+126 ± 34%, P < .01) and mechanical strength (+193 ± 38%, P < .01). To test whether ERα in VMN is involved in the regulation of bone mass, ERα was silenced using an adeno-associated viral vector. Silencing of ERα in hypothalamic VMN resulted in unchanged bone mass. In conclusion, mice lacking ERα in POMC neurons display enhanced estrogenic response on cortical bone mass and mechanical strength. We propose that the balance between inhibitory effects of central ERα activity in hypothalamic POMC neurons in ARC and stimulatory peripheral ERα-mediated effects in bone determines cortical bone mass in female mice. PMID:27254004

  13. Female Mice Lacking Estrogen Receptor-α in Hypothalamic Proopiomelanocortin (POMC) Neurons Display Enhanced Estrogenic Response on Cortical Bone Mass.

    PubMed

    Farman, H H; Windahl, S H; Westberg, L; Isaksson, H; Egecioglu, E; Schele, E; Ryberg, H; Jansson, J O; Tuukkanen, J; Koskela, A; Xie, S K; Hahner, L; Zehr, J; Clegg, D J; Lagerquist, M K; Ohlsson, C

    2016-08-01

    Estrogens are important regulators of bone mass and their effects are mainly mediated via estrogen receptor (ER)α. Central ERα exerts an inhibitory role on bone mass. ERα is highly expressed in the arcuate (ARC) and the ventromedial (VMN) nuclei in the hypothalamus. To test whether ERα in proopiomelanocortin (POMC) neurons, located in ARC, is involved in the regulation of bone mass, we used mice lacking ERα expression specifically in POMC neurons (POMC-ERα(-/-)). Female POMC-ERα(-/-) and control mice were ovariectomized (OVX) and treated with vehicle or estradiol (0.5 μg/d) for 6 weeks. As expected, estradiol treatment increased the cortical bone thickness in femur, the cortical bone mechanical strength in tibia and the trabecular bone volume fraction in both femur and vertebrae in OVX control mice. Importantly, the estrogenic responses were substantially increased in OVX POMC-ERα(-/-) mice compared with the estrogenic responses in OVX control mice for cortical bone thickness (+126 ± 34%, P < .01) and mechanical strength (+193 ± 38%, P < .01). To test whether ERα in VMN is involved in the regulation of bone mass, ERα was silenced using an adeno-associated viral vector. Silencing of ERα in hypothalamic VMN resulted in unchanged bone mass. In conclusion, mice lacking ERα in POMC neurons display enhanced estrogenic response on cortical bone mass and mechanical strength. We propose that the balance between inhibitory effects of central ERα activity in hypothalamic POMC neurons in ARC and stimulatory peripheral ERα-mediated effects in bone determines cortical bone mass in female mice. PMID:27254004

  14. Deletion of the core-H region in mice abolishes the expression of three proximal odorant receptor genes in cis

    PubMed Central

    Nishizumi, Hirofumi; Kumasaka, Kouhei; Inoue, Nobuko; Nakashima, Ai; Sakano, Hitoshi

    2007-01-01

    We have previously reported that a 2.1-kb homology (H) sequence, conserved between mouse and human, regulates the odorant receptor (OR) gene MOR28 in transgenic mice. Here, we narrowed down the essential sequences of the H to a core of 124 bp by using a transient expression system in zebrafish embryos. Transgenic experiments in mice demonstrated that the core-H sequence is sufficient to endow expression of the MOR28 minigene. Deletion and mutation analyses of the core-H region revealed two homeodomain sequences to be essential for the H enhancer activity. Targeted deletion of the core-H abolished expression of three proximal OR genes, MOR28, MOR10, and MOR83, in cis, indicating the presence of another locus control region/enhancer in the downstream region, that regulates four distal OR genes in the same MOR28 cluster. In the heterozygous mice, the H− phenotype of the mutant allele was not rescued by the wild-type H+ allele in trans. PMID:18077433

  15. Increased liver pathology in hepatitis C virus transgenic mice expressing the hepatitis B virus X protein

    SciTech Connect

    Keasler, Victor V.; Lerat, Herve; Madden, Charles R.; Finegold, Milton J.; McGarvey, Michael J.; Mohammed, Essam M.A.; Forbes, Stuart J.; Lemon, Stanley M.; Hadsell, Darryl L.; Grona, Shala J.; Hollinger, F. Blaine; Slagle, Betty L. . E-mail: bslagle@bcm.edu

    2006-04-10

    Transgenic mice expressing the full-length HCV coding sequence were crossed with mice that express the HBV X gene-encoded regulatory protein HBx (ATX mice) to test the hypothesis that HBx expression accelerates HCV-induced liver pathogenesis. At 16 months (mo) of age, hepatocellular carcinoma was identified in 21% of HCV/ATX mice, but in none of the single transgenic animals. Analysis of 8-mo animals revealed that, relative to HCV/WT mice, HCV/ATX mice had more severe steatosis, greater liver-to-body weight ratios, and a significant increase in the percentage of hepatocytes staining for proliferating cell nuclear antigen. Furthermore, primary hepatocytes from HCV, ATX, and HCV/ATX transgenic mice were more resistant to fas-mediated apoptosis than hepatocytes from nontransgenic littermates. These results indicate that HBx expression contributes to increased liver pathogenesis in HCV transgenic mice by a mechanism that involves an imbalance in hepatocyte death and regeneration within the context of severe steatosis.

  16. Effects of diet-induced obesity on secondary tumor development and plasma cytokine expression in mice

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The present study investigated the effects of diet-induced obesity on secondary tumor development and expression of plasma cytokines in mice. Three-wk old male C57BL/6 mice were fed the AIN-93G diet or a 45% fat diet (kcal %; n=25/group) for 7 wks before they were subcutaneously injected with 2.5 x ...

  17. Mice Expressing a "Hyper-Sensitive" Form of the Cannabinoid Receptor 1 (CB1) Are Neither Obese Nor Diabetic

    PubMed Central

    Andrews, Mary-Jeanette; Amin, Randa; Henderson-Redmond, Angela N.; Mackie, Ken; Czyzyk, Traci A.; Morgan, Daniel J.

    2016-01-01

    Multiple lines of evidence implicate the endocannabinoid signaling system in the modulation of metabolic disease. Genetic or pharmacological inactivation of CB1 in rodents leads to reduced body weight, resistance to diet-induced obesity, decreased intake of highly palatable food, and increased energy expenditure. Cannabinoid agonists stimulate feeding in rodents and increased levels of endocannabinoids can disrupt lipid metabolism. Therefore, the hypothesis that sustained endocannabinoid signaling can lead to obesity and diabetes was examined in this study using S426A/S430A mutant mice expressing a desensitization-resistant CB1 receptor. These mice display exaggerated and prolonged responses to acute administration of phytocannabinoids, synthetic cannabinoids, and endocannabinoids. As a consequence these mice represent a novel model for determining the effect of enhanced endocannabinoid signaling on metabolic disease. S426A/S430A mutants consumed equivalent amounts of both high fat (45%) and low fat (10%) chow control diet compared to wild-type littermate controls. S426A/S430A mutants and wild-type mice fed either high or low fat control diet displayed similar fasting blood glucose levels and normal glucose clearance following a 2 g/kg glucose challenge. Furthermore, S426A/S430A mutants and wild-type mice consumed similar amounts of chow following an overnight fast. While both THC and JZL195 significantly increased food intake two hours after injection, this increase was similar between the S426A/S430A mutant and wildtype control mice Our results indicate that S426A/S430A mutant mice expressing the desensitization-resistant form of CB1 do not exhibit differences in body weight, food intake, glucose homeostasis, or re-feeding following a fast. PMID:27501235

  18. Mice Expressing a "Hyper-Sensitive" Form of the Cannabinoid Receptor 1 (CB1) Are Neither Obese Nor Diabetic.

    PubMed

    Marcus, David J; Zee, Michael L; Davis, Brian J; Haskins, Chris P; Andrews, Mary-Jeanette; Amin, Randa; Henderson-Redmond, Angela N; Mackie, Ken; Czyzyk, Traci A; Morgan, Daniel J

    2016-01-01

    Multiple lines of evidence implicate the endocannabinoid signaling system in the modulation of metabolic disease. Genetic or pharmacological inactivation of CB1 in rodents leads to reduced body weight, resistance to diet-induced obesity, decreased intake of highly palatable food, and increased energy expenditure. Cannabinoid agonists stimulate feeding in rodents and increased levels of endocannabinoids can disrupt lipid metabolism. Therefore, the hypothesis that sustained endocannabinoid signaling can lead to obesity and diabetes was examined in this study using S426A/S430A mutant mice expressing a desensitization-resistant CB1 receptor. These mice display exaggerated and prolonged responses to acute administration of phytocannabinoids, synthetic cannabinoids, and endocannabinoids. As a consequence these mice represent a novel model for determining the effect of enhanced endocannabinoid signaling on metabolic disease. S426A/S430A mutants consumed equivalent amounts of both high fat (45%) and low fat (10%) chow control diet compared to wild-type littermate controls. S426A/S430A mutants and wild-type mice fed either high or low fat control diet displayed similar fasting blood glucose levels and normal glucose clearance following a 2 g/kg glucose challenge. Furthermore, S426A/S430A mutants and wild-type mice consumed similar amounts of chow following an overnight fast. While both THC and JZL195 significantly increased food intake two hours after injection, this increase was similar between the S426A/S430A mutant and wildtype control mice Our results indicate that S426A/S430A mutant mice expressing the desensitization-resistant form of CB1 do not exhibit differences in body weight, food intake, glucose homeostasis, or re-feeding following a fast. PMID:27501235

  19. Skeletal phenotype of growing transgenic mice that express a function-perturbing form of beta1 integrin in osteoblasts

    NASA Technical Reports Server (NTRS)

    Globus, R. K.; Amblard, D.; Nishimura, Y.; Iwaniec, U. T.; Kim, J-B; Almeida, E. A. C.; Damsky, C. D.; Wronski, T. J.; van der Meulen, M. C. H.

    2005-01-01

    Skeletal modeling entails the deposition of large amounts of extracellular matrix (ECM) to form structures tailored to withstand increasing mechanical loads during rapid growth. Specific ECM molecules bind to integrin receptors on the cell surface, thereby triggering a cascade of signaling events that affect critical cell functions. To evaluate the role of integrins during skeletal growth, transgenic mice were engineered to express a function-perturbing fragment of beta1 integrin consisting of the transmembrane domain and cytoplasmic tail under the control of the osteocalcin promoter (TG mice). Thus, transgene expression was targeted to mature cells of the osteoblast lineage, and herein we show that cultured cells resembling osteocytes from 90-day-old TG mice display impaired adhesion to collagen I, a ligand for beta1 integrin. To determine the influence of beta1 integrin on bones that are responsible for providing structural support during periods of rapid growth, we examined the phenotype of the appendicular skeleton in TG mice compared to wild type (WT) mice. According to radiographs, bones from mice of both genotypes between 14 and 90 days of age appeared similar in gross structure and density, although proximal tibiae from 35-90 days old TG mice were less curved than those of WT mice (72-92% TG/WT). Although there were only mild and transient differences in absolute bone mass and strength, once normalized to body mass, the tibial dry mass (79.1% TG/WT females), ash mass (78.5% TG/WT females), and femoral strength in torsion (71.6% TG/WT females) were reduced in TG mice compared to WT mice at 90 days of age. Similar effects of genotype on bone mass and curvature were observed in 1-year-old retired breeders, indicating that these phenotypic differences between TG and WT mice were stable well into adulthood. Effects of genotype on histomorphometric indices of cancellous bone turnover were minimal and evident only transiently during growth, but when present they

  20. The effects of HIV-1 regulatory TAT protein expression on brain reward function, response to psychostimulants and delay-dependent memory in mice.

    PubMed

    Kesby, James P; Markou, Athina; Semenova, Svetlana

    2016-10-01

    Depression and psychostimulant abuse are common comorbidities among humans with immunodeficiency virus (HIV) disease. The HIV regulatory protein TAT is one of multiple HIV-related proteins associated with HIV-induced neurotoxicity. TAT-induced dysfunction of dopamine and serotonin systems in corticolimbic brain areas may result in impaired reward function, thus, contributing to depressive symptoms and psychostimulant abuse. Transgenic mice with doxycycline-induced TAT protein expression in the brain (TAT+, TAT- control) show neuropathology resembling brain abnormalities in HIV+ humans. We evaluated brain reward function in response to TAT expression, nicotine and methamphetamine administration in TAT+ and TAT- mice using the intracranial self-stimulation procedure. We evaluated the brain dopamine and serotonin systems with high-performance liquid chromatography. The effects of TAT expression on delay-dependent working memory in TAT+ and TAT- mice using the operant delayed nonmatch-to-position task were also assessed. During doxycycline administration, reward thresholds were elevated by 20% in TAT+ mice compared with TAT- mice. After the termination of doxycycline treatment, thresholds of TAT+ mice remained significantly higher than those of TAT- mice and this was associated with changes in mesolimbic serotonin and dopamine levels. TAT+ mice showed a greater methamphetamine-induced threshold lowering compared with TAT- mice. TAT expression did not alter delay-dependent working memory. These results indicate that TAT expression in mice leads to reward deficits, a core symptom of depression, and a greater sensitivity to methamphetamine-induced reward enhancement. Our findings suggest that the TAT protein may contribute to increased depressive-like symptoms and continued methamphetamine use in HIV-positive individuals. PMID:27316905

  1. Hepatic changes in metabolic gene expression in old ghrelin and ghrelin receptor knockout mice

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Ghrelin knockout (GKO) and ghrelin receptor (growth hormone secretagogue receptor) knockout (GHSRKO) mice exhibit enhanced insulin sensitivity, but the mechanism is unclear. Insulin sensitivity declines with age and is inversely associated with accumulation of lipid in liver, a key glucoregulatory ...

  2. Sustained Dystrophin Expression Induced by Peptide-conjugated Morpholino Oligomers in the Muscles of mdx Mice

    PubMed Central

    Jearawiriyapaisarn, Natee; Moulton, Hong M; Buckley, Brian; Roberts, Jennifer; Sazani, Peter; Fucharoen, Suthat; Iversen, Patrick L; Kole, Ryszard

    2009-01-01

    Cell-penetrating peptides (CPPs), containing arginine (R), 6-aminohexanoic acid (X), and/or β-alanine (B) conjugated to phosphorodiamidate morpholino oligomers (PMOs), enhance their delivery in cell culture. In this study, the potency, functional biodistribution, and toxicity of these conjugates were evaluated in vivo, in EGFP-654 transgenic mice that ubiquitously express the aberrantly spliced EGFP-654 pre-mRNA reporter. Correct splicing and enhanced green fluorescence protein (EGFP) upregulation serve as a positive readout for peptide-PMO (PPMO) entry into cells and access to EGFP-654 pre-mRNA in the nucleus. Intraperitoneal injections of a series of PPMOs, A-N (12 mg/kg), administered once a day for four successive days resulted in splicing correction in numerous tissues. PPMO-B was highly potent in the heart, diaphragm, and quadriceps, which are key muscles in the treatment of Duchenne muscular dystrophy. We therefore investigated PPMO M23D-B, designed to force skipping of stop-codon containing dystrophin exon 23, in an mdx mouse model of the disease. Systemic delivery of M23D-B yielded persistent exon 23 skipping, yielding high and sustained dystrophin protein expression in body-wide muscles, including cardiac muscle, without detectable toxicity. The rescued dystrophin reduced serum creatinine kinase to near-wild-type levels, indicating improvement in muscle integrity. This is the first report of oligonucleotide-mediated exon skipping and dystrophin protein induction in the heart of treated animals. PMID:18545222

  3. [Allergens-induced sensitization alters airway epithelial adhesion molecules expression in mice].

    PubMed

    Zeng, Dan; Tan, Mei-Ling; Xiang, Yang; Qin, Xiao-Qun; Zhu, Li-Ming; Dai, Ai-Guo

    2015-12-25

    To explore the relationship between the epithelial adhesion molecules and immune responses of airway epithelium, we observed the expression of integrin β4 and intercellular adhesion molecule-1 (ICAM-1) in the mice airway epithelium after sensitization with allergens. BALB/c mice were sensitized with intraperitoneal injection of ovalbumin (OVA) or house dust mite (HDM) and then developed airway hyper-responsiveness as determined by barometric whole-body plethysmography. Both OVA and HDM sensitization led to increases of the number of peripheral leukocytes as well as inflammatory cells infiltration in lungs. OVA sensitized mice showed more severe inflammatory cells infiltration than HDM sensitized mice. Immunohistochemistry analysis of mice lung tissues revealed that sensitization with both allergens also led to a decrease of integrin β4 expression and an increase of ICAM-1 expression in airway epithelia. OVA sensitized mice showed a more significant increase of ICAM-1 expression compared with HDM sensitized mice. siRNA mediated silencing of integrin β4 gene in 16HBE cells resulted in an up-regulation of ICAM-1 expression. Our results indicate a possible role of airway epithelial adhesion molecules in allergen-induced airway immune responses. PMID:26701635

  4. Lack of endogenous IL-10 enhances production of proinflammatory cytokines and leads to Brucella abortus clearance in mice.

    PubMed

    Corsetti, Patrícia P; de Almeida, Leonardo A; Carvalho, Natália B; Azevedo, Vasco; Silva, Teane M A; Teixeira, Henrique C; Faria, Ana C; Oliveira, Sergio C

    2013-01-01

    IL-10 is a cytokine that regulates the balance between pathogen clearance and immunopathology. Brucella abortus is an intracellular bacterium that causes chronic disease in humans and domestic animals. Here we evaluated the contribution of IL-10 in host immune response and pathology during B. abortus infection. To assess the role of IL-10 in vivo, IL-10 knockout (KO) or 129 Sv/Ev (wild-type) mice were infected with B. abortus and the number of viable bacteria from the spleen was determined at 1, 2, 3, 6 and 14-weeks postinfection. IL-10 KO mice showed reduced bacterial loads in the spleen when compared to wild-type mice during all time points studied. Additionally, at 14-weeks postinfection IL-10 KO mice had totally cleared the infection. This clearance was preceded by an enhanced IFN-γ, TNF-α and IL-17 responses in both the serum and the spleen of IL-10 KO mice. Additionally, dendritic cells from infected IL-10 KO mice produced elevated levels of IL-12 and TNF-α compared to wild-type animals. Histopathology analysis was performed and both KO and wild-type mice developed multifocal granulomas and necrosis in the liver. However, at six-weeks postinfection reduced numbers of granulomas was detected in IL-10 KO mice compared to wild-type animals. This reduced liver pathology at later stage of infection was accompanied by increased numbers of CD4+CD25+foxp3+ T cells and expression of TGF-β in IL-10 KO splenocytes. Taken together, our findings demonstrate that IL-10 modulates the proinflammatory immune response to B. abortus infection and the lack of IL-10 increases resistance to Brucella infection. PMID:24069337

  5. Lack of Endogenous IL-10 Enhances Production of Proinflammatory Cytokines and Leads to Brucella abortus Clearance in Mice

    PubMed Central

    Corsetti, Patrícia P.; de Almeida, Leonardo A.; Carvalho, Natália B.; Azevedo, Vasco; Silva, Teane M. A.; Teixeira, Henrique C.; Faria, Ana C.; Oliveira, Sergio C.

    2013-01-01

    IL-10 is a cytokine that regulates the balance between pathogen clearance and immunopathology. Brucella abortus is an intracellular bacterium that causes chronic disease in humans and domestic animals. Here we evaluated the contribution of IL-10 in host immune response and pathology during B. abortus infection. To assess the role of IL-10 in vivo, IL-10 knockout (KO) or 129 Sv/Ev (wild-type) mice were infected with B. abortus and the number of viable bacteria from the spleen was determined at 1, 2, 3, 6 and 14-weeks postinfection. IL-10 KO mice showed reduced bacterial loads in the spleen when compared to wild-type mice during all time points studied. Additionally, at 14-weeks postinfection IL-10 KO mice had totally cleared the infection. This clearance was preceded by an enhanced IFN-γ, TNF-α and IL-17 responses in both the serum and the spleen of IL-10 KO mice. Additionally, dendritic cells from infected IL-10 KO mice produced elevated levels of IL-12 and TNF-α compared to wild-type animals. Histopathology analysis was performed and both KO and wild-type mice developed multifocal granulomas and necrosis in the liver. However, at six-weeks postinfection reduced numbers of granulomas was detected in IL-10 KO mice compared to wild-type animals. This reduced liver pathology at later stage of infection was accompanied by increased numbers of CD4+CD25+foxp3+ T cells and expression of TGF-β in IL-10 KO splenocytes. Taken together, our findings demonstrate that IL-10 modulates the proinflammatory immune response to B. abortus infection and the lack of IL-10 increases resistance to Brucella infection. PMID:24069337

  6. Expression of HIV gp120 protein increases sensitivity to the rewarding properties of methamphetamine in mice.

    PubMed

    Kesby, James P; Hubbard, David T; Markou, Athina; Semenova, Svetlana

    2014-07-01

    Methamphetamine abuse and human immunodeficiency virus (HIV) infection induce neuropathological changes in corticolimbic brain areas involved in reward and cognitive function. Little is known about the combined effects of methamphetamine and HIV infection on cognitive and reward processes. The HIV/gp120 protein induces neurodegeneration in mice, similar to HIV-induced pathology in humans. We investigated the effects of gp120 expression on associative learning, preference for methamphetamine and non-drug reinforcers, and sensitivity to the conditioned rewarding properties of methamphetamine in transgenic (tg) mice expressing HIV/gp120 protein (gp120-tg). gp120-tg mice learned the operant response for food at the same rate as non-tg mice. In the two-bottle choice procedure with restricted access to drugs, gp120-tg mice exhibited greater preference for methamphetamine and saccharin than non-tg mice, whereas preference for quinine was similar between genotypes. Under conditions of unrestricted access to methamphetamine, the mice exhibited a decreased preference for increasing methamphetamine concentrations. However, male gp120-tg mice showed a decreased preference for methamphetamine at lower concentrations than non-tg male mice. gp120-tg mice developed methamphetamine-induced conditioned place preference at lower methamphetamine doses compared with non-tg mice. No differences in methamphetamine pharmacokinetics were found between genotypes. These results indicate that gp120-tg mice exhibit no deficits in associative learning or reward/motivational function for a natural reinforcer. Interestingly, gp120 expression resulted in increased preference for methamphetamine and a highly palatable non-drug reinforcer (saccharin) and increased sensitivity to methamphetamine-induced conditioned reward. These data suggest that HIV-positive individuals may have increased sensitivity to methamphetamine, leading to high methamphetamine abuse potential in this population. PMID

  7. Expression of HIV gp120 protein increases sensitivity to the rewarding properties of methamphetamine in mice

    PubMed Central

    Kesby, James P.; Hubbard, David T.; Markou, Athina; Semenova, Svetlana

    2012-01-01

    Methamphetamine abuse and human immunodeficiency virus (HIV) infection induce neuropathological changes in corticolimbic brain areas involved in reward and cognitive function. Little is known about the combined effects of methamphetamine and HIV infection on cognitive and reward processes. The HIV/gp120 protein induces neurodegeneration in mice, similar to HIV-induced pathology in humans. We investigated the effects of gp120 expression on associative learning, preference for methamphetamine and non-drug reinforcers, and sensitivity to the conditioned rewarding properties of methamphetamine in transgenic (tg) mice expressing HIV/gp120 protein (gp120-tg). gp120-tg mice learned the operant response for food at the same rate as non-tg mice. In the two-bottle choice procedure with restricted access to drugs, gp120-tg mice exhibited greater preference for methamphetamine and saccharin than non-tg mice, whereas preference for quinine was similar between genotypes. Under conditions of unrestricted access to methamphetamine, the mice exhibited a decreased preference for increasing methamphetamine concentrations. However, male gp120-tg mice showed a decreased preference for methamphetamine at lower concentrations than non-tg male mice. gp120-tg mice developed methamphetamine-induced conditioned place preference at lower methamphetamine doses compared with non-tg mice. No differences in methamphetamine pharmacokinetics were found between genotypes. These results indicate that gp120-tg mice exhibit no deficits in associative learning or reward/motivational function for a natural reinforcer. Interestingly, gp120 expression resulted in increased preference for methamphetamine and a highly palatable non-drug reinforcer (saccharin) and increased sensitivity to methamphetamine-induced conditioned reward. These data suggest that HIV-positive individuals may have increased sensitivity to methamphetamine, leading to high methamphetamine abuse potential in this population. PMID

  8. Effect of interleukin-5 receptor-alpha short hairpin RNA-expressing vector on bone marrow eosinophilopoiesis in asthmatic mice.

    PubMed

    Mao, Hui; Wen, Fu-Qiang; Liu, Chun-Tao; Liang, Zong-An; Wang, Zeng-Li; Yin, Kai-Sheng

    2006-01-01

    Bone marrow eosinophilopoiesis induced by interleukin (IL)-5 is a major contributor to eosinophilic airway inflammation in asthma. However,research on the use of IL-5 receptor alpha (IL-5Ralpha) as the target has seldom been reported. This study was undertaken to explore the effects of inhibition of IL-5Ralpha expression through an IL-5Ralpha short hairpin RNA-expressing vector on bone marrow eosinophilopoiesis and airway inflammation in an asthmatic mouse model. An effective plasmid vector was selected that could express short hairpin RNA targeted at IL-5Ralpha (P-IL-5Ralpha). An adenovirus vector (Ad) was then constructed that was inserted in an effective template sequence (Ad-IL-5Ralpha). An animal model of asthma was established by sensitizing and challenging Balb/c mice with ovalbumin. Animals were treated intravenously with Ad-IL-5Ra and changes in bone marrow eosinophilopoiesis and airway inflammation were detected in asthmatic mice. Investigators found that P-IL-5Ra-3 targeted at the sequence of CAG CTG CCT GGT TCG TCT T markedly suppressed IL-5Ralpha expression in B lymphoma cells in vitro. In addition, Ad-IL-5Ralpha could suppress IL-5Ralpha expression in murine bone marrow cells in vitro and in vivo, and it could significantly decrease IL-5-induced eosinophilia in cultured bone marrow cells. Additional studies indicated that intravenously injected Ad-IL-5Ralpha not only selectively reduced the number of eosinophils in the bone marrow, peripheral blood, and bronchoalveolar lavage fluid, it also relieved airway inflammation in asthmatic mice. Results reported here show that blocking of IL-5Ralpha expression through RNA interference can enhance effective treatment of asthma, and that bone marrow can be used as a key targeted organ in the treatment of asthmatic mice. PMID:17276963

  9. Pre- and post-exposure safety and efficacy of attenuated rabies virus vaccines are enhanced by their expression of IFNγ

    SciTech Connect

    Barkhouse, Darryll A.; Faber, Milosz; Hooper, D. Craig

    2015-01-01

    Consistent with evidence of a strong correlation between interferon gamma (IFNγ) production and rabies virus (RABV) clearance from the CNS, we recently demonstrated that engineering a pathogenic RABV to express IFNγ highly attenuates the virus. Reasoning that IFNγ expression by RABV vaccines would enhance their safety and efficacy, we reverse-engineered two proven vaccine vectors, GAS and GASGAS, to express murine IFNγ. Mortality and morbidity were monitored during suckling mice infection, immunize/challenge experiments and mixed intracranial infections. We demonstrate that GASγ and GASγGAS are significantly attenuated in suckling mice compared to the GASGAS vaccine. GASγ better protects mice from lethal DRV4 RABV infection in both pre- and post-exposure experiments compared to GASGAS. Finally, GASγGAS reduces post-infection neurological sequelae, compared to control, during mixed intracranial infection with DRV4. These data show IFNγ expression by a vaccine vector can enhance its safety while increasing its efficacy as pre- and post-exposure treatment. - Highlights: • IFNγ expression improves attenuated rabies virus safety and immunogenicity. • IFNγ expression is safer and more immunogenic than doubling glycoprotein expression. • Co-infection with IFNγ-expressing RABV prevents wild-type rabies virus lethality. • Vaccine safety and efficacy is additive for IFNγ and double glycoprotein expression.

  10. Improved muscle-derived expression of human coagulation factor IX from a skeletal actin/CMV hybrid enhancer/promoter.

    PubMed

    Hagstrom, J N; Couto, L B; Scallan, C; Burton, M; McCleland, M L; Fields, P A; Arruda, V R; Herzog, R W; High, K A

    2000-04-15

    Hemophilia B is caused by the absence of functional coagulation factor IX (F.IX) and represents an important model for treatment of genetic diseases by gene therapy. Recent studies have shown that intramuscular injection of an adeno-associated viral (AAV) vector into mice and hemophilia B dogs results in vector dose-dependent, long-term expression of biologically active F.IX at therapeutic levels. In this study, we demonstrate that levels of expression of approximately 300 ng/mL (6% of normal human F.IX levels) can be reached by intramuscular injection of mice using a 2- to 4-fold lower vector dose (1 x 10(11) vector genomes/mouse, injected into 4 intramuscular sites) than previously described. This was accomplished through the use of an improved expression cassette that uses the cytomegalovirus (CMV) immediate early enhancer/promoter in combination with a 1.2-kilobase portion of human skeletal actin promoter. These results correlated with enhanced levels of F.IX transcript and secreted F.IX protein in transduced murine C2C12 myotubes. Systemic F.IX expression from constructs containing the CMV enhancer/promoter alone was 120 to 200 ng/mL in mice injected with 1 x 10(11) vector genomes. Muscle-specific promoters performed poorly for F.IX transgene expression in vitro and in vivo. However, the incorporation of a sequence from the alpha-skeletal actin promoter containing at least 1 muscle-specific enhancer and 1 enhancer-like element further improved muscle-derived expression of F.IX from a CMV enhancer/promoter-driven expression cassette over previously published results. These findings will allow the design of a clinical protocol for therapeutic levels of F.IX expression with lower vector doses, thus enhancing efficacy and safety of the protocol. (Blood. 2000;95:2536-2542) PMID:10753832

  11. CCAAT/enhancer binding protein {beta} deletion increases mitochondrial function and protects mice from LXR-induced hepatic steatosis

    SciTech Connect

    Rahman, Shaikh M.; Choudhury, Mahua; Janssen, Rachel C.; Baquero, Karalee C.; Miyazaki, Makoto; Friedman, Jacob E.

    2013-01-04

    Highlights: Black-Right-Pointing-Pointer LXR agonist activation increases liver TG accumulation by increasing lipogenesis. Black-Right-Pointing-Pointer C/EBP{beta}{sup -/-} mouse prevents LXR activation-mediated induction of hepatic lipogenesis. Black-Right-Pointing-Pointer C/EBP{beta} deletion increases mitochondrial transport chain function. Black-Right-Pointing-Pointer Beneficial effects of LXR activation on liver cholesterol metabolism did not change. Black-Right-Pointing-Pointer C/EBP{beta} inhibition might have important therapeutic potential. -- Abstract: Drugs designed specifically to activate liver X receptors (LXRs) have beneficial effects on lowering cholesterol metabolism and inflammation but unfortunately lead to severe hepatic steatosis. The transcription factor CCAAT/enhancer binding protein beta (C/EBP{beta}) is an important regulator of liver gene expression but little is known about its involvement in LXR-based steatosis and cholesterol metabolism. The present study investigated the role of C/EBP{beta} expression in LXR agonist (T0901317)-mediated alteration of hepatic triglyceride (TG) and lipogenesis in mice. C/EBP{beta} deletion in mice prevented LXR agonist-mediated induction of lipogenic gene expression in liver in conjunction with significant reduction of liver TG accumulation. Surprisingly, C/EBP{beta}{sup -/-} mice showed a major increase in liver mitochondrial electron chain function compared to WT mice. Furthermore, LXR activation in C/EBP{beta}{sup -/-} mice increased the expression of liver ATP-binding cassette transporter ABCG1, a gene implicated in cholesterol efflux and reducing blood levels of total and LDL-cholesterol. Together, these findings establish a central role for C/EBP{beta} in the LXR-mediated steatosis and mitochondrial function, without impairing the influence of LXR activation on lowering LDL and increasing HDL-cholesterol. Inactivation of C/EBP{beta} might therefore be an important therapeutic strategy to prevent LXR

  12. Transcriptomic plasticity in brown adipose tissue contributes to an enhanced capacity for nonshivering thermogenesis in deer mice.

    PubMed

    Velotta, Jonathan P; Jones, Jennifer; Wolf, Cole J; Cheviron, Zachary A

    2016-06-01

    For small mammals living at high altitude, aerobic heat generation (thermogenesis) is essential for survival during prolonged periods of cold, but is severely impaired under conditions of hypobaric hypoxia. Recent studies in deer mice (Peromyscus maniculatus) reveal adaptive enhancement of thermogenesis in high- compared to low-altitude populations under hypoxic cold stress, an enhancement that is attributable to modifications in the aerobic metabolism of muscles used in shivering. However, because small mammals rely heavily on nonshivering mechanisms for cold acclimatization, we tested for evidence of adaptive divergence in nonshivering thermogenesis (NST) under hypoxia. To do so, we measured NST and characterized transcriptional profiles of brown adipose tissue (BAT) in high- and low-altitude deer mice that were (i) wild-caught and acclimatized to their native altitude, and (ii) born and reared under common garden conditions at low elevation. We found that NST performance under hypoxia is enhanced in wild-caught, high-altitude deer mice, a difference that is associated with increased expression of coregulated genes that influence several physiological traits. These traits include vascularization and O2 supply to BAT, brown adipocyte proliferation and the uncoupling of oxidative phosphorylation from ATP synthesis in the generation of heat. Our results suggest that acclimatization to hypoxic cold stress is facilitated by enhancement of nonshivering heat production, which is driven by regulatory plasticity in a suite of genes that influence intersecting physiological pathways. PMID:27126783

  13. Transgenic mice expressing an intracellular fluorescent fusion of angiotensin II demonstrate renal thrombotic microangiopathy and elevated blood pressure

    PubMed Central

    Redding, K. M.; Chen, B. L.; Singh, A.; Re, R. N.; Navar, L. G.; Seth, D. M.; Sigmund, C. D.; Tang, W. W.

    2010-01-01

    We have generated transgenic mice that express angiotensin II (ANG II) fused downstream of enhanced cyan fluorescent protein, expression of which is regulated by the mouse metallothionein promoter. The fusion protein, which lacks a secretory signal, is retained intracellularly. In the present study, RT-PCR, immunoblot analyses, whole-animal fluorescent imaging, and fluorescent microscopy of murine embryonic fibroblasts confirm expression of the fusion protein in vivo and in vitro. The transgene is expressed in all tissues tested (including brain, heart, kidney, liver, lung, and testes), and radioimmunoassay of plasma samples obtained from transgenic mice indicate no increase in circulating ANG II over wild-type levels, consistent with intracellular retention of the transgene product. Kidneys from transgenic and corresponding wild-type littermates were histologically evaluated, and abnormalities in transgenic mice consistent with thrombotic microangiopathy were observed; microthrombosis was frequently observed within the glomerular capillaries and small vessels. In addition, systolic and diastolic blood pressures, measured by telemetry (n = 8 for each group), were significantly higher in transgenic mice compared with wild-type littermates. Blood pressure of line A male transgenic mice was 125 ± 1.7 over 97 ± 1.6 compared with 109 ± 1.7 over 83 ± 1.4 mmHg in wild-type littermates (systolic over diastolic). In summary, overexpression of an intracellular fluorescent fusion protein of ANG II correlates with elevated blood pressure and kidney pathology. This transgenic model may be useful to further explore the intracellular renin-angiotensin system and its implication in abnormal kidney function and hypertension. PMID:20363893

  14. FOXN1 recombinant protein enhances T-cell regeneration after hematopoietic stem cell transplantation in mice.

    PubMed

    Song, Yinhong; Su, Min; Zhu, Jing; Di, Wen; Liu, Yalan; Hu, Rong; Rood, Debra; Lai, Laijun

    2016-06-01

    A prolonged period of T-cell recovery is the major challenge in hematopoietic stem cell transplantation (HSCT). Thymic epithelial cells (TECs) are the major component of the thymic microenvironment for T-cell generation. However, TECs undergo degeneration over time. FOXN1 plays a critical role in TEC development and is required to maintain adult TECs for thymopoiesis. To investigate the potential application of FOXN1, we have cloned and expressed recombinant FOXN1 protein (rFOXN1) that was fused with cell-penetrating peptides. We show here that the rFOXN1 protein can translocate from the cell surface into the cytoplasm and nucleus. Administration of rFOXN1 into both congenic and allogeneic HSCT recipient mice increased the number of TECs, resulting in enhanced thymopoiesis that led to an increased number of functional T cells in the periphery. The increased number of TECs is due to the enhanced survival and proliferation of TECs. Our results suggest that rFOXN1 has the potential to be used in enhancing T-cell regeneration in patients following HSCT. PMID:27125859

  15. Humanized θ-Defensins (Retrocyclins) Enhance Macrophage Performance and Protect Mice from Experimental Anthrax Infections▿

    PubMed Central

    Welkos, S.; Cote, C. K.; Hahn, U.; Shastak, O.; Jedermann, J.; Bozue, J.; Jung, G.; Ruchala, P.; Pratikhya, P.; Tang, T.; Lehrer, R. I.; Beyer, W.

    2011-01-01

    Retrocyclins are humanized versions of the θ-defensin peptides expressed by the leukocytes of several nonhuman primates. Previous studies, performed in serum-free media, determined that retrocyclins 1 (RC1) and RC2 could prevent successful germination of Bacillus anthracis spores, kill vegetative B. anthracis cells, and inactivate anthrax lethal factor. We now report that retrocyclins are extensively bound by components of native mouse, human, and fetal calf sera, that heat-inactivated sera show greatly enhanced retrocyclin binding, and that native and (especially) heat-inactivated sera greatly reduce the direct activities of retrocyclins against spores and vegetative cells of B. anthracis. Nevertheless, we also found that retrocyclins protected mice challenged in vivo by subcutaneous, intraperitoneal, or intranasal instillation of B. anthracis spores. Retrocyclin 1 bound extensively to B. anthracis spores and enhanced their phagocytosis and killing by murine RAW264.7 cells. Based on the assumption that spore-bound RC1 enters phagosomes by “piggyback phagocytosis,” model calculations showed that the intraphagosomal concentration of RC1 would greatly exceed its extracellular concentration. Murine alveolar macrophages took up fluorescently labeled retrocyclin, suggesting that macrophages may also acquire extracellular RC1 directly. Overall, these data demonstrate that retrocyclins are effective in vivo against experimental murine anthrax infections and suggest that enhanced macrophage function contributes to this property. PMID:21768520

  16. Humanized theta-defensins (retrocyclins) enhance macrophage performance and protect mice from experimental anthrax infections.

    PubMed

    Welkos, S; Cote, C K; Hahn, U; Shastak, O; Jedermann, J; Bozue, J; Jung, G; Ruchala, P; Pratikhya, P; Tang, T; Lehrer, R I; Beyer, W

    2011-09-01

    Retrocyclins are humanized versions of the -defensin peptides expressed by the leukocytes of several nonhuman primates. Previous studies, performed in serum-free media, determined that retrocyclins 1 (RC1) and RC2 could prevent successful germination of Bacillus anthracis spores, kill vegetative B. anthracis cells, and inactivate anthrax lethal factor. We now report that retrocyclins are extensively bound by components of native mouse, human, and fetal calf sera, that heat-inactivated sera show greatly enhanced retrocyclin binding, and that native and (especially) heat-inactivated sera greatly reduce the direct activities of retrocyclins against spores and vegetative cells of B. anthracis. Nevertheless, we also found that retrocyclins protected mice challenged in vivo by subcutaneous, intraperitoneal, or intranasal instillation of B. anthracis spores. Retrocyclin 1 bound extensively to B. anthracis spores and enhanced their phagocytosis and killing by murine RAW264.7 cells. Based on the assumption that spore-bound RC1 enters phagosomes by "piggyback phagocytosis," model calculations showed that the intraphagosomal concentration of RC1 would greatly exceed its extracellular concentration. Murine alveolar macrophages took up fluorescently labeled retrocyclin, suggesting that macrophages may also acquire extracellular RC1 directly. Overall, these data demonstrate that retrocyclins are effective in vivo against experimental murine anthrax infections and suggest that enhanced macrophage function contributes to this property. PMID:21768520

  17. Over-Expression of CD200 Protects Mice from Dextran Sodium Sulfate Induced Colitis

    PubMed Central

    Chen, Zhiqi; Yu, Kai; Zhu, Fang; Gorczynski, Reginald

    2016-01-01

    Background and aim CD200:CD200 receptor (CD200R) interactions lead to potent immunosuppression and inhibition of autoimmune inflammation. We investigated the effect of "knockout"of CD200 or CD200R, or over-expression of CD200, on susceptibility to dextran sodium sulfate (DSS)—induced colitis, a mouse model of inflammatory bowel disease (IBD). Methods Acute or chronic colitis was induced by administration of dextran sodium sulfate (DSS) in four groups of age-matched C57BL/6 female mice: (1) CD200-transgenic mice (CD200tg); (2) wild-type (WT) mice; (3) CD200 receptor 1-deficient (CD200R1KO) mice; and (4) CD200-deficient (CD200KO) mice. The extent of colitis was determined using a histological scoring system. Colon tissues were collected for quantitative RT-PCR and Immunohistochemical staining. Supernatants from colonic explant cultures and mononuclear cells isolated from colonic tissue were used for ELISA. Results CD200KO and CD200R1KO mice showed greater sensitivity to acute colitis than WT mice, with accelerated loss of body weight, significantly higher histological scores, more severe infiltration of macrophages, neutrophils and CD3+ cells, and greater expression of macrophage-derived inflammatory cytokines, whose production was inhibited in vitro (in WT/CD200KO mouse cells) by CD200. In contrast, CD200tg mice showed less sensitivity to DSS compared with WT mice, with attenuation of all of the features seen in other groups. In a chronic colitis model, greater infiltration of Foxp3+ regulatory T (Treg) cells was seen in the colon of CD200tg mice compared to WT mice, and anti-CD25 mAb given to these mice attenuated protection. Conclusions The CD200:CD200R axis plays an immunoregulatory role in control of DSS induced colitis in mice. PMID:26841120

  18. Over-Expression of Porcine Myostatin Missense Mutant Leads to A Gender Difference in Skeletal Muscle Growth between Transgenic Male and Female Mice.

    PubMed

    Ma, Dezun; Gao, Pengfei; Qian, Lili; Wang, Qingqing; Cai, Chunbo; Jiang, Shengwang; Xiao, Gaojun; Cui, Wentao

    2015-01-01

    Myostatin, a transforming growth factor-β family member, is a negative regulator of skeletal muscle development and growth. Piedmontese cattle breeds have a missense mutation, which results in a cysteine to tyrosine substitution in the mature myostatin protein (C313Y). This loss-of-function mutation in myostatin results in a double-muscled phenotype in cattle. Myostatin propeptide is an inhibitor of myostatin activity and is considered a potential agent to stimulate muscle growth in livestock. In this study, we generated transgenic mice overexpressing porcine myostatin missense mutant (pmMS), C313Y, and wild-type porcine myostatin propeptide (ppMS), respectively, to examine their effects on muscle growth in mice. Enhanced muscle growth was observed in both pmMS and ppMS transgenic female mice and also in ppMS transgenic male mice. However, there was no enhanced muscle growth observed in pmMS transgenic male mice. To explore why there is such a big difference in muscle growth between pmMS and ppMS transgenic male mice, the expression level of androgen receptor (AR) mutant AR45 was measured by Western blot. Results indicated that AR45 expression significantly increased in pmMS transgenic male mice while it decreased dramatically in ppMS transgenic male mice. Our data demonstrate that both pmMS and ppMS act as myostatin inhibitors in the regulation of muscle growth, but the effect of pmMS in male mice is reversed by an increased AR45 expression. These results provide useful insight and basic theory to future studies on improving pork quality by genetically manipulating myostatin expression or by regulating myostatin activity. PMID:26305245

  19. Over-Expression of Porcine Myostatin Missense Mutant Leads to A Gender Difference in Skeletal Muscle Growth between Transgenic Male and Female Mice

    PubMed Central

    Ma, Dezun; Gao, Pengfei; Qian, Lili; Wang, Qingqing; Cai, Chunbo; Jiang, Shengwang; Xiao, Gaojun; Cui, Wentao

    2015-01-01

    Myostatin, a transforming growth factor-β family member, is a negative regulator of skeletal muscle development and growth. Piedmontese cattle breeds have a missense mutation, which results in a cysteine to tyrosine substitution in the mature myostatin protein (C313Y). This loss-of-function mutation in myostatin results in a double-muscled phenotype in cattle. Myostatin propeptide is an inhibitor of myostatin activity and is considered a potential agent to stimulate muscle growth in livestock. In this study, we generated transgenic mice overexpressing porcine myostatin missense mutant (pmMS), C313Y, and wild-type porcine myostatin propeptide (ppMS), respectively, to examine their effects on muscle growth in mice. Enhanced muscle growth was observed in both pmMS and ppMS transgenic female mice and also in ppMS transgenic male mice. However, there was no enhanced muscle growth observed in pmMS transgenic male mice. To explore why there is such a big difference in muscle growth between pmMS and ppMS transgenic male mice, the expression level of androgen receptor (AR) mutant AR45 was measured by Western blot. Results indicated that AR45 expression significantly increased in pmMS transgenic male mice while it decreased dramatically in ppMS transgenic male mice. Our data demonstrate that both pmMS and ppMS act as myostatin inhibitors in the regulation of muscle growth, but the effect of pmMS in male mice is reversed by an increased AR45 expression. These results provide useful insight and basic theory to future studies on improving pork quality by genetically manipulating myostatin expression or by regulating myostatin activity. PMID:26305245

  20. Smaller Dendritic Spines, Weaker Synaptic Transmission, but Enhanced Spatial Learning in Mice Lacking Shank1

    PubMed Central

    Hung, Albert Y.; Futai, Kensuke; Sala, Carlo; Valtschanoff, Juli G.; Ryu, Jubin; Woodworth, Mollie A.; Kidd, Fleur L.; Sung, Clifford C.; Miyakawa, Tsuyoshi; Bear, Mark F.; Weinberg, Richard J.; Sheng, Morgan

    2009-01-01

    Experience-dependent changes in the structure of dendritic spines may contribute to learning and memory. Encoded by three genes, the Shank family of postsynaptic scaffold proteins are abundant and enriched in the postsynaptic density (PSD) of central excitatory synapses. When expressed in cultured hippocampal neurons, Shank promotes the maturation and enlargement of dendritic spines. Recently, Shank3 has been genetically implicated in human autism, suggesting an important role for Shank proteins in normal cognitive development. Here, we report the phenotype of Shank1 knock-out mice. Shank1 mutants showed altered PSD protein composition; reduced size of dendritic spines; smaller, thinner PSDs; and weaker basal synaptic transmission. Standard measures of synaptic plasticity were normal. Behaviorally, they had increased anxiety-related behavior and impaired contextual fear memory. Remarkably, Shank1-deficient mice displayed enhanced performance in a spatial learning task; however, their long-term memory retention in this task was impaired. These results affirm the importance of Shank1 for synapse structure and function in vivo, and they highlight a differential role for Shank1 in specific cognitive processes, a feature that may be relevant to human autism spectrum disorders. PMID:18272690

  1. Ontogeny of Angiotensin Type 2 and Type 1 Receptor Expression in Mice

    PubMed Central

    Gao, Juan; Chao, Jie; Parbhu, Karma-Jaya K.; Yu, Li; Xiao, Liang; Gao, Fei; Gao, Lie

    2012-01-01

    In the current experiment, we determined AT2R and AT1R protein expression by Western blot analysis in developing normal mice. The results indicate that, (1) in all detected brain regions and in the spinal cord, adult mice exhibited significantly higher AT2R expression and lower AT1R expression in total protein extracts compared to fetuses and neonates; (2) other major organs, including heart, lung, liver, and kidney, exhibited the same expression pattern as the brain and spinal cord; (3) reciprocal changes in AT2R and AT1R expression were found in the total protein extracts from the brainstems of mice from one day prenatal to six weeks of age. There was a negative correlation between AT2R and AT1R protein expression; (4) in both membrane and cytosolic fractions from the brainstem, adult mice exhibited higher AT2R and lower AT1R expression than did fetuses and neonates; (5) in the brainstem, there were no significant differences in AT2R and AT1R mRNA levels among fetal, neonatal, and adult mice. The above results reconfirmed our previous finding in rats that adult animals have higher AT2R and lower AT1R expression compared to fetuses and neonates. These data imply an involvement of AT1R in fetal development and of AT2R in adult function. PMID:22526820

  2. Pain-related behaviors and neurochemical alterations in mice expressing sickle hemoglobin: modulation by cannabinoids

    PubMed Central

    Kohli, Divyanshoo R.; Li, Yunfang; Khasabov, Sergey G.; Gupta, Pankaj; Kehl, Lois J.; Ericson, Marna E.; Nguyen, Julia; Gupta, Vinita; Hebbel, Robert P.; Simone, Donald A.

    2010-01-01

    Sickle cell disease causes severe pain. We examined pain-related behaviors, correlative neurochemical changes, and analgesic effects of morphine and cannabinoids in transgenic mice expressing human sickle hemoglobin (HbS). Paw withdrawal threshold and withdrawal latency (to mechanical and thermal stimuli, respectively) and grip force were lower in homozygous and hemizygous Berkley mice (BERK and hBERK1, respectively) compared with control mice expressing human hemoglobin A (HbA-BERK), indicating deep/musculoskeletal and cutaneous hyperalgesia. Peripheral nerves and blood vessels were structurally altered in BERK and hBERK1 skin, with decreased expression of μ opioid receptor and increased calcitonin gene-related peptide and substance P immunoreactivity. Activators of neuropathic and inflammatory pain (p38 mitogen-activated protein kinase, STAT3, and mitogen-activated protein kinase/extracellular signal-regulated kinase) showed increased phosphorylation, with accompanying increase in COX-2, interleukin-6, and Toll-like receptor 4 in the spinal cord of hBERK1 compared with HbA-BERK. These neurochemical changes in the periphery and spinal cord may contribute to hyperalgesia in mice expressing HbS. In BERK and hBERK1, hyperalgesia was markedly attenuated by morphine and cannabinoid receptor agonist CP 55940. We show that mice expressing HbS exhibit characteristics of pain observed in sickle cell disease patients, and neurochemical changes suggestive of nociceptor and glial activation. Importantly, cannabinoids attenuate pain in mice expressing HbS. PMID:20304807

  3. Down-Regulation of Brush Border Efflux Transporter Expression in the Kidneys of Pregnant Mice

    PubMed Central

    Yacovino, Lindsay L.; Gibson, Christopher J.

    2013-01-01

    Pregnancy increases the urinary excretion of chemicals in women and rodents. It is unknown whether the enhanced clearance of drugs during pregnancy involves changes in the expression of transporters that mediate chemical secretion and reabsorption. The purpose of this study was to quantify the mRNA and protein expression of efflux transporters in kidneys from virgin and pregnant mice on gestational days 7, 11, 14, and 17 and postnatal days 1, 15, and 30 with use of quantitative polymerase chain reaction, Western blot, and immunofluorescence. Multidrug resistance protein (Mdr) 1b mRNA, multidrug resistance-associated protein (Mrp) 4 mRNA, and protein levels decreased significantly by 25–75% throughout pregnancy and lactation. Similarly, Mrp2 and multidrug and toxin extrusion transporter (Mate) 1 mRNA expression were down-regulated 20–40% during mid to late gestation but returned to control levels by postnatal day 15. In contrast, Mrp3 mRNA and protein increased 225% and 31%, respectively, at gestational day 14. Coordinated down-regulation of brush border transporters Mate1, Mrp2, and Mrp4 and up-regulation of the basolateral Mrp3 transporter would reduce chemical secretion into urine. PMID:22896729

  4. Aminoaciduria and altered renal expression of luminal amino acid transporters in mice lacking novel gene collectrin.

    PubMed

    Malakauskas, Sandra M; Quan, Hui; Fields, Timothy A; McCall, Shannon J; Yu, Ming-Jiun; Kourany, Wissam M; Frey, Campbell W; Le, Thu H

    2007-02-01

    Defects in renal proximal tubule transport manifest in a number of human diseases. Although variable in clinical presentation, disorders such as Hartnup disease, Dent's disease, and Fanconi syndrome are characterized by wasting of solutes commonly recovered by the proximal tubule. One common feature of these disorders is aminoaciduria. There are distinct classes of amino acid transporters located in the apical and basal membranes of the proximal tubules that reabsorb >95% of filtered amino acids, yet few details are known about their regulation. We present our physiological characterization of a mouse line with targeted deletion of the gene collectrin that is highly expressed in the kidney. Collectrin-deficient mice display a reduced urinary concentrating capacity due to enhanced solute clearance resulting from profound aminoaciduria. The aminoaciduria is generalized, characterized by loss of nearly every amino acid, and results in marked crystalluria. Furthermore, in the kidney, collectrin-deficient mice have decreased plasma membrane populations of amino acid transporter subtypes B(0)AT1, rBAT, and b(0,+)AT, as well as altered cellular distribution of EAAC1. Our data suggest that collectrin is a novel mediator of renal amino acid transport and may provide further insight into the pathogenesis of a number of human disease correlates. PMID:16985211

  5. Delayed cutaneous wound closure in HO-2 deficient mice despite normal HO-1 expression

    PubMed Central

    Lundvig, Ditte M S; Scharstuhl, Alwin; Cremers, Niels A J; Pennings, Sebastiaan W C; te Paske, Jeroen; van Rheden, René; van Run-van Breda, Coby; Regan, Raymond F; Russel, Frans G M; Carels, Carine E; Maltha, Jaap C; Wagener, Frank A D T G

    2014-01-01

    Impaired wound healing can lead to scarring, and aesthetical and functional problems. The cytoprotective haem oxygenase (HO) enzymes degrade haem into iron, biliverdin and carbon monoxide. HO-1 deficient mice suffer from chronic inflammatory stress and delayed cutaneous wound healing, while corneal wound healing in HO-2 deficient mice is impaired with exorbitant inflammation and absence of HO-1 expression. This study addresses the role of HO-2 in cutaneous excisional wound healing using HO-2 knockout (KO) mice. Here, we show that HO-2 deficiency also delays cutaneous wound closure compared to WT controls. In addition, we detected reduced collagen deposition and vessel density in the wounds of HO-2 KO mice compared to WT controls. Surprisingly, wound closure in HO-2 KO mice was accompanied by an inflammatory response comparable to WT mice. HO-1 induction in HO-2 deficient skin was also similar to WT controls and may explain this protection against exaggerated cutaneous inflammation but not the delayed wound closure. Proliferation and myofibroblast differentiation were similar in both two genotypes. Next, we screened for candidate genes to explain the observed delayed wound closure, and detected delayed gene and protein expression profiles of the chemokine (C-X-C) ligand-11 (CXCL-11) in wounds of HO-2 KO mice. Abnormal regulation of CXCL-11 has been linked to delayed wound healing and disturbed angiogenesis. However, whether aberrant CXCL-11 expression in HO-2 KO mice is caused by or is causing delayed wound healing needs to be further investigated. PMID:25224969

  6. Sex-linked behavioural differences in mice expressing a human insulin transgene in the medial habenula.

    PubMed

    Douhet, P; Bertaina, V; Durkin, T; Calas, A; Destrade, C

    1997-12-01

    We previously reported that a human insulin transgene was specifically expressed in the medial habenula of the adult mouse brain, and that this expression was ascribed to the delta-168 transgene. The present study analyses the possible behavioural consequences of this insulin transgene expression using measures of food intake, spontaneous activity, emotional reactivity, learning and extinction performance of an operant task. The delta-168 transgenic mice did not differ from the C57BL/6 control mice as concerns food intake, behaviour in the open field, or emotional response in an elevated plus maze. On the other hand, measures of locomotor activity in a circular corridor revealed a significantly faster decline of spontaneous locomotor activity in male as compared to female delta-168 transgenic mice. Moreover, as compared to female transgenic mice, male transgenic mice exhibited a deficit in the rate of acquisition and an acceleration of the rate of extinction of a bar press response in a Skinner box. In contrast, the behaviour of female transgenic mice did not differ from either male or female C57BL/6 control mice. The results of the present study demonstrate that the behavioural modifications observed in delta-168 transgenic mice are sex-linked and suggest that these behavioural differences result from changes in the interaction (interface) between motivational and motor mechanisms mediated via the striato-habenulo-mesencephalic system. PMID:9475633

  7. Modulations of cytochrome P450 expression in diabetic mice by berberine.

    PubMed

    Chatuphonprasert, Waranya; Nemoto, Nobuo; Sakuma, Tsutomu; Jarukamjorn, Kanokwan

    2012-03-01

    Berberine, an isoquinoline alkaloid isolated from medicinal plants such as Berberis aristata, Coptis chinesis, Coptis japonica, Coscinium fenestatun, and Hydrastis Canadensis, is widely used in Asian countries for the treatment of diabetes, hypertension, and hypercholesterolemia. Interaction between berberine and the cytochrome P450 enzymes (CYPs) has been extensively reported, but there are only a few reports of this interaction in the diabetic state. In this study, the effect of berberine on the mRNA of the CYPs in primary mouse hepatocytes and in streptozotocin (STZ)-induced diabetic mice was investigated. In primary mouse hepatocytes, berberine suppressed the induction of Cyp1a1, Cyp1a2, Cyp2e1, Cyp3a11, Cyp4a10, and Cyp4a14 mRNA expression by their prototypical inducers in a concentration-dependent fashion. However, berberine treatment alone increased the expression of Cyp2b9 and Cyp2b10 mRNA. In vivo, berberine showed the same hypoglycemic activity as metformin, an established hypoglycemic drug. The hepatic mRNA levels of Cyp1a1, Cyp2b9, Cyp2b10, Cyp3a11, Cyp4a10, and Cyp4a14 were increased in STZ-induced diabetic mice. Interestingly, berberine itself suppressed the expression of Cyp2e1, an adverse hepatic event-associated enzyme, while the expression of Cyp3a11, Cyp4a10, and Cyp4a14 were restored to normal levels by berberine. In conclusion, berberine has the potential to modify the expression of CYPs by either suppression or enhancement of CYPs' levels. Consumption of berberine as an anti-hyperglycemic compound by diabetic patients might provide an extra benefit due to its potential to restore the expression of Cyp2e1, Cyp3a, and Cyp4a to normal levels. However, an herb-drug interaction might be of concern since any berberine-containing product would definitely cause pronounced interactions based on CYP3A4 inhibition. PMID:22342832

  8. Chemokine gene expression in the brains of mice with lymphocytic choriomeningitis.

    PubMed Central

    Asensio, V C; Campbell, I L

    1997-01-01

    Chemokines are pivotal in the trafficking of leukocytes. In the present study, we examined the expression of multiple chemokine genes during the course of lymphocytic choriomeningitis (LCM) in mice. In noninfected mice, no detectable chemokine gene expression was found in the brain; however, by day 3 postinfection, the induction of a number of chemokine mRNAs was observed as follows (in order from the greatest to the least): cytokine responsive gene-2 or interferon-inducible 10-kDa protein (Crg-2/IP-10), RANTES, monocyte chemotactic protein-1 (MCP-1), macrophage inflammatory protein-1 (MIP-1beta), and MCP-3. At day 6 postinfection, the expression of these chemokine mRNAs was increased, and low expression of lymphotactin, C10, MIP-2, and MIP-1alpha mRNAs was detectable. Transcript for T-cell activation-3 was not detectable in the brain at any time following LCM virus (LCMV) infection. With some exceptions, a pattern of chemokine gene expression similar to that in the brain was observed in the peripheral organs of LCMV-infected mice. Mice that lacked expression of gamma interferon developed LCM and had a qualitatively similar but quantitatively reduced cerebral chemokine gene expression profile. In contrast, little or no chemokine gene expression was detectable in the brains of LCMV-infected athymic mice which did not develop LCM. Expression of Crg-2/IP-10 RNA was localized to predominantly resident cells of the central nervous system (CNS) and overlapped with sites of viral infection and immune cell infiltration. These findings demonstrate the expression of a number of chemokine genes in the brains of mice infected with LCMV. The pattern of chemokine gene expression in LCM may profoundly influence the characteristic phenotype and response of leukocytes in the brain and contribute to the immunopathogenesis of this fatal CNS infection. PMID:9311871

  9. 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

  10. Human α-Defensin Expression Is Not Dependent on CCAAT/Enhancer Binding Protein-ε in a Murine Model

    PubMed Central

    Glenthøj, Andreas; Dahl, Sara; Larsen, Maria T.; Cowland, Jack B.; Borregaard, Niels

    2014-01-01

    Specific granule deficiency (SGD) is a rare congenital disorder characterized by recurrent infections. The disease is caused by inactivating mutations of the CCAAT/enhancer binding protein-ε (C/EBP-ε) gene. As a consequence, specific and gelatinase granules lack most matrix proteins. Furthermore, azurophil granules contain diminished amounts of their most abundant proteins, α-defensins, also known as human neutrophil peptides (HNPs). In accordance with this, in vitro models have demonstrated induction of HNPs by C/EBP-ε. Since mice do not express myeloid defensins, they cannot per se be used to characterize the role of C/EBP-ε in controlling HNP expression in vivo. We therefore crossed a transgenic HNP-1-expressing mouse with the Cebpe-/- mouse to study the in vivo significance of C/EBP-ε for HNP-1 transcription and expression. Surprisingly, neither expression nor processing of HNP-1 was affected by lack of C/EBP-ε in these mice. Transduction of C/EBP-ε into primary bone marrow cells from HNP-1 mice induced some HNP-1 expression, but not to levels comparable to expression human cells. Taken together, our data infer that the HNP-1 of the transgenic mouse does not show an expression pattern equivalent to endogenous secondary granule proteins. This limits the use of these transgenic mice as a model for human conditions. PMID:24658030

  11. Enhanced ethanol catabolism in orphan nuclear receptor SHP-null mice.

    PubMed

    Park, Jung Eun; Lee, Mikang; Mifflin, Ryan; Lee, Yoon Kwang

    2016-05-15

    Deficiency of the orphan nuclear hormone receptor small heterodimer partner (SHP, NR0B2) protects mice from diet-induced hepatic steatosis, in part, via repression of peroxisome proliferator-activated receptor (PPAR)-γ2 (Pparg2) gene expression. Alcoholic fatty liver diseases (AFLD) share many common pathophysiological features with non-AFLD. To study the role of SHP and PPARγ2 in AFLD, we used a strategy of chronic ethanol feeding plus a single binge ethanol feeding to challenge wild-type (WT) and SHP-null (SHP(-/-)) mice with ethanol. The ethanol feeding induced liver fat accumulation and mRNA expression of hepatic Pparg2 in WT mice, which suggests that a high level of PPARγ2 is a common driving force for fat accumulation induced by ethanol or a high-fat diet. Interestingly, ethanol-fed SHP(-/-) mice displayed hepatic fat accumulation similar to that of ethanol-fed WT mice, even though their Pparg2 expression level remained lower. Mortality of SHP(-/-) mice after ethanol binge feeding was significantly reduced and their acetaldehyde dehydrogenase (Aldh2) mRNA level was higher than that of their WT counterparts. After an intoxicating dose of ethanol, SHP(-/-) mice exhibited faster blood ethanol clearance and earlier wake-up time than WT mice. Higher blood acetate, the end product of ethanol metabolism, and lower acetaldehyde levels were evident in the ethanol-challenged SHP(-/-) than WT mice. Ethanol-induced inflammatory responses and lipid peroxidation were also lower in SHP(-/-) mice. The current data show faster ethanol catabolism and extra fat storage through conversion of acetate to acetyl-CoA before its release into the circulation in this ethanol-feeding model in SHP(-/-) mice. PMID:26968209

  12. Enhanced efficacy of enzyme replacement therapy in Pompe disease through mannose-6-phosphate receptor expression in skeletal muscle.

    PubMed

    Koeberl, Dwight D; Luo, Xiaoyan; Sun, Baodong; McVie-Wylie, Alison; Dai, Jian; Li, Songtao; Banugaria, Suhrad G; Chen, Y-T; Bali, Deeksha S

    2011-06-01

    Enzyme replacement therapy (ERT) with acid α-glucosidase has become available for Pompe disease; however, the response of skeletal muscle, as opposed to the heart, has been attenuated. The poor response of skeletal muscle has been attributed to the low abundance of the cation-independent mannose-6-phosphate receptor (CI-MPR) in skeletal muscle compared to heart. To further understand the role of CI-MPR in Pompe disease, muscle-specific CI-MPR conditional knockout (KO) mice were crossed with GAA-KO (Pompe disease) mice. We evaluated the impact of CI-MPR-mediated uptake of GAA by evaluating ERT in CI-MPR-KO/GAA-KO (double KO) mice. The essential role of CI-MPR was emphasized by the lack of efficacy of ERT as demonstrated by markedly reduced biochemical correction of GAA deficiency and of glycogen accumulations in double KO mice, in comparison with the administration of the same therapeutic doses in GAA-KO mice. Clenbuterol, a selective β(2)-agonist, enhanced the CI-MPR expression in skeletal tissue and also increased efficacy from GAA therapy, thereby confirming the key role of CI-MPR with regard to enzyme replacement therapy in Pompe disease. Biochemical correction improved in both muscle and non-muscle tissues, indicating that therapy could be similarly enhanced in other lysosomal storage disorders. In summary, enhanced CI-MPR expression might improve the efficacy of enzyme replacement therapy in Pompe disease through enhancing receptor-mediated uptake of GAA. PMID:21397538

  13. Enhanced Efficacy of Enzyme Replacement Therapy in Pompe Disease Through Mannose-6-Phosphate Receptor Expression in Skeletal Muscle

    PubMed Central

    Koeberl, Dwight D.; Luo, Xiaoyan; Sun, Baodong; McVie-Wylie, Alison; Dai, Jian; Li, Songtao; Banugaria, Suhrad G.; Chen, Y-T; Bali, Deeksha S.

    2011-01-01

    Enzyme replacement therapy (ERT) with acid α-glucosidase has become available for Pompe disease; however, the response of skeletal muscle, as opposed to the heart, has been attenuated. The poor response of skeletal muscle has been attributed to the low abundance of the cation-independent mannose-6-phosphate receptor (CI-MPR) in skeletal muscle compared to heart. To further understand the role of CI-MPR in Pompe disease, muscle-specific CI-MPR conditional knockout (KO) mice were crossed with GAA-KO (Pompe disease) mice. We evaluated the impact of CI-MPR-mediated uptake of GAA by evaluating ERT in CI-MPR-KO/GAA-KO (double KO) mice. The essential role of CI-MPR was emphasized by the lack of efficacy of ERT as demonstrated by markedly reduced biochemical correction of GAA deficiency and of glycogen accumulations in double KO mice, in comparison with administration of the same therapeutic doses in GAA-KO mice. Clenbuterol, a selective β2-agonist, enhanced CI-MPR expression in skeletal tissue and also increased efficacy from GAA therapy, thereby confirming the key role of CI-MPR with regard to enzyme replacement therapy in Pompe disease. Biochemical correction improved in both muscle and non-muscle tissues, indicating that therapy could be similarly enhanced in other lysosomal storage disorders. In summary, enhanced CI-MPR expression might improve the efficacy of enzyme replacement therapy in Pompe disease through enhancing receptor-mediated uptake of GAA. PMID:21397538

  14. Fas Receptor-deficient lpr Mice are protected against Acetaminophen Hepatotoxicity due to Higher Glutathione Synthesis and Enhanced Detoxification of Oxidant Stress

    PubMed Central

    Williams, C. David; McGill, Mitchell R.; Farhood, Anwar; Jaeschke, Hartmut

    2013-01-01

    Acetaminophen (APAP) overdose is a classical model of hepatocellular necrosis; however, the involvement of the Fas receptor in the pathophysiology remains controversial. Fas receptor-deficient (lpr) and C57BL/6 mice were treated with APAP to compare the mechanisms of hepatotoxicity. Lpr mice were partially protected against APAP hepatotoxicity as indicated by reduced plasma ALT and GDH levels and liver necrosis. Hepatic Cyp2e1 protein, adduct formation and hepatic glutathione (GSH) depletion were similar, demonstrating equivalent reactive metabolite generation. There was no difference in cytokine formation or hepatic neutrophil recruitment. Interestingly, hepatic GSH recovered faster in lpr mice than in wild type animals resulting in enhanced detoxification of reactive oxygen species. Driving the increased GSH levels, mRNA induction and protein expression of glutamate-cysteine ligase (gclc) were higher in lpr mice. Inducible nitric oxide synthase (iNOS) mRNA and protein levels at 6h were significantly lower in lpr mice, which correlated with reduced nitrotyrosine staining. Heat shock protein 70 (Hsp70) mRNA levels were substantially higher in lpr mice after APAP. Conclusion: Our data suggest that the faster recovery of hepatic GSH levels during oxidant stress and peroxynitrite formation, reduced iNOS expression and enhanced induction of Hsp70 attenuated the susceptibility to APAP-induced cell death in lpr mice. PMID:23628456

  15. Cognitive-Enhancing Effect of Dianthus superbus var. Longicalycinus on Scopolamine-Induced Memory Impairment in Mice

    PubMed Central

    Weon, Jin Bae; Jung, Youn Sik; Ma, Choong Je

    2016-01-01

    Dianthus superbus (D. superbus) is a traditional crude drug used for the treatment of urethritis, carbuncles and carcinomas. The objective of this study was to confirm the cognitive enhancing effect of D. superbus in memory impairment induced mice and to elucidate the possible potential mechanism. Effect of D. superbus on scopolamine induced memory impairment on mice was evaluated using the Morris water maze and passive avoidance tests. We also investigated acetylcholinesterase (AChE) activity and brain-derived neurotropic factor (BDNF) expression in scopolamine-induced mice. HPLC-DAD analysis was performed to identify active compounds in D. superbus. The results revealed that D. superbus attenuated the learning and memory impairment induced by scopolamine. D. superbus also inhibited AChE levels in the hippocampi of the scopolamine-injected mice. Moreover, D. superbus increased BDNF expression in the hippocampus. Eight compounds were identified using HPLC-DAD analysis. The content of 4-hydroxyphenyl acetic acid was higher than contents of other compounds. These results indicated that D. superbus improved memory functioning accompanied by inhibition of AChE and upregulation of BDNF, suggesting that D. superbus may be a useful therapeutic agent for the prevention or treatment of Alzheimer’s disease. PMID:27133261

  16. Cognitive-Enhancing Effect of Dianthus superbus var. Longicalycinus on Scopolamine-Induced Memory Impairment in Mice.

    PubMed

    Weon, Jin Bae; Jung, Youn Sik; Ma, Choong Je

    2016-05-01

    Dianthus superbus (D. superbus) is a traditional crude drug used for the treatment of urethritis, carbuncles and carcinomas. The objective of this study was to confirm the cognitive enhancing effect of D. superbus in memory impairment induced mice and to elucidate the possible potential mechanism. Effect of D. superbus on scopolamine induced memory impairment on mice was evaluated using the Morris water maze and passive avoidance tests. We also investigated acetylcholinesterase (AChE) activity and brain-derived neurotropic factor (BDNF) expression in scopolamine-induced mice. HPLC-DAD analysis was performed to identify active compounds in D. superbus. The results revealed that D. superbus attenuated the learning and memory impairment induced by scopolamine. D. superbus also inhibited AChE levels in the hippocampi of the scopolamine-injected mice. Moreover, D. superbus increased BDNF expression in the hippocampus. Eight compounds were identified using HPLC-DAD analysis. The content of 4-hydroxyphenyl acetic acid was higher than contents of other compounds. These results indicated that D. superbus improved memory functioning accompanied by inhibition of AChE and upregulation of BDNF, suggesting that D. superbus may be a useful therapeutic agent for the prevention or treatment of Alzheimer's disease. PMID:27133261

  17. Enhancement of the immune response against Salmonella infection of mice by heat-killed multispecies combinations of lactic acid bacteria.

    PubMed

    Chen, Chih-Yuan; Tsen, Hau-Yang; Lin, Chun-Li; Lin, Chien-Ku; Chuang, Li-Tsen; Chen, Chin-Shuh; Chiang, Yu-Cheng

    2013-11-01

    Heat-killed lactic acid bacteria (LAB) has advantages over live LAB in that it has a long shelf-life and is therefore easy to store and transport. From four LAB strains selected by immunomodulatory activity and adherent properties, we prepared the heat-killed multispecies combination of LAB (MLAB) and the cell walls from MLAB under two conditions (100 °C for 30 min and 121 °C for 15 min). Different effects on the adherent properties of these four LAB strains were observed, depending on the heating conditions. With mouse macrophage cells, the two heat-killed MLABs (HMLABs) showed significantly higher induction activities on the production of interleukin 12 (IL-12) than their individual strains did. Heat-killed MLABs and cell-wall preparations were able to reduce the Salmonella invasion of Caco-2 and mouse macrophage cells. Feeding mice with HMLAB could inhibit the Salmonella invasion of mice significantly. For these mice, the expression level of pro-inflammatory cytokines, such as TNF-α and IL-6, in mouse serum was reduced while that of the anti-inflammatory cytokine, i.e. IL-10, was enhanced. The HMLABs developed in this study showed higher protective effect against Salmonella invasion either of Caco-2 cells or of mice, relative to the heat-killed lactobacilli, which consisted of Lactobacillus acidophilus strains selected at random. In conclusion, the HMLABs were potentially useful for the protection of mice against Salmonella infection and the induced inflammation. PMID:24000228

  18. Over-expression of hedgehog signaling is associated with epidermal tumor formation in vitamin D receptor null mice

    PubMed Central

    Teichert, Arnaud; Elalieh, Hashem; Elias, Peter; Welsh, JoEllen; Bikle, Daniel D.

    2011-01-01

    The vitamin D receptor (VDR) ligand, 1,25(OH)2D3, reduces proliferation and enhances differentiation and thus has been investigated for a role in preventing or treating cancer. Mice deficient for the VDR display a hyperproliferative response in the hair follicle and epidermis and decreased epidermal differentiation. Unlike their wild type littermates, when treated with 7,12 dimethylbenzanthracene (DMBA) or UVB, they develop skin tumors, including some characteristic of over-expression of the hedgehog (Hh) pathway. Both the epidermis and utricles of the VDR null animals over-express elements of the Hh pathway [Sonic Hedgehog (Shh, 2.02 fold), Patched1 1.58 fold, Smoothened 3.54 fold, Gli1 1.17 fold, and Gli2 1.66 fold]. This over-expression occurs at an age (11 weeks) where epidermal hyperproliferation is most visible and is spatially controlled in the epidermis. DMBA or UVB induced tumors in the VDR null mice also over-express elements of this pathway. Moreover, 1,25(OH)2D3 down-regulates the expression of some members of the Hh pathway in an epidermal explants culture system, suggesting a direct regulation by 1,25(OH)2D3. Our results suggest that increased expression of Shh in the keratinocytes of the VDR null animal activates the Hh pathway, predisposing the skin to the development of both malignant and benign epidermal neoplasms. PMID:21814234

  19. Fluoxetine prevents respiratory arrest without enhancing ventilation in DBA/1 mice.

    PubMed

    Zeng, Chang; Long, Xiaoyan; Cotten, Joseph F; Forman, Stuart A; Solt, Ken; Faingold, Carl L; Feng, Hua-Jun

    2015-04-01

    Sudden unexpected death in epilepsy (SUDEP) is a fatal epileptic event. DBA/1 mice are a relevant animal model for the study of SUDEP, as these mice exhibit seizure-induced respiratory arrest (S-IRA) leading to death, which has been observed in patients with witnessed SUDEP. Fluoxetine, a selective serotonin (5-hydroxytryptamine or 5-HT) reuptake inhibitor (SSRI), reduces S-IRA in DBA/1 mice. Given that DBA/1 mice with S-IRA can be resuscitated using a ventilator, we hypothesized that breathing stimulants can prevent S-IRA and that fluoxetine prevents S-IRA by enhancing ventilation in these mice. Spontaneous respiratory function in anesthetized or awake DBA/1 mice was examined using noninvasive plethysmography before and after administering fluoxetine or breathing stimulants, doxapram, and 5,6,7,8-tetrahydropyrido[4,3-d]pyrimidine (PK-THPP). The effects of these drugs on S-IRA in DBA/1 mice were tested. As reported previously, systemic administration of fluoxetine reduced S-IRA in awake DBA/1 mice, but fluoxetine in anesthetized and awake DBA/1 mice did not increase basal ventilation or the ventilatory response to 7% CO2. Both doxapram and PK-THPP increased ventilation in room air and in air+7% CO2 in anesthetized DBA/1 mice. However, neither of the breathing stimulants reduced the incidence of S-IRA. Our studies confirm that fluoxetine reduces S-IRA in DBA/1 mice without enhancing basal ventilation in the absence of seizures. Although breathing stimulants increased ventilation in the absence of seizures, they were ineffective in reducing S-IRA, indicating that drug-induced increases in ventilation are insufficient to compensate for S-IRA in DBA/1 mice. PMID:25771493

  20. Fibrillin-1 impairment enhances blood-brain barrier permeability and xanthoma formation in brains of apolipoprotein E-deficient mice.

    PubMed

    Van der Donckt, C; Roth, L; Vanhoutte, G; Blockx, I; Bink, D I; Ritz, K; Pintelon, I; Timmermans, J-P; Bauters, D; Martinet, W; Daemen, M J; Verhoye, M; De Meyer, G R Y

    2015-06-01

    We recently reported that apolipoprotein E (ApoE)-deficient mice with a mutation in the fibrillin-1 gene (ApoE(-/-)Fbn1(C1039G+/-)) develop accelerated atherosclerosis with enhanced inflammation, atherosclerotic plaque rupture, myocardial infarction and sudden death. In the brain, fibrillin-1 functions as an attachment protein in the basement membrane, providing structural support to the blood-brain barrier (BBB). Here, we investigated whether fibrillin-1 impairment affects the permeability of the BBB proper and the blood-cerebrospinal fluid barrier (BCSFB), and whether this leads to the accelerated accumulation of lipids (xanthomas) in the brain. ApoE(-/-) (n=61) and ApoE(-/-)Fbn1(C1039G+/-) (n=73) mice were fed a Western-type diet (WD). After 14 weeks WD, a significantly higher permeability of the BBB was observed in ApoE(-/-)Fbn1(C1039G+/-) mice compared to age-matched ApoE(-/-) mice. This was accompanied by leukocyte infiltration, enhanced expression of pro-inflammatory cytokines, matrix metalloproteinases and transforming growth factor-β, and by decreased expression of tight junction proteins claudin-5 and occludin. After 20 weeks WD, 83% of ApoE(-/-)Fbn1(C1039G+/-) mice showed xanthomas in the brain, compared to 23% of their ApoE(-/-) littermates. Xanthomas were mainly located in fibrillin-1-rich regions, such as the choroid plexus and the neocortex. Our findings demonstrate that dysfunctional fibrillin-1 impairs BBB/BCSFB integrity, facilitating peripheral leukocyte infiltration, which further degrades the BBB/BCSFB. As a consequence, lipoproteins can enter the brain, resulting in accelerated formation of xanthomas. PMID:25797463

  1. Oxytocin knockout mice demonstrate enhanced intake of sweet and nonsweet carbohydrate solutions.

    PubMed

    Sclafani, Anthony; Rinaman, Linda; Vollmer, Regis R; Amico, Janet A

    2007-05-01

    Oxytocin knockout (OT KO) mice display enhanced intake of nutritive and nonnutritive sweet solutions (i.e., sucrose and saccharin) compared with wild-type (WT) mice of the same C57BL/6 background strain. The present study further investigated the differential behavioral response of OT KO and WT mice to sucrose solutions and also examined intake preferences of OT KO and WT mice for palatable but nonsweet isocaloric solutions of carbohydrate and fat. A progressive ratio operant licking procedure demonstrated that OT KO and WT mice display a similar motivational drive to consume 10% sucrose. A series of two-bottle intake tests revealed that OT KO mice consume significantly larger amounts of both sweet and nonsweet carbohydrate solutions (i.e., sucrose, Polycose, and cornstarch) compared with WT cohorts. Intake pattern analyses revealed that OT KO mice overconsume carbohydrate solutions by initiating more drinking bouts compared with WT mice; bout sizes did not differ between the genotypes. In contrast, OT KO and WT mice did not differ in their intake of Intralipid, a palatable soybean oil emulsion. These findings indicate that the absence of OT in mice does not affect their appetitive drive to consume palatable sucrose solutions. Instead, the absence of OT may increase daily intake of palatable sweet and nonsweet solutions of carbohydrate (but not fat) by selectively blunting or masking processes that contribute to postingestive satiety. PMID:17272659

  2. HIPPOCAMPAL SPINE-ASSOCIATED Rap-SPECIFIC GTPase-ACTIVATING PROTEIN INDUCES ENHANCEMENT OF LEARNING AND MEMORY IN POSTNATALLY HYPOXIA-EXPOSED MICE

    PubMed Central

    LU, X.-J.; CHEN, X.-Q.; WENG, J.; ZHANG, H.-Y.; PAK, D. T.; LUO, J.-H.; DU, J.-Z.

    2011-01-01

    Spine-associated Rap-specific GTPase-activating protein (SPAR) is a postsynaptic protein that forms a complex with postsynaptic density (PSD)-95 and N-methyl-D-aspartate receptors (NMDARs), and morphologically regulates dendritic spines. Mild intermittent hypoxia (IH, 16.0% O2, 4 h/day for 4 weeks) is known to markedly enhance spatial learning and memory in postnatal developing mice. Here, we report that this effect is correlated with persistent increases in SPAR expression as well as long-term potentiation (LTP) in the hippocampus of IH-exposed mice. Furthermore, an infusion of SPAR antisense oligonucleotides into the dorsal hippocampus disrupted elevation of SPAR expression, preventing enhanced hippocampal LTP in IH-exposed developing mice and also reducing LTP in normoxic mice, without altering basal synaptic transmission. In SPAR antisense-treated mice, acquisition of the Morris water maze spatial learning task was impaired, as was memory retention in probe trails following training. This study provides the first evidence that SPAR is functionally required for synaptic plasticity and contributes to the IH-induced enhancement of spatial learning and memory in postnatal developing mice. PMID:19442707

  3. Salicylate Downregulates 11β-HSD1 Expression in Adipose Tissue in Obese Mice and in Humans, Mediating Insulin Sensitization

    PubMed Central

    Nixon, Mark; Wake, Deborah J.; Livingstone, Dawn E.; Stimson, Roland H.; Esteves, Cristina L.; Seckl, Jonathan R.; Chapman, Karen E.; Andrew, Ruth; Walker, Brian R.

    2012-01-01

    Recent trials show salicylates improve glycemic control in type 2 diabetes, but the mechanism is poorly understood. Expression of the glucocorticoid-generating enzyme 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) in adipose tissue is increased in vitro by proinflammatory cytokines and upregulated in obesity. 11β-HSD1 inhibition enhances insulin sensitivity. We hypothesized that salicylates downregulate 11β-HSD1 expression, contributing to their metabolic efficacy. We treated diet-induced obese (DIO) 11β-HSD1–deficient mice and C57Bl/6 mice with sodium salicylate for 4 weeks. Glucose tolerance was assessed in vivo. Tissue transcript levels were assessed by quantitative PCR and enzyme activity by incubation with 3H-steroid. Two weeks’ administration of salsalate was also investigated in a randomized double-blind placebo-controlled crossover study in 16 men, with measurement of liver 11β-HSD1 activity in vivo and adipose tissue 11β-HSD1 transcript levels ex vivo. In C57Bl/6 DIO mice, salicylate improved glucose tolerance and downregulated 11β-HSD1 mRNA and activity selectively in visceral adipose. DIO 11β-HSD1–deficient mice were resistant to these metabolic effects of salicylate. In men, salsalate reduced 11β-HSD1 expression in subcutaneous adipose, and in vitro salicylate treatment reduced adipocyte 11β-HSD1 expression and induced adiponectin expression only in the presence of 11β-HSD1 substrate. Reduced intra-adipose glucocorticoid regeneration by 11β-HSD1 is a novel mechanism that contributes to the metabolic efficacy of salicylates. PMID:22357964

  4. Developmental expression of the neuroligins and neurexins in fragile X mice.

    PubMed

    Lai, Jonathan K Y; Doering, Laurie C; Foster, Jane A

    2016-03-01

    Neuroligins and neurexins are transsynaptic proteins involved in the maturation of glutamatergic and GABAergic synapses. Research has identified synaptic proteins and function as primary contributors to the development of fragile X syndrome. Fragile X mental retardation protein (FMRP), the protein that is lacking in fragile X syndrome, binds neuroligin-1 and -3 mRNA. Using in situ hybridization, we examined temporal and spatial expression patterns of neuroligin (NLGN) and neurexin (NRXN) mRNAs in the somatosensory (S1) cortex and hippocampus in wild-type (WT) and fragile X knockout (FMR1-KO) mice during the first 5 weeks of postnatal life. Genotype-based differences in expression included increased NLGN1 mRNA in CA1 and S1 cortex, decreased NLGN2 mRNA in CA1 and dentate gyrus (DG) regions of the hippocampus, and increased NRXN3 mRNA in CA1, DG, and S1 cortex between female WT and FMR1-KO mice. In male mice, decreased expression of NRXN3 mRNA was observed in CA1 and DG regions of FMR1-KO mice. Sex differences in hippocampal expression of NLGN2, NRXN1, NRXN2, and NRXN3 mRNAs and in S1 cortex expression of NRXN3 mRNAs were observed WT mice, whereas sex differences in NLGN3, NRXN1, NRXN2, and NRXN3 mRNA expression in the hippocampus and in NLGN1, NRXN2 and NRXN3 mRNA expression in S1 cortex were detected in FMR1-KO mice. These results provide a neuroanatomical map of NLGN and NRXN expression patterns over postnatal development in WT and FMR1-KO mice. The differences in developmental trajectory of these synaptic proteins could contribute to long-term differences in CNS wiring and synaptic function. PMID:26235839

  5. Betatrophin expression is promoted in obese hyperinsulinemic type 2 but not type 1 diabetic mice.

    PubMed

    Li, EnXu; Nakata, Masanori; Shinozaki, Atsumi; Yang, Yifei; Zhang, Boyang; Yada, Toshihiko

    2016-07-30

    Regeneration of pancreatic β-cell mass benefits both type 1 and type 2 diabetic patients. A recent study identified betatrophin as a β-cell proliferation factor. However, the expressional regulation of betatrophin remains less defined. In this study, we aimed to clarify the regulation of betatrophin expression in obese type 2 vs. type 1 diabetes model animals. We experimented type 2 diabetes models, diet-induced-obesity (DIO) mice and db/db mice, and type 1 diabetes models, C57B6 mice receiving streptozotocin (STZ) or 70% pancreatectomy to destroy or remove β-cells. Serum betatrophin levels and betatrophin mRNA expressions in the liver, white adipose tissue (WAT) and brown adipose tissue (BAT) were measured. In DIO mice and db/db mice, serum betatrophin and betatrophin mRNA expressions in the liver, WAT and BAT were elevated in parallel with increases in body weight and plasma insulin. These elevated betatrophin mRNA expressions were not altered by treatment with SGLT2 inhibitor that ameliorated hyperglycemia. In pancreatectomized mice, betatrophin expression in WAT decreased in parallel with reductions in weight and insulin. In STZ-treated mice, betatrophin expressions in the liver, WAT and BAT were reduced. However, when the mouse liver slices were cultured with STZ, betatrophin expression was significantly reduced, indicating a direct action of STZ on the liver. These results indicate that the expression of betatrophin is upregulated in the liver, WAT and BAT in obese hyperinsulinemic type 2 diabetes but decreased in WAT in hypoinsulinemic type 1 diabetes, suggesting its positive correlation with body weight and plasma insulin but not blood glucose. PMID:27097546

  6. Male sterility and enhanced radiation sensitivity in TLS(-/-) mice.

    PubMed

    Kuroda, M; Sok, J; Webb, L; Baechtold, H; Urano, F; Yin, Y; Chung, P; de Rooij, D G; Akhmedov, A; Ashley, T; Ron, D

    2000-02-01

    TLS (also known as FUS) is an RNA-binding protein that contributes the N-terminal half of fusion oncoproteins implicated in the development of human liposarcomas and leukemias. Here we report that male mice homozygous for an induced mutation in TLS are sterile with a marked increase in the number of unpaired and mispaired chromosomal axes in pre-meiotic spermatocytes. Nuclear extracts from TLS(-/-) testes lack an activity capable of promoting pairing between homologous DNA sequences in vitro, and TLS(-/-) mice and embryonic fibroblasts exhibit increased sensitivity to ionizing irradiation. These results are consistent with a role for TLS in homologous DNA pairing and recombination. PMID:10654943

  7. Munc18-1 haploinsufficiency results in enhanced anxiety-like behavior as determined by heart rate responses in mice.

    PubMed

    Hager, Torben; Maroteaux, Grégoire; Pont, Paula du; Julsing, Joris; van Vliet, Rick; Stiedl, Oliver

    2014-03-01

    Heterozygous (HZ) missense mutations in the gene encoding syntaxin binding protein 1 (Stxbp1 or Munc18-1), a presynaptic protein essential for neurotransmitter release, causes early infantile epileptic encephalopathy, abnormal brain structure and mental retardation in humans. Here we investigated whether the mouse model mimics symptoms of the human phenotype. The effects of the deletion of munc18-1 were studied in HZ and wild-type (WT) mice based on heart rate (HR) and its variability (HRV) as independent measures to expand previous behavioral results of enhanced anxiety and impaired emotional learning suggesting mild cognitive impairments. HR responses were assessed during novelty exposure, during the expression and extinction of conditioned tone-dependent fear and during the diurnal phase. Novelty exposure yielded no differences in activity patterns between the two genotypes, while maximum HR differed significantly (WT: 770 bpm; HZ: 790 bpm). Retention tests after both auditory delay and trace fear conditioning showed a delayed extinction of the conditioned HR response in HZ mice compared to WT mice. Since the HR versus HRV correlation and HR dynamics assessed by nonlinear methods revealed similar function in HZ and WT mice, the higher HR responses of munc18-1 HZ mice to different emotional challenges cannot be attributed to differences in autonomic nervous system function. Thus, in contrast to the adverse consequences of deletion of a single allele of munc18-1 in humans, C57BL/6J mice show enhanced anxiety responses based on HR adjustments that extend previous results on the behavioral level without support of cognitive impairment, epileptic seizures and autonomic dysregulation. PMID:24304718

  8. Transgenic Expression of Osteoactivin/gpnmb Enhances Bone Formation In Vivo and Osteoprogenitor Differentiation Ex Vivo.

    PubMed

    Frara, Nagat; Abdelmagid, Samir M; Sondag, Gregory R; Moussa, Fouad M; Yingling, Vanessa R; Owen, Thomas A; Popoff, Steven N; Barbe, Mary F; Safadi, Fayez F

    2016-01-01

    Initial identification of osteoactivin (OA)/glycoprotein non-melanoma clone B (gpnmb) was demonstrated in an osteopetrotic rat model, where OA expression was increased threefold in mutant bones, compared to normal. OA mRNA and protein expression increase during active bone regeneration post-fracture, and primary rat osteoblasts show increased OA expression during differentiation in vitro. To further examine OA/gpnmb as an osteoinductive agent, we characterized the skeletal phenotype of transgenic mouse overexpressing OA/gpnmb under the CMV-promoter (OA-Tg). Western blot analysis showed increased OA/gpnmb in OA-Tg osteoblasts, compared to wild-type (WT). In OA-Tg mouse femurs versus WT littermates, micro-CT analysis showed increased trabecular bone volume and thickness, and cortical bone thickness; histomorphometry showed increased osteoblast numbers, bone formation and mineral apposition rates in OA-Tg mice; and biomechanical testing showed higher peak moment and stiffness. Given that OA/gpnmb is also over-expressed in osteoclasts in OA-Tg mice, we evaluated bone resorption by ELISA and histomorphometry, and observed decreased serum CTX-1 and RANK-L, and decreased osteoclast numbers in OA-Tg, compared to WT mice, indicating decreased bone remodeling in OA-Tg mice. The proliferation rate of OA-Tg osteoblasts in vitro was higher, compared to WT, as was alkaline phosphatase staining and activity, the latter indicating enhanced differentiation of OA-Tg osteoprogenitors. Quantitative RT-PCR analysis showed increased TGF-β1 and TGF-β receptors I and II expression in OA-Tg osteoblasts, compared to WT. Together, these data suggest that OA overexpression has an osteoinductive effect on bone mass in vivo and stimulates osteoprogenitor differentiation ex vivo. PMID:25899717

  9. Lipopolysaccharide-induced anti-inflammatory acute phase response is enhanced in spermidine/spermine N1-acetyltransferase (SSAT) overexpressing mice.

    PubMed

    Pirnes-Karhu, Sini; Sironen, Reijo; Alhonen, Leena; Uimari, Anne

    2012-02-01

    Bacterial lipopolysaccharide (LPS) is an effective activator of the components of innate immunity. It has been shown that polyamines and their metabolic enzymes affect the LPS-induced immune response by modulating both pro- and anti-inflammatory actions. On the other hand, LPS causes changes in cellular polyamine metabolism. In this study, the LPS-induced inflammatory response in spermidine/spermine N(1)-acetyltransferase overexpressing transgenic mice (SSAT mice) was analyzed. In liver and kidneys, LPS enhanced the activity of the polyamine biosynthetic enzyme ornithine decarboxylase and increased the intracellular putrescine content in both SSAT overexpressing and wild-type mice. In survival studies, the enhanced polyamine catabolism and concomitantly altered cellular polyamine pools in SSAT mice did not affect the LPS-induced mortality of these animals. However, in the acute phase of LPS-induced inflammatory response, the serum levels of proinflammatory cytokines interleukin-1β and interferon-γ were significantly reduced and, on the contrary, anti-inflammatory cytokine interleukin-10 was significantly increased in the sera of SSAT mice compared with the wild-type animals. In addition, hepatic acute-phase proteins C-reactive protein, haptoglobin and α(1)-acid glycoprotein were expressed in higher amounts in SSAT mice than in the wild-type animals. In summary, the study suggests that SSAT overexpression obtained in SSAT mice enhances the anti-inflammatory actions in the acute phase of LPS-induced immune response. PMID:21814792

  10. ENHANCED PRODUCTION OF CRYPTOSPORIDIUM PARVUM OOCYSTS IN IMMUNOSUPPRESSED MICE

    EPA Science Inventory

    Recently there has been an increase in the need for fresh C. parvum oocysts for engineering and biomedical research applications. In our laboratory the emphsis has shifted from the use of dairy calves to inbred C57BL/67n mice, primarily for reasons of ease of collection and proce...

  11. Enhanced Generalization of Auditory Conditioned Fear in Juvenile Mice

    ERIC Educational Resources Information Center

    Ito, Wataru; Pan, Bing-Xing; Yang, Chao; Thakur, Siddarth; Morozov, Alexei

    2009-01-01

    Increased emotionality is a characteristic of human adolescence, but its animal models are limited. Here we report that generalization of auditory conditioned fear between a conditional stimulus (CS+) and a novel auditory stimulus is stronger in 4-5-wk-old mice (juveniles) than in their 9-10-wk-old counterparts (adults), whereas nonassociative…

  12. Dietary chromium and nickel enhance UV-carcinogenesis in skin of hairless mice

    SciTech Connect

    Uddin, Ahmed N.; Burns, Fredric J.; Rossman, Toby G.; Chen, Haobin; Kluz, Thomas; Costa, Max . E-mail: costam01@nyu.edu

    2007-06-15

    The skin cancer enhancing effect of chromium (in male mice) and nickel in UVR-irradiated female Skh1 mice was investigated. The dietary vitamin E and selenomethionine were tested for prevention of chromium-enhanced skin carcinogenesis. The mice were exposed to UVR (1.0 kJ/m{sup 2} 3x weekly) for 26 weeks either alone, or combined with 2.5 or 5.0 ppm potassium chromate, or with 20, 100 or 500 ppm nickel chloride in drinking water. Vitamin E or selenomethionine was added to the lab chow for 29 weeks beginning 3 weeks before the start of UVR exposure. Both chromium and nickel significantly increased the UVR-induced skin cancer yield in mice. In male Skh1 mice, UVR alone induced 1.9 {+-} 0.4 cancers/mouse, and 2.5 or 5.0 ppm potassium chromate added to drinking water increased the yields to 5.9 {+-} 0.8 and 8.6 {+-} 0.9 cancers/mouse, respectively. In female Skh1 mice, UVR alone induced 1.7 {+-} 0.4 cancers/mouse, and the addition of 20, 100 or 500 ppm nickel chloride increased the yields to 2.8 {+-} 0.9, 5.6 {+-} 0.7 and 4.2 {+-} 1.0 cancers/mouse, respectively. Neither vitamin E nor selenomethionine reduced the cancer yield enhancement by chromium. These results confirm that chromium and nickel, while not good skin carcinogens per se, are enhancers of UVR-induced skin cancers in Skh1 mice. Data also suggest that the enhancement of UVR-induced skin cancers by chromate may not be oxidatively mediated since the antioxidant vitamin E as well as selenomethionine, found to prevent arsenite-enhanced skin carcinogenesis, failed to suppress enhancement by chromate.

  13. Procollagen C-endopeptidase Enhancer Protein 2 (PCPE2) Reduces Atherosclerosis in Mice by Enhancing Scavenger Receptor Class B1 (SR-BI)-mediated High-density Lipoprotein (HDL)-Cholesteryl Ester Uptake.

    PubMed

    Pollard, Ricquita D; Blesso, Christopher N; Zabalawi, Manal; Fulp, Brian; Gerelus, Mark; Zhu, Xuewei; Lyons, Erica W; Nuradin, Nebil; Francone, Omar L; Li, Xiang-An; Sahoo, Daisy; Thomas, Michael J; Sorci-Thomas, Mary G

    2015-06-19

    Studies in human populations have shown a significant correlation between procollagen C-endopeptidase enhancer protein 2 (PCPE2) single nucleotide polymorphisms and plasma HDL cholesterol concentrations. PCPE2, a 52-kDa glycoprotein located in the extracellular matrix, enhances the cleavage of C-terminal procollagen by bone morphogenetic protein 1 (BMP1). Our studies here focused on investigating the basis for the elevated concentration of enlarged plasma HDL in PCPE2-deficient mice to determine whether they protected against diet-induced atherosclerosis. PCPE2-deficient mice were crossed with LDL receptor-deficient mice to obtain LDLr(-/-), PCPE2(-/-) mice, which had elevated HDL levels compared with LDLr(-/-) mice with similar LDL concentrations. We found that LDLr(-/-), PCPE2(-/-) mice had significantly more neutral lipid and CD68+ infiltration in the aortic root than LDLr(-/-) mice. Surprisingly, in light of their elevated HDL levels, the extent of aortic lipid deposition in LDLr(-/-), PCPE2(-/-) mice was similar to that reported for LDLr(-/-), apoA-I(-/-) mice, which lack any apoA-I/HDL. Furthermore, LDLr(-/-), PCPE2(-/-) mice had reduced HDL apoA-I fractional clearance and macrophage to fecal reverse cholesterol transport rates compared with LDLr(-/-) mice, despite a 2-fold increase in liver SR-BI expression. PCPE2 was shown to enhance SR-BI function by increasing the rate of HDL-associated cholesteryl ester uptake, possibly by optimizing SR-BI localization and/or conformation. We conclude that PCPE2 is atheroprotective and an important component of the reverse cholesterol transport HDL system. PMID:25947382

  14. Endogenous galectin-3 expression levels modulate immune responses in galectin-3 transgenic mice.

    PubMed

    Chaudhari, Aparna D; Gude, Rajiv P; Kalraiya, Rajiv D; Chiplunkar, Shubhada V

    2015-12-01

    Galectin-3 (Gal-3), a β-galactoside-binding mammalian lectin, is involved in cancer progression and metastasis. However, there is an unmet need to identify the underlying mechanisms of cancer metastasis mediated by endogenous host galectin-3. Galectin-3 is also known to be an important regulator of immune responses. The present study was aimed at analysing how expression of endogenous galectin-3 regulates host immunity and lung metastasis in B16F10 murine melanoma model. Transgenic Gal-3(+/-) (hemizygous) and Gal-3(-/-) (null) mice exhibited decreased levels of Natural Killer (NK) cells and lower NK mediated cytotoxicity against YAC-1 tumor targets, compared to Gal-3(+/+) (wild-type) mice. On stimulation, Gal-3(+/-) and Gal-3(-/-) mice splenocytes showed increased T cell proliferation than Gal-3(+/+) mice. Intracellular calcium flux was found to be lower in activated T cells of Gal-3(-/-) mice as compared to T cells from Gal-3(+/+) and Gal-3(+/-) mice. In Gal-3(-/-) mice, serum Th1, Th2 and Th17 cytokine levels were found to be lowest, exhibiting dysregulation of pro-inflammatory and anti-inflammatory cytokines balance. Marked decrease in serum IFN-γ levels and splenic IFN-γR1 (IFN-γ Receptor 1) expressing T and NK cell percentages were observed in Gal-3(-/-) mice. On recombinant IFN-γ treatment of splenocytes in vitro, Suppressor of Cytokine Signaling (SOCS) 1 and SOCS3 protein expression was higher in Gal-3(-/-) mice compared to that in Gal-3(+/+) and Gal-3(+/-) mice; suggesting possible attenuation of Signal Transducer and Activator of Transcription (STAT) 1 mediated IFN-γ signaling in Gal-3(-/-) mice. The ability of B16F10 melanoma cells to form metastatic colonies in the lungs of Gal-3(+/+) and Gal-3(-/-) mice remained comparable, whereas it was found to be reduced in Gal-3(+/-) mice. Our data indicates that complete absence of endogenous host galectin-3 facilitates lung metastasis of B16F10 cells in mice, which may be contributed by dysregulated immune

  15. Regulation of Tyrosine Hydroxylase Expression and Phosphorylation in Dopamine Transporter-Deficient Mice.

    PubMed

    Salvatore, Michael F; Calipari, Erin S; Jones, Sara R

    2016-07-20

    Tyrosine hydroxylase (TH) and dopamine transporters (DATs) regulate dopamine (DA) neurotransmission at the biosynthesis and reuptake steps, respectively. Dysfunction or loss of these proteins occurs in impaired locomotor or addictive behavior, but little is known about the influence of DAT expression on TH function. Differences in TH phosphorylation, DA tissue content, l-DOPA biosynthesis, and DA turnover exist between the somatodendritic and terminal field compartments of nigrostriatal and mesoaccumbens pathways. We examined whether differential DAT expression affects these compartmental differences in DA regulation by comparing TH expression and phosphorylation at ser31 and ser40. In heterozygous DAT knockout (KO) (+/-) mice, DA tissue content and DA turnover were unchanged relative to wild-type mice, despite a 40% reduction in DAT protein expression. In DAT KO (-/-) mice, DA turnover increased in all DA compartments, but DA tissue content decreased (90-96%) only in terminal fields. TH protein expression and phosphorylation were differentially affected within DA pathway compartments by relative expression of DAT. TH protein decreased (∼74%), though to a significantly lesser extent than DA, in striatum and nucleus accumbens (NAc) in DAT -/- mice, with no decrease in substantia nigra or ventral tegmental area. Striatal ser31 TH phosphorylation and recovery of DA relative to TH protein expression in DAT +/- and DAT -/- mice decreased, whereas ser40 TH phosphorylation increased ∼2- to 3-fold in striatum and NAc of DAT -/- mice. These results suggest that DAT expression affects TH expression and phosphorylation largely in DA terminal field compartments, further corroborating evidence for dichotomous regulation of TH between somatodendritic and terminal field compartments of the nigrostriatal and mesoaccumbens pathways. PMID:27124386

  16. DIETARY FOLATE DEFICIENCY ENHANCES ARSENIC-INDUCED MICRONUCLEUS FORMATION IN MICE

    EPA Science Inventory


    Dietary folate deficiency enhances arsenic-induced micronucleus formation in mice.

    Folate deficiency increases background levels ofDNA damage and can enhance the mutagenicity of chemical agents. Duplicate experiments were performed to investigate the effect of dietary...

  17. Expression of SIRT1 and SIRT3 varies according to age in mice.

    PubMed

    Kwon, Youngho; Kim, Jongsik; Lee, Chae-Yeong; Kim, Hyun

    2015-03-01

    Sirtuins (SIRTs) are involved in multiple cellular processes including those related to aging, cancer, and a variety of cellular functions including cell cycle progression, DNA repair, and cellular proliferation. SIRTs have been shown to extend the yeast life span, although there is presently little known about SIRT expression in the organs of mice. In the present study, we were especially interested in identifying differences in SIRT expression between young mice and aged mice. Specifically, we investigated the expression of SIRT1 and SIRT3 in the kidney, lung, skin, adipose tissue, and spleens of 6-month-old and 24-month-old mice using immunohistochemical staining. Compared with that in younger mice, the expression of SIRT1 in 24-month-old rats was increased in kidney, lung, and spleen tissue, while that of SIRT3 was decreased in adipose, kidney, and lung tissue. The results of our study suggest that aging is associated with altered patterns of expression of SIRT1 and SIRT3. In addition, we noted that the expression patterns of SIRT1 and SIRT3 varied by organ. Taken together, the results of this study suggest the possibility that SIRTs may be involved in diseases associated with aging. PMID:25806122

  18. Expression of SIRT1 and SIRT3 varies according to age in mice

    PubMed Central

    Kwon, Youngho; Kim, Jongsik; Lee, Chae-Yeong

    2015-01-01

    Sirtuins (SIRTs) are involved in multiple cellular processes including those related to aging, cancer, and a variety of cellular functions including cell cycle progression, DNA repair, and cellular proliferation. SIRTs have been shown to extend the yeast life span, although there is presently little known about SIRT expression in the organs of mice. In the present study, we were especially interested in identifying differences in SIRT expression between young mice and aged mice. Specifically, we investigated the expression of SIRT1 and SIRT3 in the kidney, lung, skin, adipose tissue, and spleens of 6-month-old and 24-month-old mice using immunohistochemical staining. Compared with that in younger mice, the expression of SIRT1 in 24-month-old rats was increased in kidney, lung, and spleen tissue, while that of SIRT3 was decreased in adipose, kidney, and lung tissue. The results of our study suggest that aging is associated with altered patterns of expression of SIRT1 and SIRT3. In addition, we noted that the expression patterns of SIRT1 and SIRT3 varied by organ. Taken together, the results of this study suggest the possibility that SIRTs may be involved in diseases associated with aging. PMID:25806122

  19. Conditional expression of constitutively active estrogen receptor {alpha} in chondrocytes impairs longitudinal bone growth in mice

    SciTech Connect

    Ikeda, Kazuhiro; Tsukui, Tohru; Imazawa, Yukiko; Horie-Inoue, Kuniko; Inoue, Satoshi

    2012-09-07

    Highlights: Black-Right-Pointing-Pointer Conditional transgenic mice expressing constitutively active estrogen receptor {alpha} (caER{alpha}) in chondrocytes were developed. Black-Right-Pointing-Pointer Expression of caER{alpha} in chondrocytes impaired longitudinal bone growth in mice. Black-Right-Pointing-Pointer caER{alpha} affects chondrocyte proliferation and differentiation. Black-Right-Pointing-Pointer This mouse model is useful for understanding the physiological role of ER{alpha}in vivo. -- Abstract: Estrogen plays important roles in the regulation of chondrocyte proliferation and differentiation, which are essential steps for longitudinal bone growth; however, the mechanisms of estrogen action on chondrocytes have not been fully elucidated. In the present study, we generated conditional transgenic mice, designated as caER{alpha}{sup ColII}, expressing constitutively active mutant estrogen receptor (ER) {alpha} in chondrocytes, using the chondrocyte-specific type II collagen promoter-driven Cre transgenic mice. caER{alpha}{sup ColII} mice showed retardation in longitudinal growth, with short bone lengths. BrdU labeling showed reduced proliferation of hypertrophic chondrocytes in the proliferating layer of the growth plate of tibia in caER{alpha}{sup ColII} mice. In situ hybridization analysis of type X collagen revealed that the maturation of hypertrophic chondrocytes was impaired in caER{alpha}{sup ColII} mice. These results suggest that ER{alpha} is a critical regulator of chondrocyte proliferation and maturation during skeletal development, mediating longitudinal bone growth in vivo.

  20. Microbiota prevents cholesterol loss from the body by regulating host gene expression in mice

    PubMed Central

    Zhong, Chun-Yan; Sun, Wei-Wei; Ma, Yinyan; Zhu, Hongling; Yang, Pan; Wei, Hong; Zeng, Ben-Hua; Zhang, Qian; Liu, Yu; Li, Wen-Xia; Chen, Yixin; Yu, Liqing; Song, Zhi-Yuan

    2015-01-01

    We have previously observed that knockout of Niemann-Pick C1-Like 1 (NPC1L1), a cholesterol transporter essential for intestinal cholesterol absorption, reduces the output of dry stool in mice. As the food intake remains unaltered in NPC1L1-knockout (L1-KO) mice, we hypothesized that NPC1L1 deficiency may alter the gut microbiome to reduce stool output. Consistently, here we demonstrate that the phyla of fecal microbiota differ substantially between L1-KO mice and their wild-type controls. Germ-free (GF) mice have reduced stool output. Inhibition of NPC1L1 by its inhibitor ezetimibe reduces stool output in specific pathogen-free (SPF), but not GF mice. In addition, we show that GF versus SPF mice have reduced intestinal absorption and increased fecal excretion of cholesterol, particularly after treatment with ezetimibe. This negative balance of cholesterol in GF mice is associated with reduced plasma and hepatic cholesterol, and likely caused by reduced expression of NPC1L1 and increased expression of ABCG5 and ABCG8 in small intestine. Expression levels of other genes in intestine and liver largely reflect a state of cholesterol depletion and a decrease in intestinal sensing of bile acids. Altogether, our findings reveal a broad role of microbiota in regulating whole-body cholesterol homeostasis and its response to a cholesterol-lowering drug, ezetimibe. PMID:26015368

  1. Microbiota prevents cholesterol loss from the body by regulating host gene expression in mice.

    PubMed

    Zhong, Chun-Yan; Sun, Wei-Wei; Ma, Yinyan; Zhu, Hongling; Yang, Pan; Wei, Hong; Zeng, Ben-Hua; Zhang, Qian; Liu, Yu; Li, Wen-Xia; Chen, Yixin; Yu, Liqing; Song, Zhi-Yuan

    2015-01-01

    We have previously observed that knockout of Niemann-Pick C1-Like 1 (NPC1L1), a cholesterol transporter essential for intestinal cholesterol absorption, reduces the output of dry stool in mice. As the food intake remains unaltered in NPC1L1-knockout (L1-KO) mice, we hypothesized that NPC1L1 deficiency may alter the gut microbiome to reduce stool output. Consistently, here we demonstrate that the phyla of fecal microbiota differ substantially between L1-KO mice and their wild-type controls. Germ-free (GF) mice have reduced stool output. Inhibition of NPC1L1 by its inhibitor ezetimibe reduces stool output in specific pathogen-free (SPF), but not GF mice. In addition, we show that GF versus SPF mice have reduced intestinal absorption and increased fecal excretion of cholesterol, particularly after treatment with ezetimibe. This negative balance of cholesterol in GF mice is associated with reduced plasma and hepatic cholesterol, and likely caused by reduced expression of NPC1L1 and increased expression of ABCG5 and ABCG8 in small intestine. Expression levels of other genes in intestine and liver largely reflect a state of cholesterol depletion and a decrease in intestinal sensing of bile acids. Altogether, our findings reveal a broad role of microbiota in regulating whole-body cholesterol homeostasis and its response to a cholesterol-lowering drug, ezetimibe. PMID:26015368

  2. Fluoxetine Prevents Respiratory Arrest without Enhancing Ventilation in DBA/1 Mice

    PubMed Central

    Zeng, Chang; Long, Xiaoyan; Cotten, Joseph F.; Forman, Stuart A.; Solt, Ken; Faingold, Carl L.; Feng, Hua-Jun

    2015-01-01

    Sudden unexpected death in epilepsy (SUDEP) is a fatal epileptic event. The DBA/1 mouse is a relevant animal model for study of SUDEP, as these mice exhibit seizure-induced respiratory arrest (S-IRA) leading to death, which has been observed in witnessed SUDEP patients. Fluoxetine, a selective serotonin (5-hydroxytryptamine or 5-HT) reuptake inhibitor (SSRI), reduces S-IRA in DBA/1 mice. Given that DBA/1 mice with S-IRA can be resuscitated using a ventilator, we hypothesized that breathing stimulants can prevent S-IRA and that fluoxetine prevents S-IRA by enhancing ventilation in these mice. Spontaneous respiratory function in anesthetized or awake DBA/1 mice was examined using non-invasive plethysmography before and after administering fluoxetine or breathing stimulants, doxapram and 5,6,7,8-tetrahydropyrido[4,3-d]pyrimidine (PK-THPP). The effects of these drugs on S-IRA in DBA/1 mice were tested. As reported previously, systemic administration of fluoxetine reduced S-IRA in awake DBA/1 mice, but fluoxetine in anesthetized and awake DBA/1 mice did not increase basal ventilation or the ventilatory response to 7% CO2. Both doxapram and PK-THPP increased ventilation in room air and in air + 7% CO2 in anesthetized DBA/1 mice. However, neither of the breathing stimulants reduced the incidence of S-IRA. Our studies confirm that fluoxetine reduces S-IRA in DBA/1 mice, but without enhancing basal ventilation in the absence of seizures. Although breathing stimulants increased ventilation in the absence of seizures, they were ineffective in reducing S-IRA, indicating that drug-induced increases in ventilation are insufficient to compensate for S-IRA in DBA/1 mice. PMID:25771493

  3. Gene expression profile in liver of hB1F transgenic mice

    PubMed Central

    Wang, Shui-Liang; Yang, Hua; Xie, You-Hua; Wang, Yuan; Li, Jian-Zhong; Wang, Long; Wang, Zhu-Gang; Fu, Ji-Liang

    2004-01-01

    AIM: To analyze the tissue morphologic phenotype and liver gene expression profile of hB1F transgenic mice. METHODS: Transgene expression was analyzed with RT-PCR and Western blotting. For one of the transgenic mouse lines, tissue expression pattern of the transgene was also examined with immunochemical methods. Pathological analysis was used to examine the tissue morphologic phenotype of established transgenic mice. The liver gene expression profile of transgenic mice was analyzed with microchip, and some of the differentially expressed genes were verified with RT-PCR. RESULTS: The expressions of hB1F were shown in livers from 6 of 7 transgenic mouse lines. The overexpression of hB1F transgene did not cause pathological changes. Expressions of three genes were up-regulated, while down-regulation was observed for 25 genes. CONCLUSION: The overexpression of hB1F transgene may cause changes of gene expression profiles in the liver of transgenic mice. PMID:15378783

  4. Nursing frequency alters circadian patterns of mammary gene expression in lactating mice

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Milking frequency impacts lactation in dairy cattle and in rodent models of lactation. The role of circadian gene expression in this process is unknown. The hypothesis tested was that changing nursing frequency alters the circadian patterns of mammary gene expression. Mid-lactation CD1 mice were stu...

  5. Differential Fmo3 Gene Expression in Various Liver Injury Models Involving Hepatic Oxidative Stress in Mice

    PubMed Central

    Rudraiah, Swetha; Moscovitz, Jamie E.; Donepudi, Ajay C.; Campion, Sarah N.; Slitt, Angela L.; Aleksunes, Lauren M.; Manautou, José E.

    2015-01-01

    Flavin-containing monooxygenase-3 (FMO3) catalyzes metabolic reactions similar to cytochrome P450 monooxygenase however, most metabolites of FMO3 are considered non-toxic. Recent findings in our laboratory demonstrated Fmo3gene induction following toxic acetaminophen (APAP) treatment in mice.The goal of this study was to evaluate Fmo3gene expression in diverseother mouse models of hepatic oxidative stress and injury. Fmo3 gene regulation by Nrf2 was also investigated using Nrf2 knockout (Nrf2 KO) mice. In our studies, male C57BL/6J mice were treated with toxic dosesof hepatotoxicants or underwent bile duct ligation (BDL, 10d). Hepatotoxicants included APAP (400 mg/kg, 24 to 72h), alpha-naphthylisothiocyanate (ANIT; 50 mg/kg, 2 to 48h), carbontetrachloride (CCl4;10 or 30 μL/kg, 24 and 48h) and allyl alcohol (AlOH; 30 or 60 mg/kg, 6 and 24h). Because oxidative stress activates nuclear factor (erythroid-derived 2)-like 2 (Nrf2), additional studies investigated Fmo3 gene regulation by Nrf2 using Nrf2 knockout (Nrf2 KO) mice. At appropriate time-points, blood and liver samples were collected for assessment of plasma alanine aminotransferase (ALT) activity, plasma and hepatic bile acid levels, as well as liver Fmo3 mRNA and protein expression. Fmo3 mRNA expression increased significantly by 43-fold at 12h after ANIT treatment,and this increase translates to a 4-fold change in protein levels. BDL also increased Fmo3 mRNA expression by 1899-fold, but with no change in protein levels. Treatment of mice with CCl4decreased liver Fmo3gene expression, whileno change in expression was detected with AlOH treatment. Nrf2 KO mice are more susceptible to APAP (400 mg/kg, 72h) treatment compared to their wild-type (WT) counterparts, which is evidenced by greater plasma ALT activity. Fmo3 mRNA and protein expression increased in Nrf2 KO mice after APAP treatment. Collectively, not all hepatotoxicantsthat produce oxidative stress alter Fmo3gene expression. Along with APAP, toxic ANIT

  6. Diosgenin-induced cognitive enhancement in normal mice is mediated by 1,25D3-MARRS

    NASA Astrophysics Data System (ADS)

    Tohda, Chihiro; Lee, Young-A.; Goto, Yukiori; Nemere, Ilka

    2013-12-01

    We previously reported that diosgenin, a plant-derived steroidal sapogenin, improved memory and reduced axonal degeneration in an Alzheimer's disease mouse model. Diosgenin directly activated the membrane-associated rapid response steroid-binding receptor (1,25D3-MARRS) in neurons. However, 1,25D3-MARRS-mediated diosgenin signaling was only shown in vitro in the previous study. Here, we aimed to obtain in vivo evidence showing that diosgenin signaling is mediated by 1,25D3-MARRS in the mouse brain. Diosgenin treatment in normal mice enhanced object recognition memory and spike firing and cross-correlation in the medial prefrontal cortex and hippocampal CA1. In diosgenin-treated mice, axonal density and c-Fos expression was increased in the medial prefrontal and perirhinal cortices, suggesting that neuronal network activation may be enhanced. The diosgenin-induced memory enhancement and axonal growth were completely inhibited by co-treatment with a neutralizing antibody for 1,25D3-MARRS. Our in vivo data indicate that diosgenin is a memory-enhancing drug and that enhancement by diosgenin is mediated by 1,25D3-MARRS-triggered axonal growth.

  7. Ozonated sesame oil enhances cutaneous wound healing in SKH1 mice.

    PubMed

    Valacchi, Giuseppe; Lim, Yunsook; Belmonte, Giuseppe; Miracco, Clelia; Zanardi, Iacopo; Bocci, Velio; Travagli, Valter

    2011-01-01

    Ozone is well recognized as a bactericidal agent and its beneficial effect on wound healing could be a consequence of this property. Because ozone itself does not penetrate the cells but immediately reacts with polyunsaturated fatty acids, its effects should be the results of oxidative reaction. For this reason, ozonated oils could be a way to deliver ozone messengers to the skin. This paper evaluated the therapeutic effects of three different grades of ozonated sesame oil in acute cutaneous wounds made in the skin of SKH1 mice. Specifically, wound closure rate, histological parameters, and the level of key proteins such as vascular endothelial growth factors and cyclin D1 have been analyzed in relation to the peroxide level present in the ozonated oil. Treatment with moderately ozonated sesame oil--expressed as peroxide value about 1,500)--has a faster wound closure rate in the first 7 days than treatment with oil containing either lower or higher peroxide value, and even with controls. Moreover, under the same treatment, an earlier and higher response of cells involved in wound repair, a higher angiogenesis, as well as an enhanced vascular endothelial growth factors and cyclin D1 expression were observed. The present study shows the validity of ozonated sesame oil in cutaneous wound healing and emphasizes the importance of the ozonation grade. PMID:21134039

  8. Stem Cell Replacement Improves Expression of SMP30 in db/db Mice

    PubMed Central

    Li, Ming; Guo, Kequan; Taketani, Shigeru; Adachi, Yasushi; Ikehara, Susumu

    2015-01-01

    We have previously reported that replacing bone marrow stem cells may improve hyperglycemia and oxidative stress in db/db mice, a type 2 diabetic mouse model. Senescence marker protein 30 (SMP30) is an antioxidant protein that decreases with aging. However, it has not been clear whether SMP30 decreases in the livers of obese mice, and whether stem cell replacement would improve SMP30 expression in the liver. Bone marrow stem cells of db/db mice were replaced with the bone marrow stem cells of C57BL/6 mice. Plasma cytokine and insulin levels were measured, and glycogen content, expression of SMP30, and fibrosis in the liver were assessed. Our results showed that stem cell replacement increased the expression of SMP30 in the liver, resulting from decreased plasma inflammation cytokines and hyperinsulinemia in db/db mice. This is the first report that stem cell replacement increased the expression of SMP30 in the liver, and may help prevent fibrosis in the liver of db/db mice. PMID:26694363

  9. Enhanced Sympathetic Activity in Mice with Brown Adipose Tissue Transplantation (TransBATation)

    PubMed Central

    Zhu, Zheng; Spicer, Elizabeth G; Gavini, Chaitanya K; Goudjo-Ako, Ashley J; Novak, Colleen M; Shi, Haifei

    2014-01-01

    Brown adipose tissue (BAT) burns calories to produce heat, and is thus relevant to energy balance. Interscapular BAT (IBAT) of donor mice was transplanted into recipient mice (transBATation). To test whether transBATation counteracts high-fat diet (HFD)-induced obesity, some sham-operated and recipient mice were fed a HFD (HFD-sham, HFD-trans) while others remained on a standard chow (chow-sham, chow-trans). HFD-trans mice had lower body weight and fat, greater energy expenditure, but similar caloric intake compared with HFD-sham mice. We hypothesized that HFD-trans mice had elevated sympathetic activity compared with HFD-sham mice, contributing to increased energy expenditure and fuel mobilization. This was supported by findings that HFD-trans mice had greater energy expenditure during a norepinephrine challenge test and higher core temperatures after cold exposure than did HFD-sham mice, implicating enhanced whole-body metabolic response and elevated sympathetic activity. Additionally, transBATation selectively increased sympathetic drive to some, but not all, white adipose tissue depots and skeletal muscles, as well as the endogenous IBAT, heart, and liver. Collectively, transBATation confers resistance to HFD-induced obesity via increase in whole-body sympathetic activity, and differential activation of sympathetic drive to some of the tissues involved in energy expenditure and fuel mobilization. PMID:24291381

  10. Sleep fragmentation during late gestation induces metabolic perturbations and epigenetic changes in adiponectin gene expression in male adult offspring mice.

    PubMed

    Khalyfa, Abdelnaby; Mutskov, Vesco; Carreras, Alba; Khalyfa, Ahamed A; Hakim, Fahed; Gozal, David

    2014-10-01

    Sleep fragmentation (SF) is a common condition among pregnant women, particularly during late gestation. Gestational perturbations promote the emergence of adiposity and metabolic disease risk in offspring, most likely through epigenetic modifications. Adiponectin (AdipoQ) expression inversely correlates with obesity and insulin resistance. The effects of SF during late gestation on metabolic function and AdipoQ expression in visceral white adipose tissue (VWAT) of offspring mice are unknown. Male offspring mice were assessed at 24 weeks after dams were exposed to SF or control sleep during late gestation. Increased food intake, body weight, VWAT mass, and insulin resistance, with reductions in AdipoQ expression in VWAT, emerged in SF offspring. Increased DNMT3a and -b and global DNA methylation and reduced histone acetyltransferase activity and TET1, -2, and -3 expression were detected in VWAT of SF offspring. Reductions in 5-hydroxymethylcytosine and H3K4m3 and an increase in DNA 5-methylcytosine and H3K9m2 in the promoter and enhancer regions of AdipoQ emerged in adipocytes from VWAT and correlated with AdipoQ expression. SF during late gestation induces epigenetic modifications in AdipoQ in male offspring mouse VWAT adipocytes along with a metabolic syndrome-like phenotype. Thus, altered gestational environments elicited by SF impose the emergence of adverse, long-lasting metabolic consequences in the next generation. PMID:24812424

  11. A Novel mouse model of enhanced proteostasis: Full-length human heat shock factor 1 transgenic mice

    SciTech Connect

    Pierce, Anson; Wei, Rochelle; Halade, Dipti; Yoo, Si-Eun; Ran, Qitao; Richardson, Arlan

    2010-11-05

    Research highlights: {yields} Development of mouse overexpressing native human HSF1 in all tissues including CNS. {yields} HSF1 overexpression enhances heat shock response at whole-animal and cellular level. {yields} HSF1 overexpression protects from polyglutamine toxicity and favors aggresomes. {yields} HSF1 overexpression enhances proteostasis at the whole-animal and cellular level. -- Abstract: The heat shock response (HSR) is controlled by the master transcriptional regulator heat shock factor 1 (HSF1). HSF1 maintains proteostasis and resistance to stress through production of heat shock proteins (HSPs). No transgenic model exists that overexpresses HSF1 in tissues of the central nervous system (CNS). We generated a transgenic mouse overexpressing full-length non-mutant HSF1 and observed a 2-4-fold increase in HSF1 mRNA and protein expression in all tissues studied of HSF1 transgenic (HSF1{sup +/0}) mice compared to wild type (WT) littermates, including several regions of the CNS. Basal expression of HSP70 and 90 showed only mild tissue-specific changes; however, in response to forced exercise, the skeletal muscle HSR was more elevated in HSF1{sup +/0} mice compared to WT littermates and in fibroblasts following heat shock, as indicated by levels of inducible HSP70 mRNA and protein. HSF1{sup +/0} cells elicited a significantly more robust HSR in response to expression of the 82 repeat polyglutamine-YFP fusion construct (Q82YFP) and maintained proteasome-dependent processing of Q82YFP compared to WT fibroblasts. Overexpression of HSF1 was associated with fewer, but larger Q82YFP aggregates resembling aggresomes in HSF1{sup +/0} cells, and increased viability. Therefore, our data demonstrate that tissues and cells from mice overexpressing full-length non-mutant HSF1 exhibit enhanced proteostasis.

  12. Enhanced Immune Response to DNA Vaccine Encoding Bacillus anthracis PA-D4 Protects Mice against Anthrax Spore Challenge

    PubMed Central

    Kim, Na Young; Chang, Dong Suk; Kim, Yeonsu; Kim, Chang Hwan; Hur, Gyeung Haeng; Yang, Jai Myung; Shin, Sungho

    2015-01-01

    Anthrax has long been considered the most probable bioweapon-induced disease. The protective antigen (PA) of Bacillus anthracis plays a crucial role in the pathogenesis of anthrax. In the current study, we evaluated the efficiency of a genetic vaccination with the fourth domain (D4) of PA, which is responsible for initial binding of the anthrax toxin to the cellular receptor. The eukaryotic expression vector was designed with the immunoglobulin M (IgM) signal sequence encoding for PA-D4, which contains codon-optimized genes. The expression and secretion of recombinant protein was confirmed in vitro in 293T cells transfected with plasmid and detected by western blotting, confocal microscopy, and enzyme-linked immunosorbent assay (ELISA). The results revealed that PA-D4 protein can be efficiently expressed and secreted at high levels into the culture medium. When plasmid DNA was given intramuscularly to mice, a significant PA-D4-specific antibody response was induced. Importantly, high titers of antibodies were maintained for nearly 1 year. Furthermore, incorporation of the SV40 enhancer in the plasmid DNA resulted in approximately a 15-fold increase in serum antibody levels in comparison with the plasmid without enhancer. The antibodies produced were predominantly the immunoglobulin G2 (IgG2) type, indicating the predominance of the Th1 response. In addition, splenocytes collected from immunized mice produced PA-D4-specific interferon gamma (IFN-γ). The biodistribution study showed that plasmid DNA was detected in most organs and it rapidly cleared from the injection site. Finally, DNA vaccination with electroporation induced a significant increase in immunogenicity and successfully protected the mice against anthrax spore challenge. Our approach to enhancing the immune response contributes to the development of DNA vaccines against anthrax and other biothreats. PMID:26430894

  13. Enhanced Immune Response to DNA Vaccine Encoding Bacillus anthracis PA-D4 Protects Mice against Anthrax Spore Challenge.

    PubMed

    Kim, Na Young; Chang, Dong Suk; Kim, Yeonsu; Kim, Chang Hwan; Hur, Gyeung Haeng; Yang, Jai Myung; Shin, Sungho

    2015-01-01

    Anthrax has long been considered the most probable bioweapon-induced disease. The protective antigen (PA) of Bacillus anthracis plays a crucial role in the pathogenesis of anthrax. In the current study, we evaluated the efficiency of a genetic vaccination with the fourth domain (D4) of PA, which is responsible for initial binding of the anthrax toxin to the cellular receptor. The eukaryotic expression vector was designed with the immunoglobulin M (IgM) signal sequence encoding for PA-D4, which contains codon-optimized genes. The expression and secretion of recombinant protein was confirmed in vitro in 293T cells transfected with plasmid and detected by western blotting, confocal microscopy, and enzyme-linked immunosorbent assay (ELISA). The results revealed that PA-D4 protein can be efficiently expressed and secreted at high levels into the culture medium. When plasmid DNA was given intramuscularly to mice, a significant PA-D4-specific antibody response was induced. Importantly, high titers of antibodies were maintained for nearly 1 year. Furthermore, incorporation of the SV40 enhancer in the plasmid DNA resulted in approximately a 15-fold increase in serum antibody levels in comparison with the plasmid without enhancer. The antibodies produced were predominantly the immunoglobulin G2 (IgG2) type, indicating the predominance of the Th1 response. In addition, splenocytes collected from immunized mice produced PA-D4-specific interferon gamma (IFN-γ). The biodistribution study showed that plasmid DNA was detected in most organs and it rapidly cleared from the injection site. Finally, DNA vaccination with electroporation induced a significant increase in immunogenicity and successfully protected the mice against anthrax spore challenge. Our approach to enhancing the immune response contributes to the development of DNA vaccines against anthrax and other biothreats. PMID:26430894

  14. Neuropeptide S facilitates mice olfactory function through activation of cognate receptor-expressing neurons in the olfactory cortex.

    PubMed

    Shao, Yu-Feng; Zhao, Peng; Dong, Chao-Yu; Li, Jing; Kong, Xiang-Pan; Wang, Hai-Liang; Dai, Li-Rong; Hou, Yi-Ping

    2013-01-01

    Neuropeptide S (NPS) is a newly identified neuromodulator located in the brainstem and regulates various biological functions by selectively activating the NPS receptors (NPSR). High level expression of NPSR mRNA in the olfactory cortex suggests that NPS-NPSR system might be involved in the regulation of olfactory function. The present study was undertaken to investigate the effects of intracerebroventricular (i.c.v.) injection of NPS or co-injection of NPSR antagonist on the olfactory behaviors, food intake, and c-Fos expression in olfactory cortex in mice. In addition, dual-immunofluorescence was employed to identify NPS-induced Fos immunereactive (-ir) neurons that also bear NPSR. NPS (0.1-1 nmol) i.c.v. injection significantly reduced the latency to find the buried food, and increased olfactory differentiation of different odors and the total sniffing time spent in olfactory habituation/dishabituation tasks. NPS facilitated olfactory ability most at the dose of 0.5 nmol, which could be blocked by co-injection of 40 nmol NPSR antagonist [D-Val(5)]NPS. NPS administration dose-dependently inhibited food intake in fasted mice. Ex-vivo c-Fos and NPSR immunohistochemistry in the olfactory cortex revealed that, as compared with vehicle-treated mice, NPS markedly enhanced c-Fos expression in the anterior olfactory nucleus (AON), piriform cortex (Pir), ventral tenia tecta (VTT), the anterior cortical amygdaloid nucleus (ACo) and lateral entorhinal cortex (LEnt). The percentage of Fos-ir neurons that also express NPSR were 88.5% and 98.1% in the AON and Pir, respectively. The present findings demonstrated that NPS, via selective activation of the neurons bearing NPSR in the olfactory cortex, facilitates olfactory function in mice. PMID:23614017

  15. Enhanced expression of adenovirus transforming proteins.

    PubMed Central

    Gaynor, R B; Tsukamoto, A; Montell, C; Berk, A J

    1982-01-01

    Proteins encoded in regions EIA and EIB of human adenoviruses cause transformation of rodent cells. One protein from EIA also stimulates transcription of other early regions at early times in a productive infection. In the past, direct analysis of these proteins synthesized in vivo has been difficult because of the low levels produced in both transformed cells and productively infected cells. We present a simple method which leads to expression of EIA and EIB mRNAs and proteins at 30-fold greater levels than those observed during the early phase of a standard productive infection. Under these conditions, these proteins are among the most prominent translation products of infected cells. This allowed direct visualization of EIA and EIB proteins on two-dimensional gels of pulse-labeled total cell protein. Experiments with EIA and EIB mutants confirm that the identified proteins are indeed encoded in these regions. Two EIA proteins are observed, one translated from each of the major early EIA mRNAs. Both of these EIA proteins are phosphorylated. Images PMID:7143568

  16. Enhanced Tumor Formation in Mice Heterozygous for Blm Mutation

    NASA Astrophysics Data System (ADS)

    Heppner Goss, Kathleen; Risinger, Mary A.; Kordich, Jennifer J.; Sanz, Maureen M.; Straughen, Joel E.; Slovek, Lisa E.; Capobianco, Anthony J.; German, James; Boivin, Gregory P.; Groden, Joanna

    2002-09-01

    Persons with the autosomal recessive disorder Bloom syndrome are predisposed to cancers of many types due to loss-of-function mutations in the BLM gene, which encodes a recQ-like helicase. Here we show that mice heterozygous for a targeted null mutation of Blm, the murine homolog of BLM, develop lymphoma earlier than wild-type littermates in response to challenge with murine leukemia virus and develop twice the number of intestinal tumors when crossed with mice carrying a mutation in the Apctumor suppressor. These observations indicate that Blm is a modifier of tumor formation in the mouse and that Blm haploinsufficiency is associated with tumor predisposition, a finding with important implications for cancer risk in humans.

  17. Suppressed or enhanced antibody responses in vitro after BCG treatment of mice: importance of BCG viability.

    PubMed Central

    Brown, C A; Brown, I N; Sljivić, V S

    1979-01-01

    Mycobacterium bovis, BCG, is known to be capable of either enhancing or suppressing various immune responses. Using a standard technique and number of organisms, some of the parameters predetermining whether enhancement or supression will occur have been investigated. Dead BCG given intravenously into mice caused an enhancement of the antibody response in vitro to sheep erythrocytes. In contrast, the same number of viable organisms caused suppression if given intravenously but enhancement if given subcutaneously. The inclusion of 25% or more killed organisms in an intravenous inoculum of fully viable organisms changed suppression to enhancement. Treatment of BCG infected mice with streptomycin lessened the suppression but did not change it to enhancement. The possible causes of suppression are discussed. PMID:391696

  18. Changes in kinesin expression in the CNS of mice with dynein heavy chain 1 mutation.

    PubMed

    Kuźma-Kozakiewicz, Magdalena; Kaźmierczak, Beata; Usarek, Ewa; Barańczyk-Kuźma, Anna

    2013-01-01

    Dysfunction of fast axonal transport, vital for motor neurons, may lead to neurodegeneration. Anterograde transport is mediated by N-kinesins (KIFs), while retrograde transport by dynein 1 and, to a minor extent, by C-kinesins. In our earlier studies we observed changes in expression of N- and C-kinesins (KIF5A, 5C, C2) in G93ASOD1-linked mouse model of motor neuron degeneration. In the present work we analyze the profile of expression of the same kinesins in mice with a dynein 1 heavy chain mutation (Dync1h1, called Cra1), presenting similar clinical symptoms, and in Cra1/SOD1 mice with milder disease progression than SOD1 transgenics. We found significantly higher levels of mRNA for KIF5A and KIF5C but not the KIFC2 in the frontal cortex of symptomatic Cra1/+ mice (aged 365 days) compared to the wild-type controls. No changes in kinesin expression were found in the spinal cord of any age group and only mild changes in the hippocampus. The expression of kinesins in the cerebellum of the presymptomatic and symptomatic mice (aged 140 and 365 days, respectively) was much lower than in age-matched controls. In Cra1/SOD1 mice the changes in KIFs expression were similar or more severe than in the Cra1/+ groups, and they also appeared in the spinal cord. Thus, in mice with the Dync1h1 mutation, which impairs dynein 1-dependent retrograde transport, expression of kinesin mRNA is affected in various structures of the CNS and the changes are similar or milder than in mice with double Dync1h1/hSOD1G93A mutations. PMID:23460941

  19. Enhanced Susceptibility to Citrobacter rodentium Infection in MicroRNA-155-Deficient Mice

    PubMed Central

    John, Victoria; Walker, Alan W.; Hill, Jennifer L.; Abreu-Goodger, Cei; Hale, Christine; Goulding, David; Lawley, Trevor D.; Mastroeni, Pietro; Frankel, Gadi; Enright, Anton J.; Vigorito, Elena; Dougan, Gordon

    2013-01-01

    MicroRNAs (miRNAs) are small noncoding molecules that control gene expression posttranscriptionally, with microRNA-155 (miR-155) one of the first to be implicated in immune regulation. Here, we show that miR-155-deficient mice are less able to eradicate a mucosal Citrobacter rodentium infection than wild-type C57BL/6 mice. miR-155-deficient mice exhibited prolonged colonization associated with a higher C. rodentium burden in gastrointestinal tissue and spread into systemic tissues. Germinal center formation and humoral immune responses against C. rodentium were severely impaired in infected miR-155-deficient mice. A similarly susceptible phenotype was observed in μMT mice reconstituted with miR-155-deficient B cells, indicating that miR-155 is required intrinsically for mediating protection against this predominantly luminal bacterial pathogen. PMID:23264052

  20. Sasa borealis Extract Efficiently Enhanced Swimming Capacity by Improving Energy Metabolism and the Antioxidant Defense System in Mice.

    PubMed

    You, Yanghee; Kim, Kyungmi; Yoon, Ho-Geun; Choi, Kyung-Chul; Lee, Yoo-Hyun; Lee, Jeongmin; Jun, Woojin

    2015-01-01

    This study was conducted to determine the effects of 50% ethanolic extract from Sasa borealis leaves (SBE) on swimming capacity and oxidative metabolism in mice. The mice were divided into 2 groups with similar swimming times and body weights; Ex-Control and Ex-SBE were orally administered with distilled water and 250 mg/kg body weight/d of SBE. Exhaustive swimming times were prolonged by 1.5-fold in the Ex-SBE group compared to the Ex-Control. The Ex-SBE group displayed lower lactate and higher non-esterified fatty acid levels 15 min after swimming and the hepatic and muscle glycogen levels were significantly higher than that in the Ex-Control. SBE potentially enhanced mRNA expression of citrate synthase (CS), carnitine palmitoyltransferase (CPT-1), and β-hydroxyacyl coenzyme A dehydrogenase (β-HAD) in skeletal muscle. The activities and mRNA expression of catalase (CAT), glutathione peroxidase (GPx), and superoxide dismutase (SOD) were elevated in the Ex-SBE compared with the Ex-Control after exhaustive swimming. These results suggest that SBE might be used as an effective agent to enhance swimming capacity by utilization of energy substrates and might ameliorate physical exhaustion by facilitating energy-generating metabolic genes and enhancing endogenous antioxidants. PMID:26875491

  1. Altered enteroendocrine cell expression in T cell receptor alpha chain knock-out mice.

    PubMed

    Rubin, D C; Zhang, H; Qian, P; Lorenz, R G; Hutton, K; Peters, M G

    2000-10-15

    Mice lacking T cell receptor alpha chain (TCRalpha(-/-)) develop inflammation of the colon. We have examined the effect of this inflammation on the colonic epithelium by studying markers of epithelial cuff, enteroendocrine, and immune cell differentiation. Using immunohistochemical techniques, colons were compared in normal C57/BL6 and murine TCR alpha(-/-) mice aged 2 and 3 weeks and 3-11 months. TCR alpha(-/-) mice aged 3-11 months had histologic evidence of inflammation with increased expression of CD45, CD4+, CD8+, and B220+ cells and a decrease in expression of IgA+ cells. There was a decrease in the number of cholecystokinin, serotonin, and neurotensin enteroendocrine expressing cells in the colon of TCR alpha(-/-) mice. These changes were not present in 2-3-week-old suckling/weaning mice. In contrast, peptide tyrosine tyrosine (PYY), glucagon-like peptide-1, and gastrin expression did not change and small intestinal enteroendocrine cells remained unaltered. The change in colonic enteroendocrine cell expression appears to be a specific response, since only a subset of these cells was altered, and the epithelium was intact by histologic analysis. The absence of functional T cells in TCR alpha(-/-) colon has a marked effect on differentiation of a specific subpopulation of enteroendocrine cells, prior to loss of integrity of the epithelium. PMID:11054861

  2. Dual mechanisms of rapid expression of anxiety-related behavior in pilocarpine-treated epileptic mice.

    PubMed

    Otsuka, Shintaro; Ohkido, Taro; Itakura, Makoto; Watanabe, Shigeru; Yamamori, Saori; Iida, Yuuki; Saito, Masanori; Miyaoka, Hitoshi; Takahashi, Masami

    2016-07-01

    A mouse model of epilepsy was generated by inducing status epilepticus (SE) for either 1.5 or 4.5h with pilocarpine to study anxiety-related behaviors, changes in the electroencephalogram of the cerebral cortex and hippocampus, and expression of hippocampal proteins. The viability and rate of success of SE induction were high in C57BL/6N mice but not in C57BL/6J mice. C57BL/6N mice were immotile during the first 2days after SE; however, by the third day, most mice were recovered and exhibited strong anxiety-related behaviors in response to the light/dark preference test and open field test. There was a striking difference in the temporal appearance of anxiety-related behavior between the two SE durations: 1.5h SE mice exhibited strong anxiety-related behavior 3days after SE that gradually attenuated over the next few weeks, whereas 4.5h SE mice exhibited strong anxiety-related behavior 3days after SE that persisted even at nearly 1year after SE. Mice receiving both SE durations exhibited generalized seizures (GS) after SE; however, there was a marked difference in the timing and duration of GS appearance. Mice in the 4.5h SE group exhibited spontaneous GS from 4days to at least 96days after SE. In contrast, mice in the 1.5h SE group exhibited GS only within the first several days after SE; however, epileptic spike clusters continuously appeared in the cerebral cortex and hippocampus for up to twelve days after SE. Among the hippocampal proteins tested, only brain derived-neurotrophic factor (BDNF) exhibited altered expression in parallel with anxiety-related behavior. These results showed the possibility that BDNF expression in the hippocampus might cause anxiety-related behavior in adulthood. PMID:27132018

  3. Hyaluronidase Expression by an Oncolytic Adenovirus Enhances Its Intratumoral Spread and Suppresses Tumor Growth

    PubMed Central

    Guedan, Sonia; Rojas, Juan José; Gros, Alena; Mercade, Elena; Cascallo, Manel; Alemany, Ramon

    2010-01-01

    Successful virotherapy requires efficient virus spread within tumors. We tested whether the expression of hyaluronidase, an enzyme which dissociates the extracellular matrix (ECM), could enhance the intratumoral distribution of an oncolytic adenovirus and improve its therapeutic activity. As a proof of concept, we demonstrated that intratumoral coadministration of hyaluronidase in mice-bearing tumor xenografts improves the antitumor activity of an oncolytic adenovirus. Next, we constructed a replication-competent adenovirus expressing a soluble form of the human sperm hyaluronidase (PH20) under the control of the major late promoter (MLP) (AdwtRGD-PH20). Intratumoral treatment of human melanoma xenografts with AdwtRGD-PH20 resulted in degradation of hyaluronan (HA), enhanced viral distribution, and induced tumor regression in all treated tumors. Finally, the PH20 cDNA was inserted in an oncolytic adenovirus that selectively kills pRb pathway-defective tumor cells. The antitumoral activity of the novel oncolytic adenovirus expressing PH20 (ICOVIR17) was compared to that of the parental virus ICOVIR15. ICOVIR17 showed more antitumor efficacy following intratumoral and systemic administration in mice with prestablished tumors, along with an improved spread of the virus within the tumor. Importantly, a single intravenous dose of ICOVIR17 induced tumor regression in 60% of treated tumors. These results indicate that ICOVIR17 is a promising candidate for clinical testing. PMID:20442708

  4. Changes in Gene Expression Foreshadow Diet-Induced Obesity in Genetically Identical Mice

    PubMed Central

    Koza, Robert A; Nikonova, Larissa; Hogan, Jessica; Rim, Jong-Seop; Mendoza, Tamra; Faulk, Christopher; Skaf, Jihad; Kozak, Leslie P

    2006-01-01

    High phenotypic variation in diet-induced obesity in male C57BL/6J inbred mice suggests a molecular model to investigate non-genetic mechanisms of obesity. Feeding mice a high-fat diet beginning at 8 wk of age resulted in a 4-fold difference in adiposity. The phenotypes of mice characteristic of high or low gainers were evident by 6 wk of age, when mice were still on a low-fat diet; they were amplified after being switched to the high-fat diet and persisted even after the obesogenic protocol was interrupted with a calorically restricted, low-fat chow diet. Accordingly, susceptibility to diet-induced obesity in genetically identical mice is a stable phenotype that can be detected in mice shortly after weaning. Chronologically, differences in adiposity preceded those of feeding efficiency and food intake, suggesting that observed difference in leptin secretion is a factor in determining phenotypes related to food intake. Gene expression analyses of adipose tissue and hypothalamus from mice with low and high weight gain, by microarray and qRT-PCR, showed major changes in the expression of genes of Wnt signaling and tissue re-modeling in adipose tissue. In particular, elevated expression of SFRP5, an inhibitor of Wnt signaling, the imprinted gene MEST and BMP3 may be causally linked to fat mass expansion, since differences in gene expression observed in biopsies of epididymal fat at 7 wk of age (before the high-fat diet) correlated with adiposity after 8 wk on a high-fat diet. We propose that C57BL/6J mice have the phenotypic characteristics suitable for a model to investigate epigenetic mechanisms within adipose tissue that underlie diet-induced obesity. PMID:16733553

  5. Exercise-induced increase in IL-6 level enhances GLUT4 expression and insulin sensitivity in mouse skeletal muscle.

    PubMed

    Ikeda, Shin-Ichi; Tamura, Yoshifumi; Kakehi, Saori; Sanada, Hiromi; Kawamori, Ryuzo; Watada, Hirotaka

    2016-05-13

    A single bout of exercise is known to increase the insulin sensitivity of skeletal muscle; however, the underlying mechanism of this phenomenon is not fully understood. Because a single bout of exercise induces a transient increase in blood interleukin-6 (IL-6) level, we hypothesized that the enhancement of insulin sensitivity after a single bout of exercise in skeletal muscle is mediated at least in part through IL-6-dependent mechanisms. To test this hypothesis, C57BL6J mice were intravenously injected with normal IgG or an IL-6 neutralizing antibody before exercise. Twenty-four hours after a single bout of exercise, the plantaris muscle was harvested to measure insulin sensitivity and glucose transporter (GLUT)-4 expression levels by ex-vivo insulin-stimulated 2-deoxyglucose (2-DG) uptake and Western blotting, respectively. Compared with sedentary mice, mice that performed exercise showed enhanced IL-6 concentration, insulin-stimulated 2-DG uptake, and GLUT-4 expression in the plantaris muscle. The enhanced insulin sensitivity and GLUT4 expression were canceled by injection of the IL-6 neutralizing antibody before exercise. In addition, IL-6 injection increased GLUT4 expression, both in the plantaris muscle and the soleus muscle in C57BL6J mice. Furthermore, a short period of incubation with IL-6 increased GLUT4 expression in differentiated C2C12 myotubes. In summary, these results suggested that IL-6 increased GLUT4 expression in muscle and that this phenomenon may play a role in the post-exercise enhancement of insulin sensitivity in skeletal muscle. PMID:27040770

  6. Acute stress blocks the caffeine-induced enhancement of contextual memory retrieval in mice.

    PubMed

    Pierard, Chistophe; Krazem, Ali; Henkous, Nadia; Decorte, Laurence; Béracochéa, Daniel

    2015-08-15

    This study investigated in mice the dose-effect of caffeine on memory retrieval in non-stress and stress conditions. C57 Bl/6 Jico mice learned two consecutive discriminations (D1 and D2) in a four-hole board which involved either distinct contextual (CSD) or similar contextual (SSD) cues. All mice received an i.p. injection of vehicle or caffeine (8, 16 or 32mg/kg) 30min before the test session. Results showed that in non-stress conditions, the 16mg/kg caffeine dose induced a significant enhancement of D1 performance in CSD but not in SSD. Hence, we studied the effect of an acute stress (electric footshocks) administered 15min before the test session on D1 performance in caffeine-treated mice. Results showed that stress significantly decreased D1 performance in vehicle-treated controls and the memory-enhancing effect induced by the 16mg/kg caffeine dose in non-stress condition is no longer observed. Interestingly, whereas caffeine-treated mice exhibited weaker concentrations of plasma corticosterone as compared to vehicles in non-stress condition, stress significantly increased plasma corticosterone concentrations in caffeine-treated mice which reached similar level to that of controls. Overall, the acute stress blocked both the endocrinological and memory retrieval enhancing effects of caffeine. PMID:25934571

  7. Enhanced inhibitory neurotransmission in the cerebellar cortex of Atp1a3-deficient heterozygous mice

    PubMed Central

    Ikeda, Keiko; Satake, Shin'Ichiro; Onaka, Tatsushi; Sugimoto, Hiroki; Takeda, Naoki; Imoto, Keiji; Kawakami, Kiyoshi

    2013-01-01

    Dystonia is characterized by excessive involuntary and prolonged simultaneous contractions of both agonist and antagonist muscles. Although the basal ganglia have long been proposed as the primary region, recent studies indicated that the cerebellum also plays a key role in the expression of dystonia. One hereditary form of dystonia, rapid-onset dystonia with parkinsonism (RDP), is caused by loss of function mutations of the gene for the Na pump α3 subunit (ATP1A3). Little information is available on the affected brain regions and mechanism for dystonia by the mutations in RDP. The Na pump is composed of α and β subunits and maintains ionic gradients of Na+ and K+ across the cell membrane. The gradients are utilized for neurotransmitter reuptake and their alteration modulates neural excitability. To provide insight into the molecular aetiology of RDP, we generated and analysed knockout heterozygous mice (Atp1a3+/−). Atp1a3+/− showed increased symptoms of dystonia that is induced by kainate injection into the cerebellar vermis. Atp1a3 mRNA was highly expressed in Purkinje cells and molecular-layer interneurons, and its product was concentrated at Purkinje cell soma, the site of abundant vesicular γ-aminobutyric acid transporter (VGAT) signal, suggesting the presynaptic localization of the α3 subunit in the inhibitory synapse. Electrophysiological studies showed that the inhibitory neurotransmission at molecular-layer interneuron–Purkinje cell synapses was enhanced in Atp1a3+/− cerebellar cortex, and that the enhancement originated via a presynaptic mechanism. Our results shed light on the role of Atp1a3 in the inhibitory synapse, and potential involvement of inhibitory synaptic dysfunction for the pathophysiology of dystonia. PMID:23652595

  8. Gene Expression Profiles of Main Olfactory Epithelium in Adenylyl Cyclase 3 Knockout Mice

    PubMed Central

    Wang, Zhenshan; Zhou, Yanfen; Luo, Yingtao; Zhang, Jing; Zhai, Yunpeng; Yang, Dong; Zhang, Zhe; Li, Yongchao; Storm, Daniel R.; Ma, Runlin Z.

    2015-01-01

    Adenylyl Cyclase 3 (AC3) plays an important role in the olfactory sensation-signaling pathway in mice. AC3 deficiency leads to defects in olfaction. However, it is still unknown whether AC3 deficiency affects gene expression or olfactory signal transduction pathways within the main olfactory epithelium (MOE). In this study, gene microarrays were used to screen differentially expressed genes in MOE from AC3 knockout (AC3−/−) and wild-type (AC3+/+) mice. The differentially expressed genes identified were subjected to bioinformatic analysis and verified by qRT-PCR. Gene expression in the MOE from AC3−/− mice was significantly altered, compared to AC3+/+ mice. Of the 41266 gene probes, 3379 had greater than 2-fold fold change in expression levels between AC3−/− and AC3+/+ mice, accounting for 8% of the total gene probes. Of these genes, 1391 were up regulated, and 1988 were down regulated, including 425 olfactory receptor genes, 99 genes that are specifically expressed in the immature olfactory neurons, 305 genes that are specifically expressed in the mature olfactory neurons, and 155 genes that are involved in epigenetic regulation. Quantitative RT-PCR verification of the differentially expressed epigenetic regulation related genes, olfactory receptors, ion transporter related genes, neuron development and differentiation related genes, lipid metabolism and membrane protein transport etc. related genes showed that P75NTR, Hinfp, Gadd45b, and Tet3 were significantly up-regulated, while Olfr370, Olfr1414, Olfr1208, Golf, Faim2, Tsg101, Mapk10, Actl6b, H2BE, ATF5, Kirrrel2, OMP, Drd2 etc. were significantly down-regulated. In summary, AC3 may play a role in proximal olfactory signaling and play a role in the regulation of differentially expressed genes in mouse MOE. PMID:26633363

  9. Skeletal Phenotype of Transgenic Mice Expressing the Beta1 Integrin Cytoplasmic Tail In Osteoblasts

    NASA Technical Reports Server (NTRS)

    Globus, R. K.; vanderMeulen, M. C. H.; Damsky, D.; Kim, J.-B.; Amblard, D.; Amblard, D.; Nishimura, Y.; Almeida, E.; Iwaniec, U. T.; Wronski, T. J.; Dalton, Bonnie (Technical Monitor)

    2002-01-01

    To define the physiologic role of beta1 integrin in bone formation and mechanical loading, transgenic mice were generated by expressing the cytoplasmic tall and transmembrane domain of Beta1 integrin under the control of the osteocalcin promoter. In cultured cells, this truncated fragment of Beta1 can act as a dominant negative. Previously, the matrix of calvariae was shown to be abnormal in transgenic (TG) compared to wildtype (WT) mice. In this study, we analyzed appendicular bone in TG and WT, male and female mice at 14, 35, 63, 90 and 365 days old (n=8-12/gp). To assess beta1 integrin function in mechanical loading, a pilot study using hindlimb unloading by tail suspension was performed. 35d old TG and WT females were hindlimb unloaded for 4 wks (n=3-5). Body mass, bone mineral content, histomorphometric (distal femur) and biomechanical parameters were analyzed. Statistical significance (P less than.05) was defined by ANOVA using the Tukey-Kramer post-hoc test. We confirmed transgene expression by immunoprecipitating then immunoblotting bone lysates using an antibody against the beta1 tail. Body masses of TG mice at 63, 90 and 365d old were greater (16-25%) than WT. Some TG female mice at 365d appeared obese; mean abdominal fat mass was 415% greater in TG than WT mice. Tibiae were longer (5-7%) in TG than WT mice at 63 and 90d. Tibial mineral mass of 35d males was 7% lower in TG than WT mice, but at 63d was 21% higher. The % osteoblast surface in 35d TG mice was 20% higher than WT, and at 63d was 17% lower, while % osteoclast surface did not differ. In 365d mice, cancellous bone volume (125%) and endocortical mineral apposition rate (40%) were greater in TG than WT males but not females. In WT mice, hindlimb unloading caused a reduction in mineral mass of tibiae (-20%) and lumbar vertebrae (-22%) relative to normally loaded controls. Surprisingly, hindlimb unloading also caused a relative reduction (-13%) in humerus mass. The effects of hindlimb unloading on

  10. Enhanced differentiation of intraepithelial lymphocytes in the intestine of polymeric immunoglobulin receptor-deficient mice.

    PubMed

    Kato-Nagaoka, Noriko; Shimada, Shin-Ichiro; Yamakawa, Yoko; Tsujibe, Satoshi; Naito, Tomoaki; Setoyama, Hiromi; Watanabe, Yohei; Shida, Kan; Matsumoto, Satoshi; Nanno, Masanobu

    2015-09-01

    To clarify the effect of secretory IgA (sIgA) deficiency on gut homeostasis, we examined intraepithelial lymphocytes (IELs) in the small intestine (SI) of polymeric immunoglobulin receptor-deficient (pIgR(-/-) ) mice. The pIgR(-/-) mice exhibited the accumulation of CD8αβ(+) T-cell receptor (TCR)-αβ(+) IELs (CD8αβ(+) αβ-IELs) after weaning, but no increase of CD8αβ(+) γδ-IELs was detected in pIgR(-/-) TCR-β(-/-) mice compared with pIgR(+/+) TCR-β(-/-) mice. When 5-bromo-2'-deoxyuridine (BrdU) was given for 14 days, the proportion of BrdU-labelled cells in SI-IELs was not different between pIgR(+/+) mice and pIgR(-/-) mice. However, the proportion of BrdU-labelled CD8αβ(+) -IELs became higher in pIgR(-/-) mice than pIgR(+/+) mice 10 days after discontinuing BrdU-labelling. Intravenously transferred splenic T cells migrated into the intraepithelial compartments of pIgR(+/+) TCR-β(-/-) mice and pIgR(-/-) TCR-β(-/-) mice to a similar extent. In contrast, in the case of injection of immature bone marrow cells, CD8αβ(+) αβ-IELs increased much more in the SI of pIgR(-/-) TCR-β(-/-) mice than pIgR(+/+) TCR-β(-/-) mice 8 weeks after the transfer. αβ-IELs from pIgR(-/-) mice could produce more interferon-γ and interleukin-17 than those of pIgR(+/+) mice, and intestinal permeability tended to increase in the SI of pIgR(-/-) mice with aging. Taken together, these results indicate that activated CD8αβ(+) αβ-IELs preferentially accumulate in pIgR(-/-) mice through the enhanced differentiation of immature haematopoietic precursor cells, which may subsequently result in the disruption of epithelial integrity. PMID:25967857

  11. Absence of granulocyte colony-stimulating factor signaling and neutrophil development in CCAAT enhancer binding protein alpha-deficient mice.

    PubMed

    Zhang, D E; Zhang, P; Wang, N D; Hetherington, C J; Darlington, G J; Tenen, D G

    1997-01-21

    Transcription factors are master regulatory switches of differentiation, including the development of specific hematopoietic lineages from stem cells. Here we show that mice with targeted disruption of the CCAAT enhancer binding protein alpha gene (C/EBP alpha) demonstrate a selective block in differentiation of neutrophils. Mature neutrophils and eosinophils are not observed in the blood or fetal liver of mutant animals, while other hematopoietic lineages, including monocytes, are not affected. Instead, most of the white cells in the peripheral blood of mutant mice had the appearance of myeloid blasts. We also observed a selective loss of expression of a critical gene target of CCAAT enhancer binding protein alpha, the granulocyte colony-stimulating factor receptor. As a result, multipotential myeloid progenitors from the mutant fetal liver are unable to respond to granulocyte colony-stimulating factor signaling, although they are capable of forming granulocyte-macrophage and macrophage colonies in methylcellulose in response to other growth factors. Finally, we demonstrate that the lack of granulocyte development results from a defect intrinsic to the hematopoietic system; transplanted fetal liver from mutant mice can reconstitute lymphoid but not neutrophilic cells in irradiated recipients. These studies suggest a model by which transcription factors can direct the differentiation of multipotential precursors through activation of expression of a specific growth factor receptor, allowing proliferation and differentiation in response to a specific extracellular signal. In addition, the c/ebp alpha -/- mice may be useful in understanding the mechanisms involved in acute myelogenous leukemia, in which a block in differentiation of myeloid precursors is a key feature of the disease. PMID:9012825

  12. Reduced Expression of MYC Increases Longevity and Enhances Healthspan

    PubMed Central

    Hofmann, Jeffrey W.; Zhao, Xiaoai; De Cecco, Marco; Peterson, Abigail L.; Pagliaroli, Luca; Manivannan, Jayameenakshi; Hubbard, Gene B.; Ikeno, Yuji; Zhang, Yongqing; Feng, Bin; Li, Xiaxi; Serre, Thomas; Qi, Wenbo; Van Remmen, Holly; Miller, Richard A.; Bath, Kevin G.; de Cabo, Rafael; Xu, Haiyan; Neretti, Nicola; Sedivy, John M.

    2015-01-01

    SUMMARY MYC is a highly pleiotropic transcription factor whose deregulation promotes cancer. In contrast, we find that Myc haploinsufficient (Myc+/−) mice exhibit increased lifespan. They show resistance to several age-associated pathologies, including osteoporosis, cardiac fibrosis and immunosenescence. They also appear to be more active, with a higher metabolic rate and healthier lipid metabolism. Transcriptomic analysis reveals a gene expression signature enriched for metabolic and immune processes. The ancestral role of MYC as a regulator of ribosome biogenesis is reflected in reduced protein translation, which is inversely correlated with longevity. We also observe changes in nutrient and energy sensing pathways, including reduced serum IGF-1, increased AMPK activity, and decreased AKT, TOR and S6K activities. In contrast to observations in other longevity models, Myc+/− mice do not show improvements in stress management pathways. Our findings indicate that MYC activity has a significant impact on longevity and multiple aspects of mammalian healthspan. PMID:25619689

  13. Reduced expression of MYC increases longevity and enhances healthspan.

    PubMed

    Hofmann, Jeffrey W; Zhao, Xiaoai; De Cecco, Marco; Peterson, Abigail L; Pagliaroli, Luca; Manivannan, Jayameenakshi; Hubbard, Gene B; Ikeno, Yuji; Zhang, Yongqing; Feng, Bin; Li, Xiaxi; Serre, Thomas; Qi, Wenbo; Van Remmen, Holly; Miller, Richard A; Bath, Kevin G; de Cabo, Rafael; Xu, Haiyan; Neretti, Nicola; Sedivy, John M

    2015-01-29

    MYC is a highly pleiotropic transcription factor whose deregulation promotes cancer. In contrast, we find that Myc haploinsufficient (Myc(+/-)) mice exhibit increased lifespan. They show resistance to several age-associated pathologies, including osteoporosis, cardiac fibrosis, and immunosenescence. They also appear to be more active, with a higher metabolic rate and healthier lipid metabolism. Transcriptomic analysis reveals a gene expression signature enriched for metabolic and immune processes. The ancestral role of MYC as a regulator of ribosome biogenesis is reflected in reduced protein translation, which is inversely correlated with longevity. We also observe changes in nutrient and energy sensing pathways, including reduced serum IGF-1, increased AMPK activity, and decreased AKT, TOR, and S6K activities. In contrast to observations in other longevity models, Myc(+/-) mice do not show improvements in stress management pathways. Our findings indicate that MYC activity has a significant impact on longevity and multiple aspects of mammalian healthspan. PMID:25619689

  14. Copper transport during lactation in transgenic mice expressing the human ATP7A protein

    PubMed Central

    Llanos, Roxana M.; Michalczyk, Agnes A.; Freestone, David J.; Currie, Scott; Linder, Maria C.; Ackland, M. Leigh; Mercer, Julian F.B.

    2008-01-01

    Both copper transporting ATPases, ATP7A and ATP7B, are expressed in mammary epithelial cells but their role in copper delivery to milk has not been clarified. We investigated the role of ATP7A in delivery of copper to milk using transgenic mice that over-express human ATP7A. In mammary gland of transgenic mice, human ATP7A protein was 10- to 20-fold higher than in control mice, and was localized to the basolateral membrane of mammary epithelial cells in lactating mice. The copper concentration in the mammary gland of transgenic dams and stomach contents of transgenic pups was significantly reduced compared to non-transgenic mice. The mRNA levels of endogenous Atp7a, Atp7b, and Ctr1 copper transporters in the mammary gland were not altered by the expression of the ATP7A transgene, and the protein levels of Atp7b and ceruloplasmin were similar in transgenic and non-transgenic mice. These data suggest that ATP7A plays a role in removing excess copper from the mammary epithelial cells rather than supplying copper to milk. PMID:18515074

  15. 11β-Hydroxysteroid Dehydrogenase Type 1 Is Expressed in Neutrophils and Restrains an Inflammatory Response in Male Mice

    PubMed Central

    Coutinho, Agnes E.; Kipari, Tiina M. J.; Zhang, Zhenguang; Esteves, Cristina L.; Lucas, Christopher D.; Gilmour, James S.; Webster, Scott P.; Walker, Brian R.; Hughes, Jeremy; Savill, John S.; Seckl, Jonathan R.; Rossi, Adriano G.

    2016-01-01

    Endogenous glucocorticoid action within cells is enhanced by prereceptor metabolism by 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1), which converts intrinsically inert cortisone and 11-dehydrocorticosterone into active cortisol and corticosterone, respectively. 11β-HSD1 is highly expressed in immune cells elicited to the mouse peritoneum during thioglycollate-induced peritonitis and is down-regulated as the inflammation resolves. During inflammation, 11β-HSD1-deficient mice show enhanced recruitment of inflammatory cells and delayed acquisition of macrophage phagocytic capacity. However, the key cells in which 11β-HSD1 exerts these effects remain unknown. Here we have identified neutrophils (CD11b+,Ly6G+,7/4+ cells) as the thioglycollate-recruited cells that most highly express 11β-HSD1 and show dynamic regulation of 11β-HSD1 in these cells during an inflammatory response. Flow cytometry showed high expression of 11β-HSD1 in peritoneal neutrophils early during inflammation, declining at later states. In contrast, expression in blood neutrophils continued to increase during inflammation. Ablation of monocytes/macrophages by treatment of CD11b-diphtheria-toxin receptor transgenic mice with diphtheria toxin prior to thioglycollate injection had no significant effect on 11β-HSD1 activity in peritoneal cells, consistent with neutrophils being the predominant 11β-HSD1 expressing cell type at this time. Similar to genetic deficiency in 11β-HSD1, acute inhibition of 11β-HSD1 activity during thioglycollate-induced peritonitis augmented inflammatory cell recruitment to the peritoneum. These data suggest that neutrophil 11β-HSD1 increases during inflammation to contribute to the restraining effect of glucocorticoids upon neutrophil-mediated inflammation. In human neutrophils, lipopolysaccharide activation increased 11β-HSD1 expression, suggesting the antiinflammatory effects of 11β-HSD1 in neutrophils may be conserved in humans. PMID:27145012

  16. 11β-Hydroxysteroid Dehydrogenase Type 1 Is Expressed in Neutrophils and Restrains an Inflammatory Response in Male Mice.

    PubMed

    Coutinho, Agnes E; Kipari, Tiina M J; Zhang, Zhenguang; Esteves, Cristina L; Lucas, Christopher D; Gilmour, James S; Webster, Scott P; Walker, Brian R; Hughes, Jeremy; Savill, John S; Seckl, Jonathan R; Rossi, Adriano G; Chapman, Karen E

    2016-07-01

    Endogenous glucocorticoid action within cells is enhanced by prereceptor metabolism by 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1), which converts intrinsically inert cortisone and 11-dehydrocorticosterone into active cortisol and corticosterone, respectively. 11β-HSD1 is highly expressed in immune cells elicited to the mouse peritoneum during thioglycollate-induced peritonitis and is down-regulated as the inflammation resolves. During inflammation, 11β-HSD1-deficient mice show enhanced recruitment of inflammatory cells and delayed acquisition of macrophage phagocytic capacity. However, the key cells in which 11β-HSD1 exerts these effects remain unknown. Here we have identified neutrophils (CD11b(+),Ly6G(+),7/4(+) cells) as the thioglycollate-recruited cells that most highly express 11β-HSD1 and show dynamic regulation of 11β-HSD1 in these cells during an inflammatory response. Flow cytometry showed high expression of 11β-HSD1 in peritoneal neutrophils early during inflammation, declining at later states. In contrast, expression in blood neutrophils continued to increase during inflammation. Ablation of monocytes/macrophages by treatment of CD11b-diphtheria-toxin receptor transgenic mice with diphtheria toxin prior to thioglycollate injection had no significant effect on 11β-HSD1 activity in peritoneal cells, consistent with neutrophils being the predominant 11β-HSD1 expressing cell type at this time. Similar to genetic deficiency in 11β-HSD1, acute inhibition of 11β-HSD1 activity during thioglycollate-induced peritonitis augmented inflammatory cell recruitment to the peritoneum. These data suggest that neutrophil 11β-HSD1 increases during inflammation to contribute to the restraining effect of glucocorticoids upon neutrophil-mediated inflammation. In human neutrophils, lipopolysaccharide activation increased 11β-HSD1 expression, suggesting the antiinflammatory effects of 11β-HSD1 in neutrophils may be conserved in humans. PMID:27145012

  17. Loss of AND-34/BCAR3 expression in mice results in rupture of the adult lens

    PubMed Central

    Near, Richard I.; Smith, Richard S.; Toselli, Paul A.; Freddo, Thomas F.; Bloom, Alexander B.; Vanden Borre, Pierre; Seldin, David C.

    2009-01-01

    Purpose AND-34/BCAR3 (Breast Cancer Anti-Estrogen Resistance 3) associates with the focal adhesion adaptor protein, p130CAS/BCAR1. Expression of AND-34 regulates epithelial cell growth pattern, motility, and growth factor dependence. We sought to establish the effects of the loss of AND-34 expression in a mammalian organism. Methods AND-34−/− mice were generated by homologous recombination. Histopathology, in situ hybridization, and western blotting were performed on murine tissues. Results Western analyses confirmed total loss of expression in AND-34−/− splenic lymphocytes. Mice lacking AND-34 are fertile and have normal longevity. While AND-34 is widely expressed in wild type mice, histologic analysis of multiple organs in AND-34−/− mice is unremarkable and analyses of lymphocyte development show no overt changes. A small percentage of AND-34−/− mice show distinctive small white eye lesions resulting from the migration of ruptured cortical lens tissue into the anterior chamber. Following initial vacuolization and liquefaction of the lens cortex first observed at postnatal day three, posterior lens rupture occurs in all AND-34−/− mice, beginning as early as three weeks and seen in all mice at three months. Western blot analysis and in situ hybridization confirmed the presence of AND-34 RNA and protein in lens epithelial cells, particularly at the lens equator. Prior data link AND-34 expression to the activation of Akt signaling. While Akt Ser 473 phosphorylation was readily detectable in AND-34+/+ lens epithelial cells, it was markedly reduced in the AND-34−/− lens epithelium. Basal levels of p130Cas phosphorylation were higher in AND-34+/+ than in AND-34−/− lens epithelium. Conclusions These results demonstrate the loss of AND-34 dysregulates focal adhesion complex signaling in lens epithelial cells and suggest that AND-34-mediated signaling is required for maintenance of the structural integrity of the adult ocular lens. PMID:19365570

  18. Learning to cope with stress modulates anterior cingulate cortex stargazin expression in monkeys and mice.

    PubMed

    Lee, Alex G; Capanzana, Roxanne; Brockhurst, Jacqueline; Cheng, Michelle Y; Buckmaster, Christine L; Absher, Devin; Schatzberg, Alan F; Lyons, David M

    2016-05-01

    Intermittent mildly stressful situations provide opportunities to learn, practice, and improve coping with gains in subsequent emotion regulation. Here we investigate the effects of learning to cope with stress on anterior cingulate cortex gene expression in monkeys and mice. Anterior cingulate cortex is involved in learning, memory, cognitive control, and emotion regulation. Monkeys and mice were randomized to either stress coping or no-stress treatment conditions. Profiles of gene expression were acquired with HumanHT-12v4.0 Expression BeadChip arrays adapted for monkeys. Three genes identified in monkeys by arrays were then assessed in mice by quantitative real-time polymerase chain reaction. Expression of a key gene (PEMT) involved in acetylcholine biosynthesis was increased in monkeys by coping but this result was not verified in mice. Another gene (SPRY2) that encodes a negative regulator of neurotrophic factor signaling was decreased in monkeys by coping but this result was only partly verified in mice. The CACNG2 gene that encodes stargazin (also called TARP gamma-2) was increased by coping in monkeys as well as mice randomized to coping with or without subsequent behavioral tests of emotionality. As evidence of coping effects distinct from repeated stress exposures per se, increased stargazin expression induced by coping correlated with diminished emotionality in mice. Stargazin modulates glutamate receptor signaling and plays a role in synaptic plasticity. Molecular mechanisms of synaptic plasticity that mediate learning and memory in the context of coping with stress may provide novel targets for new treatments of disorders in human mental health. PMID:27003116

  19. INDUCTION OF EARLY GROWTH RESPONSE GENE 2 EXPRESSION IN THE FOREBRAIN OF MICE PERFORMING AN ATTENTION-SET-SHIFTING TASK

    PubMed Central

    DeSteno, Deirdre A.; Schmauss, Claudia

    2008-01-01

    Early growth response (egr) genes encode transcription factors that are induced by stimuli that cause synaptic plasticity. Here we show that the expression of one member of this family, egr-2, is induced in the orbital frontal cortex (OFC) and medial prefrontal cortex (mPFC) of mice performing an attention-set-shifting task (ASST). The ASST is a series of two-choice perceptual discriminations between different odors and textures. Within the OFC and mPFC, different subregions exhibited egr-2 induction in response to different test-related features. In the medial OFC and the anterior cingulate subregion of the mPFC, egr-2 induction occurred in response to exposure to the novel odor stimulus. In the ventrolateral OFC and the pre- and infralimbic mPFC, additional egr-2 induction occurred during the associative learning phase of the ASST. In the infralimbic mPFC, further egr-2 induction occurred when mice performed set-shifting and reversal learning phases of the ASST. Mice with enhanced set-shifting performance exhibited decreased egr-2 induction in the mPFC indicating that the magnitude of egr-2 induction correlates with the magnitude of attentional demand. This decrease was largest in the infralimbic mPFC suggesting further that egr-2 induction in this region plays a role in the attentional control during set-shifting. In contrast to egr-2, neither egr-1 nor egr-3 expression was altered in ASST-tested mice, and no egr-2 induction occurred in mice that performed a spatial working memory task. These findings suggest a specific role of egr-2-mediated transcriptional activation in cognitive functions associated with attention. PMID:18280047

  20. Attenuation of experimental colitis in glutathione peroxidase 1 and catalase double knockout mice through enhancing regulatory T cell function.

    PubMed

    Kim, Hyung-Ran; Lee, Anbok; Choi, Eun-Jeong; Kie, Jeong-Hae; Lim, Woosung; Lee, Hyeon Kook; Moon, Byung-In; Seoh, Ju-Young

    2014-01-01

    Reactive oxygen species (ROS) have been implicated in the progression of inflammatory diseases including inflammatory bowel diseases (IBD). Meanwhile, several studies suggested the protective role of ROS in immune-mediated inflammatory diseases, and it was recently reported that dextran sodium sulfate (DSS)-induced colitis was attenuated in mice with an elevated level of ROS due to deficiency of peroxiredoxin II. Regulatory T cells (Tregs) are critical in the prevention of IBD and Treg function was reported to be closely associated with ROS level, but it has been investigated only in lowered levels of ROS so far. In the present study, in order to clarify the relationship between ROS level and Treg function, and their role in the pathogenesis of IBD, we investigated mice with an elevated level of ROS due to deficiency of both glutathione peroxidase (GPx)-1 and catalase (Cat) for the susceptibility of DSS-induced colitis in association with Treg function. The results showed that DSS-induced colitis was attenuated and Tregs were hyperfunctional in GPx1-/- × Cat-/- mice. In vivo administration of N-acetylcysteine (NAC) aggravated DSS-induced colitis and decreased Treg function to the level comparable to WT mice. Attenuated Th17 cell differentiation from naïve CD4+ cells as well as impaired production of IL-6 and IL-17A by splenocytes upon stimulation suggested anti-inflammatory tendency of GPx1-/- × Cat-/- mice. Suppression of Stat3 activation in association with enhancement of indoleamine 2,3-dioxygenase and FoxP3 expression might be involved in the immunosuppressive mechanism of GPx1-/- × Cat-/- mice. Taken together, it is implied that ROS level is critical in the regulation of Treg function, and IBD may be attenuated in appropriately elevated levels of ROS. PMID:24743300

  1. Impaired activity-dependent FMRP translation and enhanced mGluR-dependent LTD in Fragile X premutation mice.

    PubMed

    Iliff, Adam J; Renoux, Abigail J; Krans, Amy; Usdin, Karen; Sutton, Michael A; Todd, Peter K

    2013-03-15

    Fragile X premutation-associated disorders, including Fragile X-associated Tremor Ataxia Syndrome, result from unmethylated CGG repeat expansions in the 5' untranslated region (UTR) of the FMR1 gene. Premutation-sized repeats increase FMR1 transcription but impair rapid translation of the Fragile X mental retardation protein (FMRP), which is absent in Fragile X Syndrome (FXS). Normally, FMRP binds to RNA and regulates metabotropic glutamate receptor (mGluR)-mediated synaptic translation, allowing for dendritic synthesis of several proteins. FMRP itself is also synthesized at synapses in response to mGluR activation. However, the role of activity-dependent translation of FMRP in synaptic plasticity and Fragile X-premutation-associated disorders is unknown. To investigate this question, we utilized a CGG knock-in mouse model of the Fragile X premutation with 120-150 CGG repeats in the mouse Fmr1 5' UTR. These mice exhibit increased Fmr1 mRNA production but impaired FMRP translational efficiency, leading to a modest reduction in basal FMRP expression. Cultured hippocampal neurons and synaptoneurosomes derived from CGG KI mice demonstrate impaired FMRP translation in response to the group I mGluR agonist 3,5-dihydroxyphenylglycine. Electrophysiological analysis reveals enhanced mGluR-mediated long-term depression (mGluR-LTD) at CA3-CA1 synapses in acute hippocampal slices prepared from CGG KI mice relative to wild-type littermates, similar to Fmr1 knockout mice. However, unlike mGluR-LTD in mice completely lacking FMRP, mGluR-LTD in CGG knock-in mice remains dependent on new protein synthesis. These studies demonstrate partially overlapping synaptic plasticity phenotypes in mouse models of FXS and Fragile X premutation disorders and support a role for activity-dependent synthesis of FMRP in enduring forms of synaptic plasticity. PMID:23250915

  2. Morphine Enhances Hepatitis C Virus (HCV) Replicon Expression

    PubMed Central

    Li, Yuan; Zhang, Ting; Douglas, Steven D.; Lai, Jian-Ping; Xiao, Wei-Dong; Pleasure, David E.; Ho, Wen-Zhe

    2003-01-01

    Little information is available regarding whether substance abuse enhances hepatitis C virus (HCV) replication and promotes HCV disease progression. We investigated whether morphine alters HCV mRNA expression in HCV replicon-containing liver cells. Morphine significantly increased HCV mRNA expression, an effect which could be abolished by either of the opioid receptor antagonists, naltrexone or β-funaltrexamine. Investigation of the mechanism responsible for this enhancement of HCV replicon expression demonstrated that morphine activated NF-κB promoter and that caffeic acid phenethyl ester, a specific inhibitor of the activation of NF-κB, blocked morphine-activated HCV RNA expression. In addition, morphine compromised the anti-HCV effect of interferon alpha (IFN-α). Our in vitro data indicate that morphine may play an important role as a positive regulator of HCV replication in human hepatic cells and may compromise IFN-α therapy. PMID:12937158

  3. Tenomodulin expression in the periodontal ligament enhances cellular adhesion.

    PubMed

    Komiyama, Yuske; Ohba, Shinsuke; Shimohata, Nobuyuki; Nakajima, Keiji; Hojo, Hironori; Yano, Fumiko; Takato, Tsuyoshi; Docheva, Denitsa; Shukunami, Chisa; Hiraki, Yuji; Chung, Ung-Il

    2013-01-01

    Tenomodulin (Tnmd) is a type II transmembrane protein characteristically expressed in dense connective tissues such as tendons and ligaments. Its expression in the periodontal ligament (PDL) has also been demonstrated, though the timing and function remain unclear. We investigated the expression of Tnmd during murine tooth eruption and explored its biological functions in vitro. Tnmd expression was related to the time of eruption when occlusal force was transferred to the teeth and surrounding tissues. Tnmd overexpression enhanced cell adhesion in NIH3T3 and human PDL cells. In addition, Tnmd-knockout fibroblasts showed decreased cell adhesion. In the extracellular portions of Tnmd, the BRICHOS domain or CS region was found to be responsible for Tnmd-mediated enhancement of cell adhesion. These results suggest that Tnmd acts on the maturation or maintenance of the PDL by positively regulating cell adhesion via its BRICHOS domain. PMID:23593173

  4. Tenomodulin Expression in the Periodontal Ligament Enhances Cellular Adhesion

    PubMed Central

    Komiyama, Yuske; Ohba, Shinsuke; Shimohata, Nobuyuki; Nakajima, Keiji; Hojo, Hironori; Yano, Fumiko; Takato, Tsuyoshi; Docheva, Denitsa; Shukunami, Chisa; Hiraki, Yuji; Chung, Ung-il

    2013-01-01

    Tenomodulin (Tnmd) is a type II transmembrane protein characteristically expressed in dense connective tissues such as tendons and ligaments. Its expression in the periodontal ligament (PDL) has also been demonstrated, though the timing and function remain unclear. We investigated the expression of Tnmd during murine tooth eruption and explored its biological functions in vitro. Tnmd expression was related to the time of eruption when occlusal force was transferred to the teeth and surrounding tissues. Tnmd overexpression enhanced cell adhesion in NIH3T3 and human PDL cells. In addition, Tnmd-knockout fibroblasts showed decreased cell adhesion. In the extracellular portions of Tnmd, the BRICHOS domain or CS region was found to be responsible for Tnmd-mediated enhancement of cell adhesion. These results suggest that Tnmd acts on the maturation or maintenance of the PDL by positively regulating cell adhesion via its BRICHOS domain. PMID:23593173

  5. Effects of Unloading and Reloading on Expressions of Skelatal Muscle Membrane Proteins in Mice

    NASA Astrophysics Data System (ADS)

    Ohno, Y.; Ikuta, A.; Goto, A.; Sugiura, T.; Ohira, Y.; Yoshioka, T.; Goto, K.

    2013-02-01

    Effects of unloading and reloading on the expression levels of tripartite motif-containing 72 (TRIM72) and caveolin-3 (Cav-3) of soleus muscle in mice were investigated. Male C57BL/6J mice (11-week old) were randomly assigned to control and hindlimb-suspended groups. Some of mice in hindlimb-suspended group were subjected to continuous hindlimb suspension (HS) for 2 weeks with or without 7 days of ambulation recovery. Following HS, the muscle weight and protein expression levels of TRIM72 and Cav-3 in soleus were decreased. On the other hand, the gradual increases in muscle mass, TRIM72 and Cav-3 were observed after reloading following HS. Therefore, it was suggested that mechanical loading played a key role in a regulatory system for protein expressions of TRIM72 and Cav-3.

  6. Differential gene expression in high- and low-active inbred mice.

    PubMed

    Dawes, Michelle; Moore-Harrison, Trudy; Hamilton, Alicia T; Ceaser, Tyrone; Kochan, Kelli J; Riggs, Penny K; Lightfoot, J Timothy

    2014-01-01

    Numerous candidate genes have been suggested in the recent literature with proposed roles in regulation of voluntary physical activity, with little evidence of these genes' functional roles. This study compared the haplotype structure and expression profile in skeletal muscle and brain of inherently high- (C57L/J) and low- (C3H/HeJ) active mice. Expression of nine candidate genes [Actn2, Actn3, Casq1, Drd2, Lepr, Mc4r, Mstn, Papss2, and Glut4 (a.k.a. Slc2a4)] was evaluated via RT-qPCR. SNPs were observed in regions of Actn2, Casq1, Drd2, Lepr, and Papss2; however, no SNPs were located in coding sequences or associated with any known regulatory sequences. In mice exposed to a running wheel, Casq1 (P = 0.0003) and Mstn (P = 0.002) transcript levels in the soleus were higher in the low-active mice. However, when these genes were evaluated in naïve animals, differential expression was not observed, demonstrating a training effect. Among naïve mice, no genes in either tissue exhibited differential expression between strains. Considering that no obvious SNP mechanisms were determined or differential expression was observed, our results indicate that genomic structural variation or gene expression data alone is not adequate to establish any of these genes' candidacy or causality in relation to regulation of physical activity. PMID:24551844

  7. Identification of proteins highly expressed in uterine fluid from mice with hydrometra.

    PubMed

    Antonson, Per; Nalvarte, Ivan; Varshney, Mukesh; Xu, Li; Windahl, Sara H; Humire, Patricia; Ohlsson, Claes; Gustafsson, Jan-Åke; Dahlman-Wright, Karin

    2015-10-30

    Estrogen receptor alpha (ERα) is an important regulator of the estrous cycle and mice with global ERα deletion, as well as some conditional knockout mouse lines, have an interruption in the estrous cycle. In this study we observed that conditional ERα knockout mice where the Cre gene is regulated by the rat insulin promoter (RIP), RIP-Cre/ERα(KO) mice, have a 3.7-fold increase in serum 17β-estradiol levels, blocked estrous cycle, and develop a fluid-filled uterus (hydrometra). Using a proteomics approach, we identified three proteins, lactoferrin, complement C3 and chitinase 3-like protein 1 (CHI3L1), as highly expressed proteins in hydrometra fluid. The mRNA levels of the corresponding genes were more than 50-fold higher in RIP-Cre/ERα(KO) uterus compared to controls. High expression of CHI3L1 in the uterine fluid was not reflected as elevated levels in the serum. The high expression of lactoferrin, complement C3 and CHI3L1 in the uterine fluid, in association with elevated estrogen levels, prompted us to address if the expression of these genes is related to reproduction. However, gonadotropin treatment of mice reduced the uterine expression of these genes in a model of in vitro fertilization. Our findings identify lactoferrin, complement C3 and CHI3L1 as highly expressed proteins in hydrometra fluid in association with chronically elevated serum estradiol levels. PMID:26393907

  8. Enhancer Runaway and the Evolution of Diploid Gene Expression

    PubMed Central

    Fyon, Frédéric; Cailleau, Aurélie; Lenormand, Thomas

    2015-01-01

    Evidence is mounting that the evolution of gene expression plays a major role in adaptation and speciation. Understanding the evolution of gene regulatory regions is indeed an essential step in linking genotypes and phenotypes and in understanding the molecular mechanisms underlying evolutionary change. The common view is that expression traits (protein folding, expression timing, tissue localization and concentration) are under natural selection at the individual level. Here, we use a theoretical approach to show that, in addition, in diploid organisms, enhancer strength (i.e., the ability of enhancers to activate transcription) may increase in a runaway process due to competition for expression between homologous enhancer alleles. These alleles may be viewed as self-promoting genetic elements, as they spread without conferring a benefit at the individual level. They gain a selective advantage by getting associated to better genetic backgrounds: deleterious mutations are more efficiently purged when linked to stronger enhancers. This process, which has been entirely overlooked so far, may help understand the observed overrepresentation of cis-acting regulatory changes in between-species phenotypic differences, and sheds a new light on investigating the contribution of gene expression evolution to adaptation. PMID:26561855

  9. Activation of mitochondrial function and Hb expression in non-haematopoietic cells by an EPO inducer ameliorates ischaemic diseases in mice

    PubMed Central

    Hsu, Pei-Lun; Horng, Lin-Yea; Peng, Kang-Yung; Wu, Chia-Ling; Sung, Hui-Ching; Wu, Rong-Tsun

    2013-01-01

    Background and Purpose Many organs suffer from ischaemic injuries that reduce their ability to generate sufficient energy, which is required for functional maintenance and repair. Erythropoietin (EPO) ameliorates ischaemic injuries by pleiotropic effects. The aim of this study was to investigate the effect and mechanism of a small molecule EH-201, and found it as a potent EPO inducer and its effect in non-haematopoietic cells for therapeutic potential in ischemic disorders. Experimental Approach Mice kidney slices, primary hepatocytes, primary cardiomyocytes and C2C12 myoblasts were treated with EH-201. The effects of this treatment on EPO, Hb expression and mitochondrial biogenesis were analysed. In vivo, doxorubicin-induced cardiomyopathic mice were treated with EH-201. The mice were subjected to an endurance test, electrocardiography and echocardiography, and a histological examination of the isolated hearts was performed. EH-201 was also administered to cisplatin-induced nephropathic mice. Key Results In non-haematopoietic cells, EH-201 was potent at inducing EPO. EH-201 also stimulated mitochondrial biogenesis and enhanced the expression of Hb by a mechanism dependent on EPO-mediated signalling. In mechanistic studies, using EPO and EPO receptor-neutralizing antibodies, we confirmed that EH-201 enhances EPO-EPOR autocrine activity. EH-201 robustly increased the endurance performance activity of healthy and cardiomyopathic mice during hypoxic stress, enhanced myocardial mitochondrial biogenesis and Hb expression, and also improved cardiac function. EH-201 ameliorated anaemia and renal dysfunction in nephropathic mice. Conclusions and Implications The enhancement and recovery of cellular functions through the stimulation of mitochondrial activity and Hb production in non-haematopoietic cells by an inducer of endogenous EPO has potential as a therapeutic strategy for ischaemic diseases. PMID:23530756

  10. Immune responses elicited in mice with recombinant Lactococcus lactis expressing F4 fimbrial adhesin FaeG by oral immunization.

    PubMed

    Liu, Shujie; Li, Yongming; Xu, Ziwei; Wang, Yicheng

    2010-08-01

    Enterotoxigenic Escherichia coli (ETEC) is a major pathogenic agent causing piglet diarrhea. The major subunit and adhesin FaeG of F4(+) ETEC is an important virulence factor with strong immunogenicity. To determine whether Lactococcus lactis (L. lactis) could effectively deliver FaeG to the mucosal immune system, recombinant L. lactis expressing FaeG was constructed, and immune responses in mice following oral route delivery of recombinant L. lactis were explored. The production of FaeG expressed in L. lactis was up to approximately 10% of soluble whole-cell proteins, and recombinant FaeG (rFaeG) possessed good immunoreactivity by Western blot analysis. Oral immunization with recombinant L. lactis expressing FaeG induced F4-specific mucosal and systemic immune responses in the mice. In addition, high dose recombinant L. lactis or co-administration of high dose recombinant L. lactis with CTB enhanced the immune responses. These results suggested that L. lactis expressing FaeG was a promising candidate vaccine against ETEC. PMID:20532816

  11. Enhanced antibody responses induced by Candida albicans in mice.

    PubMed

    Cutler, J E; Lloyd, R K

    1982-12-01

    Candida albicans may immunopotentiate antibody responses in mice to antigens unrelated to the fungus. This effect occurred best with cell-associated, rather than soluble, antigens. When dead yeasts, cell walls, or a water-soluble candidal polysaccharide were used, immunopotentiation was most dramatic when the antigen and fungal materials were given concomitantly via an intraperitoneal injection. However, mice infected with viable yeasts several days before antigen administration also developed heightened responses to the antigen. The mechanism of the C. albicans-induced adjuvanticity was not defined, but the effect seemed to correlate with induction of inflammation. The presence of C. albicans or other inflammatory agents in the peritoneal cavity caused a more rapid uptake of particulate antigen by the liver. The relationship between this event and immunopotentiation is not known. These studies demonstrate that C. albicans may have profound effects on host immune responses. Because immunological aberrations are commonly found in patients with candidiasis it may be important to determine whether some of these aberrations result from, rather than precede candidiasis. PMID:6185421

  12. Suberoylanilide Hydroxamic Acid Restores Estrogen Reduced-cTnI Expression in Neonatal Hearts of Mice.

    PubMed

    Peng, Chang; Luo, Xiaomei; Xing, Qianlu; Sun, Huichao; Huang, Xupei

    2016-10-01

    Diastolic cardiac dysfunction can be caused by abnormality in cTnI expression during cardiogenesis. In this study, we investigated the effects of estrogen on the abnormal expression of cTnI in the hearts of neonatal mice and its potential epigenetic mechanisms. We then evaluated suberoylanilide hydroxamic acid (SAHA), a HDAC inhibitor, as a new target treatment of diastolic cardiac dysfunction. Postnatal day 0.5 C57BL/6 mice were injected with estrogen for 1 week, then the hearts of 7-day-old neonatal mice were retrieved for examination. The activities of HDAC and HAT were assayed by colorimetry, and the interaction of cTnI with HDAC5 in mice hearts were examined using chromatin immunoprecipitation assays. The expression of cTnI was tested by quantitative real-time RT-PCR and Western blot. Estrogen treated groups displayed a significantly increased HDAC activity in the hearts of neonatal mice while HAT activity remained unchanged. Additionally, HDAC5 was higher at the cTnI promoter, as compared to the saline treated control groups. The acetylation of histone H3K9ac on cTnI promoter significantly decreased in the hearts of neonatal mice treated with estrogen, and the expression of cTnI at transcriptional and protein levels also decreased. SAHA was shown to increase the acetylation of histone H3K9ac and upregulate the expression of cTnI. The data demonstrated that SAHA can correct cTnI expression abnormality caused by estrogen through inhibiting the binding of HDAC5 to the promoter of cTnI. J. Cell. Biochem. 117: 2377-2384, 2016. © 2016 Wiley Periodicals, Inc. PMID:27379430

  13. Human SUMO fusion systems enhance protein expression and solubility.

    PubMed

    Wang, Zhongyuan; Li, Haolong; Guan, Wei; Ling, Haili; Wang, Zhiyong; Mu, Tianyang; Shuler, Franklin D; Fang, Xuexun

    2010-10-01

    A major challenge associated with recombinant protein production in Escherichia coli is generation of large quantities of soluble, functional protein. Yeast SUMO (small ubiquitin-related modifier), has been shown to enhance heterologous protein expression and solubility as fusion tag, however, the effects of human SUMOs on protein expression have not been investigated. Here we describe the use of human SUMO1 and SUMO2 as a useful gene fusion technology. Human SUMO1 and SUMO2 fusion expression vectors were constructed and tested in His-tag and ubiquitin fusion expression systems. Two difficult-to-express model proteins, matrix metalloprotease-13 (MMP13) and enhanced green fluorescence protein (eGFP) were fused to the C-terminus of the human SUMO1 and SUMO2 expression vectors. These constructs were expressed in E. coli and evaluation of MMP13 and eGFP expression and solubility was conducted. We found that both SUMO1 and SUMO2 had the ability to enhance the solubility of MMP13 and eGFP, with the SUMO2 tag having a more significant effect. Since fusion tags produce varying quantities of soluble proteins, we assessed the effect of SUMO2 coupled with ubiquitin (Ub). SUMO2-ubiquitin and ubiquitin-SUMO2 fusion expression plasmids were constructed with eGFP as a passenger protein. Following expression in E. coli, both plasmids could improve eGFP expression and solubility similar to the SUMO2 fusion and better than the ubiquitin fusion. The sequential order of SUMO2 and ubiquitin had little effect on expression and solubility of eGFP. Purification of eGFP from the gene fusion product, SUMO2-ubiquitin-eGFP, involved cleavage by a deubiquitinase (Usp2-cc) and Ni-Sepharose column chromatography. The eGFP protein was purified to high homogeneity. In summary, human SUMO1 and SUMO2 are useful gene fusion technologies enhancing the expression, solubility and purification of model heterologous proteins. PMID:20457256

  14. Enhanced phosphorylation of cyclic AMP response element binding protein in Brain of mice following repetitive hypoxic exposure

    SciTech Connect

    Gao Yanan; Gao Ge; Long Caixia; Han Song; Zu Pengyu; Fang Li . E-mail: lfang@utmb.edu; Li Junfa . E-mail: junfali@cpums.edu.cn

    2006-02-10

    Cerebral ischemic/hypoxic preconditioning (I/HPC) is a phenomenon of endogenous protection that renders Brain tolerant to sustained ischemia/hypoxia. This profound protection induced by I/HPC makes it an attractive target for developing potential clinical therapeutic approaches. However, the molecular mechanism of I/HPC is unclear. Cyclic AMP (cAMP) response element binding protein (CREB), a selective nuclear transcriptional factor, plays a key role in the neuronal functions. Phosphorylation of CREB on Ser-133 may facilitate its transcriptional activity in response to various stresses. In the current study, we observed the changes in CREB phosphorylation (Ser-133) and protein expression in Brain of auto-hypoxia-induced HPC mice by using Western blot analysis. We found that the levels of phosphorylated CREB (Ser-133), but not protein expression of CREB, increased significantly (p < 0.05) in the hippocampus and the frontal cortex of mice after repetitive hypoxic exposure (H2-H4, n = 6 for each group), when compared to that of the normoxic (H0, n = 6) or hypoxic exposure once group (H1, n = 6). In addition, a significant enhancement (p < 0.05) of CREB phosphorylation (Ser-133) could also be found in the nuclear extracts from the whole hippocampus of hypoxic preconditioned mice (H2-H4, n = 6 for each group). These results suggest that the phosphorylation of CREB might be involved in the development of cerebral hypoxic preconditioning.

  15. Reintroducing domesticated wild mice to sociality induces adaptive transgenerational effects on MUP expression.

    PubMed

    Nelson, Adam C; Cauceglia, Joseph W; Merkley, Seth D; Youngson, Neil A; Oler, Andrew J; Nelson, Randy J; Cairns, Bradley R; Whitelaw, Emma; Potts, Wayne K

    2013-12-01

    When brought into captivity, wild animals can adapt to domestication within 10 generations. Such adaptations may decrease fitness in natural conditions. Many selective pressures are disrupted in captivity, including social behavioral networks. Although lack of sociality in captivity appears to mediate domestication, the underlying mechanisms are not well understood. Additionally, determining the contribution of genetic inheritance vs. transgenerational effects during relaxed selection may provide insight into the flexibility of adaptation. When wild-derived mice kept under laboratory conditions for eight generations were reintroduced to sociality and promiscuity (free mate choice), they adapted within two generations. Fitness assessments between this promiscuous lineage and a monogamous laboratory lineage revealed male-specific effects. Promiscuous-line males had deficits in viability, but a striking advantage in attracting mates, and their scent marks were also more attractive to females. Here, we investigate mechanistic details underlying this olfactory signal and identify a role of major urinary protein (MUP) pheromones. Promiscuous-line males inherit higher MUP expression than monogamous-line males through transgenerational inheritance. Sociality-driven maternal and paternal effects reveal intriguing conflicts among parents and offspring over pheromone expression. MUP up-regulation is not driven by hormone-driven transduction pathways, but rather is associated with reduction in DNA methylation of a CpG dinucleotide in the promoter. This reduction in methylation could enhance transcription by promoting the binding of transcription factor USF1 (upstream stimulatory factor 1). Finally, we experimentally demonstrate that increased MUP expression is a female attractant. These results identify molecular mechanisms guiding domestication and adaptive responses to fluctuating sociality. PMID:24248373

  16. ICAM-1–expressing neutrophils exhibit enhanced effector functions in murine models of endotoxemia

    PubMed Central

    Woodfin, Abigail; Beyrau, Martina; Voisin, Mathieu-Benoit; Ma, Bin; Whiteford, James R.; Hordijk, Peter L.; Hogg, Nancy

    2016-01-01

    Intracellular adhesion molecule-1 (ICAM-1) is a transmembrane glycoprotein expressed on the cell surface of numerous cell types such as endothelial and epithelial cells, vascular smooth muscle cells, and certain leukocyte subsets. With respect to the latter, ICAM-1 has been detected on neutrophils in several clinical and experimental settings, but little is known about the regulation of expression or function of neutrophil ICAM-1. In this study, we report on the de novo induction of ICAM-1 on the cell surface of murine neutrophils by lipopolysaccharide (LPS), tumor necrosis factor, and zymosan particles in vitro. The induction of neutrophil ICAM-1 was associated with enhanced phagocytosis of zymosan particles and reactive oxygen species (ROS) generation. Conversely, neutrophils from ICAM-1–deficient mice were defective in these effector functions. Mechanistically, ICAM-1–mediated intracellular signaling appeared to support neutrophil ROS generation and phagocytosis. In vivo, LPS-induced inflammation in the mouse cremaster muscle and peritoneal cavity led to ICAM-1 expression on intravascular and locally transmigrated neutrophils. The use of chimeric mice deficient in ICAM-1 on myeloid cells demonstrated that neutrophil ICAM-1 was not required for local neutrophil transmigration, but supported optimal intravascular and extravascular phagocytosis of zymosan particles. Collectively, the present results shed light on regulation of expression and function of ICAM-1 on neutrophils and identify it as an additional regulator of neutrophil effector responses in host defense. PMID:26647392

  17. ICAM-1-expressing neutrophils exhibit enhanced effector functions in murine models of endotoxemia.

    PubMed

    Woodfin, Abigail; Beyrau, Martina; Voisin, Mathieu-Benoit; Ma, Bin; Whiteford, James R; Hordijk, Peter L; Hogg, Nancy; Nourshargh, Sussan

    2016-02-18

    Intracellular adhesion molecule-1 (ICAM-1) is a transmembrane glycoprotein expressed on the cell surface of numerous cell types such as endothelial and epithelial cells, vascular smooth muscle cells, and certain leukocyte subsets. With respect to the latter, ICAM-1 has been detected on neutrophils in several clinical and experimental settings, but little is known about the regulation of expression or function of neutrophil ICAM-1. In this study, we report on the de novo induction of ICAM-1 on the cell surface of murine neutrophils by lipopolysaccharide (LPS), tumor necrosis factor, and zymosan particles in vitro. The induction of neutrophil ICAM-1 was associated with enhanced phagocytosis of zymosan particles and reactive oxygen species (ROS) generation. Conversely, neutrophils from ICAM-1-deficient mice were defective in these effector functions. Mechanistically, ICAM-1-mediated intracellular signaling appeared to support neutrophil ROS generation and phagocytosis. In vivo, LPS-induced inflammation in the mouse cremaster muscle and peritoneal cavity led to ICAM-1 expression on intravascular and locally transmigrated neutrophils. The use of chimeric mice deficient in ICAM-1 on myeloid cells demonstrated that neutrophil ICAM-1 was not required for local neutrophil transmigration, but supported optimal intravascular and extravascular phagocytosis of zymosan particles. Collectively, the present results shed light on regulation of expression and function of ICAM-1 on neutrophils and identify it as an additional regulator of neutrophil effector responses in host defense. PMID:26647392

  18. Subcutaneous or oral immunization of mice with Lactococcus lactis expressing F4 fimbrial adhesin FaeG.

    PubMed

    Liu, Shujie; Li, Yongming; Xu, Ziwei; Wang, Yicheng

    2013-01-01

    Enterotoxigenic Escherichia coli (ETEC) is one of the most common causes of diarrhea in neonatal and postweaning piglets. Fimbrial adhesion of ETEC has been considered an important colonization factor with antigenicity. To safely and effectively deliver the F4 (K88) fimbrial adhesin FaeG to the immune system, we have previously constructed the secretory expression vector pNZ8112-faeG, and FaeG was produced in cytoplasmic form in Lactococcus lactis. In this work, BALB/c mice were immunized with recombinant L. lactis to further determine the immunogenicity of recombinant FaeG (rFaeG) via the subcutaneous or oral route. Subcutaneous immunization in mice with recombinant L. lactis induced a significant increase in the F4-specific serum IgG titer and the number of antibody-secreting cells (ASCs) in the spleen. Oral immunization of mice with recombinant L. lactis induced mucosal and systemic F4-specific immune responses and increased the number of ASCs in the spleen, mesenteric lymph nodes and Peyer's patches. High-dose (2.8 × 10(11) CFU) recombinant strains and adjuvant cholera toxin B subunit enhanced specific mucosal immune responses. The results suggest the feasibility of delivering rFaeG expressed in L. lactis to the immune system in order to induce an F4-specific immune response. PMID:23386358

  19. Transgenic mice over-expressing carbonic anhydrase I showed aggravated joint inflammation and tissue destruction

    PubMed Central

    2012-01-01

    Background Studies have demonstrated that carbonic anhydrase I (CA1) stimulates calcium salt precipitation and cell calcification, which is an essential step in new bone formation. Our study had reported that CA1 encoding gene has a strong association with rheumatoid arthritis (RA) and ankylosing spondylitis (AS), two rheumatic diseases with abnormal new bone formation and bone resorption in joints. This study investigated the effect of CA1 on joint inflammation and tissue destruction in transgenic mice that over-express CA1 (CA1-Tg). Methods CA1-Tg was generated with C57BL/6J mice by conventional methods. CA1-Tg was treated with collagen-II to induce arthritis (CIA). Wild-type mice, CA1-Tg treated with bovine serum albumin (BSA) and transgenic mice over-expressing PADI4 (PADI4-Tg), a gene known to be involved in rheumatoid arthritis, were used as controls. Histochemistry and X-ray radiographic assay were used to examine joint destruction. Western blotting and real time-PCR were used to examine CA1 expression. Results CIA was observed in 60% of CA1-Tg, 20% of PADI4-Tg and 20% of wild-type mice after collagen injections. No CIA was found in CA1-Tg mice that received injections of BSA. The arthritic score was 5.5 ± 0.84 in the CA1-Tgs but the score was less than 2 in the injected wild-type mice and the PADI4-Tgs. The thickness of the hind paws in the CA1-Tgs was 3.46 ± 0.11 mm, which was thicker than that of PADI4-Tgs (2.23 ± 0.08 mm), wild-type mice (2.08 ± 0.06 mm) and BSA-treated CA1-Tgs (2.04 ± 0.07 mm). Histochemistry showed obvious inflammation, synovial hyperplasia and bone destruction in the joints of CA1-Tg that was not detected in PADI4-Tgs or wild-type mice. X-ray assays showed bone fusion in the paws and spines of CA1-Tg mice. Conclusion Over-expression of CA1 may aggravate joint inflammation and tissue destruction in the transgenic mice. PMID:23256642

  20. ZBP-89 Regulates Expression of Tryptophan Hydroxylase I and Mucosal Defense Against Salmonella Typhimurium in Mice

    PubMed Central

    Essien, Bryan; Grasberger, Helmut; Romain, Rachael D.; Law, David J.; Veniaminova, Natalia A.; Saqui-Salces, Milena; El-Zaatari, Mohamad; Tessier, Arthur; Hayes, Michael M.; Yang, Alexander C.; Merchant, Juanita L.

    2013-01-01

    Background & Aims ZBP-89 (also ZNF148 or Zfp148) is a butyrate-inducible zinc finger transcription factor that binds to GC-rich DNA elements. Deletion of the N-terminal domain is sufficient to increase mucosal susceptibility to chemical injury and inflammation. We investigated whether conditional deletion of ZBP-89 from the intestinal and colonic epithelium of mice increases their susceptibility to pathogens such as Salmonella typhimurium. Methods We generated mice with a conditional null allele of Zfp148 (ZBP-89FL/FL), using homologous recombination to flank Zfp148 with LoxP sites (ZBP-89FL/FL), and then breeding the resulting mice with those that express VillinCre. We used microarray analysis to compare gene expression patterns in colonic mucosa between ZBP-89FL/FL and C57BL/6 wild-type mice (controls). Mice were gavaged with 2 isogenic strains of S typhimurium after administration of streptomycin. Results Microarray analysis revealed that the colonic mucosa of ZBP-89FL/FL mice had reduced levels of tryptophan hydroxylase 1 (Tph1) mRNA, encoding the rate-limiting enzyme in enterochromaffin cell serotonin (5HT) biosynthesis. DNA affinity precipitation demonstrated direct binding of ZBP-89 to the mouse Tph1 promoter, which was required for its basal and butyrate-inducible expression. ZBP-89FL/FL mice did not increase mucosal levels of 5HT in response to S typhimurium infection and succumbed to the infection 2 days before control mice. The ΔhilA isogenic mutant of S typhimurium lacks this butyrate-regulated locus and stimulated, rather than suppressed, expression of Tph1 approximately 50-fold in control, but not ZBP-89FL/FL mice, correlating with fecal levels of butyrate. Conclusions ZBP-89 is required for butyrate-induced expression of the Tph1 gene and subsequent production of 5HT in response to bacterial infection in mice. Reductions in epithelial ZBP-89 increase susceptibility to colitis and sepsis following infection with S typhimurium, partly due to reduced

  1. MRP-1 expression levels determine strain-specific susceptibility to sodium arsenic-induced renal injury between C57BL/6 and BALB/c mice

    SciTech Connect

    Kimura, Akihiko; Ishida, Yuko; Wada, Takashi; Yokoyama, Hitoshi; Mukaida, Naofumi; Kondo, Toshikazu . E-mail: kondot@wakayama-med.ac.jp

    2005-02-15

    To clarify the pathophysiological mechanism underlying acute renal injury caused by acute exposure to arsenic, we subcutaneously injected both BALB/c and C57BL/6 mice with sodium arsenite (NaAs; 13.5 mg/kg). BALB/c mice exhibited exaggerated elevation of serum blood urea nitrogen (BUN) and creatinine (CRE) levels, compared with C57BL/6 mice. Moreover, half of BALB/c mice died by 24 h, whereas all C57BL/6 mice survived. Histopathological examination on kidney revealed severe hemorrhages, acute tubular necrosis, neutrophil infiltration, cast formation, and disappearance of PAS-positive brush borders in BALB/c mice, later than 10 h. These pathological changes were remarkably attenuated in C57BL/6 mice, accompanied with lower intrarenal arsenic concentrations, compared with BALB/c mice. Among heavy metal inducible proteins including multidrug resistance-associated protein (MRP)-1, multidrug resistance gene (MDR)-1, metallothionein (MT)-1, and arsenite inducible, cysteine- and histidine-rich RNA-associated protein (AIRAP), intrarenal MDR-1, MT-1, and AIRAP gene expression was enhanced to a similar extent in both strains, whereas NaAs challenge augmented intrarenal MRP-1 mRNA and protein expression levels in C57BL/6 but not BALB/c mice. Moreover, the administration of a specific inhibitor of MRP-1, MK-571, significantly exaggerated acute renal injury in C57BL/6 mice. Thus, MRP-1 is crucially involved in arsenic efflux and eventually prevention of acute renal injury upon acute exposure to NaAs.

  2. Serotonin- and Dopamine-Related Gene Expression in db/db Mice Islets and in MIN6 β-Cells Treated with Palmitate and Oleate

    PubMed Central

    Cataldo, L. R.; Olmos, P.; Galgani, J. E.; Valenzuela, R.; Aranda, E.; Cortés, V. A.; Santos, J. L.

    2016-01-01

    High circulating nonesterified fatty acids (NEFAs) concentration, often reported in diabetes, leads to impaired glucose-stimulated insulin secretion (GSIS) through not yet well-defined mechanisms. Serotonin and dopamine might contribute to NEFA-dependent β-cell dysfunction, since extracellular signal of these monoamines decreases GSIS. Moreover, palmitate-treated β-cells may enhance the expression of the serotonin receptor Htr2c, affecting insulin secretion. Additionally, the expression of monoamine-oxidase type B (Maob) seems to be lower in islets from humans and mice with diabetes compared to nondiabetic islets, which may lead to increased monoamine concentrations. We assessed the expression of serotonin- and dopamine-related genes in islets from db/db and wild-type (WT) mice. In addition, the effect of palmitate and oleate on the expression of such genes, 5HT content, and GSIS in MIN6 β-cell was determined. Lower Maob expression was found in islets from db/db versus WT mice and in MIN6 β-cells in response to palmitate and oleate treatment compared to vehicle. Reduced 5HT content and impaired GSIS in response to palmitate (−25%; p < 0.0001) and oleate (−43%; p < 0.0001) were detected in MIN6 β-cells. In conclusion, known defects of GSIS in islets from db/db mice and MIN6 β-cells treated with NEFAs are accompanied by reduced Maob expression and reduced 5HT content. PMID:27366756

  3. Slug expression enhances tumor formation in a non-invasive rectal cancer model

    PubMed Central

    Camp, E. Ramsay; Findlay, Victoria J.; Vaena, Silvia G.; Walsh, Jarret; Lewin, David N.; Turner, David P.; Watson, Dennis K

    2011-01-01

    Background Epithelial-to-mesenchymal transition (EMT) is a series of molecular changes allowing epithelial cancer cells to acquire properties of mesenchymal cells: increased motility and invasion and protection from apoptosis. Transcriptional regulators such as Slug mediate EMT, working in part to repress E-cadherin transcription. We report a novel, non-invasive in vivo rectal cancer model to explore the role of Slug in colorectal cancer (CRC) tumor development. Methods For the generation of DLD-1 cells overexpressing Slug (Slug DLD-1), a Slug or empty (Empty DLD-1) pCMV-3Tag-1 (kanamycin resistant) vector was used for transfection. Cells were evaluated for Slug and E-cadherin expression, and cell migration and invasion. For the in vivo study, colon cancer cells (parental DLD-1, Slug DLD-1, empty DLD-1, and HCT-116) were submucosally injected into the posterior rectum of nude mice using endoscopic guidance. After 28 days, tumors were harvested and tissue was analyzed. Results Slug expression in our panel of colon cancer cell lines was inversely correlated with E-cadherin expression and enhanced migration/invasion. Slug DLD-1 cells demonstrated a 21-fold increased Slug and 19-fold decreased E-cadherin expression compared with empty DLD-1. Similarly, the Slug DLD-1 cells had significantly enhanced cellular migration and invasion. In the orthotopic rectal cancer model, Slug DLD-1 cells formed rectal tumors in 9/10 (90%) of the mice (mean volume = 458 mm3) compared with only 1/10 (10%) with empty DLD-1 cells. Conclusion Slug mediates EMT with enhanced in vivo rectal tumor formation. Our non-invasive in vivo model enables researchers to explore the molecular consequences of altered genes in a clinically relevant rectal cancer in an effort to develop novel therapeutic approaches for patients with rectal cancer. PMID:21470622

  4. Macrophage function in alloxan diabetic mice: expression of adhesion molecules, generation of monokines and oxygen and NO radicals

    PubMed Central

    Ptak, W; Klimek, M; Bryniarski, K; Ptak, M; Majcher, P

    1998-01-01

    The increased incidence of bacterial and mycotic infections in poorly controlled diabetic patients or animals is frequently attributed to impaired activities of professional phagocytes (granulocytes, macrophages) in hypoinsulinaemic milieu. We measured production of monokines (IL-6 and tumour necrosis factor-alpha (TNF-α)), active NO and reactive oxygen intermediates (ROIs), as well as expression of several cell surface adhesion molecules (Mac-1, -2 and -3, intercellular adhesion molecule-1 (ICAM-1) and FcγRII), by thioglycollate medium-induced peritoneal macrophages of normoglycaemic and alloxan diabetic CBA/J mice (blood glucose level in the range 300 or 500 mg/dl). Macrophages of animals with moderate diabetes (300 mg/dl) produced significantly more IL-6 and TNF-α and ROIs than cells of control mice and showed an increased expression of all cell surface molecules, except Mac-3. NO/NO2 production was not affected. Administration of insulin restored enhanced values to normal levels, except for the production of ROIs which remained unusually high. We conclude that two separate mechanisms influence macrophage physiology in diabetes—lack of saturation of insulin receptors on macrophages and an indirect effect due to formation of advanced glycosylation endproducts (AGE) on their surfaces. The latter is possibly responsible for increased generation of ROIs, since it cannot be down-regulated by prolonged insulin treatment. How the increased activity of macrophages of moderately diabetic mice (enhanced production of proinflammatory monokines and oxygen radicals as well as expression of molecules) is related to their ability to kill bacteria is now under investigation. PMID:9764597

  5. Expression and immune recognition of SV40 Tag in transgenic mice that develop metastatic osteosarcomas.

    PubMed

    Marton, I; Johnson, S E; Damjanov, I; Currier, K S; Sundberg, J P; Knowles, B B

    2000-04-01

    Mature adult mice of the C57BL/6-TgN(Amy1TAg)501Knw transgenic mouse lineage, 501, containing a liver alpha-amylase promoted-SV40 Tag hybrid gene, routinely develop SV40 Tag-induced metastatic osteosarcomas. This form of alpha-amylase was known to be expressed in the liver, salivary glands, pancreas, and fat. Cells in the normal rib adjacent to the periosteum also express alpha-amylase suggesting that transgene expression is correctly targeted to generate osteosarcomas. 501 mice express SV40 Tag in the salivary glands but do not develop abnormalities in these organs by the time of their death from SV40-induced osteosarcomas. Mice of the C57BL/6 strain make a strong and effective anti-tumor immune response to SV40 Tag immunization. However, immunization of 501 mice with SV40 Tag early in life does not alter or prevent SV40 Tag-induced osteosarcomagenesis. 501 mice mount a significantly less effective cytotoxic T-lymphocyte response following SV40 Tag immunization while 501 osteosarcoma-derived cells are fully susceptible to SV40 Tag-specific T-cell lysis. This suggests that partial tolerance, not loss of antigen presentation by tumor cells, characterizes this mouse model of endogenous bone tumor development. To determine whether the immune recognition of endogenous SV40 Tag could influence tumorigenesis, the metastatic potential and time of death from tumor was investigated in CD4-null mutant 501 mice and beta-2 microglobulin-null mutant 501 mice. The size and number of metastases in these strains and longevity of these strains varied. We suggest that components of both the innate and adaptive immune response control tumor appearance and progression. PMID:10951695

  6. Maternal High-Fat Feeding Increases Placental Lipoprotein Lipase Activity by Reducing SIRT1 Expression in Mice.

    PubMed

    Qiao, Liping; Guo, Zhuyu; Bosco, Chris; Guidotti, Stefano; Wang, Yunfeng; Wang, Mingyong; Parast, Mana; Schaack, Jerome; Hay, William W; Moore, Thomas R; Shao, Jianhua

    2015-09-01

    This study investigated how maternal overnutrition and obesity regulate expression and activation of proteins that facilitate lipid transport in the placenta. To create a maternal overnutrition and obesity model, primiparous C57BL/6 mice were fed a high-fat (HF) diet throughout gestation. Fetuses from HF-fed dams had significantly increased serum levels of free fatty acid and body fat. Despite no significant difference in placental weight, lipoprotein lipase (LPL) protein levels and activity were remarkably elevated in placentas from HF-fed dams. Increased triglyceride content and mRNA levels of CD36, VLDLr, FABP3, FABPpm, and GPAT2 and -3 were also found in placentas from HF-fed dams. Although both peroxisome proliferator-activated receptor-γ (PPARγ) and CCAAT/enhancer binding protein-α protein levels were significantly increased in placentas of the HF group, only PPARγ exhibited a stimulative effect on LPL expression in cultured JEG-3 human trophoblasts. Maternal HF feeding remarkably decreased SIRT1 expression in placentas. Through use of an SIRT1 activator and inhibitor and cultured trophoblasts, an inhibitory effect of SIRT1 on LPL expression was demonstrated. We also found that SIRT1 suppresses PPARγ expression in trophoblasts. Most importantly, inhibition of PPARγ abolished the SIRT1-mediated regulatory effect on LPL expression. Together, these results indicate that maternal overnutrition induces LPL expression in trophoblasts by reducing the inhibitory effect of SIRT1 on PPARγ. PMID:25948680

  7. Clemastine Enhances Myelination in the Prefrontal Cortex and Rescues Behavioral Changes in Socially Isolated Mice

    PubMed Central

    Dupree, Jeffrey L.; Gacias, Mar; Frawley, Rebecca; Sikder, Tamjeed; Naik, Payal; Casaccia, Patrizia

    2016-01-01

    Altered myelin structure and oligodendrocyte function have been shown to correlate with cognitive and motor dysfunction and deficits in social behavior. We and others have previously demonstrated that social isolation in mice induced behavioral, transcriptional, and ultrastructural changes in oligodendrocytes of the prefrontal cortex (PFC). However, whether enhancing myelination and oligodendrocyte differentiation could be beneficial in reversing such changes remains unexplored. To test this hypothesis, we orally administered clemastine, an antimuscarinic compound that has been shown to enhance oligodendrocyte differentiation and myelination in vitro, for 2 weeks in adult mice following social isolation. Clemastine successfully reversed social avoidance behavior in mice undergoing prolonged social isolation. Impaired myelination was rescued by oral clemastine treatment, and was associated with enhanced oligodendrocyte progenitor differentiation and epigenetic changes. Clemastine induced higher levels of repressive histone methylation (H3K9me3), a marker for heterochromatin, in oligodendrocytes, but not neurons, of the PFC. This was consistent with the capability of clemastine in elevating H3K9 histone methyltransferases activity in cultured primary mouse oligodendrocytes, an effect that could be antagonized by cotreatment with muscarine. Our data suggest that promoting adult myelination is a potential strategy for reversing depressive-like social behavior. SIGNIFICANCE STATEMENT Oligodendrocyte development and myelination are highly dynamic processes influenced by experience and neuronal activity. However, whether enhancing myelination and oligodendrocyte differentiation is beneficial to treat depressive-like behavior has been unexplored. Mice undergoing prolonged social isolation display impaired myelination in the prefrontal cortex. Clemastine, a Food and Drug Administration-approved antimuscarinic compound that has been shown to enhance myelination under

  8. Clemastine Enhances Myelination in the Prefrontal Cortex and Rescues Behavioral Changes in Socially Isolated Mice.

    PubMed

    Liu, Jia; Dupree, Jeffrey L; Gacias, Mar; Frawley, Rebecca; Sikder, Tamjeed; Naik, Payal; Casaccia, Patrizia

    2016-01-20

    Altered myelin structure and oligodendrocyte function have been shown to correlate with cognitive and motor dysfunction and deficits in social behavior. We and others have previously demonstrated that social isolation in mice induced behavioral, transcriptional, and ultrastructural changes in oligodendrocytes of the prefrontal cortex (PFC). However, whether enhancing myelination and oligodendrocyte differentiation could be beneficial in reversing such changes remains unexplored. To test this hypothesis, we orally administered clemastine, an antimuscarinic compound that has been shown to enhance oligodendrocyte differentiation and myelination in vitro, for 2 weeks in adult mice following social isolation. Clemastine successfully reversed social avoidance behavior in mice undergoing prolonged social isolation. Impaired myelination was rescued by oral clemastine treatment, and was associated with enhanced oligodendrocyte progenitor differentiation and epigenetic changes. Clemastine induced higher levels of repressive histone methylation (H3K9me3), a marker for heterochromatin, in oligodendrocytes, but not neurons, of the PFC. This was consistent with the capability of clemastine in elevating H3K9 histone methyltransferases activity in cultured primary mouse oligodendrocytes, an effect that could be antagonized by cotreatment with muscarine. Our data suggest that promoting adult myelination is a potential strategy for reversing depressive-like social behavior. Significance statement: Oligodendrocyte development and myelination are highly dynamic processes influenced by experience and neuronal activity. However, whether enhancing myelination and oligodendrocyte differentiation is beneficial to treat depressive-like behavior has been unexplored. Mice undergoing prolonged social isolation display impaired myelination in the prefrontal cortex. Clemastine, a Food and Drug Administration-approved antimuscarinic compound that has been shown to enhance myelination under

  9. Enhancing hepatic mitochondrial fatty acid oxidation stimulates eating in food-deprived mice

    PubMed Central

    Mansouri, Abdelhak; Pacheco-López, Gustavo; Ramachandran, Deepti; Arnold, Myrtha; Leitner, Claudia; Prip-Buus, Carina; Langhans, Wolfgang

    2014-01-01

    Hepatic fatty acid oxidation (FAO) has long been implicated in the control of eating. Nevertheless, direct evidence for a causal relationship between changes in hepatic FAO and changes in food intake is still missing. Here we tested whether increasing hepatic FAO via adenovirus-mediated expression of a mutated form of the key regulatory enzyme of mitochondrial FAO carnitine palmitoyltransferase 1A (CPT1mt), which is active but insensitive to inhibition by malonyl-CoA, affects eating and metabolism in mice. CPT1mt expression increased hepatocellular CPT1 protein levels. This resulted in an increase in circulating ketone body levels in fasted CPT1mt-expressing mice, suggesting an increase in hepatic FAO. These mice did not show any significant changes in cumulative food intake, energy expenditure, or respiratory quotient after 4-h food deprivation. After 24-h food deprivation, however, the CPT1mt-expressing mice displayed increased food intake. Thus expression of CPT1mt in the liver increases hepatic FAO capacity, but does not inhibit eating. Rather, it may even stimulate eating after prolonged food deprivation. These data do not support the hypothesis that an increase in hepatic FAO decreases food intake. PMID:25427767

  10. Intron V, not intron I of human thrombopoietin, improves expression in the milk of transgenic mice regulated by goat beta-casein promoter.

    PubMed

    Li, Yan; Hao, Hu; Zhou, Mingqian; Zhou, Hongwei; Ye, Jianbin; Ning, Lijun; Ning, Yunshan

    2015-01-01

    Introns near 5' end of genes generally enhance gene expression because of an enhancer /a promoter within their sequence or as intron-mediated enhancement. Surprisingly, our previous experiments found that the vector containing the last intron (intron V) of human thromobopoietin (hTPO) expressed higher hTPO in cos-1 cell than the vector containing intron I regulated by cytomegalovirus promoter. Moreover, regulated by 1.0 kb rat whey acidic protein promoter, hTPO expression was higher in transgenic mice generated by intron V-TPOcDNA than in transgenic mice generated by TPOcDNA and TPOgDNA. However, it is unknown whether the enhancement of hTPO expression by intron I is decreased by uAUG7 at 5'-UTR of hTPO in vivo. Currently, we constructed vectors regulated by stronger 6.5 kb β-casein promoter, including pTPOGA (containing TPOcDNA), pTPOGB (containing TUR-TPOcDNA, TUR including exon1, intron I and non-coding exon2 of hTPO gene), pTPOGC (containing ΔTUR-TPOcDNA, nucleotides of TUR from uAUG7 to physiological AUG were deleted), pTPOGD (containing intron V-TPOcDNA) and pTPOGE (containing TPOgDNA), to evaluate the effect of intron I on hTPO expression and to further verify whether intron V enhances hTPO expression in the milk of transgenic mice. The results demonstrated that intron V, not intron I improved hTPO expression. PMID:26527459

  11. Intron V, not intron I of human thrombopoietin, improves expression in the milk of transgenic mice regulated by goat beta-casein promoter

    PubMed Central

    Li, Yan; Hao, Hu; Zhou, Mingqian; Zhou, Hongwei; Ye, Jianbin; Ning, Lijun; Ning, Yunshan

    2015-01-01

    Introns near 5′ end of genes generally enhance gene expression because of an enhancer /a promoter within their sequence or as intron-mediated enhancement. Surprisingly, our previous experiments found that the vector containing the last intron (intron V) of human thromobopoietin (hTPO) expressed higher hTPO in cos-1 cell than the vector containing intron I regulated by cytomegalovirus promoter. Moreover, regulated by 1.0 kb rat whey acidic protein promoter, hTPO expression was higher in transgenic mice generated by intron V-TPOcDNA than in transgenic mice generated by TPOcDNA and TPOgDNA. However, it is unknown whether the enhancement of hTPO expression by intron I is decreased by uAUG7 at 5′-UTR of hTPO in vivo. Currently, we constructed vectors regulated by stronger 6.5kb β-casein promoter, including pTPOGA (containing TPOcDNA), pTPOGB (containing TUR-TPOcDNA, TUR including exon1, intron I and non-coding exon2 of hTPO gene), pTPOGC (containing ΔTUR-TPOcDNA, nucleotides of TUR from uAUG7 to physiological AUG were deleted), pTPOGD (containing intron V-TPOcDNA) and pTPOGE (containing TPOgDNA), to evaluate the effect of intron I on hTPO expression and to further verify whether intron V enhances hTPO expression in the milk of transgenic mice. The results demonstrated that intron V, not intron I improved hTPO expression. PMID:26527459

  12. Deletion of IL-33R attenuates VEGF expression and enhances necrosis in mammary carcinoma

    PubMed Central

    Pejnovic, Nada N.; Mitrovic, Slobodanka L. J.; Arsenijevic, Nebojsa N.; Simovic Markovic, Bojana J.; Lukic, Miodrag L.

    2016-01-01

    Interleukin-33 (IL-33)/IL-33 receptor (IL-33R, ST2) signaling pathway promotes mammary cancer growth and metastasis by inhibiting anti-tumor immunity. However, the role of IL-33/IL-33R axis in neoangiogenesis and tumor necrosis is not elucidated. Therefore, the aim of this study was to investigate the role of IL-33/IL-33R axis in mammary tumor necrosis. Deletion of IL-33R (ST2) gene in BALB/c mice enhanced tumor necrosis and attenuated tumor growth in 4T1 breast cancer model, which was associated with markedly decreased expression of vascular endothelial growth factor (VEGF) and IL-33 in mammary tumor cells. We next analyzed IL-33, IL-33R and VEGF expression and microvascular density (MVD) in breast tumors from 40 female patients with absent or present tumor necrosis. We found significantly higher expression of IL-33, IL-33R and VEGF in breast cancer tissues with absent tumor necrosis. Both, IL-33 and IL-33R expression correlated with VEGF expression in tumor cells. Further, VEGF expression positively correlated with MVD in perinecrotic zone. Taking together, our data indicate that IL-33/IL-33R pathway is critically involved in mammary tumor growth by facilitating expression of pro-angiogenic VEGF in tumor cells and attenuating tumor necrosis. These data add an unidentified mechanism by which IL-33/IL-33R axis facilitates tumor growth. PMID:26919112

  13. Expression profiling pre-diabetic mice to uncover drugs with clinical application to type 1 diabetes.

    PubMed

    Pang, Dimeng; Irvine, Katharine M; Mehdi, Ahmed M; Thomas, Helen E; Harris, Mark; Hamilton-Williams, Emma E; Thomas, Ranjeny

    2015-08-01

    In the NOD mouse model of type 1 diabetes (T1D), genetically identical mice in the same environment develop diabetes at different rates. Similar heterogeneity in the rate of progression to T1D exists in humans, but the underlying mechanisms are unclear. Here, we aimed to discover peripheral blood (PB) genes in NOD mice predicting insulitis severity and rate of progression to diabetes. We then wished to use these genes to mine existing databases to identify drugs effective in diabetes. In a longitudinal study, we analyzed gene expression in PB samples from NOD.CD45.2 mice at 10 weeks of age, then scored pancreatic insulitis at 14 weeks or determined age of diabetes onset. In a multilinear regression model, Tnf and Tgfb mRNA expression in PB predicted insulitis score (R (2)=0.56, P=0.01). Expression of these genes did not predict age of diabetes onset. However, by expression-profiling PB genes in 10-week-old NOD.CD45.2 mice, we found a signature of upregulated genes that predicted delayed or no diabetes. Major associated pathways included chromatin organization, cellular protein location and regulation of nitrogen compounds and RNA. In a clinical cohort, three of these genes were differentially expressed between first-degree relatives, T1D patients and controls. Bioinformatic analysis of differentially expressed genes in NOD.CD45.2 PB identified drugs that are predicted to delay or prevent diabetes. Of these drugs, 11 overlapped with drugs predicted to induce a human 'non-progressor' expression profile. These data demonstrate that disease heterogeneity in diabetes-prone mice can be exploited to mine novel clinical T1D biomarkers and drug targets. PMID:26366287

  14. Expression profiling pre-diabetic mice to uncover drugs with clinical application to type 1 diabetes

    PubMed Central

    Pang, Dimeng; Irvine, Katharine M; Mehdi, Ahmed M; Thomas, Helen E; Harris, Mark; Hamilton-Williams, Emma E; Thomas, Ranjeny

    2015-01-01

    In the NOD mouse model of type 1 diabetes (T1D), genetically identical mice in the same environment develop diabetes at different rates. Similar heterogeneity in the rate of progression to T1D exists in humans, but the underlying mechanisms are unclear. Here, we aimed to discover peripheral blood (PB) genes in NOD mice predicting insulitis severity and rate of progression to diabetes. We then wished to use these genes to mine existing databases to identify drugs effective in diabetes. In a longitudinal study, we analyzed gene expression in PB samples from NOD.CD45.2 mice at 10 weeks of age, then scored pancreatic insulitis at 14 weeks or determined age of diabetes onset. In a multilinear regression model, Tnf and Tgfb mRNA expression in PB predicted insulitis score (R2=0.56, P=0.01). Expression of these genes did not predict age of diabetes onset. However, by expression-profiling PB genes in 10-week-old NOD.CD45.2 mice, we found a signature of upregulated genes that predicted delayed or no diabetes. Major associated pathways included chromatin organization, cellular protein location and regulation of nitrogen compounds and RNA. In a clinical cohort, three of these genes were differentially expressed between first-degree relatives, T1D patients and controls. Bioinformatic analysis of differentially expressed genes in NOD.CD45.2 PB identified drugs that are predicted to delay or prevent diabetes. Of these drugs, 11 overlapped with drugs predicted to induce a human ‘non-progressor' expression profile. These data demonstrate that disease heterogeneity in diabetes-prone mice can be exploited to mine novel clinical T1D biomarkers and drug targets. PMID:26366287

  15. Memory-Enhancing Effects of the Crude Extract of Polygala tenuifolia on Aged Mice.

    PubMed

    Li, Zongyang; Liu, Yamin; Wang, Liwei; Liu, Xinmin; Chang, Qi; Guo, Zhi; Liao, Yonghong; Pan, Ruile; Fan, Tai-Ping

    2014-01-01

    Learning and memory disorders arise from distinct age-associated processes, and aging animals are often used as a model of memory impairment. The root of Polygala tenuifolia has been commonly used in some Asian countries as memory enhancer and its memory improvement has been reported in various animal models. However, there is less research to verify its effect on memory functions in aged animals. Herein, the memory-enhancing effects of the crude extract of Polygala tenuifolia (EPT) on normal aged mice were assessed by Morris water maze (MWM) and step-down passive avoidance tests. In MWM tests, the impaired spatial memory of the aged mice was partly reversed by EPT (100 and 200 mg/kg; P < 0.05) as compared with the aged control mice. In step-down tests, the nonspatial memory of the aged mice was improved by EPT (100 and 200 mg/kg; P < 0.05). Additionally, EPT could increase superoxide dismutase (SOD) and catalase (CAT) activities, inhibit monoamine oxidase (MAO) and acetyl cholinesterase (AChE) activities, and decrease the levels of malondialdehyde (MDA) in the brain tissue of the aged mice. The results showed that EPT improved memory functions of the aged mice probably via its antioxidant properties and via decreasing the activities of MAO and AChE. PMID:24744810

  16. Memory-Enhancing Effects of the Crude Extract of Polygala tenuifolia on Aged Mice

    PubMed Central

    Li, Zongyang; Liu, Yamin; Wang, Liwei; Liu, Xinmin; Chang, Qi; Guo, Zhi; Liao, Yonghong; Pan, Ruile; Fan, Tai-Ping

    2014-01-01

    Learning and memory disorders arise from distinct age-associated processes, and aging animals are often used as a model of memory impairment. The root of Polygala tenuifolia has been commonly used in some Asian countries as memory enhancer and its memory improvement has been reported in various animal models. However, there is less research to verify its effect on memory functions in aged animals. Herein, the memory-enhancing effects of the crude extract of Polygala tenuifolia (EPT) on normal aged mice were assessed by Morris water maze (MWM) and step-down passive avoidance tests. In MWM tests, the impaired spatial memory of the aged mice was partly reversed by EPT (100 and 200 mg/kg; P < 0.05) as compared with the aged control mice. In step-down tests, the nonspatial memory of the aged mice was improved by EPT (100 and 200 mg/kg; P < 0.05). Additionally, EPT could increase superoxide dismutase (SOD) and catalase (CAT) activities, inhibit monoamine oxidase (MAO) and acetyl cholinesterase (AChE) activities, and decrease the levels of malondialdehyde (MDA) in the brain tissue of the aged mice. The results showed that EPT improved memory functions of the aged mice probably via its antioxidant properties and via decreasing the activities of MAO and AChE. PMID:24744810

  17. Enhanced resistance to acute infection with Trypanosoma cruzi in mice treated with an interferon inducer.

    PubMed Central

    James, S L; Kipnis, T L; Sher, A; Hoff, R

    1982-01-01

    For an exploration of the effects of interferon-inducible resistance mechanisms in acute American trypanosomiasis, the synthetic interferon inducer tilerone hydrochloride was administered to mice of the C57BL/6J strain, which is highly resistant to Trypanosoma cruzi, 18 to 24 h before infection with a potentially lethal dose of bloodstream trypomastigotes. Although all of the control mice died within 30 days of the acute infection, approximately 50% of the tilerone-treated animals were able to survive indefinitely (P less than 0.05). The tilerone-treated mice demonstrated significant levels of serum interferon and splenic natural killer cells at the time of infection. Macrophages isolated from the peritoneal cavities of tilerone-treated C57BL/6J mice appeared to kill significant numbers of trypanosomes during 2 to 3 days of in vitro culture, indicating that activated macrophages may contribute to the enhanced resistance to T. cruzi infection in these mice. Beige mice treated with tilerone did not survive T. cruzi infection as well as tilerone-treated heterozygotes did, suggesting a role for natural killer cells in interferon-induced resistance. These results suggest that interferon or effector mechanisms enhanced by interferon induction can play a significant role in influencing resistance to T. cruzi infection. PMID:6173326

  18. Environmental enrichment improves novel object recognition and enhances agonistic behavior in male mice.

    PubMed

    Mesa-Gresa, Patricia; Pérez-Martinez, Asunción; Redolat, Rosa

    2013-01-01

    Environmental enrichment (EE) is an experimental paradigm in which rodents are housed in complex environments containing objects that provide stimulation, the effects of which are expected to improve the welfare of these subjects. EE has been shown to considerably improve learning and memory in rodents. However, knowledge about the effects of EE on social interaction is generally limited and rather controversial. Thus, our aim was to evaluate both novel object recognition and agonistic behavior in NMRI mice receiving EE, hypothesizing enhanced cognition and slightly enhanced agonistic interaction upon EE rearing. During a 4-week period half the mice (n = 16) were exposed to EE and the other half (n = 16) remained in a standard environment (SE). On PND 56-57, animals performed the object recognition test, in which recognition memory was measured using a discrimination index. The social interaction test consisted of an encounter between an experimental animal and a standard opponent. Results indicated that EE mice explored the new object for longer periods than SE animals (P < .05). During social encounters, EE mice devoted more time to sociability and agonistic behavior (P < .05) than their non-EE counterparts. In conclusion, EE has been shown to improve object recognition and increase agonistic behavior in adolescent/early adulthood mice. In the future we intend to extend this study on a longitudinal basis in order to assess in more depth the effect of EE and the consistency of the above-mentioned observations in NMRI mice. PMID:23588702

  19. Effect of Hyperoxia on Retinoid Metabolism and Retinoid Receptor Expression in the Lungs of Newborn Mice

    PubMed Central

    Chen, Hsing-Jin; Chiang, Bor-Luen

    2015-01-01

    Background Preterm newborns that receive oxygen therapy often develop bronchopulmonary dysplasia (BPD), which is abnormal lung development characterized by impaired alveologenesis. Oxygen-mediated injury is thought to disrupt normal lung growth and development. However, the mechanism of hyperoxia-induced BPD has not been extensively investigated. We established a neonatal mouse model to investigate the effects of normobaric hyperoxia on retinoid metabolism and retinoid receptor expression. Methods Newborn mice were exposed to hyperoxic or normoxic conditions for 15 days. The concentration of retinol and retinyl palmitate in the lung was measured by HPLC to gauge retinoid metabolism. Retinoid receptor mRNA levels were assessed by real-time PCR. Proliferation and retinoid receptor expression in A549 cells were assessed in the presence and absence of exogenous vitamin A. Results Hyperoxia significantly reduced the body and lung weight of neonatal mice. Hyperoxia also downregulated expression of RARα, RARγ, and RXRγ in the lungs of neonatal mice. In vitro, hyperoxia inhibited proliferation and expression of retinoid receptors in A549 cells. Conclusion Hyperoxia disrupted retinoid receptor expression in neonatal mice. PMID:26509921

  20. Enhanced subliminal emotional responses to dynamic facial expressions.

    PubMed

    Sato, Wataru; Kubota, Yasutaka; Toichi, Motomi

    2014-01-01

    Emotional processing without conscious awareness plays an important role in human social interaction. Several behavioral studies reported that subliminal presentation of photographs of emotional facial expressions induces unconscious emotional processing. However, it was difficult to elicit strong and robust effects using this method. We hypothesized that dynamic presentations of facial expressions would enhance subliminal emotional effects and tested this hypothesis with two experiments. Fearful or happy facial expressions were presented dynamically or statically in either the left or the right visual field for 20 (Experiment 1) and 30 (Experiment 2) ms. Nonsense target ideographs were then presented, and participants reported their preference for them. The results consistently showed that dynamic presentations of emotional facial expressions induced more evident emotional biases toward subsequent targets than did static ones. These results indicate that dynamic presentations of emotional facial expressions induce more evident unconscious emotional processing. PMID:25250001

  1. Heterozygous L1-deficient mice express an autism-like phenotype.

    PubMed

    Sauce, Bruno; Wass, Christopher; Netrakanti, Meera; Saylor, Joshua; Schachner, Melitta; Matzel, Louis D

    2015-10-01

    The L1CAM (L1) gene encodes a cell adhesion molecule that contributes to several important processes in the developing and adult nervous system, including neuronal migration, survival, and plasticity. In humans and mice, mutations in the X chromosome-linked gene L1 cause severe neurological defects in males. L1 heterozygous female mice with one functional copy of the L1 gene show complex morphological features that are different from L1 fully-deficient and wild-type littermate mice. However, almost no information is available on the behavior of L1 heterozygous mice and humans. Here, we investigated the behavior of heterozygous female mice in which the L1 gene is constitutively inactivated. These mice were compared to wild-type littermate females. Animals were assessed in five categories of behavioral tests: five tests for anxiety/stress/exploration, four tests for motor abilities, two tests for spatial learning, three tests for social behavior, and three tests for repetitive behavior. We found that L1 heterozygous mice express an autism-like phenotype, comprised of reduced social behaviors and excessive self-grooming (a repetitive behavior also typical in animal models of autism). L1 heterozygous mice also exhibited an increase in sensitivity to light, assessed by a reluctance to enter the lighted areas of novel environments. However, levels of anxiety, stress, motor abilities, and spatial learning in L1 heterozygous mice were similar to those of wild-type mice. These observations raise the possibility that using molecules known to trigger L1 functions may become valuable in the treatment of autism in humans. PMID:26079769

  2. Developmental Changes of ENaC Expression and Function in the Inner Ear of Pendrin Knock-Out Mice as a Perspective on the Development of Endolymphatic Hydrops

    PubMed Central

    Kim, Bo Gyung; Kim, Jin Young; Kim, Hee Nam; Bok, Jinwoong; Namkung, Wan

    2014-01-01

    Pendrin mutations cause enlarged vestibular aqueducts and various degrees of sensorineural hearing loss. The selective abolition of pendrin causes dilation of the membranous labyrinth known as endolymphatic hydrops, loss of the endocochlear potential, and consequently loss of hearing function. Because Na+ transport is one of the most important driving forces for fluid transport, the epithelial Na+ channel (ENaC) is believed to play an important role in fluid volume regulation in the inner ear. Therefore, the dysfunction of Na+ transport through ENaC by the acidification of endolymph in Pendred syndrome is one of the potential causes of endolymphatic hydrops. We investigated the changes of ENaC expression and function during the development of the pendrin knock-out mouse. In the cochlea, the expression of β and γENaC was significantly increased at P56 in Pds−/− mice compared with Pds+/+ mice. In the vestibule, the expression of βENaC was significantly increased at P56, and γENaC expression significantly increased from P6 to P56 in Pds−/− mice. The ENaC-dependent trans-epithelial current was not significantly different between Pds+/+ and Pds−/− mice in Reissner’s membrane or the saccular extramacular roof epithelium at P0, but the current was significantly increased in Pds−/− mice at P56 compared with Pds+/+ mice. These findings indicate that the expression and function of ENaC were enhanced in Pds−/− mice after the development of endolymphatic hydrops as a compensatory mechanism. This result provides insight into the role of Na+ transport in the development and regulation of endolymphatic hydrops due to pendrin mutations. PMID:24752462

  3. Developmental changes of ENaC expression and function in the inner ear of pendrin knock-out mice as a perspective on the development of endolymphatic hydrops.

    PubMed

    Kim, Bo Gyung; Kim, Jin Young; Kim, Hee Nam; Bok, Jinwoong; Namkung, Wan; Choi, Jae Young; Kim, Sung Huhn

    2014-01-01

    Pendrin mutations cause enlarged vestibular aqueducts and various degrees of sensorineural hearing loss. The selective abolition of pendrin causes dilation of the membranous labyrinth known as endolymphatic hydrops, loss of the endocochlear potential, and consequently loss of hearing function. Because Na+ transport is one of the most important driving forces for fluid transport, the epithelial Na+ channel (ENaC) is believed to play an important role in fluid volume regulation in the inner ear. Therefore, the dysfunction of Na+ transport through ENaC by the acidification of endolymph in Pendred syndrome is one of the potential causes of endolymphatic hydrops. We investigated the changes of ENaC expression and function during the development of the pendrin knock-out mouse. In the cochlea, the expression of β and γENaC was significantly increased at P56 in Pds-/- mice compared with Pds+/+ mice. In the vestibule, the expression of βENaC was significantly increased at P56, and γENaC expression significantly increased from P6 to P56 in Pds-/- mice. The ENaC-dependent trans-epithelial current was not significantly different between Pds+/+ and Pds-/- mice in Reissner's membrane or the saccular extramacular roof epithelium at P0, but the current was significantly increased in Pds-/- mice at P56 compared with Pds+/+ mice. These findings indicate that the expression and function of ENaC were enhanced in Pds-/- mice after the development of endolymphatic hydrops as a compensatory mechanism. This result provides insight into the role of Na+ transport in the development and regulation of endolymphatic hydrops due to pendrin mutations. PMID:24752462

  4. Hyperoxia increases hepatic arginase expression and ornithine production in mice

    SciTech Connect

    Malleske, Daniel T.; Rogers, Lynette K.; Velluci, Sean M.; Young, Tamara L.; Park, Min S.; Long, Donald W.; Welty, Stephen E.; Smith, Charles V.; Nelin, Leif D. . E-mail: NelinL@pediatrics.ohio-state.edu

    2006-08-15

    Hyperoxic exposure affects the levels and activities of some hepatic proteins. We tested the hypothesis that hyperoxic exposure would result in greater hepatic .NO concentrations. C3H/HeN mice were exposed to >95% O{sub 2} for 72 or 96 h and compared to room air-breathing controls. In contrast to our working hypothesis, exposure to >95% O{sub 2} for 96 h decreased hepatic nitrite/nitrate NO {sub X} concentrations (10.9 {+-} 2.2 nmol/g liver versus 19.3 {+-} 2.4 nmol/g liver in room air, P < 0.05). The hepatic levels of endothelial NO synthase (eNOS) and inducible NOS (iNOS) proteins were not different among the groups. The arginases, which convert L-arginine to urea and L-ornithine, may affect hepatic NOS activities by decreasing L-arginine bioavailability. Hepatic ornithine concentrations were greater in hyperoxic animals than in controls (318 {+-} 18 nmol/g liver in room air, and 539 {+-} 64, and 475 {+-} 40 at 72 and 96 h of hyperoxia, respectively, P < 0.01). Hepatic arginase I protein levels were greater in hyperoxic animals than in controls. Hepatic carbamoyl phosphate synthetase (CPS) protein levels and activities were not different among groups. These results indicate that increases in hepatic levels of arginase I in mice exposed to hyperoxia may diminish .NO production, as reflected by lower liver levels of NO {sub X}. The resultant greater hepatic ornithine concentrations may represent a mechanism to facilitate tissue repair, by favoring the production of polyamines and/or proline.

  5. GFAP expression and social deficits in transgenic mice overexpressing human sAPPα.

    PubMed

    Bailey, Antoinette R; Hou, Huayan; Song, Min; Obregon, Demian F; Portis, Samantha; Barger, Steven; Shytle, Doug; Stock, Saundra; Mori, Takashi; Sanberg, Paul G; Murphy, Tanya; Tan, Jun

    2013-09-01

    Autistic individuals display impaired social interactions and language, and restricted, stereotyped behaviors. Elevated levels of secreted amyloid precursor protein-alpha (sAPPα), the product of α-secretase cleavage of APP, are found in the plasma of some individuals with autism. The sAPPα protein is neurotrophic and neuroprotective and recently showed a correlation to glial differentiation in human neural stem cells (NSCs) via the IL-6 pathway. Considering evidence of gliosis in postmortem autistic brains, we hypothesized that subsets of patients with autism would exhibit elevations in CNS sAPPα and mice generated to mimic this observation would display markers suggestive of gliosis and autism-like behavior. Elevations in sAPPα levels were observed in brains of autistic patients compared to controls. Transgenic mice engineered to overexpress human sAPPα (TgsAPPα mice) displayed hypoactivity, impaired sociability, increased brain glial fibrillary acidic protein (GFAP) expression, and altered Notch1 and IL-6 levels. NSCs isolated from TgsAPPα mice, and those derived from wild-type mice treated with sAPPα, displayed suppressed β-tubulin III and elevated GFAP expression. These results suggest that elevations in brain sAPPα levels are observed in subsets of individuals with autism and TgsAPPα mice display signs suggestive of gliosis and behavioral impairment. PMID:23840007

  6. GFAP expression and social deficits in transgenic mice overexpressing human sAPPα

    PubMed Central

    Bailey, Antoinette R; Hou, Huayan; Song, Min; Obregon, Demian F; Portis, Samantha; Barger, Steven; Shytle, Doug; Stock, Saundra; Mori, Takashi; Sanberg, Paul G; Murphy, Tanya; Tan, Jun

    2013-01-01

    Autistic individuals display impaired social interactions and language, and restricted, stereotyped behaviors. Elevated levels of secreted amyloid precursor protein-alpha (sAPPα), the product of α-secretase cleavage of APP, are found in the plasma of some individuals with autism. The sAPPα protein is neurotrophic and neuroprotective and recently showed a correlation to glial differentiation in human neural stem cells (NSCs) via the IL-6 pathway. Considering evidence of gliosis in postmortem autistic brains, we hypothesized that subsets of patients with autism would exhibit elevations in CNS sAPPα and mice generated to mimic this observation would display markers suggestive of gliosis and autism-like behavior. Elevations in sAPPα levels were observed in brains of autistic patients compared to controls. Transgenic mice engineered to overexpress human sAPPα (TgsAPPα mice) displayed hypoactivity, impaired sociability, increased brain glial fibrillary acidic protein (GFAP) expression, and altered Notch1 and IL-6 levels. NSCs isolated from TgsAPPα mice, and those derived from wild-type mice treated with sAPPα, displayed suppressed β-tubulin III and elevated GFAP expression. These results suggest that elevations in brain sAPPα levels are observed in subsets of individuals with autism and TgsAPPα mice display signs suggestive of gliosis and behavioral impairment. PMID:23840007

  7. Dysregulation of temperature and liver cytokine gene expression in immunodeficient wasted mice

    SciTech Connect

    Libertin, C.R.; Ling-Indeck, L.; Weaver, P.; Chang-Liu, Chin-Mei; Strezoska, V.; Heckert, B.; Woloschak, G.E. |

    1995-04-25

    Wasted mice bear the spontaneous autosomal recessive mutation wst/wst; this genotype is associated with weight loss beginning at 21 days of age, neurologic dysfunction, immunodeficiency at mucosal sites, and increased sensitivity to the killing effects of ionizing radiation. The pathology underlying the disease symptoms is unknown. Experiments reported here were designed to examine thermoregulation and liver expression of specific cytokines in wasted mice and in littermate and parental controls. Our experiments found that wasted mice begin to show a drop in body temperature at 21-23 days following birth, continuing until death at the age of 28 days. Concomitant with that, livers from wasted mice expressed increased amounts of mRNAs specific for cytokines IL,6 and IL-1, the acute phase reactant C-reactive protein, c-jun, and apoptosis-associated Rp-8 when compared to littermate and parental control animals. Levels of {beta}-transforming growth factor (TGF), c-fos, proliferating cell nuclear antigen (PCNA), and ornithine amino transferase (OAT) transcripts were the same in livers from wasted mice and controls. These results suggest a relationship between an acute phase reactant response in wasted mice and temperature dysregulation.

  8. Accentuated response to phenylhydrazine and erythropoietin in mice genetically impaired for their GATA-1 expression (GATA-1(low) mice).

    PubMed

    Vannucchi, A M; Bianchi, L; Cellai, C; Paoletti, F; Carrai, V; Calzolari, A; Centurione, L; Lorenzini, R; Carta, C; Alfani, E; Sanchez, M; Migliaccio, G; Migliaccio, A R

    2001-05-15

    The response of mice genetically unable to up-regulate GATA-1 expression (GATA-1(low) mice) to acute (phenylhydrazine [PHZ]-induced anemia) and chronic (in vivo treatment for 5 days with 10 U erythropoietin [EPO] per mouse) erythroid stimuli was investigated. Adult GATA-1(low) mice are profoundly thrombocytopenic (platelet counts [x 10(9)/L] 82.0 +/- 28.0 vs 840 +/- 170.0 of their control littermates, P <.001) but have a normal hematocrit (Hct) (approximately.47 proportion of 1.0 [47%]). The spleens of these mutants are 2.5-fold larger than normal and contain 5-fold more megakaryocytic (4A5(+)), erythroid (TER-119(+)), and bipotent (erythroid/megakaryocytic, TER-119(+)/4A5(+)) precursor cells. Both the marrow and the spleen of these animals contain higher frequencies of burst-forming units-erythroid (BFU-E)- and colony-forming units-erythroid (CFU-E)-derived colonies (2-fold and 6-fold, respectively) than their normal littermates. The GATA-1(low) mice recover 2 days faster from the PHZ-induced anemia than their normal littermates (P <.01). In response to EPO, the Hct of the GATA-1(low) mice raised to.68 proportion of 1.0 (68%) vs the.55 proportion of 1.0 (55%) reached by the controls (P <.01). Both the GATA-1(low) and the normal mice respond to PHZ and EPO with similar (2- to 3-fold) increases in size and cellularity of the spleen (increases are limited mostly to cells, both progenitor and precursor, of the erythroid lineage). However, in spite of the similar relative cellular increases, the increases of all these cell populations are significantly higher, in absolute cell numbers, in the mutant than in the wild-type mice. In conclusion, the GATA-1(low) mutation increases the magnitude of the response to erythroid stimuli as a consequence of the expansion of the erythroid progenitor cells in their spleen. PMID:11342429

  9. FOLATE DEFICIENCY ENHANCES ARSENIC-INDUCED GENOTOXICITY IN MICE

    EPA Science Inventory

    Folate deficiency increases background levels of DNA damage and can enhance the mutagenicity of chemical agents. Duplicate experiments were performed to investigate the effect of dietary folate deficiency on arsenic induction of micronuclei (MN) in peripheral blood cells. Male C5...

  10. Resistance to Malaria by Enhanced Phagocytosis of Erythrocytes in LMP7-deficient Mice

    PubMed Central

    Duan, Xuefeng; Imai, Takashi; Chou, Bin; Tu, Liping; Himeno, Kunisuke; Suzue, Kazutomo; Hirai, Makoto; Taniguchi, Tomoyo; Okada, Hiroko; Shimokawa, Chikako; Hisaeda, Hajime

    2013-01-01

    General cellular functions of proteasomes occur through protein degradation, whereas the specific function of immunoproteasomes is the optimization of antigen processing associated with MHC class I. We and others previously reported that deficiency in subunits of immunoproteasomes impaired the activation of antigen-specific CD8+ T cells, resulting in higher susceptibility to tumor and infections. We demonstrated that CD8+ T cells contributed to protection against malaria parasites. In this study, we evaluated the role of immunoproteasomes in the course of infection with rodent malaria parasites. Unexpectedly, Plasmodium yoelii infection of mice deficient in LMP7, a catalytic subunit of immunoproteasomes, showed lower parasite growth in the early phase of infection and lower lethality compared with control mice. The protective characteristics of LMP7-deficient mice were not associated with enhanced immune responses, as the mutant mice showed comparable or diminished activation of innate and acquired immunity. The remarkable difference was observed in erythrocytes instead of immune responses. Parasitized red blood cells (pRBCs) purified from LMP7-deficient mice were more susceptible to phagocytosis by macrophages compared with those from wild-type mice. The susceptibility of pRBC to phagocytosis appeared to correlate with deformity of the membrane structures that were only observed after infection. Our results suggest that RBCs of LMP7-deficient mice were more likely to deform in response to infection with malaria parasites, presumably resulting in higher susceptibility to phagocytosis and in the partial resistance to malaria. PMID:23527234

  11. F. novicida-Infected A. castellanii Does Not Enhance Bacterial Virulence in Mice

    PubMed Central

    Ozanic, Mateja; Gobin, Ivana; Brezovec, Martin; Marecic, Valentina; Trobonjaca, Zlatko; Abu Kwaik, Yousef; Santic, Marina

    2016-01-01

    Francisella tularensis is a facultative intracellular bacterium that causes tularemia in humans and animals. Epidemiology of tularemia worldwide is often associated with water-borne transmission, which includes mosquitoes and amoebae as the potential host reservoirs of the bacteria in water environment. In vitro studies showed intracellular replication of F. tularensis within Acanthamoeba castellanii and Hartmanella vermiformis cells. While infection of amoeba by Legionella pneumophila has been shown to enhance infectivity of L. pneumophila the role of F. tularensis-infected protozoa in the pathogenesis of tularemia is not known. We used 6 h coculture of A. castellanii and F. novicida for investigation of the effect of inhaled amoeba on the pathogenesis of tularemia on in vivo model. Balb/c mice were infected intratracheally with F. novicida or with F. novicida-infected A. castellanii. Surprisingly, infection with F. novicida-infected A. castellanii did not lead to bronchopneumonia in Balb/c mice, and Francisella did not disseminate into the liver and spleen. Upon inhalation, F. novicida infects a variety of host cells, though neutrophils are the predominant cells early during infection in the lung infiltrates of pulmonary tularemia. The numbers of neutrophils in the lungs of Balb/c mice were significantly lower in the infection of mice with F. novicida-infected A. castellanii in comparison to group of mice infected only with F. novicida. These results demonstrate that following inoculation of mice with F. novicida-infected A. castellanii, mice did not develop tularemia. PMID:27242974

  12. Anodal transcranial direct current stimulation boosts synaptic plasticity and memory in mice via epigenetic regulation of Bdnf expression

    PubMed Central

    Podda, Maria Vittoria; Cocco, Sara; Mastrodonato, Alessia; Fusco, Salvatore; Leone, Lucia; Barbati, Saviana Antonella; Colussi, Claudia; Ripoli, Cristian; Grassi, Claudio

    2016-01-01

    The effects of transcranial direct current stimulation (tDCS) on brain functions and the underlying molecular mechanisms are yet largely unknown. Here we report that mice subjected to 20-min anodal tDCS exhibited one-week lasting increases in hippocampal LTP, learning and memory. These effects were associated with enhanced: i) acetylation of brain-derived neurotrophic factor (Bdnf) promoter I; ii) expression of Bdnf exons I and IX; iii) Bdnf protein levels. The hippocampi of stimulated mice also exhibited enhanced CREB phosphorylation, pCREB binding to Bdnf promoter I and recruitment of CBP on the same regulatory sequence. Inhibition of acetylation and blockade of TrkB receptors hindered tDCS effects at molecular, electrophysiological and behavioral levels. Collectively, our findings suggest that anodal tDCS increases hippocampal LTP and memory via chromatin remodeling of Bdnf regulatory sequences leading to increased expression of this gene, and support the therapeutic potential of tDCS for brain diseases associated with impaired neuroplasticity. PMID:26908001

  13. Genome-wide association study of behavioral, physiological and gene expression traits in outbred CFW mice.

    PubMed

    Parker, Clarissa C; Gopalakrishnan, Shyam; Carbonetto, Peter; Gonzales, Natalia M; Leung, Emily; Park, Yeonhee J; Aryee, Emmanuel; Davis, Joe; Blizard, David A; Ackert-Bicknell, Cheryl L; Lionikas, Arimantas; Pritchard, Jonathan K; Palmer, Abraham A

    2016-08-01

    Although mice are the most widely used mammalian model organism, genetic studies have suffered from limited mapping resolution due to extensive linkage disequilibrium (LD) that is characteristic of crosses among inbred strains. Carworth Farms White (CFW) mice are a commercially available outbred mouse population that exhibit rapid LD decay in comparison to other available mouse populations. We performed a genome-wide association study (GWAS) of behavioral, physiological and gene expression phenotypes using 1,200 male CFW mice. We used genotyping by sequencing (GBS) to obtain genotypes at 92,734 SNPs. We also measured gene expression using RNA sequencing in three brain regions. Our study identified numerous behavioral, physiological and expression quantitative trait loci (QTLs). We integrated the behavioral QTL and eQTL results to implicate specific genes, including Azi2 in sensitivity to methamphetamine and Zmynd11 in anxiety-like behavior. The combination of CFW mice, GBS and RNA sequencing constitutes a powerful approach to GWAS in mice. PMID:27376237

  14. Leptin deficiency leads to the regulation of kinin receptors expression in mice.

    PubMed

    Abe, Karina Camasmie; Mori, Marcelo Alves da Silva; Pesquero, Joao Bosco

    2007-02-01

    Kinins are vasoactive and pro-inflammatory peptides generated by the cleavage of the kininogen by kallikreins. Two kinin receptors have been described and denominated B1 and B2. Obesity frequently accompanies other pathologies, such as diabetes and hypertention. The clustering of these pathologies is usually known as "metabolic syndrome". Mice lacking leptin gene (ob/ob) are severely obese and hyperphagic. Using quantitative RT-PCR analysis of B1 and B2 mRNAs expression, we described for the first time a correlation between the kallikrein-kinin system (KKS) and severe obesity in mice. The ob/ob mice presented lower expression of B2 mRNA in the white adipose tissue (WAT) and hypothalamus, both primary sites for neuroendocrine regulation of the energetic metabolism. B1 mRNA, however, is overexpressed in these tissues of ob/ob mice. An upregulation of the B1 mRNA has also been seen in liver, abdominal aorta and stomach fundus. However, different from the lean mice, the expression of the B1 mRNA in brown adipose tissue (BAT) and heart is completely abolished. Our data show that kinin receptors are differently modulated in distinct tissues in obesity. These findings suggest a connection between the KKS and obesity, and suggest that kinin receptors could be involved in the ethiopathogenesis of the metabolic syndrome. PMID:17184856

  15. Increased expression of FGF1-mediated signaling molecules in adipose tissue of obese mice.

    PubMed

    Choi, Youngshim; Jang, Suhyeon; Choi, Myung-Sook; Ryoo, Zae Young; Park, Taesun

    2016-06-01

    Fibroblast growth factors (FGFs) are pleiotropic growth factors that control cell proliferation, migration, and differentiation. Herein, we evaluated whether visceral adiposity of mice is accompanied by the alteration of signaling molecules mediated by fibroblast growth factor receptor 1 (FGFR1) induced by using two different male C57BL/6J mice models of obesity namely high-fat diet (HFD)-induced obesity for 12 weeks or mice with genetic deletion of leptin (ob/ob). Both HFD-fed and ob/ob mice exhibited significantly higher messenger RNA (mRNA) levels of FGF1, cyclin D (cycD), transcription factor E2F1, peroxisome proliferator-activated receptor-gamma 2 (PPAR-γ2), CCAAT-enhancer-binding protein alpha (C/EBPα), and adipocyte protein 2 (aP2) genes in their epididymal adipose tissues compared to those of the normal diet (ND)-fed and lean control mice, respectively. In addition, immunoblot analyses of the epididymal adipose tissues revealed that both mice exposed to HFD and ob/ob mice exhibited elevated phosphorylation of FGFR1, extracellular-signal-regulated kinase (ERK), and retinoblastoma (Rb) proteins. These data support the notion that FGF1-mediated signaling represents an important signaling cascade related to adipogenesis, at least partially, among other known signaling pathways. These new findings regarding the molecular mechanisms controlling adipose tissue plasticity provide a novel insight about the functional network with potential therapeutic application against obesity. PMID:26847131

  16. Coxsackievirus B 1-induced polymyositis. Lack of disease expression in nu/nu mice.

    PubMed Central

    Ytterberg, S R; Mahowald, M L; Messner, R P

    1987-01-01

    Chronic inflammatory myositis similar to human polymyositis occurs in mice after infection with a strain of Coxsackievirus B 1 (CVB 1). To investigate the role of T cells in the pathogenesis of this disorder, we compared disease expression in T cell-deficient athymic nude (nu/nu) mice and heterozygotes (nu/+) with normal T cell function. Acute infectious myositis occurred in nu/nu and nu/+ mice. Chronic (greater than 21 d postinfection) weakness and myositis, however, developed only in nu/+. Resistance to disease in nu/nu mice was not explained by insusceptibility to infection; the amount of virus lethal for 50% of mice and virus replication were comparable in both groups. Additionally, anti-CVB 1 antibody production was similar in both groups. Reconstitution of infected nu/nu mice with spleen cells from normal mice resulted in disease. These results demonstrate that chronic weakness after infection with this virus is not simply a sequela of acute myonecrosis and suggest that T cells play a pivotal role in the pathogenesis of chronic myositis. Images PMID:3038960

  17. Impaired pulmonary host defense in mice lacking expression of the CXC chemokine lungkine.

    PubMed

    Chen, S C; Mehrad, B; Deng, J C; Vassileva, G; Manfra, D J; Cook, D N; Wiekowski, M T; Zlotnik, A; Standiford, T J; Lira, S A

    2001-03-01

    Lungkine (CXCL15) is a novel CXC chemokine that is highly expressed in the adult mouse lung. To determine the biologic function of Lungkine, we generated Lungkine null mice by targeted gene disruption. These mice did not differ from wild-type mice in their hematocrits or in the relative number of cells in leukocyte populations of peripheral blood or other tissues, including lung and bone marrow. However, Lungkine null mice were more susceptible to Klebsiella pneumonia infection, with a decreased survival and increased lung bacterial burden compared with infected wild-type mice. Histologic analysis of the lung and assessment of leukocytes in the bronchioalveolar lavage revealed that neutrophil numbers were normal in the lung parenchyma, but reduced in the airspace. The production of other neutrophil chemoattractants in the Lungkine null mice did not differ from that in wild-type mice, and neutrophil migration into other tissues was normal. Taken together, these findings demonstrate that Lungkine is an important mediator of neutrophil migration from the lung parenchyma into the airspace. PMID:11207292

  18. Impaired growth of pancreatic exocrine cells in transgenic mice expressing human activin {beta}E subunit

    SciTech Connect

    Hashimoto, Osamu . E-mail: ohashim@vmas.kitasato-u.ac.jp; Ushiro, Yuuki; Sekiyama, Kazunari; Yamaguchi, Osamu; Yoshioka, Kazuki; Mutoh, Ken-Ichiro; Hasegawa, Yoshihisa

    2006-03-10

    Activins, TGF-{beta} superfamily members, have multiple functions in a variety of cells and tissues. Recently, additional activin {beta} subunit genes, {beta}C and {beta}E, have been identified. To explore the role of activin E, we created transgenic mice overexpressing human activin {beta}E subunit. There were pronounced differences in the pancreata of the transgenic animals as compared with their wild-type counterparts. Pancreatic weight, expressed relative to total body weight, was significantly reduced. Histologically, adipose replacement of acini in the exocrine pancreas was observed. There was a significant decrease in the number of PCNA-positive cells in the acinar cells, indicating reduced proliferation in the exocrine pancreas of the transgenic mice. However, quantitative pancreatic morphometry showed that the total number and mass of the islets of the transgenic mice were comparable with those of the nontransgenic control mice. Our findings suggest a role for activin E in regulating the proliferation of pancreatic exocrine cells.

  19. Humanized TLR7/8 expression drives proliferative multisystemic histiocytosis in C57BL/6 mice.

    PubMed

    Snyder, Jessica M; Treuting, Piper M; Nagy, Lee; Yam, Cathy; Yi, Jaehun; Brasfield, Alicia; Nguyen, Lisa Phuong Anh; Hajjar, Adeline M

    2014-01-01

    A humanized TLR7/TLR8 transgenic mouse line was engineered for studies using TLR7/8 ligands as vaccine adjuvants. The mice developed a spontaneous immune-mediated phenotype prior to six months of age characterized by runting, lethargy, blepharitis, and corneal ulceration. Histological examination revealed a marked, multisystemic histiocytic infiltrate that effaced normal architecture. The histological changes were distinct from those previously reported in mouse models of systemic lupus erythematosus. When the mice were crossed with MyD88-/- mice, which prevented toll-like receptor signaling, the inflammatory phenotype resolved. Illness may be caused by constitutive activation of human TLR7 or TLR8 in the bacterial artificial chromosome positive mice as increased TLR7 and TLR8 expression or activation has previously been implicated in autoimmune disease. PMID:25229618

  20. Chronic Inflammatory Liver Disease in Mice Expressing a CD28-specific Ligand

    PubMed Central

    Corse, Emily; Gottschalk, Rachel A.; Park, Joon Seok; Sepulveda, Manuel A.; Loke, P’ng; Sullivan, Timothy J.; Johnson, Linda K.; Allison, James P.

    2012-01-01

    Inflammation of the normally tolerant liver microenvironment precedes development of chronic liver disease. Study of the pathogenesis of autoimmune liver diseases such as autoimmune hepatitis (AIH) has been hampered by a lack of autochthonous chronic animal models. Through our studies of T cell costimulation, we generated transgenic mice expressing a ligand specific for the CD28 receptor, which normally shares ligands with the related inhibitory receptor CTLA-4. The mice spontaneously develop chronic inflammatory liver disease with several pathologies found in AIH including elevated serum aminotransferases in the context of normal alkaline phosphatase and bilirubin levels, lymphocytic inflammation, focal necrosis, oval cell hyperplasia, and fibrosis. The prevalence of IFN-γ-producing CD8+ T cells in the livers of transgenic mice suggests a role for autoimmune cytotoxicity in the chronic disease state. The CD28 ligand-specific transgenic mice will facilitate evaluation of CD8+ T cell function in liver disease pathologies found in AIH. PMID:23248264

  1. The expression of thioredoxin-1 in acute epinephrine stressed mice.

    PubMed

    Jia, Jin-Jing; Zeng, Xian-Si; Li, Kun; Ma, Li-Fang; Chen, Lei; Song, Xin-Qiang

    2016-09-01

    Stress, a state of perceived threat to homeostasis, regulates a panel of important physiological functions. The human mind and body respond to stress by activating the sympathetic nervous system and secreting the catecholamines epinephrine and norepinephrine in the "fight-or-flight" response. However, the protective mechanism of acute stress is still unknown. In the present study, an acute stress mouse model was constructed by intraperitoneal injection of epinephrine (0.2 mg kg(-1)) for 4 h. Epinephrine treatment induced heat shock 70(Hsp70) expression in the stress responsive tissues, such as the cortex, hippocampus, thymus, and kidney. Further, the expression of thioredoxin-1(Trx-1), a cytoprotective protein, was also upregulated in these stress responsive tissues. In addition, the phosphorylation of cAMP-response element binding protein (CREB), a transcription factor of Trx-1, was increased after treatment with epinephrine. The block of CREB activation by H89 inhibited the acute epinephrine stress-induced Trx-1 and Hsp70 expression. Taken together, our data suggest that acute stimuli of epinephrine induced Trx-1 expression through activating CREB and may represent a protective role against stress. PMID:27511023

  2. Exposure to diesel exhaust up-regulates iNOS expression in ApoE knockout mice

    SciTech Connect

    Bai Ni; Kido, Takashi; Kavanagh, Terrance J.; Kaufman, Joel D.; Rosenfeld, Michael E.; Breemen, Cornelis van; Eeden, Stephan F. van

    2011-09-01

    Traffic related particulate matter air pollution is a risk factor for cardiovascular events; however, the biological mechanisms are unclear. We hypothesize that diesel exhaust (DE) inhalation induces up-regulation of inducible nitric oxide synthase (iNOS), which is known to contribute to vascular dysfunction, progression of atherosclerosis and ultimately cardiovascular morbidity and mortality. Methods: ApoE knockout mice (30-week) were exposed to DE (at 200 {mu}g/m{sup 3} of particulate matter) or filtered-air (control) for 7 weeks (6 h/day, 5 days/week). iNOS expression in the blood vessels and heart was evaluated by immunohistochemistry and western blotting analysis. To examine iNOS activity, thoracic aortae were mounted in a wire myograph, and vasoconstriction stimulated by phenylephrine (PE) was measured with and without the presence of the specific inhibitor for iNOS (1400 W). NF-{kappa}B (p65) activity was examined by ELISA. The mRNA expression of iNOS and NF-{kappa}B (p65) was determined by real-time PCR. Results: DE exposure significantly enhanced iNOS expression in the thoracic aorta (4-fold) and heart (1.5 fold). DE exposure significantly attenuated PE-stimulated vasoconstriction by {approx} 20%, which was partly reversed by 1400 W. The mRNA expression of iNOS and NF-{kappa}B was significantly augmented after DE exposure. NF-{kappa}B activity was enhanced 2-fold after DE inhalation, and the augmented NF-{kappa}B activity was positively correlated with iNOS expression (R{sup 2} = 0.5998). Conclusions: We show that exposure to DE increases iNOS expression and activity possibly via NF-{kappa}B-mediated pathway. We suspect that DE exposure-caused up-regulation of iNOS contributes to vascular dysfunction and atherogenesis, which could ultimately lead to urban air pollution-associated cardiovascular morbidity and mortality. - Highlights: > Exposed ApoE knockout mice (30-week) to diesel exhaust (DE) for 7 weeks. > Examine iNOS expression and activity in the

  3. In vivo imaging of clock gene expression in multiple tissues of freely moving mice.

    PubMed

    Hamada, Toshiyuki; Sutherland, Kenneth; Ishikawa, Masayori; Miyamoto, Naoki; Honma, Sato; Shirato, Hiroki; Honma, Ken-Ichi

    2016-01-01

    Clock genes are expressed throughout the body, although how they oscillate in unrestrained animals is not known. Here, we show an in vivo imaging technique that enables long-term simultaneous imaging of multiple tissues. We use dual-focal 3D tracking and signal-intensity calibration to follow gene expression in a target area. We measure circadian rhythms of clock genes in the olfactory bulb, right and left ears and cortices, and the skin. In addition, the kinetic relationship between gene expression and physiological responses to experimental cues is monitored. Under stable conditions gene expression is in phase in all tissues. In response to a long-duration light pulse, the olfactory bulb shifts faster than other tissues. In Cry1(-/-) Cry2(-/-) arrhythmic mice circadian oscillation is absent in all tissues. Thus, our system successfully tracks circadian rhythms in clock genes in multiple tissues in unrestrained mice. PMID:27285820

  4. Ursolic acid enhances pentobarbital-induced sleeping behaviors via GABAergic neurotransmission in mice.

    PubMed

    Jeon, Se Jin; Park, Ho Jae; Gao, Qingtao; Pena, Irene Joy Dela; Park, Se Jin; Lee, Hyung Eun; Woo, Hyun; Kim, Hee Jin; Cheong, Jae Hoon; Hong, Eunyoung; Ryu, Jong Hoon

    2015-09-01

    Prunella vulgaris is widely used as a herbal medicine for cancers, inflammatory diseases, and other infections. Although it has long been used, few studies have examined its effects on central nervous system function. Here, we first observed that ethanolic extracts of P. vulgaris (EEPV) prolonged pentobarbital-induced sleep duration in mice. It is known that EEPV consists of many active components including triterpenoid (ursolic acid and oleanolic acid), which have many biological activities. Therefore, we evaluated which EEPV components induced sleep extension in pentobarbital-mediated sleeping model in mice. Surprisingly, despite their structural similarity and other common functions such as anti-inflammation, anti-cancer, and tissue protection, only ursolic acid enhanced sleep duration in pentobarbital-treated mice. These results were attenuated by bicuculline treatment, which is a GABAA receptor antagonist. The present results suggest that ursolic acid from P. vulgaris enhances sleep duration through GABAA receptor activation and could be a therapeutic candidate for insomnia treatment. PMID:26102564

  5. Taste information derived from T1R-expressing taste cells in mice.

    PubMed

    Yoshida, Ryusuke; Ninomiya, Yuzo

    2016-03-01

    The taste system of animals is used to detect valuable nutrients and harmful compounds in foods. In humans and mice, sweet, bitter, salty, sour and umami tastes are considered the five basic taste qualities. Sweet and umami tastes are mediated by G-protein-coupled receptors, belonging to the T1R (taste receptor type 1) family. This family consists of three members (T1R1, T1R2 and T1R3). They function as sweet or umami taste receptors by forming heterodimeric complexes, T1R1+T1R3 (umami) or T1R2+T1R3 (sweet). Receptors for each of the basic tastes are thought to be expressed exclusively in taste bud cells. Sweet (T1R2+T1R3-expressing) taste cells were thought to be segregated from umami (T1R1+T1R3-expressing) taste cells in taste buds. However, recent studies have revealed that a significant portion of taste cells in mice expressed all T1R subunits and responded to both sweet and umami compounds. This suggests that sweet and umami taste cells may not be segregated. Mice are able to discriminate between sweet and umami tastes, and both tastes contribute to behavioural preferences for sweet or umami compounds. There is growing evidence that T1R3 is also involved in behavioural avoidance of calcium tastes in mice, which implies that there may be a further population of T1R-expressing taste cells that mediate aversion to calcium taste. Therefore the simple view of detection and segregation of sweet and umami tastes by T1R-expressing taste cells, in mice, is now open to re-examination. PMID:26912569

  6. Gene expression microarray analysis of the sciatic nerve of mice with diabetic neuropathy.

    PubMed

    Zhang, Lei; Qu, Shen; Liang, Aibin; Jiang, Hong; Wang, Hao

    2015-02-01

    The present study aimed to explore novel target genes that regulate the development of diabetic neuropathy (DN) by analyzing gene expression profiles in the sciatic nerve of infected mice. The GSE11343 microarray dataset, which was downloaded from Gene Expression Omnibus, included data on 4 control samples and 5 samples from mice with diabetes induced by streptozotocin (STZ), 5 samples from normal mice treated with rosiglitazone (Rosi) and 5 samples from mice with diabetes induced by STZ and treated with Rosi. Differentially expressed genes (DEGs) between the different groups were identified using the substitution augmentation modification redefinition (SAMR) model. The Gene Ontology (GO) term and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses were performed using the Database for Annotation, Visualization and Integrated Discovery (DAVID). Regulatory and protein‑protein interaction networks were searched using BioCarta and STRING, respectively. The protein structures of potential regulatory genes were predicted using the SYBYL program. Compared with the controls, 1,384 DEGs were identified in the mice with STZ-induced diabetes and 7 DEGs were identified in the mice treated with Rosi. There were 518 DEGs identified between the mice in the STZ + Rosi and STZ groups. We identified 45 GO items, and the calmodulin nerve phosphatase and chemokine signaling pathways were identified as the main pathways. Three genes [myristoylated alanine-rich protein kinase C substrate (Marcks), GLI pathogenesis-related 2 (Glipr2) and centrosomal protein 170 kDa (Cep170)] were found to be co-regulated by both STZ and Rosi, the protein structure of which was predicted and certain binding activity to Rosi was docked. Our study demonstrates that the Marcks, Glipr2 and Cep170 genes may be underlying drug targets in the treatment of DN. PMID:25435094

  7. Gene expression microarray analysis of the sciatic nerve of mice with diabetic neuropathy

    PubMed Central

    ZHANG, LEI; QU, SHEN; LIANG, AIBIN; JIANG, HONG; WANG, HAO

    2015-01-01

    The present study aimed to explore novel target genes that regulate the development of diabetic neuropathy (DN) by analyzing gene expression profiles in the sciatic nerve of infected mice. The GSE11343 microarray dataset, which was downloaded from Gene Expression Omnibus, included data on 4 control samples and 5 samples from mice with diabetes induced by streptozotocin (STZ), 5 samples from normal mice treated with rosiglitazone (Rosi) and 5 samples from mice with diabetes induced by STZ and treated with Rosi. Differentially expressed genes (DEGs) between the different groups were identified using the substitution augmentation modification redefinition (SAMR) model. The Gene Ontology (GO) term and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses were performed using the Database for Annotation, Visualization and Integrated Discovery (DAVID). Regulatory and protein-protein interaction networks were searched using BioCarta and STRING, respectively. The protein structures of potential regulatory genes were predicted using the SYBYL program. Compared with the controls, 1,384 DEGs were identified in the mice with STZ-induced diabetes and 7 DEGs were identified in the mice treated with Rosi. There were 518 DEGs identified between the mice in the STZ + Rosi and STZ groups. We identified 45 GO items, and the calmodulin nerve phosphatase and chemokine signaling pathways were identified as the main pathways. Three genes [myristoylated alanine-rich protein kinase C substrate (Marcks), GLI pathogenesis-related 2 (Glipr2) and centrosomal protein 170 kDa (Cep170)] were found to be co-regulated by both STZ and Rosi, the protein structure of which was predicted and certain binding activity to Rosi was docked. Our study demonstrates that the Marcks, Glipr2 and Cep170 genes may be underlying drug targets in the treatment of DN. PMID:25435094

  8. Visualization of Signaling Molecules During Neutrophil Recruitment in Transgenic Mice Expressing FRET Biosensors.

    PubMed

    Mizuno, Rei; Kamioka, Yuji; Sakai, Yoshiharu; Matsuda, Michiyuki

    2016-01-01

    A number of chemical mediators regulate neutrophil recruitment to inflammatory sites either positively or negatively. Although the actions of each chemical mediator on the intracellular signaling networks controlling cell migration have been studied with neutrophils cultured in vitro, how such chemical mediators act cooperatively or counteractively in vivo remains largely unknown. To understand the mechanisms regulating neutrophil recruitment to the inflamed intestine in vivo, we recently generated transgenic mice expressing biosensors based on FRET (Förster resonance energy transfer) and set up two-photon excitation microscopy to observe the gastrointestinal tract in living mice. By measuring FRET in neutrophils, we showed activity changes of protein kinases in the neutrophils recruited to inflamed intestines. In this chapter, we describe the protocol used to visualize the protein kinase activities in neutrophils of the inflamed intestine of transgenic mice expressing the FRET biosensors. PMID:27246030

  9. Electroacupuncture Reduces Aβ Production and BACE1 Expression in SAMP8 Mice

    PubMed Central

    Dong, Wei-Guo; Wang, Feng; Chen, Ye; Zheng, Xue-Hua; Xie, Yong-Cai; Guo, Wan-Qing; Shi, Hong

    2015-01-01

    Electroacupuncture (EA) has been reported to have beneficial effects on Alzheimer’s disease (AD). BACE1 (β-site amyloid precursor protein-cleaving enzyme 1) is involved in the abnormal production of amyloid-β plaque (Aβ), a hallmark of AD pathophysiology. Thus, the present study investigated the effects of EA on memory impairment, Aβ production, and BACE1 expression in senescence-accelerated mouse prone 8 (SAMP8) mice. We found that EA improved spatial learning and memory impairment of SAMP8 mice. Furthermore, EA attenuated Aβ production and repressed the expression of BACE1 in the hippocampus of SAMP8 mice. Taken together, our results suggest that EA could have a potential therapeutic application in AD and that BACE1 may be an important target of EA in the treatment of AD. PMID:26283960

  10. Systemic inflammation enhances surgery-induced cognitive dysfunction in mice.

    PubMed

    Fidalgo, António Rei; Cibelli, Mario; White, John P M; Nagy, Istvan; Maze, Mervyn; Ma, Daqing

    2011-07-01

    The activation of the immune system, by either lipopolysaccharide (LPS) administration or surgical trauma, has been shown to be capable of affecting hippocampal function, causing memory impairment. Here, we examined the extent to which LPS-induced infection may aggravate impairment of memory function following orthopaedic surgery. Hippocampal memory function impairment was assessed using fear-conditioning tasks, while IL-1β levels in plasma and hippocampus were measured using ELISA. LPS-induced inflammation disrupted hippocampal memory consolidation as evidenced by reduced contextual freezing time exhibited by infected mice. Likewise, surgery caused hippocampal-dependent memory impairment, which was associated with increased levels of IL-1β both in plasma and hippocampus. However, a sub-pyrogenic dose of LPS alone failed to impair memory function. This dose of LPS, when administered prior to surgery, exacerbated surgery-induced cognitive dysfunction as evidenced by further reduction of contextual freezing time. Also, it caused a concomitant additional increase in the levels of IL-1β in both plasma and hippocampus of those animals. Our data suggest that sub-clinical infection may sensitise the immune system augmenting the severity of post-operative cognitive dysfunction. PMID:21575676

  11. Subhepatotoxic exposure to arsenic enhances lipopolysaccharide-induced liver injury in mice

    PubMed Central

    Arteel, Gavin E.; Guo, Luping; Schlierf, Thomas; Beier, Juliane I; Kaiser, J. Phillip; Chen, Theresa S; Liu, Marsha; Conklin, Daniel P.; Miller, Heather L.; von Montfort, Claudia; States, J. Christopher

    2008-01-01

    Exposure to arsenic via drinking water is a serious health concern in the US. Whereas studies have identified arsenic alone as an independent risk factor for liver disease, concentrations of arsenic required to damage this organ are generally higher than found in the US water supply. The purpose of the current study was to test the hypothesis that arsenic (at subhepatotoxic doses) may also sensitize the liver to a second hepatotoxin. To test this hypothesis, the effect of chronic exposure to arsenic on liver damage caused by acute lipopolysaccharide (LPS) was determined in mice. Male C57Bl/6J mice (4-6 weeks) were exposed to arsenic (49 ppm as sodium arsenite in drinking water). After 7 months of exposure, animals were injected with LPS (10 mg/kg i.p.) and sacrificed 24 h later. Arsenic alone caused no overt hepatotoxicity, as determined by plasma enzymes and histology. In contrast, arsenic exposure dramatically enhanced liver damage caused by LPS, increasing the number and size of necroinflammatory foci. This effect of arsenic was coupled with increases in indices of oxidative stress (4-HNE adducts, depletion of GSH and methionine pools). The number of apoptotic (TUNEL) hepatocytes was similar in the LPS and arsenic/LPS groups. In contrast, arsenic pre-exposure blunted the increase in proliferating (PCNA) hepatocytes caused by LPS; this change in the balance between cell death and proliferation was coupled with a robust loss of liver weight in the arsenic/LPS compared to the LPS alone group. The impairment of proliferation after LPS caused by arsenic was also coupled with alterations in the expression of key mediators of cell cycle progression (p27, p21, CDK6 and Cyclin D1). Taken together, these results suggest that arsenic, at doses that are not overtly hepatotoxic per se, significantly enhances LPS-induced liver injury. These results further suggest that arsenic levels in the drinking water may be a risk modifier for the development of chronic liver diseases

  12. HemoHIM enhances the therapeutic efficacy of ionizing radiation treatment in tumor-bearing mice.

    PubMed

    Park, Hae-Ran; Ju, Eun-Jin; Jo, Sung-Kee; Jung, Uhee; Kim, Sung-Ho

    2010-02-01

    Although radiotherapy is commonly used for a variety of cancers, radiotherapy alone does not achieve a satisfactory therapeutic outcome. In this study, we examined the possibility that HemoHIM can enhance the anticancer effects of ionizing radiation (IR) in melanoma-bearing mice. The HemoHIM was prepared by adding the ethanol-insoluble fraction to the total water extract of a mixture of three edible herbs-Angelica Radix, Cnidium Rhizoma, and Paeonia Radix. Anticancer effects of HemoHIM were evaluated in melanoma-bearing mice exposed to IR. IR treatment (5 Gy at 7 days after melanoma cell injection) reduced the weight of the solid tumors, and HemoHIM supplementation with IR enhanced the decreases in tumor weight (P < .03). In the melanoma-bearing mice treated with IR, HemoHIM administration also increased the activity of natural killer cells and cytotoxic T cells, although the proportions of these cells in spleen were not different. In addition, HemoHIM administration increased the interleukin-2 and tumor necrosis factor-alpha secretion from lymphocytes stimulated with concanavalin A, which seemed to contribute to the enhanced efficacy of HemoHIM in tumor-bearing mice treated with IR. In conclusion, HemoHIM may be a beneficial supplement during radiotherapy for enhancing the antitumor efficacy. PMID:20136435

  13. ENHANCED MORTALITY AND LIVER DAMAGE IN VIRUS-INFECTED MICE EXPOSED TO PARAXYLENE

    EPA Science Inventory

    This study assessed effects of p-xylene exposure on mice concurrently infected with murine cytomegalovirus (MCMV). ossible effects included: enhanced infection due to p-xylene induced immune suppression; increased p-xylene toxicity due to viral suppression of cytochrome P-450 (P-...

  14. SYNTHETIC PARTICLES ENHANCE AIRWAY RESPONSES TO OVALBUMIN ANTIGEN IN BALB/C MICE

    EPA Science Inventory

    SYNTHETIC PARTICLES ENHANCE AIRWAY RESPONSES TO OVALBUMIN ANTIGEN IN BALB/CJ MICE. S H Gavett, N Haykal-Coates, and M I Gilmour. Experimental Toxicology Division, NHEERL, ORD, USEPA, Research Triangle Park, NC

    Levels of airborne particulate matter (PM) are positively c...

  15. SYNTHETIC COPPER-CONTAINING PARTICLES ENHANCE ALLERGIC AIRWAY RESPONSES IN MICE

    EPA Science Inventory

    SYNTHETIC COPPER-CONTAINING PARTICLES ENHANCE ALLERGIC AIRWAY RESPONSES IN MICE. SH Gavett, MI Gilmour, and N Haykal-Coates. National Health and Environ Effects Research Lab, USEPA, Res Triangle Park, NC USA
    Respiratory morbidity and mortality associated with increases in ...

  16. Expression of Truncated PITX3 in the Developing Lens Leads to Microphthalmia and Aphakia in Mice

    PubMed Central

    Wada, Kenta; Matsushima, Yoshibumi; Tada, Tomoki; Hasegawa, Sayaka; Obara, Yo; Yoshizawa, Yasuhiro; Takahashi, Gou; Hiai, Hiroshi; Shimanuki, Midori; Suzuki, Sari; Saitou, Junichi; Yamamoto, Naoki; Ichikawa, Masumi; Watanabe, Kei; Kikkawa, Yoshiaki

    2014-01-01

    Microphthalmia is a severe ocular disorder, and this condition is typically caused by mutations in transcription factors that are involved in eye development. Mice carrying mutations in these transcription factors would be useful tools for defining the mechanisms underlying developmental eye disorders. We discovered a new spontaneous recessive microphthalmos mouse mutant in the Japanese wild-derived inbred strain KOR1/Stm. The homozygous mutant mice were histologically characterized as microphthalmic by the absence of crystallin in the lens, a condition referred to as aphakia. By positional cloning, we identified the nonsense mutation c.444C>A outside the genomic region that encodes the homeodomain of the paired-like homeodomain transcription factor 3 gene (Pitx3) as the mutation responsible for the microphthalmia and aphakia. We examined Pitx3 mRNA expression of mutant mice during embryonic stages using RT-PCR and found that the expression levels are higher than in wild-type mice. Pitx3 over-expression in the lens during developmental stages was also confirmed at the protein level in the microphthalmos mutants via immunohistochemical analyses. Although lens fiber differentiation was not observed in the mutants, strong PITX3 protein signals were observed in the lens vesicles of the mutant lens. Thus, we speculated that abnormal PITX3, which lacks the C-terminus (including the OAR domain) as a result of the nonsense mutation, is expressed in mutant lenses. We showed that the expression of the downstream genes Foxe3, Prox1, and Mip was altered because of the Pitx3 mutation, with large reductions in the lens vesicles in the mutants. Similar profiles were observed by immunohistochemical analysis of these proteins. The expression profiles of crystallins were also altered in the mutants. Therefore, we speculated that the microphthalmos/aphakia in this mutant is caused by the expression of truncated PITX3, resulting in the abnormal expression of downstream targets and

  17. Enhanced corticosteroid signaling alters synaptic plasticity in the dentate gyrus in mice lacking the fragile X mental retardation protein.

    PubMed

    Ghilan, M; Hryciw, B N; Brocardo, P S; Bostrom, C A; Gil-Mohapel, J; Christie, B R

    2015-05-01

    The fragile X mental retardation protein (FMRP) is an important regulator of protein translation, and a lack of FMRP expression leads to a cognitive disorder known as fragile X syndrome (FXS). Clinical symptoms characterizing FXS include learning impairments and heightened anxiety in response to stressful situations. Here, we report that, in response to acute stress, mice lacking FMRP show a faster elevation of corticosterone and a more immediate impairment in N-methyl-d-aspartate receptor (NMDAR) dependent long-term potentiation (LTP) in the dentate gyrus (DG). These stress-induced LTP impairments were rescued by administering the glucocorticoid receptor (GR) antagonist RU38486. Administration of RU38486 also enhanced LTP in Fmr1(-/y) mice in the absence of acute stress to wild-type levels, and this enhancement was blocked by application of the NMDAR antagonist 2-amino-5-phosphonopentanoic acid. These results suggest that a loss of FMPR results in enhanced GR signaling that may adversely affect NMDAR dependent synaptic plasticity in the DG. PMID:25731748

  18. Altered NK Cell Development and Enhanced NK Cell-Mediated Resistance to MCMV in NKG2D-Deficient Mice

    PubMed Central

    Zafirova, Biljana; Mandarić, Sanja; Antulov, Ronald; Krmpotić, Astrid; Jonsson, Helena; Yokoyama, Wayne M.; Jonjić, Stipan; Polić, Bojan

    2009-01-01

    Summary NKG2D is a potent activating receptor on NK cells which acts as a molecular sensor for stressed cells expressing NKG2D ligands such as infected or tumor transformed cells. Although NKG2D is expressed on NK cell precursors, its role in NK cell development is still not known. We have generated NKG2D-deficient mice by targeting the Klrk1 locus. Here we provide evidence for an important regulatory role of NKG2D in the development of NK cells. The absence of NKG2D causes faster division of NK cells, perturbation in size of some NK cell subpopulations and their augmented sensitivity to apoptosis. As expected, NKG2D−/− NK cells are less responsive to tumor targets expressing NKG2D ligands. NKG2D−/− mice, however, show an enhanced NK cell-mediated resistance to MCMV infection as a consequence of NK cell dysregulation. Altogether, these findings provide evidence for yet unknown regulatory function of NKG2D in NK cell physiology. PMID:19631564