Cross-platform single cell analysis of kidney development shows stromal cells express Gdnf.

PubMed

Magella, Bliss; Adam, Mike; Potter, Andrew S; Venkatasubramanian, Meenakshi; Chetal, Kashish; Hay, Stuart B; Salomonis, Nathan; Potter, S Steven

2018-02-01

The developing kidney provides a useful model for study of the principles of organogenesis. In this report we use three independent platforms, Drop-Seq, Chromium 10x Genomics and Fluidigm C1, to carry out single cell RNA-Seq (scRNA-Seq) analysis of the E14.5 mouse kidney. Using the software AltAnalyze, in conjunction with the unsupervised approach ICGS, we were unable to identify and confirm the presence of 16 distinct cell populations during this stage of active nephrogenesis. Using a novel integrative supervised computational strategy, we were able to successfully harmonize and compare the cell profiles across all three technological platforms. Analysis of possible cross compartment receptor/ligand interactions identified the nephrogenic zone stroma as a source of GDNF. This was unexpected because the cap mesenchyme nephron progenitors had been thought to be the sole source of GDNF, which is a key driver of branching morphogenesis of the collecting duct system. The expression of Gdnf by stromal cells was validated in several ways, including Gdnf in situ hybridization combined with immunohistochemistry for SIX2, and marker of nephron progenitors, and MEIS1, a marker of stromal cells. Finally, the single cell gene expression profiles generated in this study confirmed and extended previous work showing the presence of multilineage priming during kidney development. Nephron progenitors showed stochastic expression of genes associated with multiple potential differentiation lineages. Copyright © 2017 Elsevier Inc. All rights reserved.

Age and lesion-induced increases of GDNF transgene expression in brain following intracerebral injections of DNA nanoparticles.

PubMed

Yurek, D M; Hasselrot, U; Cass, W A; Sesenoglu-Laird, O; Padegimas, L; Cooper, M J

2015-01-22

In previous studies that used compacted DNA nanoparticles (DNP) to transfect cells in the brain, we observed higher transgene expression in the denervated striatum when compared to transgene expression in the intact striatum. We also observed that long-term transgene expression occurred in astrocytes as well as neurons. Based on these findings, we hypothesized that the higher transgene expression observed in the denervated striatum may be a function of increased gliosis. Several aging studies have also reported an increase of gliosis as a function of normal aging. In this study we used DNPs that encoded for human glial cell line-derived neurotrophic factor (hGDNF) and either a non-specific human polyubiquitin C (UbC) or an astrocyte-specific human glial fibrillary acidic protein (GFAP) promoter. The DNPs were injected intracerebrally into the denervated or intact striatum of young, middle-aged or aged rats, and glial cell line-derived neurotrophic factor (GDNF) transgene expression was subsequently quantified in brain tissue samples. The results of our studies confirmed our earlier finding that transgene expression was higher in the denervated striatum when compared to intact striatum for DNPs incorporating either promoter. In addition, we observed significantly higher transgene expression in the denervated striatum of old rats when compared to young rats following injections of both types of DNPs. Stereological analysis of GFAP+ cells in the striatum confirmed an increase of GFAP+ cells in the denervated striatum when compared to the intact striatum and also an age-related increase; importantly, increases in GFAP+ cells closely matched the increases in GDNF transgene levels. Thus neurodegeneration and aging may lay a foundation that is actually beneficial for this particular type of gene therapy while other gene therapy techniques that target neurons are actually targeting cells that are decreasing as the disease progresses. Copyright © 2014 IBRO. Published by

Noribogaine, but not 18-MC, exhibits similar actions as ibogaine on GDNF expression and ethanol self-administration

PubMed Central

Carnicella, Sebastien; He, Dao-Yao; Yowell, Quinn V.; Glick, Stanley D.; Ron, Dorit

2013-01-01

Ibogaine is a naturally occurring alkaloid that has been reported to decrease various adverse phenotypes associated with exposure to drugs of abuse and alcohol in human and rodent models. Unfortunately, ibogaine cannot be used as a medication to treat addiction because of severe side effects. Previously, we reported that the desirable actions of ibogaine to reduce self-administration of, and relapse to, alcohol consumption are mediated via the upregulation of the expression of the glial cell line-derived neurotrophic factor (GDNF) in the midbrain ventral tegmental area (VTA), and the consequent activation of the GDNF pathway. The ibogaine metabolite, noribogaine, and a synthetic derivative of ibogaine, 18-Methoxycoronaridine (18-MC), possess a similar anti-addictive profile as ibogaine in rodent models, but without some of its adverse side effects. Here, we determined whether noribogaine and/or 18-MC, like ibogaine, increase GDNF expression, and whether their site of action to reduce alcohol consumption is the VTA. We used SH-SY5Y cells as a cell culture model and found that noribogaine, like ibogaine, but not 18-MC, induces a robust increase in GDNF mRNA levels. Next, we tested the effect of intra-VTA infusion of noribogaine and 18-MC on rat operant alcohol self-administration and found that noribogaine, but not 18-MC, in the VTA decreases responding for alcohol. Together, our results suggest that noribogaine and 18-MC have different mechanisms and sites of action. PMID:21040239

Role of trophic factors GDNF, IGF-1 and VEGF in major depressive disorder: A comprehensive review of human studies

PubMed Central

Sharma, Ajaykumar N.; da Costa e Silva, Bruno Fernando Borges; Soares, Jair C.; Carvalho, André F.; Quevedo, Joao

2016-01-01

Rationale The neurotrophin hypothesis of major depressive disorder (MDD) postulates that this illness results from aberrant neurogenesis in brain regions that regulates emotion and memory. Notwithstanding this theory has primarily implicated BDNF in the neurobiology of MDD. Recent evidence suggests that other trophic factors namely GDNF, VEGF and IGF-1 may also be involved. Purpose The present review aimed to critically summarize evidence regarding changes in GDNF, IGF-1 and VEGF in individuals with MDD compared to healthy controls. In addition, we also evaluated the role of these mediators as potential treatment response biomarkers for MDD. Methods A comprehensive review of original studies studies measuring peripheral, central or mRNA levels of GDNF, IGF-1 or VEGF in patients with MDD was conducted. The PubMed/MEDLINE database was searched for peer-reviewed studies published in English through June 2nd, 2015. Results Most studies reported a reduction in peripheral GDNF and its mRNA levels in MDD patients versus controls. In contrast, IGF-1 levels in MDD patients compared to controls were discrepant across studies. Finally, most studies reported high peripheral VEGF levels and mRNA expression in MDD patients compared to healthy controls. Conclusions GDNF, IGF-1 and VEGF levels and their mRNA expression appear to be differentially altered in MDD patients compared to healthy individuals, indicating that these molecules might play an important role in the pathophysiology of depression and antidepressant action of therapeutic interventions. PMID:26956384

Effects of Hindlimb Unweighting on MBP and GDNF Expression and Morphology in Rat Dorsal Root Ganglia Neurons.

PubMed

Zhang, Heng; Ren, Ning-Tao; Zhou, Fang-Qiang; Li, Jie; Lei, Wei; Liu, Ning; Bi, Long; Wu, Zi-Xiang; Zhang, Ran; Zhang, Yong-Gang; Cui, Geng

2016-09-01

With the development of technology and space exploration, studies on long-duration space flights have shown that microgravity induces damage to multiple organs, including the dorsal root ganglia (DRG). However, very little is known about the effects of long-term microgravity on DRG neurons. This study investigated the effects of microgravity on lumbar 5 (L5) DRG neurons in rats using the hindlimb unweighting (HU) model. Male (M) and female (F) Sprague-Dawley rats were randomly divided into M- and F-control (CON) groups and M- and F-HU groups, respectively (n = 10). At the end of HU treatment for 4 weeks, morphological changes were detected. Myelin basic protein (MBP) and degenerated myelin basic protein (dgen-MBP) expressions were analyzed by immunofluorescence and western blot assays. Glial cell line-derived neurotrophic factor (GDNF) protein and mRNA expressions were also analyzed by immunohistochemistry, western blot, and RT-PCR analysis, respectively. Compared with the corresponding CON groups, the HU groups exhibited slightly loose junctions between DRG neurons, some separated ganglion cells and satellite cells, and lightly stained Nissl bodies that were of smaller size and had a scattered distribution. High levels of dgen-MBP and low MBP expressions were appeared and GDNF expressions were significantly decreased in both HU groups. Changes were more pronounced in the F-HU group than in the M-HU group. In conclusion, HU treatment induced damage of L5 DRG neurons, which was correlated with decreased total MBP protein expression, increased dgen-MBP expression, and reduced GDNF protein and mRNA expression. Importantly, these changes were more severe in F-HU rats compared with M-HU rats.

Noribogaine, but not 18-MC, exhibits similar actions as ibogaine on GDNF expression and ethanol self-administration.

PubMed

Carnicella, Sebastien; He, Dao-Yao; Yowell, Quinn V; Glick, Stanley D; Ron, Dorit

2010-10-01

Ibogaine is a naturally occurring alkaloid that has been reported to decrease various adverse phenotypes associated with exposure to drugs of abuse and alcohol in human and rodent models. Unfortunately, ibogaine cannot be used as a medication to treat addiction because of severe side effects. Previously, we reported that the desirable actions of ibogaine to reduce self-administration of, and relapse to, alcohol consumption are mediated via the upregulation of the expression of the glial cell line-derived neurotrophic factor (GDNF) in the midbrain ventral tegmental area (VTA), and the consequent activation of the GDNF pathway. The ibogaine metabolite, noribogaine, and a synthetic derivative of ibogaine, 18-Methoxycoronaridine (18-MC), possess a similar anti-addictive profile as ibogaine in rodent models, but without some of its adverse side effects. Here, we determined whether noribogaine and/or 18-MC, like ibogaine, increase GDNF expression, and whether their site of action to reduce alcohol consumption is the VTA. We used SH-SY5Y cells as a cell culture model and found that noribogaine, like ibogaine, but not 18-MC, induces a robust increase in GDNF mRNA levels. Next, we tested the effect of intra-VTA infusion of noribogaine and 18-MC on rat operant alcohol self-administration and found that noribogaine, but not 18-MC, in the VTA decreases responding for alcohol. Together, our results suggest that noribogaine and 18-MC have different mechanisms and sites of action. © 2010 The Authors, Addiction Biology © 2010 Society for the Study of Addiction.

Conserved and non-conserved characteristics of porcine glial cell line-derived neurotrophic factor expressed in the testis.

PubMed

Kakiuchi, Kazue; Taniguchi, Kazumi; Kubota, Hiroshi

2018-05-16

Glial cell line-derived neurotrophic factor (GDNF) is essential for the self-renewal and proliferation of spermatogonial stem cells (SSCs) in mice, rats, and rabbits. Although the key extrinsic factors essential for spermatogonial proliferation in other mammals have not been determined, GDNF is one of the potential candidates. In this study, we isolated porcine GDNF (pGDNF) cDNAs from neonatal testis and generated recombinant pGDNF to investigate its biological activity on gonocytes/undifferentiated spermatogonia, including SSCs. In porcine testis, long and short forms of GDNF transcripts, the counterparts of pre-(α)pro and pre-(β)pro GDNF identified in humans and rodents, were expressed. The two transcripts encode identical mature proteins. Recombinant pGDNF supported proliferation of murine SSCs in culture, and their stem cell activity was confirmed by a transplantation assay. Subsequently, porcine gonocytes/undifferentiated spermatogonia were cultured with pGDNF; however, pGDNF did not affect their proliferation. Furthermore, GDNF expression was localised to the vascular smooth muscle cells, and its cognate receptor GFRA1 expression was negligible during spermatogonial proliferation in the testes. These results indicate that although pGDNF retains structural similarity with those of other mammals and conserves the biological activity on the self-renewal of murine SSCs, porcine SSCs likely require extrinsic factors other than GDNF for their proliferation.

Functional Neuroprotection and Efficient Regulation of GDNF Using Destabilizing Domains in a Rodent Model of Parkinson's Disease

PubMed Central

Quintino, Luis; Manfré, Giuseppe; Wettergren, Erika Elgstrand; Namislo, Angrit; Isaksson, Christina; Lundberg, Cecilia

2013-01-01

Glial cell line–derived neurotrophic factor (GDNF) has great potential to treat Parkinson's disease (PD). However, constitutive expression of GDNF can over time lead to side effects. Therefore, it would be useful to regulate GDNF expression. Recently, a new gene inducible system using destabilizing domains (DD) from E. coli dihydrofolate reductase (DHFR) has been developed and characterized. The advantage of this novel DD is that it is regulated by trimethoprim (TMP), a well-characterized drug that crosses the blood–brain barrier and can therefore be used to regulate gene expression in the brain. We have adapted this system to regulate expression of GDNF. A C-terminal fusion of GDNF and a DD with an additional furin cleavage site was able to be efficiently regulated in vitro, properly processed and was able to bind to canonical GDNF receptors, inducing a signaling cascade response in target cells. In vivo characterization of the protein showed that it could be efficiently induced by TMP and it was only functional when gene expression was turned on. Further characterization in a rodent model of PD showed that the regulated GDNF protected neurons, improved motor behavior of animals and was efficiently regulated in a pathological setting. PMID:23881415

GDNF is a novel ethanol-responsive gene in the VTA: Implications for the development and persistence of excessive drinking

PubMed Central

Ahmadiantehrani, Somayeh; Barak, Segev; Ron, Dorit

2012-01-01

Glial cell line-derived neurotrophic factor (GDNF) is a potent inhibitor of ethanol consumption and relapse (Carnicella et al., 2008; Carnicella and Ron, 2009; Carnicella et al., 2009c; Barak et al., 2011a), and GDNF heterozygous knockout mice display increased reward sensitivity to ethanol, and consume more ethanol after a period of abstinence, than their wild-type littermates (Carnicella et al., 2009b). Here, we tested whether ethanol alters GDNF expression in the ventral tegmental area (VTA; GDNF’s site of action) and/or the nucleus accumbens (NAc; the main source of GDNF), and if so, determine the role of the endogenous growth factor in the regulation of ethanol consumption. Systemic administration of ethanol increased GDNF expression and protein levels in the VTA, but not the NAc. Additionally, GDNF levels were elevated after an ethanol-drinking session in rats that consumed ethanol in the intermittent-access two-bottle choice procedure for 1 week, but not 7 weeks. Deprivation following 7 weeks of excessive ethanol intake reduced GDNF levels, while a short ethanol binge drinking period following deprivation upregulated GDNF expression. Importantly, knockdown of GDNF within the VTA using adenovirus expressing short hairpin RNA facilitated the escalation of ethanol drinking by ethanol-naïve rats, but not by rats with a history of excessive ethanol consumption. These results suggest that during initial ethanol-drinking experiences, GDNF in the VTA is increased and protects against the development of excessive ethanol intake. However, the growth factor’s protective response to ethanol breaks down after protracted excessive ethanol intake and withdrawal, resulting in persistent, excessive ethanol consumption. PMID:23298382

Alterations in BDNF (brain derived neurotrophic factor) and GDNF (glial cell line-derived neurotrophic factor) serum levels in bipolar disorder: The role of lithium.

PubMed

Tunca, Zeliha; Ozerdem, Aysegul; Ceylan, Deniz; Yalçın, Yaprak; Can, Güneş; Resmi, Halil; Akan, Pınar; Ergör, Gül; Aydemir, Omer; Cengisiz, Cengiz; Kerim, Doyuran

2014-09-01

Brain-derived neurotrophic factor (BDNF) has been consistently reported to be decreased in mania or depression in bipolar disorders. Evidence suggests that Glial cell line-derived neurotrophic factor (GDNF) has a role in the pathogenesis of mood disorders. Whether GDNF and BDNF act in the same way across different episodes in bipolar disorders is unclear. BDNF and GDNF serum levels were measured simultaneously by enzyme-linked immunosorbent assay (ELISA) method in 96 patients diagnosed with bipolar disorder according to DSM-IV (37 euthymic, 33 manic, 26 depressed) in comparison to 61 healthy volunteers. SCID- I and SCID-non patient version were used for clinical evaluation of the patients and healthy volunteers respectively. Correlations between the two trophic factor levels, and medication dose, duration and serum levels of lithium or valproate were studied across different episodes of illness. Patients had significantly lower BDNF levels during mania and depression compared to euthymic patients and healthy controls. GDNF levels were not distinctive. However GDNF/BDNF ratio was higher in manic state compared to euthymia and healthy controls. Significant negative correlation was observed between BDNF and GDNF levels in euthymic patients. While BDNF levels correlated positively, GDNF levels correlated negatively with lithium levels. Regression analysis confirmed that lithium levels predicted only GDNF levels positively in mania, and negatively in euthymia. Small sample size in different episodes and drug-free patients was the limitation of thestudy. Current data suggests that lithium exerts its therapeutic action by an inverse effect on BDNF and GDNF levels, possibly by up-regulating BDNF and down-regulating GDNF to achieve euthymia. Copyright © 2014 Elsevier B.V. All rights reserved.

GDNF-based therapies, GDNF-producing interneurons, and trophic support of the dopaminergic nigrostriatal pathway. Implications for Parkinson’s disease

PubMed Central

d’Anglemont de Tassigny, Xavier; Pascual, Alberto; López-Barneo, José

2015-01-01

The glial cell line-derived neurotrophic factor (GDNF) is a well-established trophic agent for dopaminergic (DA) neurons in vitro and in vivo. GDNF is necessary for maintenance of neuronal morphological and neurochemical phenotype and protects DA neurons from toxic damage. Numerous studies on animal models of Parkinson’s disease (PD) have reported beneficial effects of GDNF on nigrostriatal DA neuron survival. However, translation of these observations to the clinical setting has been hampered so far by side effects associated with the chronic continuous intra-striatal infusion of recombinant GDNF. In addition, double blind and placebo-controlled clinical trials have not reported any clinically relevant effect of GDNF on PD patients. In the past few years, experiments with conditional Gdnf knockout mice have suggested that GDNF is necessary for maintenance of DA neurons in adulthood. In parallel, new methodologies for exogenous GDNF delivery have been developed. Recently, it has been shown that a small population of scattered, electrically interconnected, parvalbumin positive (PV+) GABAergic interneurons is responsible for most of the GDNF produced in the rodent striatum. In addition, cholinergic striatal interneurons appear to be also involved in the modulation of striatal GDNF. In this review, we summarize current knowledge on brain GDNF delivery, homeostasis, and its effects on nigrostriatal DA neurons. Special attention is paid to the therapeutic potential of endogenous GDNF stimulation in PD. PMID:25762899

Neurotrophic Factors NGF, GDNF and NTN Selectively Modulate HSV1 and HSV2 Lytic Infection and Reactivation in Primary Adult Sensory and Autonomic Neurons

PubMed Central

Yanez, Andy A.; Harrell, Telvin; Sriranganathan, Heather J.; Ives, Angela M.; Bertke, Andrea S.

2017-01-01

Herpes simplex viruses (HSV1 and HSV2) establish latency in peripheral ganglia after ocular or genital infection, and can reactivate to produce different patterns and frequencies of recurrent disease. Previous studies showed that nerve growth factor (NGF) maintains HSV1 latency in embryonic sympathetic and sensory neurons. However, adult sensory neurons are no longer dependent on NGF for survival, some populations cease expression of NGF receptors postnatally, and the viruses preferentially establish latency in different populations of sensory neurons responsive to other neurotrophic factors (NTFs). Thus, NGF may not maintain latency in adult sensory neurons. To identify NTFs important for maintaining HSV1 and HSV2 latency in adult neurons, we investigated acute and latently-infected primary adult sensory trigeminal (TG) and sympathetic superior cervical ganglia (SCG) after NTF removal. NGF and glial cell line-derived neurotrophic factor (GDNF) deprivation induced HSV1 reactivation in adult sympathetic neurons. In adult sensory neurons, however, neurturin (NTN) and GDNF deprivation induced HSV1 and HSV2 reactivation, respectively, while NGF deprivation had no effects. Furthermore, HSV1 and HSV2 preferentially reactivated from neurons expressing GFRα2 and GFRα1, the high affinity receptors for NTN and GDNF, respectively. Thus, NTN and GDNF play a critical role in selective maintenance of HSV1 and HSV2 latency in primary adult sensory neurons. PMID:28178213

Neurotrophic Factors NGF, GDNF and NTN Selectively Modulate HSV1 and HSV2 Lytic Infection and Reactivation in Primary Adult Sensory and Autonomic Neurons.

PubMed

Yanez, Andy A; Harrell, Telvin; Sriranganathan, Heather J; Ives, Angela M; Bertke, Andrea S

2017-02-07

Herpes simplex viruses (HSV1 and HSV2) establish latency in peripheral ganglia after ocular or genital infection, and can reactivate to produce different patterns and frequencies of recurrent disease. Previous studies showed that nerve growth factor (NGF) maintains HSV1 latency in embryonic sympathetic and sensory neurons. However, adult sensory neurons are no longer dependent on NGF for survival, some populations cease expression of NGF receptors postnatally, and the viruses preferentially establish latency in different populations of sensory neurons responsive to other neurotrophic factors (NTFs). Thus, NGF may not maintain latency in adult sensory neurons. To identify NTFs important for maintaining HSV1 and HSV2 latency in adult neurons, we investigated acute and latently-infected primary adult sensory trigeminal (TG) and sympathetic superior cervical ganglia (SCG) after NTF removal. NGF and glial cell line-derived neurotrophic factor (GDNF) deprivation induced HSV1 reactivation in adult sympathetic neurons. In adult sensory neurons, however, neurturin (NTN) and GDNF deprivation induced HSV1 and HSV2 reactivation, respectively, while NGF deprivation had no effects. Furthermore, HSV1 and HSV2 preferentially reactivated from neurons expressing GFRα2 and GFRα1, the high affinity receptors for NTN and GDNF, respectively. Thus, NTN and GDNF play a critical role in selective maintenance of HSV1 and HSV2 latency in primary adult sensory neurons.

Combining glial cell line-derived neurotrophic factor gene delivery (AdGDNF) with L-arginine decreases contusion size but not behavioral deficits after traumatic brain injury.

PubMed

Degeorge, M L; Marlowe, D; Werner, E; Soderstrom, K E; Stock, M; Mueller, A; Bohn, M C; Kozlowski, D A

2011-07-27

Our laboratory has previously demonstrated that viral administration of glial cell line-derived neurotrophic factor (AdGDNF), one week prior to a controlled cortical impact (CCI) over the forelimb sensorimotor cortex of the rat (FL-SMC) is neuroprotective, but does not significantly enhance recovery of sensorimotor function. One possible explanation for this discrepancy is that although protected, neurons may not have been functional due to enduring metabolic deficiencies. Additionally, metabolic events following TBI may interfere with expression of therapeutic proteins administered to the injured brain via gene therapy. The current study focused on enhancing the metabolic function of the brain by increasing cerebral blood flow (CBF) with l-arginine in conjunction with administration of AdGDNF immediately following CCI. An adenoviral vector harboring human GDNF was injected unilaterally into FL-SMC of the rat immediately following a unilateral CCI over the FL-SMC. Within 30min of the CCI and AdGDNF injections, some animals were injected with l-arginine (i.v.). Tests of forelimb function and asymmetry were administered for 4weeks post-injury. Animals were sacrificed and contusion size and GDNF protein expression measured. This study demonstrated that rats treated with AdGDNF and l-arginine post-CCI had a significantly smaller contusion than injured rats who did not receive any treatment, or injured rats treated with either AdGDNF or l-arginine alone. Nevertheless, no amelioration of behavioral deficits was seen. These findings suggest that AdGDNF alone following a CCI was not therapeutic and although combining it with l-arginine decreased contusion size, it did not enhance behavioral recovery. Copyright © 2011 Elsevier B.V. All rights reserved.

Targeting the Gdnf Gene in peritubular myoid cells disrupts undifferentiated spermatogonial cell development

PubMed Central

Chen, Liang-Yu; Willis, William D.; Eddy, Edward M.

2016-01-01

Spermatogonial stem cells (SSCs) are a subpopulation of undifferentiated spermatogonia located in a niche at the base of the seminiferous epithelium delimited by Sertoli cells and peritubular myoid (PM) cells. SSCs self-renew or differentiate into spermatogonia that proliferate to give rise to spermatocytes and maintain spermatogenesis. Glial cell line-derived neurotrophic factor (GDNF) is essential for this process. Sertoli cells produce GDNF and other growth factors and are commonly thought to be responsible for regulating SSC development, but limited attention has been paid to the role of PM cells in this process. A conditional knockout (cKO) of the androgen receptor gene in PM cells resulted in male infertility. We found that testosterone (T) induces GDNF expression in mouse PM cells in vitro and neonatal spermatogonia (including SSCs) co-cultured with T-treated PM cells were able to colonize testes of germ cell-depleted mice after transplantation. This strongly suggested that T-regulated production of GDNF by PM cells is required for spermatogonial development, but PM cells might produce other factors in vitro that are responsible. In this study, we tested the hypothesis that production of GDNF by PM cells is essential for spermatogonial development by generating mice with a cKO of the Gdnf gene in PM cells. The cKO males sired up to two litters but became infertile due to collapse of spermatogenesis and loss of undifferentiated spermatogonia. These studies show for the first time, to our knowledge, that the production of GDNF by PM cells is essential for undifferentiated spermatogonial cell development in vivo. PMID:26831079

Overexpression of GDNF in the uninjured DRG exerts analgesic effects on neuropathic pain following segmental spinal nerve ligation in mice.

PubMed

Takasu, Kumiko; Sakai, Atsushi; Hanawa, Hideki; Shimada, Takashi; Suzuki, Hidenori

2011-11-01

Glial cell line-derived neurotrophic factor (GDNF), a survival-promoting factor for a subset of nociceptive small-diameter neurons, has been shown to exert analgesic effects on neuropathic pain. However, its detailed mechanisms of action are still unknown. In the present study, we investigated the site-specific analgesic effects of GDNF in the neuropathic pain state using lentiviral vector-mediated GDNF overexpression in mice with left fifth lumbar (L5) spinal nerve ligation (SNL) as a neuropathic pain model. A lentiviral vector expressing both GDNF and enhanced green fluorescent protein (EGFP) was constructed and injected into the left dorsal spinal cord, uninjured fourth lumbar (L4) dorsal root ganglion (DRG), injured L5 DRG, or plantar skin of mice. In SNL mice, injection of the GDNF-EGFP-expressing lentivirus into the dorsal spinal cord or uninjured L4 DRG partially but significantly reduced the mechanical allodynia in association with an increase in GDNF protein expression in each virus injection site, whereas injection into the injured L5 DRG or plantar skin had no effects. These results suggest that GDNF exerts its analgesic effects in the neuropathic pain state by acting on the central terminals of uninjured DRG neurons and/or on the spinal cells targeted by the uninjured DRG neurons. This article shows that GDNF exerts its analgesic effects on neuropathic pain by acting on the central terminals of uninjured DRG neurons and/or on the spinal cells targeted by these neurons. Therefore, research focusing on these GDNF-dependent neurons in the uninjured DRG would provide a new strategy for treating neuropathic pain. Copyright © 2011 American Pain Society. Published by Elsevier Inc. All rights reserved.

Ex vivo delivery of GDNF maintains motor function and prevents neuronal loss in a transgenic mouse model of Huntington's disease.

PubMed

Ebert, Allison D; Barber, Amelia E; Heins, Brittany M; Svendsen, Clive N

2010-07-01

Huntington's disease (HD) is an autosomal dominant disorder caused by expansion of polyglutamine repeats in the huntingtin gene leading to loss of striatal and cortical neurons followed by deficits in cognition and choreic movements. Growth factor delivery to the brain has shown promise in various models of neurodegenerative diseases, including HD, by reducing neuronal death and thus limiting motor impairment. Here we used mouse neural progenitor cells (mNPCs) as growth factor delivery vehicles in the N171-82Q transgenic mouse model of HD. mNPCs derived from the developing mouse striatum were isolated and infected with lentivirus expressing either glial cell line-derived neurotrophic factor (GDNF) or green fluorescent protein (GFP). Next, mNPCs(GDNF) or mNPCs(GFP) were transplanted bilaterally into the striatum of pre-symptomatic N171-82Q mice. We found that mNPCs(GDNF), but not mNPCs(GFP), maintained rotarod function and increased striatal neuron survival out to 3months post-transplantation. Importantly, histological analysis showed GDNF expression through the duration of the experiment. Our data show that mNPCs(GDNF) can survive transplantation, secrete GDNF for several weeks and are able to maintain motor function in this model of HD. Copyright 2010 Elsevier Inc. All rights reserved.

Melatonin promotes goat spermatogonia stem cells (SSCs) proliferation by stimulating glial cell line-derived neurotrophic factor (GDNF) production in Sertoli cells.

PubMed

Niu, Bowen; Li, Bo; Wu, Chongyang; Wu, Jiang; Yan, Yuan; Shang, Rui; Bai, Chunling; Li, Guangpeng; Hua, Jinlian

2016-11-22

Melatonin has been reported to be an important endogenous hormone for regulating neurogenesis, immunityand the biological clock. Recently, the effects of melatonin on neural stem cells (NSCs), mesenchymal stem cells(MSCs), and induced pluripotent stem cells(iPSCs) have been reported; however, the effects of melatonin on spermatogonia stem cells (SSCs) are not clear. Here, 1μM and 1nM melatonin was added to medium when goat SSCs were cultured in vitro, the results showed that melatonin could increase the formation and size of SSC colonies. Real-time quantitative PCR (QRT-PCR) and western blot analysis showed that the expression levels of SSC proliferation and self-renewal markers were up-regulated. Meanwhile, QRT-PCR results showed that melatonin inhibit the mRNA expression level of SSC differentiation markers. ELISA analysis showed an obvious increase in the concentration of GDNF (a niche factor secreted by Sertoli cells) in the medium when treated with melatonin. Meanwhile, the phosphorylation level of AKT, a downstream of GDNF-GFRa1-RET pathway was activated. In conclusion, melatonin promotes goat SSC proliferation by stimulating GDNF production in Sertoli cells.

Autoregulation of glial cell line-derived neurotrophic factor expression: implications for the long-lasting actions of the anti-addiction drug, Ibogaine.

PubMed

He, Dao-Yao; Ron, Dorit

2006-11-01

We recently showed that the up-regulation of the glial cell line-derived neurotrophic factor (GDNF) pathway in the midbrain, is the molecular mechanism by which the putative anti-addiction drug Ibogaine mediates its desirable action of reducing alcohol consumption. Human reports and studies in rodents have shown that a single administration of Ibogaine results in a long-lasting reduction of drug craving (humans) and drug and alcohol intake (rodents). Here we determine whether, and how, Ibogaine exerts its long-lasting actions on GDNF expression and signaling. Using the dopaminergic-like SHSY5Y cell line as a culture model, we observed that short-term Ibogaine exposure results in a sustained increase in GDNF expression that is mediated via the induction of a long-lasting autoregulatory cycle by which GDNF positively regulates its own expression. We show that the initial exposure of cells to Ibogaine or GDNF results in an increase in GDNF mRNA, leading to protein expression and to the corresponding activation of the GDNF signaling pathway. This, in turn, leads to a further increase in the mRNA level of the growth factor. The identification of a GDNF-mediated, autoregulatory long-lasting feedback loop could have important implications for GDNF's potential value as a treatment for addiction and neurodegenerative diseases.

Glial Cell Line-Derived Neurotrophic Factor (GDNF) serum level in women with schizophrenia and depression, correlation with clinical and metabolic parameters.

PubMed

Skibinska, Maria; Kapelski, Pawel; Pawlak, Joanna; Rajewska-Rager, Aleksandra; Dmitrzak-Weglarz, Monika; Szczepankiewicz, Aleksandra; Czerski, Piotr; Twarowska-Hauser, Joanna

2017-10-01

Neurotrophic factors have been implicated in neuropsychiatric disorders, including schizophrenia and depression. Glial Cell Line-Derived Neurotrophic Factor (GDNF) promotes development, differentiation, and protection of dopaminergic, serotonergic, GABAergic and noradrenergic neurons as well as glial cells in different brain regions. This study examined serum levels of GDNF in schizophrenia and depression and its correlation with metabolic parameters during 8 weeks of treatment. Serum GDNF level, fasting serum glucose and lipid profile were measured at baseline and week 8 in 133 women: 55 with schizophrenia, 30 with a first episode depression and 48 healthy controls. The severity of the symptoms was evaluated using Positive and Negative Syndrome Scale (PANSS), 17-item Hamilton Depression Rating Scale (HDRS) and Beck Depression Inventory (BDI). There was statistically significant higher GDNF level in schizophrenia at baseline when compared with week 8. Correlations of GDNF with PANSS in schizophrenia and cholesterol level in depression have also been detected. To our knowledge, this is the first study which correlates GDNF levels with metabolic parameters. Our results show no differences in GDNF serum level between schizophrenia, a first depressive episode, and healthy controls. GDNF serum level did not correlate with metabolic parameters except for total cholesterol in depression. Copyright © 2017 Elsevier B.V. All rights reserved.

Cross-link regulation of precursor N-cadherin and FGFR1 by GDNF increases U251MG cell viability.

PubMed

Tang, Chuan-Xi; Gu, Yan-Xia; Liu, Xin-Feng; Tong, Shu-Yan; Ayanlaja, Abiola A; Gao, Yue; Ji, Guang-Quan; Xiong, Ye; Huang, Lin-Yan; Gao, Dian-Shuai

2018-07-01

Glial cell line-derived neurotrophic factor (GDNF) is considered to be involved in the development of glioma. However, uncovering the underlying mechanism of the proliferation of glioma cells is a challenging work in progress. We have identified the binding of the precursor of N-cadherin (proN-cadherin) and GDNF on the cell membrane in previous studies. In the present study, we observed increased U251 Malignant glioma (U251MG) cell viability by exogenous GDNF (50 ng/ml). We also confirmed that the high expression of the proN-cadherin was stimulated by exogenous GDNF. Concurrently, we affirmed that lower expression of proN-cadherin correlated with reduced glioma cell viability. Additionally, we observed glioma cell U251MG viability as the phosphorylation level of FGFR1 at Y653 and Y654 was increased after exogenous GDNF treatment, which led to increased interaction between proN-cadherin and FGFR1 (pY653+Y654). Our experiments presented a new mechanism adopted by GDNF supporting glioma development and indicated a possible therapeutic potential via the inhibition of proN-cadherin/FGFR1 interaction.

Somatotropinomas, but not nonfunctioning pituitary adenomas, maintain a functional apoptotic RET/Pit1/ARF/p53 pathway that is blocked by excess GDNF.

PubMed

Diaz-Rodriguez, Esther; Garcia-Rendueles, Angela R; Ibáñez-Costa, Alejandro; Gutierrez-Pascual, Ester; Garcia-Lavandeira, Montserrat; Leal, Alfonso; Japon, Miguel A; Soto, Alfonso; Venegas, Eva; Tinahones, Francisco J; Garcia-Arnes, Juan A; Benito, Pedro; Angeles Galvez, Maria; Jimenez-Reina, Luis; Bernabeu, Ignacio; Dieguez, Carlos; Luque, Raul M; Castaño, Justo P; Alvarez, Clara V

2014-11-01

Acromegaly is caused by somatotroph cell adenomas (somatotropinomas [ACROs]), which secrete GH. Human and rodent somatotroph cells express the RET receptor. In rodents, when normal somatotrophs are deprived of the RET ligand, GDNF (Glial Cell Derived Neurotrophic Factor), RET is processed intracellularly to induce overexpression of Pit1 [Transcription factor (gene : POUF1) essential for transcription of Pituitary hormones GH, PRL and TSHb], which in turn leads to p19Arf/p53-dependent apoptosis. Our purpose was to ascertain whether human ACROs maintain the RET/Pit1/p14ARF/p53/apoptosis pathway, relative to nonfunctioning pituitary adenomas (NFPAs). Apoptosis in the absence and presence of GDNF was studied in primary cultures of 8 ACROs and 3 NFPAs. Parallel protein extracts were analyzed for expression of RET, Pit1, p19Arf, p53, and phospho-Akt. When GDNF deprived, ACRO cells, but not NFPAs, presented marked level of apoptosis that was prevented in the presence of GDNF. Apoptosis was accompanied by RET processing, Pit1 accumulation, and p14ARF and p53 induction. GDNF prevented all these effects via activation of phospho-AKT. Overexpression of human Pit1 (hPit1) directly induced p19Arf/p53 and apoptosis in a pituitary cell line. Using in silico studies, 2 CCAAT/enhancer binding protein alpha (cEBPα) consensus-binding sites were found to be 100% conserved in mouse, rat, and hPit1 promoters. Deletion of 1 cEBPα site prevented the RET-induced increase in hPit1 promoter expression. TaqMan qRT-PCR (real time RT-PCR) for RET, Pit1, Arf, TP53, GDNF, steroidogenic factor 1, and GH was performed in RNA from whole ACRO and NFPA tumors. ACRO but not NFPA adenomas express RET and Pit1. GDNF expression in the tumors was positively correlated with RET and negatively correlated with p53. In conclusion, ACROs maintain an active RET/Pit1/p14Arf/p53/apoptosis pathway that is inhibited by GDNF. Disruption of GDNF's survival function might constitute a new therapeutic route in

Effect of GDNF on depressive-like behavior, spatial learning and key genes of the brain dopamine system in genetically predisposed to behavioral disorders mouse strains.

PubMed

Naumenko, Vladimir S; Kondaurova, Elena M; Bazovkina, Daria V; Tsybko, Anton S; Ilchibaeva, Tatyana V; Khotskin, Nikita V; Semenova, Alina A; Popova, Nina K

2014-11-01

The effect of glial cell line-derived neurotrophic factor (GDNF) on behavior and brain dopamine system in predisposed to depressive-like behavior ASC (Antidepressant Sensitive Cataleptics) mice in comparison with the parental "nondepressive" CBA mice was studied. In 7days after administration (800ng, i.c.v.) GDNF decreased escape latency time and the path traveled to reach hidden platform in Morris water maze in ASC mice. GDNF enhanced depressive-like behavioral traits in both "nondepressive" CBA and "depressive" ASC mice. In CBA mice, GDNF decreased functional response to agonists of D1 (chloro-APB hydrobromide) and D2 (sumanirole maleate) receptors in tail suspension test, reduced D2 receptor gene expression in the substantia nigra and increased monoamine oxydase A (MAO A) gene expression in the striatum. GDNF increased D1 and D2 receptor genes expression in the nucleus accumbens of ASC mice but failed to alter expression of catechol-O-methyltransferase, dopamine transporter, MAO B and tyrosine hydroxylase genes in both investigated mouse strains. Thus, GDNF produced long-term genotype-dependent effect on behavior and the brain dopamine system. GDNF pretreatment (1) reduced D1 and D2 receptors functional responses and D2 receptor gene expression in s. nigra of CBA mice; (2) increased D1 and D2 receptor genes expression in n. accumbens of ASC mice and (3) improved spatial learning in ASC mice. GDNF enhanced depressive-like behavior both in CBA and ASC mice. The data suggest that genetically defined variance in the cross-talk between GDNF and brain dopamine system contributes to the variability of GDNF-induced responses and might be responsible for controversial GDNF effects. Copyright © 2014 Elsevier B.V. All rights reserved.

GDNF facilitates differentiation of the adult dentate gyrus-derived neural precursor cells into astrocytes via STAT3

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

Boku, Shuken, E-mail: shuboku@med.hokudai.ac.jp; Nakagawa, Shin; Takamura, Naoki

2013-05-17

Highlights: •GDNF has no effect on ADP proliferation and apoptosis. •GDNF increases ADP differentiation into astrocyte. •A specific inhibitor of STAT3 decreases the astrogliogenic effect of GDNF. •STAT3 knockdown by lentiviral shRNA vector also decreases the astrogliogenic effect of GDNF. •GDNF increases the phosphorylation of STAT3. -- Abstract: While the pro-neurogenic actions of antidepressants in the adult hippocampal dentate gyrus (DG) are thought to be one of the mechanisms through which antidepressants exert their therapeutic actions, antidepressants do not increase proliferation of neural precursor cells derived from the adult DG. Because previous studies showed that antidepressants increase the expression andmore » secretion of glial cell line-derived neurotrophic factor (GDNF) in C6 glioma cells derived from rat astrocytes and GDNF increases neurogenesis in adult DG in vivo, we investigated the effects of GDNF on the proliferation, differentiation and apoptosis of cultured neural precursor cells derived from the adult DG. Data showed that GDNF facilitated the differentiation of neural precursor cells into astrocytes but had no effect on their proliferation or apoptosis. Moreover, GDNF increased the phosphorylation of STAT3, and both a specific inhibitor of STAT3 and lentiviral shRNA for STAT3 decreased their differentiation into astrocytes. Taken together, our findings suggest that GDNF facilitates astrogliogenesis from neural precursor cells in adult DG through activating STAT3 and that this action might indirectly affect neurogenesis.« less

Aminochrome decreases NGF, GDNF and induces neuroinflammation in organotypic midbrain slice cultures.

PubMed

de Araújo, Fillipe M; Ferreira, Rafael S; Souza, Cleide S; Dos Santos, Cleonice Creusa; Rodrigues, Tácio L R S; E Silva, Juliana Helena C; Gasparotto, Juciano; Gelain, Daniel Pens; El-Bachá, Ramon S; D Costa, Maria de Fátima; Fonseca, José Claudio M; Segura-Aguilar, Juan; Costa, Silvia L; Silva, Victor Diogenes A

2018-05-01

Recent evidence shows that aminochrome induces glial activation related to neuroinflammation. This dopamine derived molecule induces formation and stabilization of alpha-synuclein oligomers, mitochondria dysfunction, oxidative stress, dysfunction of proteasomal and lysosomal systems, endoplasmic reticulum stress and disruption of the microtubule network, but until now there has been no evidence of effects on production of cytokines and neurotrophic factors, that are mechanisms involved in neuronal loss in Parkinson's disease (PD). This study examines the potential role of aminochrome on the regulation of NGF, GDNF, TNF-α and IL-1β production and microglial activation in organotypic midbrain slice cultures from P8 - P9 Wistar rats. We demonstrated aminochrome (25 μM, for 24 h) induced reduction of GFAP expression, reduction of NGF and GDNF mRNA levels, morphological changes in Iba1 + cells, and increase of both TNF-α, IL-1β mRNA and protein levels. Moreover, aminochrome (25 μM, for 48 h) induced morphological changes in the edge of slices and reduction of TH expression. These results demonstrate neuroinflammation, as well as negative regulation of neurotrophic factors (GDNF and NGF), may be involved in aminochrome-induced neurodegeneration, and they contribute to a better understanding of PD pathogenesis. Copyright © 2018 Elsevier B.V. All rights reserved.

Augmentation of the Ascending Component of the Peristaltic Reflex and Substance P Release by Glial Cell Line-Derived Neurotrophic Factor (GDNF)

PubMed Central

Grider, JR; Heuckeroth, RO; Kuemmerle, JF; Murthy, KS

2010-01-01

Glial cell line derived neurotrophic factor (GDNF) is present in adult gut although its role in the mature enteric nervous system is not well defined. The aim of the present study was to examine the role of GDNF as neuromodulator of the ascending phase of the peristaltic reflex. Colonic segments were prepared as flat sheets and placed in compartmented chambers so as to separate the sensory and motor limbs of the reflex. Ascending contraction was measured in the orad compartment and mucosal stroking stimuli (2-8 strokes) were applied in the caudad compartment. GDNF and substance P release were measured and the effects of GDNF and GDNF antibody on contraction and release were determined. Mice with reduced levels of GDNF (Gdnf+/-) and wild type littermates were also examined. GDNF was released in a stimulus-dependent manner into the orad motor but not caudad sensory compartment. Addition of GDNF to the orad motor but not caudad sensory compartment augmented ascending contraction and substance P release. Conversely, addition of GDNF antibody to the orad motor but not caudad sensory compartment reduced ascending contraction and substance P release. Similarly, the ascending contraction and substance P release into the orad motor compartment was reduced in Gdnf+/- mice as compared to wild type littermates. The results suggest that endogenous GDNF is released during the ascending contraction component of the peristaltic reflex where it acts as a neuromodulator to augment substance P release from motor neurons thereby augmenting contraction of circular muscle orad to the site of stimulation. PMID:20331804

Glial cell line-derived neurotrophic factor (GDNF) induces neuritogenesis in the cochlear spiral ganglion via neural cell adhesion molecule (NCAM)

PubMed Central

Euteneuer, Sara; Yang, Kuo H.; Chavez, Eduardo; Leichtle, Anke; Loers, Gabriele; Olshansky, Adel; Pak, Kwang; Schachner, Melitta; Ryan, Allen F.

2013-01-01

Glial cell line-derived neurotrophic factor (GDNF) increases survival and neurite extension of spiral ganglion neurons (SGNs), the primary neurons of the auditory system, via yet unknown signaling mechanisms. In other cell types, signaling is achieved by the GPI-linked GDNF family receptor α1 (GFRα1) via recruitment of transmembrane receptors: Ret (re-arranged during transformation) and/or NCAM (neural cell adhesion molecule). Here we show that GDNF enhances neuritogenesis in organotypic cultures of spiral ganglia from 5-day-old rats and mice. Addition of GFRα1-Fc increases this effect. GDNF/GFRα1-Fc stimulation activates intracellular PI3K/Akt and MEK/Erk signaling cascades as detected by Western blot analysis of cultures prepared from rats at postnatal days 5 (P5, before the onset of hearing) and 20 (P20, after the onset of hearing). Both cascades mediate GDNF stimulation of neuritogenesis, since application of the Akt inhibitor Wortmannin or the Erk inhibitor U0126 abolished GDNF/GFRα1-Fc stimulated neuritogenesis in P5 rats. Since cultures of P5 NCAM-deficient mice failed to respond by neuritogenesis to GDNF/GFRα1-Fc, we conclude that NCAM serves as a receptor for GDNF signaling responsible for neuritogenesis in early postnatal spiral ganglion. PMID:23262364

Effects of Forskolin on Trefoil factor 1 expression in cultured ventral mesencephalic dopaminergic neurons.

PubMed

Jensen, P; Ducray, A D; Widmer, H R; Meyer, M

2015-12-03

Trefoil factor 1 (TFF1) belongs to a family of secreted peptides that are mainly expressed in the gastrointestinal tract. Notably, TFF1 has been suggested to operate as a neuropeptide, however, its specific cellular expression, regulation and function remain largely unknown. We have previously shown that TFF1 is expressed in developing and adult rat ventral mesencephalic tyrosine hydroxylase-immunoreactive (TH-ir) dopaminergic neurons. Here, we investigated the expression of TFF1 in rat ventral mesencephalic dopaminergic neurons (embryonic day 14) grown in culture for 5, 7 or 10 days in the absence (controls) or presence of either glial cell line-derived neurotrophic factor (GDNF), Forskolin or the combination. No TFF1-ir cells were identified at day 5 and only a few at day 7, whereas TH was markedly expressed at both time points. At day 10, several TFF1-ir cells were detected, and their numbers were significantly increased after the addition of GDNF (2.2-fold) or Forskolin (4.1-fold) compared to controls. Furthermore, the combination of GDNF and Forskolin had an additive effect and increased the number of TFF1-ir cells by 5.6-fold compared to controls. TFF1 expression was restricted to neuronal cells, and the percentage of TH/TFF1 co-expressing cells was increased to the same extent in GDNF and Forskolin-treated cultures (4-fold) as compared to controls. Interestingly, the combination of GDNF and Forskolin resulted in a significantly increased co-expression (8-fold) of TH/TFF1, which could indicate that GDNF and Forskolin targeted different subpopulations of TH/TFF1 neurons. Short-term treatment with Forskolin resulted in an increased number of TFF1-ir cells, and this effect was significantly reduced by the MEK1 inhibitor PD98059 or the protein kinase A (PKA) inhibitor H89, suggesting that Forskolin induced TFF1 expression through diverse signaling pathways. In conclusion, distinct populations of cultured dopaminergic neurons express TFF1, and their numbers can be

In vivo dynamics of GFRα1-positive spermatogonia stimulated by GDNF signals using a bead transplantation assay

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

Uchida, Aya; Kishi, Kasane; Aiyama, Yoshimi

In mouse testes, spermatogonial stem cells (SSCs), a subpopulation of GFRα1 (GDNF family receptor-α1)-positive spermatogonia, are widely distributed along the convoluted seminiferous tubules. The proliferation and differentiation of the SSCs are regulated in part by local expression of GDNF (glial cell-derived neurotorphic factor), one of major niche factors for SSCs. However, the in vivo dynamics of the GDNF-stimulated GFRα1-positive spermatogonia remains unclear. Here, we developed a simple method for transplanting DiI-labeled and GDNF-soaked beads into the mouse testicular interstitium. By using this method, we examined the dynamics of GFRα1-positive spermatogonia in the tubular walls close to the transplanted GDNF-soaked beads. Themore » bead-derived GDNF signals were able to induce the stratified aggregate formation of GFRα1-positive undifferentiated spermatogonia by day 3 post-transplantation. Each aggregate consisted of tightly compacted A{sub single} and marginal A{sub paired}–A{sub aligned} GFRα1-positive spermatogonia and was surrounded by A{sub aligned} GFRα1-negative spermatogonia at more advanced stages. These data not only provide in vivo evidence for the inductive roles of GDNF in forming a rapid aggregation of GFRα1-positive spermatogonia but also indicate the usefulness of this in vivo assay system of various growth factors for the stem/progenitor spermatogonia in mammalian spermatogenesis. - Highlights: • A novel bead transplantation assay was developed to examine the in vivo effects of growth factors on spermatogonia. • A rapid aggregation of GFRα1-positive spermatogonia was induced by the transplanted GDNF-soaked beads. • Tightly-compacted A{sub single} and marginal A{sub paired}–A{sub aligned} spermatogonia were formed in each GFRα1-positive aggregate.« less

The glial cell line-derived neurotrophic factor (GDNF) does not acutely change acetylcholine release in developing and adult neuromuscular junction.

PubMed

Garcia, Neus; Santafé, Manel M; Tomàs, Marta; Lanuza, Maria A; Besalduch, Nuria; Priego, Merche; Tomàs, Josep

2010-08-16

We use immunocytochemistry to show that the trophic molecule glial cell line-derived neurotrophic factor (GDNF) and its receptor GDNF family receptor alpha-1 (GFRalpha-1) are present in both neonatal (P6) and adult (P45) rodent neuromuscular junctions (NMJ) colocalized with several synaptic markers. However, incubation with exogenous GDNF (10-200ng/ml, 1-3h), does not affect spontaneous ACh release. Moreover, GDNF does not change the size of the evoked ACh release from the weak and the strong axonal inputs on dually innervated postnatal endplates nor in the most developed singly-innervated synapses at P6 and P45. Our findings indicate that GDNF (unlike neurotrophins) does not acutely modulate transmitter release during the developmental process of synapse elimination nor as the NMJ matures. Copyright 2010 Elsevier Ireland Ltd. All rights reserved.

[Participation of GDNF, LIMK1 signal pathways and heat shock proteins in processes of Drosophila learning and memory formation].

PubMed

Nikitina, E A; Medvedeva, A V; Dolgaia, Iu F; Korochkin, L I; Pavlova, G V; Savvateeva-Popova, E V

2012-01-01

Molecular mechanisms of the synapse and dendrite maintenance and their disturbance in psychiatric and neurodegenerative diseases (ND) are intensively studied in searching for target genes of therapeutic actions. It is suggested that glia, alongside with well-studied pre- and postsynaptic neurons, is the third, poorly studied partner in synaptic transmission (the tripartite synapse) that is involved in the positive feedback between the first two partners. This bidirectional coupling between presynaptic neurons and their postsynaptic targets involve neurotrophins (NTF), such as glial cell-derived neurotrophic factor (GDNF) that is produced LIM kinase 1 (LIMK1, the key enzyme of actin remodeling). The cytoplasmic domain of neuregulins interacts with LIMK1. Since neurons and axons that do not receive a sufficient NTF amount are at risk of degeneration and synapse elimination, GDNF seems to be the best studied factor of the ND therapy. The delivery of GDNF stem cells to the neurodegeneration locus is very efficient. There has been proposed a new approach based on use of Drosophila heat shock (hs) promoter. This promoter responds to the mammalian body temperature as to the shock factor resulting in the constant expression of the GDNF gene. The Drosophila models allow studying any given component of the bidirectional communication between pre- and postsynaptic neurons in development of the main diagnostic ND symptom, such as defective memory resulted from synaptic atrophy. In the present study we used the Drosophila stocks imitating different disturbances of the nervous system: Canton-S (wild type), GDNF (transgenic flies that carry human glial-cell-line derived nerve factor (GDNF) gene under hs promoter), l(1)ts403 with dusturbance of HSPs mRNA extranuclear transport, a defect of intracellular stress report, and agn(ts3) mutation in LIMK1 gene. We have revealed functional connections at the behavioral level (learning/memory) depending on the GDNF and LIMK1 brain

Recombinant GDNF: Tetanus toxin fragment C fusion protein produced from insect cells

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

Li, Jianhong; Chian, Ru-Ju; Ay, Ilknur

2009-07-31

Glial cell line-derived neurotrophic factor (GDNF) has potent survival-promoting effects on CNS motor neurons in experimental animals. Its therapeutic efficacy in humans, however, may have been limited by poor bioavailability to the brain and spinal cord. With a view toward improving delivery of GDNF to CNS motor neurons in vivo, we generated a recombinant fusion protein comprised of rat GDNF linked to the non-toxic, neuron-binding fragment of tetanus toxin. Recombinant GDNF:TTC produced from insect cells was a soluble homodimer like wild-type GDNF and was bi-functional with respect to GDNF and TTC activity. Like recombinant rat GDNF, the fusion protein increasedmore » levels of immunoreactive phosphoAkt in treated NB41A3-hGFR{alpha}-1 neuroblastoma cells. Like TTC, GDNF:TTC bound to immobilized ganglioside GT1b in vitro with high affinity and selectivity. These results support further testing of recombinant GDNF:TTC as a non-viral vector to improve delivery of GDNF to brain and spinal cord in vivo.« less

Glial cell line-derived neurotrophic factor protects against high-fat diet-induced hepatic steatosis by suppressing hepatic PPAR-γ expression.

PubMed

Mwangi, Simon Musyoka; Peng, Sophia; Nezami, Behtash Ghazi; Thorn, Natalie; Farris, Alton B; Jain, Sanjay; Laroui, Hamed; Merlin, Didier; Anania, Frank; Srinivasan, Shanthi

2016-01-15

Glial cell line-derived neurotrophic factor (GDNF) protects against high-fat diet (HFD)-induced hepatic steatosis in mice, however, the mechanisms involved are not known. In this study we investigated the effects of GDNF overexpression and nanoparticle delivery of GDNF in mice on hepatic steatosis and fibrosis and the expression of genes involved in the regulation of hepatic lipid uptake and de novo lipogenesis. Transgenic overexpression of GDNF in liver and other metabolically active tissues was protective against HFD-induced hepatic steatosis. Mice overexpressing GDNF had significantly reduced P62/sequestosome 1 protein levels suggestive of accelerated autophagic clearance. They also had significantly reduced peroxisome proliferator-activated receptor-γ (PPAR-γ) and CD36 gene expression and protein levels, and lower expression of mRNA coding for enzymes involved in de novo lipogenesis. GDNF-loaded nanoparticles were protective against short-term HFD-induced hepatic steatosis and attenuated liver fibrosis in mice with long-standing HFD-induced hepatic steatosis. They also suppressed the liver expression of steatosis-associated genes. In vitro, GDNF suppressed triglyceride accumulation in Hep G2 cells through enhanced p38 mitogen-activated protein kinase-dependent signaling and inhibition of PPAR-γ gene promoter activity. These results show that GDNF acts directly in the liver to protect against HFD-induced cellular stress and that GDNF may have a role in the treatment of nonalcoholic fatty liver disease.

IB4(+) nociceptors mediate persistent muscle pain induced by GDNF.

PubMed

Alvarez, Pedro; Chen, Xiaojie; Bogen, Oliver; Green, Paul G; Levine, Jon D

2012-11-01

Skeletal muscle is a well-known source of glial cell line-derived neurotrophic factor (GDNF), which can produce mechanical hyperalgesia. Since some neuromuscular diseases are associated with both increased release of GDNF and intense muscle pain, we explored the role of GDNF as an endogenous mediator in muscle pain. Intramuscularly injected GDNF induced a dose-dependent (0.1-10 ng/20 μl) persistent (up to 3 wk) mechanical hyperalgesia in the rat. Once hyperalgesia subsided, injection of prostaglandin E(2) at the site induced a prolonged mechanical hyperalgesia (>72 h) compared with naïve rats (<4 h; hyperalgesic priming). Selective neurotoxic destruction of IB4(+) nociceptors attenuated both GDNF hyperalgesia and hyperalgesic priming. Ergonomic muscular injury induced by eccentric exercise or mechanical vibration increased muscle GDNF levels at 24 h, a time point where rats also exhibited marked muscle hyperalgesia. Intrathecal antisense oligodeoxynucleotides to mRNA encoding GFRα1, the canonical binding receptor for GDNF, reversibly inhibited eccentric exercise- and mechanical vibration-induced muscle hyperalgesia. Finally, electrophysiological recordings from nociceptors innervating the gastrocnemius muscle in anesthetized rats, revealed significant increase in response to sustained mechanical stimulation after local GDNF injection. In conclusion, these data indicate that GDNF plays a role as an endogenous mediator in acute and induction of chronic muscle pain, an effect likely to be produced by GDNF action at GFRα1 receptors located in IB4(+) nociceptors.

TGFβ induces GDNF responsiveness in neurons by recruitment of GFRα1 to the plasma membrane

PubMed Central

Peterziel, H.; Unsicker, K.; Krieglstein, K.

2002-01-01

We have previously shown that the neurotrophic effect of glial cell line–derived neurotrophic factor (GDNF) in vitro and in vivo requires the presence of transforming growth factor (TGF)β. Using primary neurons (chick E8 ciliary) we show that the combination of GDNF plus TGFβ promotes survival, whereas the single factors do not. This cooperative effect is inhibited by blocking the extracellular signal-regulated kinase (ERK)/MAPK pathway, but not by interfering with the PI3 kinase signaling cascade. Although there is no functional GDNF signaling in the absence of TGFβ, pretreatment with TGFβ confers GDNF responsiveness to the cells. This is not due to upregulation of GDNF receptors mRNA and protein, but to TGFβ-induced recruitment of the glycosyl-phosphatidylinositol-anchored GDNF receptor (GFR)α1 to the plasma membrane. This is supported by the fact that GDNF in the presence of a soluble GFRα1 can promote survival in the absence of TGFβ. Our data suggest that TGFβ is involved in GFRα1 membrane translocation, thereby permitting GDNF signaling and neurotrophic effects. PMID:12370242

The timing and location of glial cell line-derived neurotrophic factor expression determine enteric nervous system structure and function.

PubMed

Wang, Hongtao; Hughes, Inna; Planer, William; Parsadanian, Alexander; Grider, John R; Vohra, Bhupinder P S; Keller-Peck, Cynthia; Heuckeroth, Robert O

2010-01-27

Ret signaling is critical for formation of the enteric nervous system (ENS) because Ret activation promotes ENS precursor survival, proliferation, and migration and provides trophic support for mature enteric neurons. Although these roles are well established, we now provide evidence that increasing levels of the Ret ligand glial cell line-derived neurotrophic factor (GDNF) in mice causes alterations in ENS structure and function that are critically dependent on the time and location of increased GDNF availability. This is demonstrated using two different strains of transgenic mice and by injecting newborn mice with GDNF. Furthermore, because different subclasses of ENS precursors withdraw from the cell cycle at different times during development, increases in GDNF at specific times alter the ratio of neuronal subclasses in the mature ENS. In addition, we confirm that esophageal neurons are GDNF responsive and demonstrate that the location of GDNF production influences neuronal process projection for NADPH diaphorase-expressing, but not acetylcholinesterase-, choline acetyltransferase-, or tryptophan hydroxylase-expressing, small bowel myenteric neurons. We further demonstrate that changes in GDNF availability influence intestinal function in vitro and in vivo. Thus, changes in GDNF expression can create a wide variety of alterations in ENS structure and function and may in part contribute to human motility disorders.

Mechanisms of impaired nephrogenesis with fetal growth restriction: altered renal transcription and growth factor expression

PubMed Central

Abdel-Hakeem, Ahmed K; Henry, Tasmia Q; Magee, Thomas R; Desai, Mina; Ross, Michael; Mansano, Roy; Torday, John; Nast, Cynthia C.

2010-01-01

Objective Maternal food restriction during pregnancy results in growth restricted newborns and reduced glomerular number, contributing to programmed offspring hypertension. We investigated whether reduced nephrogenesis may be programmed by dysregulation of factors controlling ureteric bud branching and mesenchyme to epithelial transformation. Study Design 10 to 20 days gestation, Sprague Dawley pregnant rats (n=6/group) received ad libitum food; FR rats were 50% food restricted. At embryonic day 20, mRNA and protein expression of WT1, Pax2, FGF2, GDNF, cRET, WNT4, WNT11, BMP4, BMP7, and FGF7 were determined by real-time PCR and Western blotting. Results Maternal FR resulted in up-regulated mRNA expression for WT1, FGF2, and BMP7 whereas Pax2, GDNF, FGF7, BMP4, WNT4, and WNT11 mRNAs were down-regulated. Protein expression was concordant for WT1, GDNF, Pax2, FGF7, BMP4 and WNT4. Conclusion Maternal FR altered gene expression of fetal renal transcription and growth factors, and likely contributes to development of offspring hypertension. PMID:18639218

Glial cell-derived neurotrophic factor alleviates sepsis-induced neuromuscular dysfunction by decreasing the expression of γ- and α7-nicotinic acetylcholine receptors in an experimental rat model of neuromyopathy.

PubMed

Wang, Xin; Min, Su; Xie, Fei; Yang, Jun; Li, Liang; Chen, Jingyuan

2018-02-05

Sepsis-induced neuromuscular dysfunction results from up-regulation of the expression of γ- and α7-nicotinic acetylcholine receptors (nAChR). Although glial cell derived neurotrophic factor (GDNF) has been implicated in repairing and supporting neurons, little is known about the effects of GDNF on demyelination of nerves in sepsis. In this study, we tested the hypothesis that GDNF could alleviate sepsis-induced neuromuscular dysfunction by decreasing the expression of γ- and α7-nAChR in an experimental rat model of neuromyopathy. Rats were randomly divided into a sham group and a sepsis group. Levels of inflammatory factors, muscle function, and nicotinic acetylcholine receptors were tested in rats after cecal ligation and puncture (CLP). At 24 h after CLP, GDNF was injected around the sciatic nerve of sepsis rats, cytokines were detected by enzyme-linked immunosorbent assay (ELISA), and immunofluorescence staining was used to detect the expression of nAChRs. GDNF and its downstream effector (Erk1/2 and GFR-α), neuregulin-1 (NRG-1) and γ- and α7-nAChR were measured using Western blot analysis. The expression of GDNF reached a minimum at 24 h after CLP. Compared with the sham group, the release of cytokines and the expression of γ- and α7-nAChR were significantly increased in the sepsis group. The administration of GDNF significantly alleviated sepsis-induced neuromuscular dysfunction, as well as reducing the expression of γ- and α7-nAChR. In addition, the expression of Erk1/2, GFR-α, NRG-1 were significantly increased after GDNF treatment. GDNF administration may improve patient outcomes by reducing the demyelination of nerves and the expression of γ- and α7-nAChR. Copyright © 2018. Published by Elsevier Inc.

Non-viral gene therapy for GDNF production in RCS rat: the crucial role of the plasmid dose.

PubMed

Touchard, E; Heiduschka, P; Berdugo, M; Kowalczuk, L; Bigey, P; Chahory, S; Gandolphe, C; Jeanny, J-C; Behar-Cohen, F

2012-09-01

Glial cell line-derived neurotrophic factor (GDNF) is one of the candidate molecules among neurotrophic factors proposed for a potential treatment of retinitis pigmentosa (RP). It must be administered repeatedly or through sustained releasing systems to exert prolonged neuroprotective effects. In the dystrophic Royal College of Surgeon's (RCS) rat model of RP, we found that endogenous GDNF levels dropped during retinal degeneration time course, opening a therapeutic window for GDNF supplementation. We showed that after a single electrotransfer of 30 μg of GDNF-encoding plasmid in the rat ciliary muscle, GDNF was produced for at least 7 months. Morphometric, electroretinographic and optokinetic analyses highlighted that this continuous release of GDNF delayed photoreceptors (PRs) as well as retinal functions loss until at least 70 days of age in RCS rats. Unexpectedly, increasing the GDNF secretion level accelerated PR degeneration and the loss of electrophysiological responses. This is the first report: (i) demonstrating the efficacy of GDNF delivery through non-viral gene therapy in RP; (ii) establishing the efficacy of intravitreal administration of GDNF in RP associated with a mutation in the retinal pigment epithelium; and (iii) warning against potential toxic effects of GDNF within the eye/retina.

GDNF-RET signaling in ER-positive breast cancers is a key determinant of response and resistance to aromatase inhibitors

PubMed Central

Morandi, Andrea; Martin, Lesley-Ann; Gao, Qiong; Pancholi, Sunil; Mackay, Alan; Robertson, David; Zvelebil, Marketa; Dowsett, Mitch; Plaza-Menacho, Ivan; Isacke, Clare M.

2013-01-01

Most breast cancers at diagnosis are estrogen receptor (ER)-positive and depend on estrogen for growth and survival. Blocking estrogen biosynthesis by aromatase inhibitors (AI) has therefore become a first-line endocrine therapy for post-menopausal women with ER-positive breast cancers. Despite providing substantial improvements in patient outcome, AI resistance remains a major clinical challenge. The receptor tyrosine kinase RET and its co-receptor GFRα1 are upregulated in a subset of ER-positive breast cancers, and the RET ligand, glial-derived neurotrophic factor (GDNF) is upregulated by inflammatory cytokines. Here we report the findings of a multidisciplinary strategy to address the impact of GDNF-RET signaling in the response to AI treatment. In breast cancer cells in 2D and 3D culture, GDNF-mediated RET signaling is enhanced in a model of AI resistance. Further, GDNF-RET signaling promoted the survival of AI-resistant cells and elicited resistance in AI-sensitive cells. Both these effects were selectively reverted by the RET kinase inhibitor NVP-BBT594. Gene expression profiling in ER-positive cancers defined a proliferation-independent GDNF-response signature that prognosed poor patient outcome and, more importantly, predicted poor response to AI treatment with the development of resistance. We validated these findings by demonstrating increased RET protein expression levels in an independent cohort of AI-resistant patient specimens. Together, our results establish GDNF-RET signaling as a rational therapeutic target to combat or delay the onset of AI resistance in breast cancer. PMID:23650283

AAV2-mediated gene transfer of GDNF to the striatum of MPTP monkeys enhances the survival and outgrowth of co-implanted fetal dopamine neurons

PubMed Central

Elsworth, JD; Redmond, DE; Leranth, C; Bjugstad, KB; Sladek, JR; Collier, TJ; Foti, SB; Samulski, RJ; Vives, KP; Roth, RH

2009-01-01

Neural transplantation offers the potential of treating Parkinson’s disease by grafting fetal dopamine neurons to depleted regions of the brain. However, clinical studies of neural grafting in Parkinson’s disease have produced only modest improvements. One of the main reasons for this is the low survival rate of transplanted neurons. The inadequate supply of critical neurotrophic factors in the adult brain is likely to be a major cause of early cell death and restricted outgrowth of fetal grafts placed into the mature striatum. Glial derived neurotrophic factor (GDNF) is a potent neurotrophic factor that is crucial to the survival, outgrowth and maintenance of dopamine neurons, and so is a candidate for protecting grafted fetal dopamine neurons in the adult brain. We found that implantation of adeno-associated virus type 2 encoding GDNF (AAV2-GDNF) in the normal monkey caudate nucleus induced over-expression of GDNF that persisted for at least 6 months after injection. In a 6-month within-animal controlled study, AAV2-GDNF enhanced the survival of fetal dopamine neurons by 4-fold, and increased the outgrowth of grafted fetal dopamine neurons by almost 3-fold in the caudate nucleus of MPTP-treated monkeys, compared with control grafts in the other caudate nucleus. Thus, the addition of GDNF gene therapy to neural transplantation may be a useful strategy to improve treatment for Parkinson’s disease. PMID:18346734

[Effects of the scalp acupuncture at baihui (DU20) through qubin (GB7) on the expressions of GDNF VEGF in the brain tissue of rats with acute intracerebral hemorrhage].

PubMed

Zhang, Guo-Wei; Zou, Wei; Liu, Fang

2012-09-01

To observe the effects of the scalp acupuncture at Baihui (DU20) through Qubin (GB7) on the expressions of glial cell line-derived neurotrophic factor (GDNF) and vascular endothelial growth factor (VEGF) in the brain tissue of rats with acute intracerebral hemorrhage (AICH). Totally 150 healthy male Wistar rats were used to prepare the intracerebral hemorrhage (ICH) model. They were randomly divided into the model group, the acupuncture group, and the Western medicine group, 50 in each group. Rats in each group were then randomly divided into five subgroups, i. e., 6 h, day 1, day 2, day 3, and day 7, 10 in each subgroup. Another 10 normal rats were recruited as the blank control group. Rats in the acupuncture group were acupunctured at Baihui (DU20) through Qubin (GB7) on the lesion side. Rats in the Western medicine group were administered with aniracetam diluent 1 mL by gastrogavage, 3 times daily. Rats in the model group were bound the same way as those in the acupuncture group 30 min daily, and then administered with normal saline 1 mL by gastrogavage, 3 times daily. The expressions of GDNF and VEGF in the brain tissue of the rats in each group were detected using in situ hybridization and immunohistochemical methods. Compared with the blank control group, the number of GDNF positive cells in the model group increased at 6 h-3 days, and the number of VEGF positive cells in the model group increased at each time point, showing statistical difference (P<0.01). The numbers of GDNF positive cells increased in the acupuncture group at each time point, showing statistical difference when compared with those of the model group and the Western medicine group (P<0.01). There was no statistical difference between the model group and the Western medicine group at each time point (P>0.05). Compared with the model group, the number of VEGF positive cells in the acupuncture group decreased at 6 h -3 days. The VEGF positive cells increased at day 7 and were higher than that

The gene coding for glial cell line derived neurotrophic factor (GDNF) maps to chromosome 5p12-p13.1

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

Schindelhauer, D.; Schuffenhauer, S.; Meitinger, T.

1995-08-10

The gene coding for glial cell line derived neurotrophic factor (GDNF) has biological properties that may have potential as a treatment for Parkinson`s and motoneuron diseases. Using the NIGMS Mapping Panel 2, we have localized the GDNF gene to human chromosome 5p12-p13.1. Large NruI and NotI fragments on chromosome 5 will facilitate the construction of a long-range map of the region. 26 refs., 1 fig., 1 tab.

Dampened Amphetamine-Stimulated Behavior and Altered Dopamine Transporter Function in the Absence of Brain GDNF.

PubMed

Kopra, Jaakko J; Panhelainen, Anne; Af Bjerkén, Sara; Porokuokka, Lauriina L; Varendi, Kärt; Olfat, Soophie; Montonen, Heidi; Piepponen, T Petteri; Saarma, Mart; Andressoo, Jaan-Olle

2017-02-08

Midbrain dopamine neuron dysfunction contributes to various psychiatric and neurological diseases, including drug addiction and Parkinson's disease. Because of its well established dopaminotrophic effects, the therapeutic potential of glial cell line-derived neurotrophic factor (GDNF) has been studied extensively in various disorders with disturbed dopamine homeostasis. However, the outcomes from preclinical and clinical studies vary, highlighting a need for a better understanding of the physiological role of GDNF on striatal dopaminergic function. Nevertheless, the current lack of appropriate animal models has limited this understanding. Therefore, we have generated novel mouse models to study conditional Gdnf deletion in the CNS during embryonic development and reduction of striatal GDNF levels in adult mice via AAV-Cre delivery. We found that both of these mice have reduced amphetamine-induced locomotor response and striatal dopamine efflux. Embryonic GDNF deletion in the CNS did not affect striatal dopamine levels or dopamine release, but dopamine reuptake was increased due to increased levels of both total and synaptic membrane-associated dopamine transporters. Collectively, these results suggest that endogenous GDNF plays an important role in regulating the function of dopamine transporters in the striatum. SIGNIFICANCE STATEMENT Delivery of ectopic glial cell line-derived neurotrophic factor (GDNF) promotes the function, plasticity, and survival of midbrain dopaminergic neurons, the dysfunction of which contributes to various neurological and psychiatric diseases. However, how the deletion or reduction of GDNF in the CNS affects the function of dopaminergic neurons has remained unknown. Using conditional Gdnf knock-out mice, we found that endogenous GDNF affects striatal dopamine homeostasis and regulates amphetamine-induced behaviors by regulating the level and function of dopamine transporters. These data regarding the physiological role of GDNF are

Synergistic Effects of GDNF and VEGF on Lifespan and Disease Progression in a Familial ALS Rat Model

PubMed Central

Krakora, Dan; Mulcrone, Patrick; Meyer, Michael; Lewis, Christina; Bernau, Ksenija; Gowing, Genevieve; Zimprich, Chad; Aebischer, Patrick; Svendsen, Clive N; Suzuki, Masatoshi

2013-01-01

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease characterized by the progressive loss of motor neurons in the brain and spinal cord. We have recently shown that human mesenchymal stem cells (hMSCs) modified to release glial cell line-derived neurotrophic factor (GDNF) decrease disease progression in a rat model of ALS when delivered to skeletal muscle. In the current study, we determined whether or not this effect could be enhanced by delivering GDNF in concert with other trophic factors. hMSC engineered to secrete GDNF (hMSC-GDNF), vascular endothelial growth factor (hMSC-VEGF), insulin-like growth factor-I (hMSC-IGF-I), or brain-derived neurotrophic factor (hMSC-BDNF), were prepared and transplanted bilaterally into three muscle groups. hMSC-GDNF and hMSC-VEGF prolonged survival and slowed the loss of motor function, but hMSC-IGF-I and hMSC-BDNF did not have any effect. We then tested the efficacy of a combined ex vivo delivery of GDNF and VEGF in extending survival and protecting neuromuscular junctions (NMJs) and motor neurons. Interestingly, the combined delivery of these neurotrophic factors showed a strong synergistic effect. These studies further support ex vivo gene therapy approaches for ALS that target skeletal muscle. PMID:23712039

Glial-derived neurotropic factor and RET gene expression in normal human anterior pituitary cell types and in pituitary tumors.

PubMed

Japón, Miguel A; Urbano, Angel G; Sáez, Carmen; Segura, Dolores I; Cerro, Alfonso Leal; Diéguez, Carlos; Alvarez, Clara V

2002-04-01

Glial-derived neurotropic factor (GDNF) signaling is mediated through a 2-component system consisting of the so-called GDNF receptor-alpha (GFRalpha1), which binds to GDNF. This complex activates the tyrosine kinase receptor RET. In this paper we demonstrate GDNF, GFRalpha1, and RET mRNA and protein expression in the human anterior pituitary gland. Double immunohistochemistry of anterior pituitary sections showed GDNF immunoreactivity in more than 95% of somatotrophs and to a lesser extent in corticotrophs (20%); it was almost absent in the remaining cell types. Also, although more than 95% of somatotrophs were stained for RET, no positive immunostaining could be detected in other cell types. Furthermore, we have looked for GDNF and RET in human pituitary adenomas of various hormonal phenotypes. Strong positive immunostaining was found for c-RET in all of the GH-secreting adenomas screened as well as in 50% of ACTH-producing adenomas. Positive immunostaining for GDNF was found in all of the GH-secreting adenomas and in 10% of the corticotropinomas. Lastly, we found strong positive immunostaining for GFRalpha1 in 90% of the somatotropinomas and 50% of the corticotropinomas as well as in 1 of 8 prolactinomas and 1 of 13 nonfunctioning adenomas. All of the remaining pituitary tumors screened were negative for RET, GDNF, and GFRalpha1. This study indicates that GDNF may well be acting in the regulation of somatotroph cell growth and/or cell function in the normal human anterior pituitary gland. The expression of RET in all of the somatotropinomas and in 50% of the ACTH-producing tumors implies that GDNF and RET could be involved in the pathogenesis of pituitary tumors.

Biology of GDNF and its receptors - Relevance for disorders of the central nervous system.

PubMed

Ibáñez, Carlos F; Andressoo, Jaan-Olle

2017-01-01

A targeted effort to identify novel neurotrophic factors for midbrain dopaminergic neurons resulted in the isolation of GDNF (glial cell line-derived neurotrophic factor) from the supernatant of a rat glial cell line in 1993. Over two decades and 1200 papers later, the GDNF ligand family and their different receptor systems are now recognized as one of the major neurotrophic networks in the nervous system, important for the development, maintenance and function of a variety of neurons and glial cells. The many ways in which the four members of the GDNF ligand family can signal and function allow these factors to take part in the control of multiple types of processes, from neuronal survival to axon guidance and synapse formation in the developing nervous system, to synaptic function and regenerative responses in the adult. In this review, we will briefly summarize basic aspects of GDNF signaling mechanisms and receptor systems and then review our current knowledge of the physiology of GDNF activities in the central nervous system, with an eye to its relevance for neurodegenerative and neuropsychiatric diseases. Copyright © 2016 Elsevier Inc. All rights reserved.

GDNF family receptor α-1 in the catfish: Possible implication to brain dopaminergic activity.

PubMed

Mamta, Sajwan-Khatri; Senthilkumaran, Balasubramanian

2018-05-31

Glial cell line-derived neurotrophic factor (GDNF)is a potent trophic factor that preferentially binds to GDNF family receptor α-1 (GFRα-1)by regulating dopaminergic (DA-ergic) neuronsin brain. Present study aimed to evaluate the significance of GFRα-1 expression during early brain development in catfish. Initially, the full-length cDNA of GFRα-1 was cloned from adult brain which showed high homology with other vertebrate counterparts. Quantitative PCR analysis of tissue distribution revealed ubiquitous expression of GFRα-1 in the tissues analyzed with high levels in female brain and ovary. Significant high expression was evident in brain at 75 and 100 days post hatch females than the respective age-match males. Expression of GFRα-1 was high in brain during the spawning phase when compared to other reproductive phases. Localization of GFRα-1 revealed its presence in preoptic area-hypothalamus which correlated well with the expression profile in discrete areas of brain in adult catfish. Transient silencing of GFRα-1through siRNA lowered expression levels of GFRα-1, which further down regulated the expression of certain brain-specific genes. Expression of GFRα-1 in brain declined significantly upon treatment with the 1-methyl-1,2,3,6-tetrahydropyridinecausing neurodegeneration which further correlated with catecholamines (CA), L-3,4-dihydroxyphenylalanine, DA and norepinephrine levels. Taken together, GFRα-1 plausibly entrains gonadotropin-releasing hormone and gonadotropin axiseither directly or indirectly, at least by partially targeting CA-ergic activity. Copyright © 2018 Elsevier Inc. All rights reserved.

Intervertebral disc degeneration-induced expression of pain-related molecules: glial cell-derived neurotropic factor as a key factor.

PubMed

Jung, Woon-Won; Kim, Hyun-Sook; Shon, Jong-Ryeul; Lee, Min; Lee, Sang-Heon; Sul, Donggeun; Na, Heung Sik; Kim, Joo Han; Kim, Byung-Jo

2011-10-01

Discogenic low back pain has been shown to develop into chronic intractable pain due to an unknown pathogenesis. To study the mechanism of discogenic pain, we analyzed the serial expression of pain-related molecules in the dorsal root ganglia (DRG) and thalamus using a newly developed rat model of disc degeneration. Ten microliters of complete Freund's adjuvant was injected into the L5-6 disc of male Sprague-Dawley rats for 10 minutes using a 26-gauge needle. Using a behavioral test, rats with significant pain were selected and subsequently serial gene expression of pain-related molecules in the DRG and the thalamus was analyzed by reverse transcriptase polymerase chain reaction. The expression of tumor necrosis factor-α and interleukin-1β significantly increased at 4 and 8 weeks in the DRG of rats with pain. Furthermore, interleukin-6 was significantly increased at 4 weeks in the DRG; however, these cytokines did not show a significant change in the thalamus. Calcitonin gene-related peptide and substance P were significantly increased in DRG at 4 and 8 weeks and in the thalamus at 2 and 4 weeks. The level of nerve growth factor-β did not significantly increase in the DRG or thalamus, whereas glial cell line-derived neurotropic factor (GDNF) was significantly increased at 2 weeks and was sustained through 8 weeks in both the DRG and thalamus. The disc degeneration rat model described herein led to significant pain of a chronic nature. The gradual and persistent increase of GDNF in both the thalamus and DRG suggests that GDNF might be a key factor in the development of intractable, chronic discogenic pain.

Intermittent convection-enhanced delivery of GDNF into rhesus monkey putamen: absence of local or cerebellar toxicity.

PubMed

Luz, Matthias; Allen, Philip C; Bringas, John; Boiko, Chris; Stockinger, Diane E; Nikula, Kristen J; Lewis, Owen; Woolley, Max; Fibiger, H Christian; Bankiewicz, Krystof; Mohr, Erich

2018-05-22

Glial cell line-derived neurotrophic factor (GDNF) has demonstrated neurorestorative and neuroprotective effects in rodent and nonhuman primate models of Parkinson's disease. However, continuous intraputamenal infusion of GDNF (100 µg/day) resulted in multifocal cerebellar Purkinje cell loss in a 6-month toxicity study in rhesus monkeys. It was hypothesized that continuous leakage of GDNF into the cerebrospinal fluid compartment during the infusions led to down-regulation of GDNF receptors on Purkinje cells, and that subsequent acute withdrawal of GDNF then mediated the observed cerebellar lesions. Here we present the results of a 9-month toxicity study in which rhesus monkeys received intermittent intraputamenal infusions via convection-enhanced delivery. Animals were treated with GDNF (87.1 µg; N = 14) or vehicle (N = 6) once every 4 weeks for a total of 40 weeks (11 treatments). Four of the GDNF-treated animals were utilized in a satellite study assessing the impact of concomitant catheter repositioning prior to treatment. In the main study, eight animals (5 GDNF, 3 control) were euthanized at the end of the treatment period, along with the four satellite study animals, while the remaining eight animals (5 GDNF, 3 control) were euthanized at the end of a 12-week recovery period. There were no GDNF-related adverse effects and in particular, no GDNF-related microscopic findings in the brain, spinal cord, dorsal root ganglia, or trigeminal ganglia. Therefore, 87.1 µg/4 weeks is considered the no observed adverse effect level for GDNF in rhesus monkeys receiving intermittent, convection-enhanced delivery of GDNF for 9 months.

Post-treatment effects of local GDNF administration to the inner ears of deafened guinea pigs.

PubMed

Fransson, Anette; Maruyama, Jun; Miller, Josef M; Ulfendahl, Mats

2010-09-01

For patients with profound hearing loss, a cochlear implant is the only treatment available today. The function of a cochlear implant depends in part on the function and survival of spiral ganglion neurons. Following deafferentation, glial cell-derived neurotrophic factor (GDNF) is known to affect spiral ganglion neuron survival. The purpose of this study was to assess delayed GDNF treatment after deafening, the effects of cessation of GDNF treatment, and the effects of subsequent antioxidants on responsiveness and survival of the spiral ganglion neurons. Three-week deafened (by local neomycin administration) guinea pigs were implanted in the scala tympani with a combined cochlear implant electrode and cannula. GDNF (1 μg/mL) or artificial perilymph was then delivered for 4 weeks, following which the animals received systemic ascorbic acid  +  Trolox or saline for an additional 4 weeks. Thresholds for electrically-evoked auditory brain stem responses (eABRs) were significantly elevated at 3 weeks with deafness, stabilized with GDNF, and showed no change with GDNF cessation and treatment with antioxidants or saline. The populations of spiral ganglion neurons were reduced with deafness (by 40% at 3 weeks and 70% at 11 weeks), and rescued from cell death by GDNF with no further reduction at 8 weeks following 4 weeks of cessation of GDNF treatment equally in both the antioxidant- and saline-treated groups. Local growth factor treatment of the deaf ear may prevent deterioration in electrical responsiveness and rescue auditory nerve cells from death; these effects outlast the period of treatment, and may enhance the benefits of cochlear implant therapy for the deaf.

Effect of Removal of Spermatogonial Stem Cells (SSCs) from In Vitro Culture on Gene Expression of Niche Factors in Bovine

PubMed Central

Akbarinejad, Vahid; Tajik, Parviz; Movahedin, Mansoureh; Youssefi, Reza

2016-01-01

Background: Niche cells, regulating Spermatogonial Stem Cells (SSCs) fate are believed to have a reciprocal communication with SSCs. The present study was conducted to evaluate the effect of SSC elimination on the gene expression of Glial cell line-Derived Neurotrophic Factor (GDNF), Fibroblast Growth Factor 2 (FGF2) and Kit Ligand (KITLG), which are the main growth factors regulating SSCs development and secreted by niche cells, primarily Sertoli cells. Methods: Following isolation, bovine testicular cells were cultured for 12 days on extracellular matrix-coated plates. In the germ cell-removed group, the SSCs were removed from the in vitro culture using differential plating; however, in the control group, no intervention in the culture was performed. Colony formation of SSCs was evaluated using an inverted microscope. The gene expression of growth factors and spermatogonia markers were assessed using quantitative real time PCR. Results: SSCs colonies were developed in the control group but they were rarely observed in the germ cell-removed group; moreover, the expression of spermatogonia markers was detected in the control group while it was not observed in the germ cell-removed group, substantiating the success of SSCs removal. The expression of Gdnf and Fgf2 was greater in the germ cell-removed than control group (p<0.05), whereas the expression of Kitlg was lower in the germ cell-removed than control group (p< 0.05). Conclusion: In conclusion, the results revealed that niche cells respond to SSCs removal by upregulation of GDNF and FGF2, and downregulation of KITLG in order to stimulate self-renewal and arrest differentiation. PMID:27563426

The In Vitro Differentiation of GDNF Gene-Engineered Amniotic Fluid-Derived Stem Cells into Renal Tubular Epithelial-Like Cells.

PubMed

Lu, Ying; Wang, Zhuojun; Chen, Lu; Wang, Jia; Li, Shulin; Liu, Caixia; Sun, Dong

2018-05-01

Amniotic fluid is an alternative source of stem cells, and human amniotic fluid-derived stem cells (AFSCs) obtained from a small amount of amniotic fluid collected during the second trimester represent a novel source for use in regenerative medicine. These AFSCs are characterized by lower diversity, a higher proliferation rate, and a wider differentiation capability than adult mesenchymal stem cells. AFSCs are selected based on the cell surface marker c-kit receptor (CD117) using immunomagnetic sorting. Glial cell line-derived neurotrophic factor (GDNF) is expressed during early kidney development and regulates the proliferation and differentiation of stem cells in vitro. In this study, c-kit-sorted AFSCs were induced toward osteogenic or adipogenic differentiation. AFSCs engineered via the insertion of GDNF were cocultured with mouse renal tubular epithelial cells (mRTECs), which were preconditioned by hypoxia-reoxygenation in vitro. After coculture for 8 days, AFSCs differentiation into epithelial-like cells was evaluated by performing immunofluorescence, flow cytometry, and quantitative real-time polymerase chain reaction to identify cells expressing the renal epithelial markers, cytokeratin 18 (CK18), E-cadherin, aquaporin-1 (AQP1), and paired box 2 gene (Pax2). The GDNF gene enhanced AFSCs differentiation into RTECs. AFSCs possess self-renewal ability and multiple differentiation potential and thus represent a new source of stem cells.

History of Glial Cell Line-Derived Neurotrophic Factor (GDNF) and Its Use for Spinal Cord Injury Repair.

PubMed

Walker, Melissa J; Xu, Xiao-Ming

2018-06-13

Following an initial mechanical insult, traumatic spinal cord injury (SCI) induces a secondary wave of injury, resulting in a toxic lesion environment inhibitory to axonal regeneration. This review focuses on the glial cell line-derived neurotrophic factor (GDNF) and its application, in combination with other factors and cell transplantations, for repairing the injured spinal cord. As studies of recent decades strongly suggest that combinational treatment approaches hold the greatest therapeutic potential for the central nervous system (CNS) trauma, future directions of combinational therapies will also be discussed.

GDNF-secreting mesenchymal stem cells provide localized neuroprotection in an inflammation-driven rat model of Parkinson's disease.

PubMed

Hoban, D B; Howard, L; Dowd, E

2015-09-10

Constraints involving the delivery method of glial cell line-derived neurotrophic factor (GDNF) have hampered its efficacy as a neuroprotectant in Parkinson's disease. Ex vivo gene therapy, in which suitable cells, such as bone marrow-derived mesenchymal stem cells (MSCs), are genetically engineered to overexpress GDNF (GDNF-MSCs) prior to transplantation may be more beneficial than direct brain infusion of the neurotrophin. Previously, GDNF-MSCs have been assessed in the commonly employed 6-hydroxydopamine neurotoxic model of Parkinson's disease. In this study however, we used an emerging inflammatory model of Parkinson's disease (the lipopolysaccharide (LPS) model) to assess the ability of transplanted GDNF-MSCs to protect against LPS-induced neuroinflammation, neurodegeneration and behavioral impairment. Thirty male Sprague-Dawley rats were used in this experiment. Rats were performance matched based on baseline motor function tests into three groups (LPS lesion only, LPS lesion+GFP-MSCs, LPS lesion+GDNF-MSCs; n=10/group). Both cell groups received a unilateral intra-striatal transplant of either 200,000 GDNF-MSCs or 200,000 GFP-MSCs (as a control). One day post-transplantation, all rats received a unilateral intra-nigral infusion of LPS (10 μg in 2 μl sterile saline). Rats were sacrificed by transcardial perfusion-fixation and their brains were used for post mortem quantitative immunohistochemistry. Injection of LPS into the substantia nigra induced a pronounced local inflammatory response which resulted in 20% loss of nigrostriatal dopaminergic neurons and impaired contralateral motor function. Following transplantation of GDNF-MSCs to the striatum, dense areas of TH-positive staining directly proximal to the transplant site were observed. Most importantly, this effect was observed only in the GDNF-MSC transplanted group and not the GFP-MSC transplanted group demonstrating protection and/or sprouting of the dopaminergic terminals induced by the secreted GDNF

Naringin protects the nigrostriatal dopaminergic projection through induction of GDNF in a neurotoxin model of Parkinson's disease.

PubMed

Leem, Eunju; Nam, Jin Han; Jeon, Min-Tae; Shin, Won-Ho; Won, So-Yoon; Park, Sang-Joon; Choi, Myung-Sook; Jin, Byung Kwan; Jung, Un Ju; Kim, Sang Ryong

2014-07-01

This study investigated the effect of naringin, a major flavonoid in grapefruit and citrus fruits, on the degeneration of the nigrostriatal dopaminergic (DA) projection in a neurotoxin model of Parkinson's disease (PD) in vivo and the potential underlying mechanisms focusing on the induction of glia-derived neurotrophic factor (GDNF), well known as an important neurotrophic factor involved in the survival of adult DA neurons. 1-Methyl-4-phenylpyridinium (MPP(+)) was unilaterally injected into the medial forebrain bundle of rat brains for a neurotoxin model of PD in the presence or absence of naringin by daily intraperitoneal injection. To ascertain whether naringin-induced GDNF contributes to neuroprotection, we further investigated the effects of intranigral injection of neutralizing antibodies against GDNF in the MPP(+) rat model of PD. Our observations demonstrate that naringin could increase the level of GDNF in DA neurons, contributing to neuroprotection in the MPP(+) rat model of PD, with activation of mammalian target of rapamycin complex 1. Moreover, naringin could attenuate the level of tumor necrosis factor-α in microglia increased by MPP(+)-induced neurotoxicity in the substantia nigra. These results indicate that naringin could impart to DA neurons the important ability to produce GDNF as a therapeutic agent against PD with anti-inflammatory effects, suggesting that naringin is a beneficial natural product for the prevention of DA degeneration in the adult brain. Copyright © 2014 Elsevier Inc. All rights reserved.

Pharmacokinetics and safety in rhesus monkeys of a monoclonal antibody-GDNF fusion protein for targeted blood-brain barrier delivery.

PubMed

Pardridge, William M; Boado, Ruben J

2009-10-01

Glial-derived neurotrophic factor (GDNF) is a potential therapy for stroke, Parkinson's disease, or drug addiction. However, GDNF does not cross the blood-brain barrier (BBB). GDNF is re-engineered as a fusion protein with a chimeric monoclonal antibody (MAb) to the human insulin receptor (HIR), which acts as a molecular Trojan horse to deliver the GDNF across the BBB. The pharmacokinetics (PK), toxicology, and safety pharmacology of the HIRMAb-GDNF fusion protein were investigated in Rhesus monkeys. The fusion protein was administered as an intravenous injection at doses up to 50 mg/kg over a 60 h period to 56 Rhesus monkeys. The plasma concentration of the HIRMAb-GDNF fusion protein was measured with a 2-site sandwich ELISA. No adverse events were observed in a 2-week terminal toxicology study, and no neuropathologic changes were observed. The PK analysis showed a linear relationship between plasma AUC and dose, a large systemic volume of distribution, as well as high clearance rates of 8-10 mL/kg/min. A no-observable-adverse-effect level is established in the Rhesus monkey for the acute administration of the HIRMAb-GDNF fusion protein. The fusion protein targeting the insulin receptor has a PK profile similar to a classical small molecule.

Effects of delayed treatment with combined GDNF and continuous electrical stimulation on spiral ganglion cell survival in deafened guinea pigs.

PubMed

Scheper, Verena; Paasche, Gerrit; Miller, Josef M; Warnecke, Athanasia; Berkingali, Nurdanat; Lenarz, Thomas; Stöver, Timo

2009-05-01

Electrical stimulation (ES) of spiral ganglion cells (SGC) via a cochlear implant is the standard treatment for profound sensor neural hearing loss. However, loss of hair cells as the morphological correlate of sensor neural hearing loss leads to deafferentation and death of SGC. Although immediate treatment with ES or glial cell line-derived neurotrophic factor (GDNF) can prevent degeneration of SGC, only few studies address the effectiveness of delayed treatment. We hypothesize that both interventions have a synergistic effect and that even delayed treatment would protect SGC. Therefore, an electrode connected to a pump was implanted into the left cochlea of guinea pigs 3 weeks after deafening. The contralateral untreated cochleae served as deafened intraindividual controls. Four groups were set up. Control animals received intracochlear infusion of artificial perilymph (AP/-). The experimental groups consisted of animals treated with AP in addition to continuous ES (AP/ES) or treated with GDNF alone (GDNF/-) or GDNF combined with continuous ES (GDNF/ES). Acoustically and electrically evoked auditory brain stem responses were recorded. All animals were killed 48 days after deafening; their cochleae were histologically evaluated. Survival of SGC increased significantly in the GDNF/- and AP/ES group compared with the AP/- group. A highly significant increase in SGC density was observed in the GDNF/ES group compared with the control group. Additionally, animals in the GDNF/ES group showed reduced EABR thresholds. Thus, delayed treatment with GDNF and ES can protect SGC from degeneration and may improve the benefits of cochlear implants.

Dendrobium alkaloids prevent Aβ25–35-induced neuronal and synaptic loss via promoting neurotrophic factors expression in mice

PubMed Central

Nie, Jing; Tian, Yong; Zhang, Yu; Lu, Yan-Liu; Li, Li-Sheng

2016-01-01

Background Neuronal and synaptic loss is the most important risk factor for cognitive impairment. Inhibiting neuronal apoptosis and preventing synaptic loss are promising therapeutic approaches for Alzheimer’s disease (AD). In this study, we investigate the protective effects of Dendrobium alkaloids (DNLA), a Chinese medicinal herb extract, on β-amyloid peptide segment 25–35 (Aβ25-35)-induced neuron and synaptic loss in mice. Method Aβ25–35(10 µg) was injected into the bilateral ventricles of male mice followed by an oral administration of DNLA (40 mg/kg) for 19 days. The Morris water maze was used for evaluating the ability of spatial learning and memory function of mice. The morphological changes were examined via H&E staining and Nissl staining. TUNEL staining was used to check the neuronal apoptosis. The ultrastructure changes of neurons were observed under electron microscope. Western blot was used to evaluate the protein expression levels of ciliary neurotrophic factor (CNTF), glial cell line-derived neurotrophic factor (GDNF), and brain-derived neurotrophic factor (BDNF) in the hippocampus and cortex. Results DNLA significantly attenuated Aβ25–35-induced spatial learning and memory impairments in mice. DNLA prevented Aβ25–35-induced neuronal loss in the hippocampus and cortex, increased the number of Nissl bodies, improved the ultrastructural injury of neurons and increased the number of synapses in neurons. Furthermore, DNLA increased the protein expression of neurotrophic factors BDNF, CNTF and GDNF in the hippocampus and cortex. Conclusions DNLA can prevent neuronal apoptosis and synaptic loss. This effect is mediated at least in part via increasing the expression of BDNF, GDNF and CNTF in the hippocampus and cortex; improving Aβ-induced spatial learning and memory impairment in mice. PMID:27994964

Neurturin and GDNF promote proliferation and survival of enteric neuron and glial progenitors in vitro.

PubMed

Heuckeroth, R O; Lampe, P A; Johnson, E M; Milbrandt, J

1998-08-01

Signaling through the c-Ret tyrosine kinase and the endothelin B receptor pathways is known to be critical for development of the enteric nervous system. To clarify the role of these receptors in enteric nervous system development, the effect of ligands for these receptors was examined on rat enteric neuron precursors in fully defined medium in primary culture. In this culture system, dividing Ret-positive cells differentiate, cluster into ganglia containing neurons and enteric glia, and create extensive networks reminiscent of the enteric plexus established in vivo. Glial cell-line-derived neurotrophic factor (GDNF) and neurturin both potently support survival and proliferation of enteric neuron precursors in this system. Addition of either neurturin or GDNF to these cultures increased the number of both neurons and enteric glia. Persephin, a third GDNF family member, shares many properties with neurturin and GDNF in the central nervous system and in kidney development. By contrast, persephin does not promote enteric neuron precursor proliferation or survival in these cultures. Endothelin-3 also does not increase the number of enteric neurons or glia in these cultures. Copyright 1998 Academic Press.

Human neural stem cells survive long term in the midbrain of dopamine-depleted monkeys after GDNF overexpression and project neurites toward an appropriate target.

PubMed

Wakeman, Dustin R; Redmond, D Eugene; Dodiya, Hemraj B; Sladek, John R; Leranth, Csaba; Teng, Yang D; Samulski, R Jude; Snyder, Evan Y

2014-06-01

Transplanted multipotent human fetal neural stem cells (hfNSCs) significantly improved the function of parkinsonian monkeys in a prior study primarily by neuroprotection, with only 3%-5% of cells expressing a dopamine (DA) phenotype. In this paper, we sought to determine whether further manipulation of the neural microenvironment by overexpression of a developmentally critical molecule, glial cell-derived neurotrophic factor (GDNF), in the host striatum could enhance DA differentiation of hfNSCs injected into the substantia nigra and elicit growth of their axons to the GDNF-expressing target. hfNSCs were transplanted into the midbrain of 10 green monkeys exposed to 1-methyl-4-phenyl-1,2,3,6-tetrahydro-pyridine. GDNF was delivered concomitantly to the striatum via an adeno-associated virus serotype 5 vector, and the fate of grafted cells was assessed after 11 months. Donor cells remained predominantly within the midbrain at the injection site and sprouted numerous neurofilament-immunoreactive fibers that appeared to course rostrally toward the striatum in parallel with tyrosine hydroxylase-immunoreactive fibers from the host substantia nigra but did not mature into DA neurons. This work suggests that hfNSCs can generate neurons that project long fibers in the adult primate brain. However, in the absence of region-specific signals and despite GDNF overexpression, hfNSCs did not differentiate into mature DA neurons in large numbers. It is encouraging, however, that the adult primate brain appeared to retain axonal guidance cues. We believe that transplantation of stem cells, specifically instructed ex vivo to yield DA neurons, could lead to reconstruction of some portion of the nigrostriatal pathway and prove beneficial for the parkinsonian condition. ©AlphaMed Press.

NANOS2 acts downstream of glial cell line-derived neurotrophic factor signaling to suppress differentiation of spermatogonial stem cells.

PubMed

Sada, Aiko; Hasegawa, Kazuteru; Pin, Pui Han; Saga, Yumiko

2012-02-01

Stem cells are maintained by both stem cell-extrinsic niche signals and stem cell-intrinsic factors. During murine spermatogenesis, glial cell line-derived neurotrophic factor (GDNF) signal emanated from Sertoli cells and germ cell-intrinsic factor NANOS2 represent key regulators for the maintenance of spermatogonial stem cells. However, it remains unclear how these factors intersect in stem cells to control their cellular state. Here, we show that GDNF signaling is essential to maintain NANOS2 expression, and overexpression of Nanos2 can alleviate the stem cell loss phenotype caused by the depletion of Gfra1, a receptor for GDNF. By using an inducible Cre-loxP system, we show that NANOS2 expression is downregulated upon the conditional knockout (cKO) of Gfra1, while ectopic expression of Nanos2 in GFRA1-negative spermatogonia does not induce de novo GFRA1 expression. Furthermore, overexpression of Nanos2 in the Gfra1-cKO testes prevents precocious differentiation of the Gfra1-knockout stem cells and partially rescues the stem cell loss phenotypes of Gfra1-deficient mice, indicating that the stem cell differentiation can be suppressed by NANOS2 even in the absence of GDNF signaling. Taken together, we suggest that NANOS2 acts downstream of GDNF signaling to maintain undifferentiated state of spermatogonial stem cells. Copyright © 2011 AlphaMed Press.

Mutation and deletion analysis of GFRα-1, encoding the co-receptor for the GDNF/RET complex, in human brain tumours

PubMed Central

Gimm, O; Gössling, A; Marsh, D J; Dahia, P L M; Mulligan, L M; Deimling, A von; Eng, C

1999-01-01

Glial cell line-derived neurotrophic factor (GDNF) plays a key role in the control of vertebrate neuron survival and differentiation in both the central and peripheral nervous systems. GDNF preferentially binds to GFRα-1 which then interacts with the receptor tyrosine kinase RET. We investigated a panel of 36 independent cases of mainly advanced sporadic brain tumours for the presence of mutations in GDNF and GFRα-1. No mutations were found in the coding region of GDNF. We identified six previously described GFRα-1 polymorphisms, two of which lead to an amino acid change. In 15 of 36 brain tumours, all polymorphic variants appeared to be homozygous. Of these 15 tumours, one also had a rare, apparently homozygous, sequence variant at codon 361. Because of the rarity of the combination of homozygous sequence variants, analysis for hemizygous deletion was pursued in the 15 samples and loss of heterozygosity was found in 11 tumours. Our data suggest that intragenic point mutations of GDNF or GFRα-1 are not a common aetiologic event in brain tumours. However, either deletion of GFRα-1 and/or nearby genes may contribute to the pathogenesis of these tumours. © 1999 Cancer Research Campaign PMID:10408842

GDNF Gene Is Associated With Tourette Syndrome in a Family Study

PubMed Central

Huertas-Fernández, Ismael; Gómez-Garre, Pilar; Madruga-Garrido, Marcos; Bernal-Bernal, Inmaculada; Bonilla-Toribio, Marta; Martín-Rodríguez, Juan Francisco; Cáceres-Redondo, María Teresa; Vargas-González, Laura; Carrillo, Fátima; Pascual, Alberto; Tischfield, Jay A.; King, Robert A.; Heiman, Gary A.; Mir, Pablo

2016-01-01

Background Tourette syndrome is a disorder characterized by persistent motor and vocal tics, and frequently accompanied by the comorbidities attention deficit hyperactivity disorder and obsessive-compulsive disorder. Impaired synaptic neurotransmission has been implicated in its pathogenesis. Our aim was to investigate the association of 28 candidate genes, including genes related to synaptic neurotransmission and neurotrophic factors, with Tourette syndrome. Methods We genotyped 506 polymorphisms in a discovery cohort from the United States composed of 112 families and 47 unrelated singletons with Tourette syndrome (201 cases and 253 controls). Genes containing significant polymorphisms were imputed to fine-map the signal(s) to potential causal variants. Allelic analyses in Tourette syndrome cases were performed to check the role in attention deficit hyperactivity disorder and obsessive-compulsive disorder comorbidities. Target polymorphisms were further studied in a replication cohort from southern Spain composed of 37 families and three unrelated singletons (44 cases and 73 controls). Results The polymorphism rs3096140 in glial cell line–derived neurotrophic factor gene (GDNF) was significant in the discovery cohort after correction (P = 1.5 × 10−4). No linkage disequilibrium was found between rs3096140 and other functional variants in the gene. We selected rs3096140 as target polymorphism, and the association was confirmed in the replication cohort (P = 0.01). No association with any comorbidity was found. Conclusions As a conclusion, a common genetic variant in GDNF is associated with Tourette syndrome. A defect in the production of GDNF could compromise the survival of parvalbumin interneurons, thus altering the excitatory/inhibitory balance in the corticostriatal circuitry. Validation of this variant in other family cohorts is necessary. PMID:26096985

GDNF gene is associated with tourette syndrome in a family study.

PubMed

Huertas-Fernández, Ismael; Gómez-Garre, Pilar; Madruga-Garrido, Marcos; Bernal-Bernal, Inmaculada; Bonilla-Toribio, Marta; Martín-Rodríguez, Juan Francisco; Cáceres-Redondo, María Teresa; Vargas-González, Laura; Carrillo, Fátima; Pascual, Alberto; Tischfield, Jay A; King, Robert A; Heiman, Gary A; Mir, Pablo

2015-07-01

Tourette syndrome is a disorder characterized by persistent motor and vocal tics, and frequently accompanied by the comorbidities attention deficit hyperactivity disorder and obsessive-compulsive disorder. Impaired synaptic neurotransmission has been implicated in its pathogenesis. Our aim was to investigate the association of 28 candidate genes, including genes related to synaptic neurotransmission and neurotrophic factors, with Tourette syndrome. We genotyped 506 polymorphisms in a discovery cohort from the United States composed of 112 families and 47 unrelated singletons with Tourette syndrome (201 cases and 253 controls). Genes containing significant polymorphisms were imputed to fine-map the signal(s) to potential causal variants. Allelic analyses in Tourette syndrome cases were performed to check the role in attention deficit hyperactivity disorder and obsessive-compulsive disorder comorbidities. Target polymorphisms were further studied in a replication cohort from southern Spain composed of 37 families and three unrelated singletons (44 cases and 73 controls). The polymorphism rs3096140 in glial cell line-derived neurotrophic factor gene (GDNF) was significant in the discovery cohort after correction (P = 1.5 × 10(-4) ). No linkage disequilibrium was found between rs3096140 and other functional variants in the gene. We selected rs3096140 as target polymorphism, and the association was confirmed in the replication cohort (P = 0.01). No association with any comorbidity was found. As a conclusion, a common genetic variant in GDNF is associated with Tourette syndrome. A defect in the production of GDNF could compromise the survival of parvalbumin interneurons, thus altering the excitatory/inhibitory balance in the corticostriatal circuitry. Validation of this variant in other family cohorts is necessary. © 2015 International Parkinson and Movement Disorder Society.

The Neuroprotective Effects of Flaxseed Oil Supplementation on Functional Motor Recovery in a Model of Ischemic Brain Stroke: Upregulation of BDNF and GDNF.

PubMed

Bagheri, Abolqasem; Talei, Sahand; Hassanzadeh, Negar; Mokhtari, Tahmineh; Akbari, Mohammad; Malek, Fatemeh; Jameie, Seyed Behnamedin; Sadeghi, Yousef; Hassanzadeh, Gholamreza

2017-12-01

Cerebral ischemic stroke is a common leading cause of disability. Flaxseed is a richest plant-based source of antioxidants. In this study, the effects of flaxseed oil (FSO) pretreatment on functional motor recovery and gene expression and protein content of neurotrophic factors in motor cortex area in rat model of brain ischemia/reperfusion (I/R) were assessed. Transient middle cerebral artery occlusion (tMCAo) in rats was used as model brain I/R. Rats (6 in each group) were randomly divided into four groups of Control (Co+normal saline [NS]), Sham (Sh+NS), tMCAo+NS and tMCAo+FSO. After three weeks of pretreatment with vehicle or FSO (0.2 ml~800 mg/kg body weight), the rats were operated in sham and ischemic groups. Ischemia was induced for 1 h and then reperfused. After 24 h of reperfusion, neurological examination was performed, and animals were sacrificed, and their brains were used for molecular and histopathological studies. FSO significantly improved the functional motor recovery compared with tMCAo+NS group (P<0.05). A significant reduction in brain-derived neurotrophic factor (BDNF) and glial cell-derived neurotrophic factor (GDNF) mRNAs and protein levels were observed in the tMCAo+NS group compared with Co+NS and Sh+NS group (P<0.05). A significant increase of BDNF and GDNF mRNAs and proteins was recorded in the tMCAo+FSO group compared with Co+NS, Sh+NS and tMCAO+NS groups (P<0.05). The results of the current study demonstrated that pretreatment with FSO had neuroprotective effects on motor cortex area following cerebral ischemic stroke by increasing the neurotrophic factors (BDNF, GDNF).

Ultrasound-triggered effects of the microbubbles coupled to GDNF- and Nurr1-loaded PEGylated liposomes in a rat model of Parkinson's disease.

PubMed

Yue, Peijian; Gao, Lin; Wang, Xuejing; Ding, Xuebing; Teng, Junfang

2018-06-01

The purpose of this study was to investigate ultrasound-triggered effects of the glial cell line-derived neurotrophic factor (GDNF) + nuclear receptor-related factor 1 (Nurr1)-polyethylene glycol (PEG)ylated liposomes-coupled microbubbles (PLs-GDNF + Nurr1-MBs) on behavioral impairment and neuron loss in a rat model of Parkinson's disease (PD). The unloaded PEGylated liposomes-coupled microbubbles (PLs-MBs) were characterized for zeta potential, particle size, and concentration. 6-hydroxydopamine (6-OHDA) was used to establish the PD rat model. Rotational, climbing pole, and suspension tests were used to detect behavioral impairment. The immunohistochemical staining of tyrosine hydroxylase (TH) and dopamine transporter (DAT) was used to assess the neuron loss. Western blot and quantitative real-time PCR (qRT-PCR) analysis were used to measure the expression levels of GDNF and Nurr1. The particle size of PLs-MBs was gradually increased, while the concentration and absolute zeta potential were gradually decreased as the time prolongs. 6-OHDA increased amphetamine-induced rotations and loss of dopaminergic neurons as compared to sham group. Interestingly, PLs-GDNF-MBs or PLs-Nurr1-MBs decreased rotations and increased the TH and DAT immunoreactivity. Combined of both genes resulted in a robust reduction in the rotations and a greater increase of the dopaminergic neurons. The delivery of PLs-GDNF + Nurr1-MBs into the brains using magnetic resonance imaging (MRI)-guided focused ultrasound may be more efficacious for the treatment of PD than the single treatment. © 2017 Wiley Periodicals, Inc.

Transplantation of bone marrow mononuclear cells modulates hippocampal expression of growth factors in chronically epileptic animals.

PubMed

Zanirati, Gabriele; Azevedo, Pamella Nunes; Marinowic, Daniel Rodrigo; Rodrigues, Felipe; de Oliveira Dias, Ana Christina; Venturin, Gianina Teribele; Greggio, Samuel; Simão, Fabrício; DaCosta, Jaderson Costa

2015-05-01

In previous studies, transplantation of bone marrow mononuclear cells (BMMCs) in epileptic animals has been found to be neuroprotective. However, the mechanism by which the BMMCs act remains unclear. We hypothesize that BMMCs may provide neuroprotection to the epileptic brain through trophic support. To test our hypothesis, we studied the temporal expression of neurotrophins after BMMC transplantation in the epileptic rat hippocampus. Chronically epileptic rats were intravenously transplanted with 1 × 10(7) BMMCs isolated from GFP transgenic mice. Expression levels of BDNF, GDNF, NGF, VEGF, and TGF-β1, and their receptors, were evaluated by ELISA and/or qRT-PCR analysis. Our data revealed increased protein expression of BDNF, GDNF, NGF, and VEGF and reduced levels of TGF-β1 in the hippocampus of transplanted epileptic animals. Additionally, an increase in the mRNA expression of BDNF, GDNF, and VEGF, a reduction in TGF-β1, and a decrease in mRNA levels of the TrkA and TGFR-β1 receptors were also observed. The gain provided by transplanted BMMCs in the epileptic brain may be related to the ability of these cells in modulating the network of neurotrophins and angiogenic signals. © 2015 John Wiley & Sons Ltd.

Effect of glial cell line-derived neurotrophic factor on behavior and key members of the brain serotonin system in mouse strains genetically predisposed to behavioral disorders.

PubMed

Naumenko, Vladimir S; Bazovkina, Daria V; Semenova, Alina A; Tsybko, Anton S; Il'chibaeva, Tatyana V; Kondaurova, Elena M; Popova, Nina K

2013-12-01

The effect of glial cell line-derived neurotrophic factor (GDNF) on behavior and on the serotonin (5-HT) system of a mouse strain predisposed to depressive-like behavior, ASC/Icg (Antidepressant Sensitive Cataleptics), in comparison with the parental "nondepressive" CBA/Lac mice was studied. Within 7 days after acute administration, GDNF (800 ng, i.c.v.) decreased cataleptic immobility but increased depressive-like behavioral traits in both investigated mouse strains and produced anxiolytic effects in ASC mice. The expression of the gene encoding the key enzyme for 5-HT biosynthesis in the brain, tryptophan hydroxylase-2 (Tph-2), and 5-HT1A receptor gene in the midbrain as well as 5-HT2A receptor gene in the frontal cortex were increased in GDNF-treated ASC mice. At the same time, GDNF decreased 5-HT1A and 5-HT2A receptor gene expression in the hippocampus of ASC mice. GDNF failed to change Tph2, 5-HT1A , or 5-HT2A receptor mRNA levels in CBA mice as well as 5-HT transporter gene expression and 5-HT1A and 5-HT2A receptor functional activity in both investigated mouse strains. The results show 1) a GDNF-induced increase in the expression of key genes of the brain 5-HT system, Tph2, 5-HT1A , and 5-HT2A receptors, and 2) significant genotype-dependent differences in the 5-HT system response to GDNF treatment. The data suggest that genetically defined cross-talk between neurotrophic factors and the brain 5-HT system underlies the variability in behavioral response to GDNF. Copyright © 2013 Wiley Periodicals, Inc.

Precursor N-cadherin mediates glial cell line-derived neurotrophic factor-promoted human malignant glioma

PubMed Central

Zhu, Shuang; Zhang, Baole; Qin, Yuxia; Yao, Ruiqin; Zhou, Hao; Gao, Dian Shuai

2017-01-01

As the most prevalent primary brain tumor, gliomas are highly metastatic, invasive and are characteristic of high levels of glial cell-line derived neurotrophic factor (GDNF). GDNF is an important factor for invasive glioma cell growth; however, the underlying mechanism involved is unclear. In this study, we affirm a significantly higher expression of the precursor of N-cadherin (proN-cadherin) in most gliomas compared with normal brain tissues. Our findings reveal that GDNF interacts with the extracellular domain of proN-cadherin, which suggests that proN-cadherin mediates GDNF-induced glioma cell migration and invasion. We hypothesize that proN-cadherin might cause homotypic adhesion loss within neighboring cells and at the same time promote heterotypic adhesion within the extracellular matrix (ECM) through a certain mechanism. This study also demonstrates that the interaction between GDNF and proN-cadherin activates specific intracellular signaling pathways; furthermore, GDNF promoted the secretion of matrix metalloproteinase-9 (MMP-9), which degrades the ECM via proN-cadherin. To reach the future goal of developing novel therapies of glioma, this study, reveals a unique mechanism of glioma cell migration and invasion. PMID:28212546

Precursor N-cadherin mediates glial cell line-derived neurotrophic factor-promoted human malignant glioma.

PubMed

Xiong, Ye; Liu, Liyun; Zhu, Shuang; Zhang, Baole; Qin, Yuxia; Yao, Ruiqin; Zhou, Hao; Gao, Dian Shuai

2017-04-11

As the most prevalent primary brain tumor, gliomas are highly metastatic, invasive and are characteristic of high levels of glial cell-line derived neurotrophic factor (GDNF). GDNF is an important factor for invasive glioma cell growth; however, the underlying mechanism involved is unclear. In this study, we affirm a significantly higher expression of the precursor of N-cadherin (proN-cadherin) in most gliomas compared with normal brain tissues. Our findings reveal that GDNF interacts with the extracellular domain of proN-cadherin, which suggests that proN-cadherin mediates GDNF-induced glioma cell migration and invasion. We hypothesize that proN-cadherin might cause homotypic adhesion loss within neighboring cells and at the same time promote heterotypic adhesion within the extracellular matrix (ECM) through a certain mechanism. This study also demonstrates that the interaction between GDNF and proN-cadherin activates specific intracellular signaling pathways; furthermore, GDNF promoted the secretion of matrix metalloproteinase-9 (MMP-9), which degrades the ECM via proN-cadherin. To reach the future goal of developing novel therapies of glioma, this study, reveals a unique mechanism of glioma cell migration and invasion.

TITER AND PRODUCT AFFECTS THE DISTRIBUTION OF GENE EXPRESSION AFTER INTRAPUTAMINAL CONVECTION-ENHANCED DELIVERY

PubMed Central

Emborg, Marina E.; Hurley, Samuel A.; Joers, Valerie; Tromp, Do P.M.; Swanson, Christine R.; Ohshima-Hosoyama, Sachiko; Bondarenko, Viktorya; Cummisford, Kyle; Sonnemans, Marc; Hermening, Stephan; Blits, Bas; Alexander, Andrew L.

2014-01-01

Background Efficacy and safety of intracerebral gene therapy for brain disorders, like Parkinson’s disease, depends on appropriate distribution of gene expression. Objectives To assess if the distribution of gene expression is affected by vector titer and protein type. Methods Four adult macaque monkeys seronegative for adeno-associated virus 5 (AAV5) received in the right and left ventral postcommisural putamen 30μl inoculation of a high or low titer suspension of AAV5 encoding glial derived neurotrophic factor (GDNF) or green fluorescent protein (GFP). Inoculations were performed using convection enhanced delivery and intraoperative MRI (IMRI). Results IMRI confirmed targeting and infusion cloud irradiating from the catheter tip into surrounding area. Postmortem analysis six weeks after surgery revealed GFP and GDNF expression ipsilateral to the injection side that had a titer-dependent distribution. GFP and GDNF expression was also observed in fibers in the Substantia Nigra (SN) pars reticulata (pr), demonstrating anterograde transport. Few GFP-positive neurons were present in the SN pars compacta (pc), possibly by direct retrograde transport of the vector. GDNF was present in many SNpc and SNpr neurons. Conclusions After controlling for target and infusate volume, intracerebral distribution of gene product is affected by vector titer and product biology. PMID:24943657

Regulation of GluR2 promoter activity by neurotrophic factors via a neuron-restrictive silencer element.

PubMed

Brené, S; Messer, C; Okado, H; Hartley, M; Heinemann, S F; Nestler, E J

2000-05-01

The AMPA glutamate receptor subunit GluR2, which plays a critical role in regulation of AMPA channel function, shows altered levels of expression in vivo after several chronic perturbations. To evaluate the possibility that transcriptional mechanisms are involved, we studied a 1254-nucleotide fragment of the 5'-promoter region of the mouse GluR2 gene in neural-derived cell lines. We focused on regulation of GluR2 promoter activity by two neurotrophic factors, which are known to be altered in vivo in some of the same systems that show GluR2 regulation. Glial-cell line derived neurotrophic factor (GDNF) and brain-derived neurotrophic factor (BDNF) both induced GluR2 promoter activity. This was associated with increased expression of endogenous GluR2 immunoreactivity in the cells as measured by Western blotting. The effect of GDNF and BDNF appeared to be mediated via a NRSE (neuron-restrictive silencer element) present within the GluR2 promoter. The response to these neurotrophic factors was lost upon mutating or deleting this site, but not several other putative response elements present within the promoter. Moreover, overexpression of REST (restrictive element silencer transcription factor; also referred to as NRSF or neuron restrictive silencer factor), which is known to act on NRSEs in other genes to repress gene expression, blocked the ability of GDNF to induce GluR2 promoter activity. However, GDNF did not alter endogenous levels of REST in the cells. Together, these findings suggest that GluR2 expression can be regulated by neurotrophic factors via an apparently novel mechanism involving the NRSE present within the GluR2 gene promoter.

Direct muscle delivery of GDNF with human mesenchymal stem cells improves motor neuron survival and function in a rat model of familial ALS

PubMed Central

Suzuki, Masatoshi; McHugh, Jacalyn; Tork, Craig; Shelley, Brandon; Hayes, Antonio; Bellantuono, Ilaria; Aebischer, Patrick; Svendsen, Clive N.

2008-01-01

Amyotrophic Lateral Sclerosis (ALS) is a fatal neurodegenerative disease in which there is a progressive loss of motor neurons and their connections to muscle leading to paralysis. To maintain muscle connections in a rat model of familial ALS, we performed intramuscular transplantation with human mesenchymal stem cells (hMSC) as “Trojan horses” to deliver growth factors to the terminals of motor neurons as well as the skeletal muscles. hMSC engineered to secrete glial cell line derived neurotrophic factor (hMSC-GDNF) were transplanted bilaterally into three muscle groups. The cells survived within the muscle, released GDNF, and significantly increased the number of neuromuscular connections and motor neuron cell bodies in the spinal cord at mid stages of the disease. Furthermore, intramuscular transplantation with hMSC-GDNF could ameliorate motor neuron loss within the spinal cord which connected to the limb muscles with transplants. While disease onset was similar in all animals, hMSC-GDNF significantly delayed disease progression, increasing overall lifespan by up to 28 days, which is one of the longest effects on survival noted for this rat model of familial ALS. This pre-clinical data provides a novel and practical approach towards ex vivo gene therapy for ALS. PMID:18797452

Direct muscle delivery of GDNF with human mesenchymal stem cells improves motor neuron survival and function in a rat model of familial ALS.

PubMed

Suzuki, Masatoshi; McHugh, Jacalyn; Tork, Craig; Shelley, Brandon; Hayes, Antonio; Bellantuono, Ilaria; Aebischer, Patrick; Svendsen, Clive N

2008-12-01

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease in which there is a progressive loss of motor neurons and their connections to muscle, leading to paralysis. In order to maintain muscle connections in a rat model of familial ALS (FALS), we performed intramuscular transplantation with human mesenchymal stem cells (hMSCs) used as "Trojan horses" to deliver growth factors to the terminals of motor neurons and to the skeletal muscles. hMSCs engineered to secrete glial cell line-derived neurotrophic factor (hMSC-GDNF) were transplanted bilaterally into three muscle groups. The cells survived within the muscle, released GDNF, and significantly increased the number of neuromuscular connections and motor neuron cell bodies in the spinal cord at mid-stages of the disease. Further, intramuscular transplantation with hMSC-GDNF was found to ameliorate motor neuron loss within the spinal cord where it connects with the limb muscles receiving transplants. While disease onset was similar in all the animals, hMSC-GDNF significantly delayed disease progression, increasing overall lifespan by up to 28 days, which is one of the largest effects on survival noted for this rat model of FALS. This preclinical data provides a novel and practical approach toward ex vivo gene therapy for ALS.

Brain stem slice conditioned medium contains endogenous BDNF and GDNF that affect neural crest boundary cap cells in co-culture.

PubMed

Kaiser, Andreas; Kale, Ajay; Novozhilova, Ekaterina; Siratirakun, Piyaporn; Aquino, Jorge B; Thonabulsombat, Charoensri; Ernfors, Patrik; Olivius, Petri

2014-05-30

Conditioned medium (CM), made by collecting medium after a few days in cell culture and then re-using it to further stimulate other cells, is a known experimental concept since the 1950s. Our group has explored this technique to stimulate the performance of cells in culture in general, and to evaluate stem- and progenitor cell aptitude for auditory nerve repair enhancement in particular. As compared to other mediums, all primary endpoints in our published experimental settings have weighed in favor of conditioned culture medium, where we have shown that conditioned culture medium has a stimulatory effect on cell survival. In order to explore the reasons for this improved survival we set out to analyze the conditioned culture medium. We utilized ELISA kits to investigate whether brain stem (BS) slice CM contains any significant amounts of brain-derived neurotrophic factor (BDNF) and glial cell derived neurotrophic factor (GDNF). We further looked for a donor cell with progenitor characteristics that would be receptive to BDNF and GDNF. We chose the well-documented boundary cap (BC) progenitor cells to be tested in our in vitro co-culture setting together with cochlear nucleus (CN) of the BS. The results show that BS CM contains BDNF and GDNF and that survival of BC cells, as well as BC cell differentiation into neurons, were enhanced when BS CM were used. Altogether, we conclude that BC cells transplanted into a BDNF and GDNF rich environment could be suitable for treatment of a traumatized or degenerated auditory nerve. Copyright © 2014 Elsevier B.V. All rights reserved.

Neuroprotective effects of electroacupuncture on hypoxic-ischemic encephalopathy in newborn rats are associated with increased expression of GDNF-RET and protein kinase B.

PubMed

Xu, Tao; Xu, Neng-Gui; Yang, Zhong-Hua; Wan, Yan-Zhen; Wu, Qing-Long; Huang, Kang-Bai

2016-06-01

To explore the neuroprotective effects of electroacupuncture (EA) on hypoxic-ischemic encephalopathy (HIE) and to further investigate the role of glial cell line-derived neurotrophic factor (GDNF) family receptor member RET (rearranged during transfection) and its key downstream phosphatidylinositol 3 kinase (PI-3K)/protein kinase B (Akt) pathway in the process. A total of 220 seven-day-old SD rats (of either sex, from 22 broods) were randomly divided into two groups, one (30 rats) for sham-surgery group and the other (190 rats) for HIE model group. The HIE model was established using the left common carotid artery ligation method in combination with hypoxic treatment. The successfully established rats were randomly divided into five groups, including control model group, EA group, sham-EA group, antagonist group and antagonist plus electroacupuncture group, with 35 rats in each group. Baihui (GV 20), Dazhui (GV 14), Quchi (LI 11) and Yongquan (KI 1) acupoints were chosen for acupuncture. EA was performed at Baihui and Quchi for 10 min once a day for continuous 1, 3, 7 and 21 days, respectively. The rats were then killed after the operation and injured cerebral cortex was taken for the measurement of neurologic damage by hematoxylin-eosin (HE) staining and the degenerative changes of cortical ultrastructure by transmission electron microscopy. RET mRNA level and Akt protein level were detected by real-time reverse-transcription polymerase chain reaction (RT-PCR) and western blot analysis, respectively. EA could ameliorate neurologic damage of the first somatic sensory area (S1Tr) and alleviate the degenerative changes of ultrastructure of cortical neurons in rats subjected to HIE. And the longer acupuncture treatment lasted, the better its therapeutic effect would be. This was accompanied by gradually increased expression of GDNF family receptor RET at the mRNA level and its downstream signaling Akt at the protein level in the ischemic cortex. EA has

Glial cell line-derived neurotrophic factor-dependent RET activation can be mediated by two different cell-surface accessory proteins

PubMed Central

Sanicola, M.; Hession, C.; Worley, D.; Carmillo, P.; Ehrenfels, C.; Walus, L.; Robinson, S.; Jaworski, G.; Wei, H.; Tizard, R.; Whitty, A.; Pepinsky, R. B.; Cate, R. L.

1997-01-01

Glial cell line-derived neurotrophic factor (GDNF)-dependent activation of the tyrosine kinase receptor RET is necessary for kidney and enteric neuron development, and mutations in RET are associated with human diseases. Activation of RET by GDNF has been shown to require an accessory component, GDNFR-α (RETL1). We report the isolation and characterization of rat and human cDNAs for a novel cell-surface associated accessory protein, RETL2, that shares 49% identity with RETL1. Both RETL1 and RETL2 can mediate GDNF dependent phosphorylation of RET, but they exhibit different patterns of expression in fetal and adult tissues. The most striking differences in expression observed were in the adult central and peripheral nervous systems. In addition, the mechanisms by which the two accessory proteins facilitate the activation of RET by GDNF are quite distinct. In vitro binding experiments with soluble forms of RET, RETL1 and RETL2 demonstrate that while RETL1 binds GDNF tightly to form a membrane-associated complex which can then interact with RET, RETL2 only forms a high affinity complex with GDNF in the presence of RET. This strong RET dependence of the binding of RETL2 to GDNF was confirmed by FACS analysis on RETL1 and RETL2 expressing cells. Together with the recent discovery of a GDNF related protein, neurturin, these data raise the possibility that RETL1 and RETL2 have distinctive roles during development and in the nervous system of the adult. RETL1 and RETL2 represent new candidate susceptibility genes and/or modifier loci for RET-associated diseases. PMID:9177201

Functional regeneration of the transected recurrent laryngeal nerve using a collagen scaffold loaded with laminin and laminin-binding BDNF and GDNF

PubMed Central

Wang, Baoxin; Yuan, Junjie; Chen, Xinwei; Xu, Jiafeng; Li, Yu; Dong, Pin

2016-01-01

Recurrent laryngeal nerve (RLN) injury remains a challenge due to the lack of effective treatments. In this study, we established a new drug delivery system consisting of a tube of Heal-All Oral Cavity Repair Membrane loaded with laminin and neurotrophic factors and tested its ability to promote functional recovery following RLN injury. We created recombinant fusion proteins consisting of brain-derived neurotrophic factor (BDNF) and glial cell line-derived neurotrophic factor (GDNF) fused to laminin-binding domains (LBDs) in order to prevent neurotrophin diffusion. LBD-BDNF, LBD-GDNF, and laminin were injected into a collagen tube that was fitted to the ends of the transected RLN in rats. Functional recovery was assessed 4, 8, and 12 weeks after injury. Although vocal fold movement was not restored until 12 weeks after injury, animals treated with the collagen tube loaded with laminin, LBD-BDNF and LBD-GDNF showed improved recovery in vocalisation, arytenoid cartilage angles, compound muscle action potentials and regenerated fibre area compared to animals treated by autologous nerve grafting (p < 0.05). These results demonstrate the drug delivery system induced nerve regeneration following RLN transection that was superior to that induced by autologus nerve grafting. It may have potential applications in nerve regeneration of RLN transection injury. PMID:27558932

The production of glial cell line-derived neurotrophic factor by human sertoli cells is substantially reduced in sertoli cell-only testes.

PubMed

Singh, D; Paduch, D A; Schlegel, P N; Orwig, K E; Mielnik, A; Bolyakov, A; Wright, W W

2017-05-01

Do human Sertoli cells in testes that exhibit the Sertoli cell-only (SCO) phenotype produce substantially less glial cell line-derived neurotrophic factor (GDNF) than Sertoli cells in normal testes? In human SCO testes, both the amounts of GDNF mRNA per testis and the concentration of GDNF protein per Sertoli cell are markedly reduced as compared to normal testes. In vivo, GDNF is required to sustain the numbers and function of mouse spermatogonial stem cells (SSCs) and their immediate progeny, transit-amplifying progenitor spermatogonia. GDNF is expressed in the human testis, and the ligand-binding domain of the GDNF receptor, GFRA1, has been detected on human SSCs. The numbers and/or function of these stem cells are markedly reduced in some infertile men, resulting in the SCO histological phenotype. We determined the numbers of human spermatogonia per mm2 of seminiferous tubule surface that express GFRA1 and/or UCHL1, another marker of human SSCs. We measured GFRA1 mRNA expression in order to document the reduced numbers and/or function of SSCs in SCO testes. We quantified GDNF mRNA in testes of humans and mice, a species with GDNF-dependent SSCs. We also compared GDNF mRNA expression in human testes with normal spermatogenesis to that in testes exhibiting the SCO phenotype. As controls, we also measured transcripts encoding two other Sertoli cell products, kit ligand (KITL) and clusterin (CLU). Finally, we compared the amounts of GDNF per Sertoli cell in normal and SCO testes. Normal human testes were obtained from beating heart organ donors. Biopsies of testes from men who were infertile due to maturation arrest or the SCO phenotype were obtained as part of standard care during micro-testicular surgical sperm extraction. Cells expressing GFRA1, UCHL1 or both on whole mounts of normal human seminiferous tubules were identified by immunohistochemistry and confocal microscopy and their numbers were determined by image analysis. Human GDNF mRNA and GFRA1 mRNA were

Muscle-Derived GDNF: A Gene Therapeutic Approach for Preserving Motor Neuron Function in ALS

DTIC Science & Technology

2015-08-01

using AAV at UW - Madison . Although we prepared our animal colony for diaphragm injections of AAV-GDNF, we decided to postpone the experiments based on...data obtained by Dr. Masatoshi Suzuki (University of Wisconsin- Madison ) with AAV2/6-GDNF. As methods used in the current study were identical to the...study animals (figure 4C), no overt relationship was observed. Figure 4- Motor neuron quantification in the lumbar spinal cord of animals treated

Prevention of Memory Impairment and Neurotrophic Factors Increased by Lithium in Wistar Rats Submitted to Pneumococcal Meningitis Model

PubMed Central

Simões, Lutiana R.; Abreu, Roberta R. E. S.; Goularte, Jéssica A.; Collodel, Allan; Giridharan, Vijayasree Vayalanellore

2017-01-01

The aim of this study was to investigate the effects of lithium on brain-derived neurotrophic factor (BDNF), nerve growth factor (NGF), and glial cell line-derived neurotrophic factor (GDNF) expression in the hippocampus and on memory in experimental pneumococcal meningitis. The mood-stabilizer lithium is known as a neuroprotective agent with many effects on the brain. In this study, animals received either artificial cerebrospinal fluid or Streptococcus pneumoniae suspension at a concentration of 5 × 109 CFU/mL. Eighteen hours after induction, all animals received ceftriaxone. The animals received saline or lithium (47.5 mg/kg) or tamoxifen (1 mg/kg) as adjuvant treatment, and they were separated into six groups: control/saline, control/lithium, control/tamoxifen, meningitis/saline, meningitis/lithium, and meningitis/tamoxifen. Ten days after meningitis induction, animals were subjected to open-field habituation and the step-down inhibitory avoidance tasks. Immediately after these tasks, the animals were killed and their hippocampus was removed to evaluate the expression of BDNF, NGF, and GDNF. In the meningitis group, treatment with lithium and tamoxifen resulted in improvement in memory. Meningitis group showed decreased expression of BDNF and GDNF in the hippocampus while lithium reestablished the neurotrophin expression. Lithium was able to prevent memory impairment and reestablishes hippocampal neurotrophin expression in experimental pneumococcal meningitis. PMID:29200666

GDNF and NGF family members and receptors in human fetal and adult spinal cord and dorsal root ganglia.

PubMed

Josephson, A; Widenfalk, J; Trifunovski, A; Widmer, H R; Olson, L; Spenger, C

2001-11-12

We describe the expression of mRNA encoding ligands and receptors of members of the GDNF family and members of the neurotrophin family in the adult human spinal cord and dorsal root ganglia (DRG). Fetal human spinal cord and ganglia were investigated for the presence of ligands and receptors of the neurotrophin family. Tissues were collected from human organ donors and after routine elective abortions. Messenger RNA was found encoding RET, GFR alpha-1, BDNF, trkB, and trkC in the adult human spinal cord and BDNF, NT-3, p75, trkB, and trkC in the fetal human spinal cord. The percentage of adult human DRG cells expressing p75, trkA, trkB, or trkC was 57, 46, 29, and 24%, respectively, and that of DRG cells expressing RET, GFR alpha-1, GFR alpha-2, or GFR alpha-3 was 79, 20, 51, and 32%, respectively. GFR alpha-2 was expressed selectively in small, GFR alpha-3 principally in small and GFR alpha-1 and RET in both large and small adult human DRG neurons. p75 and trkB were expressed by a wide range of DRG neurons while trkA was expressed in most small diameter and trkC primarily in large DRG neurons. Fetal DRG cells were positive for the same probes as adult DRG cells except for NT-3, which was only found in fetal DRG cells. Messenger RNA species only expressed at detectable levels in fetal but not adult spinal cord tissues included GDNF, GFR alpha-2, NT-3, and p75. Notably, GFR alpha-2, which is expressed in the adult rat spinal cord, was not found in the adult human spinal cord. Copyright 2001 Wiley-Liss, Inc.

Glial-derived neurotrophic factor is essential for blood-nerve barrier functional recovery in an experimental murine model of traumatic peripheral neuropathy.

PubMed

Dong, Chaoling; Helton, E Scott; Zhou, Ping; Ouyang, Xuan; d'Anglemont de Tassigny, Xavier; Pascual, Alberto; López-Barneo, José; Ubogu, Eroboghene E

2018-06-18

There is emerging evidence that glial-derived neurotrophic factor (GDNF) is a potent inducer of restrictive barrier function in tight junction-forming microvascular endothelium and epithelium, including the human blood-nerve barrier (BNB) in vitro. We sought to determine the role of GDNF in restoring BNB function in vivo by evaluating sciatic nerve horseradish peroxidase (HRP) permeability in tamoxifen-inducible GDNF conditional knockout (CKO) adult mice following non-transecting crush injury via electron microscopy, with appropriate wildtype (WT) and heterozygous (HET) littermate controls. A total of 24 age-, genotype- and sex-matched mice >12 weeks of age were injected with 30 mg/kg HRP via tail vein injection 7 or 14 days following unilateral sciatic nerve crush, and both sciatic nerves were harvested 30 minutes later for morphometric assessment by light and electron microscopy. The number and percentage of HRP-permeable endoneurial microvessels were ascertained to determine the effect of GDNF in restoring barrier function in vivo. Following sciatic nerve crush, there was significant upregulation in GDNF protein expression in WT and HET mice that was abrogated in CKO mice. GDNF significantly restored sciatic nerve BNB HRP impermeability to near normal levels by day 7, with complete restoration seen by day 14 in WT and HET mice. A significant recovery lag was observed in CKO mice. This effect was independent on VE-Cadherin or claudin-5 expression on endoneurial microvessels. These results imply an important role of GDNF in restoring restrictive BNB function in vivo, suggesting a potential strategy to re-establish the restrictive endoneurial microenvironment following traumatic peripheral neuropathies.

High-frequency stimulation of the globus pallidus interna nucleus modulates GFRα1 gene expression in the basal ganglia.

PubMed

Ho, Duncun Xun Kiat; Tan, Yong Chee; Tan, Jiayi; Too, Heng Phon; Ng, Wai Hoe

2014-04-01

Deep brain stimulation (DBS) is an established therapy for movement disorders such as Parkinson's disease (PD). Although the efficacy of DBS is clear, its precise molecular mechanism remains unknown. The glial cell line derived factor (GDNF) family of ligands has been shown to confer neuroprotective effects on dopaminergic neurons, and putaminal infusion of GDNF have been investigated in PD patients with promising results. Despite the potential therapeutic role of GDNF in alleviating motor symptoms, there is no data on the effects of electrical stimulation on GDNF-family receptor (GFR) expression in the basal ganglia structures. Here, we report the effects of electrical stimulation on GFRα1 isoforms, particularly GFRα1a and GFRα1b. Wistar rats underwent 2 hours of high frequency stimulation (HFS) at the globus pallidus interna nucleus. A control group was subjected to a similar procedure but without stimulation. The HFS group, sacrificed 24 hours after treatment, had a threefold decrease in mRNA expression level of GFRα1b (p=0.037), but the expression level reverted to normal 72 hours after stimulation. Our preliminary data reveal the acute effects of HFS on splice isoforms of GFRα1, and suggest that HFS may modulate the splice isoforms of GFRα1a and GFRα1b to varying degrees. Going forward, elucidating the interactions between HFS and GFR may shed new insights into the complexity of GDNF signaling in the nervous system and lead to better design of clinical trials using these signaling pathways to halt disease progression in PD and other neurodegenerative diseases. Copyright © 2013 Elsevier Ltd. All rights reserved.

Retinal ganglion cells survival in a glaucoma model by GDNF/Vit E PLGA microspheres prepared according to a novel microencapsulation procedure.

PubMed

Checa-Casalengua, Patricia; Jiang, Caihui; Bravo-Osuna, Irene; Tucker, Budd A; Molina-Martínez, Irene T; Young, Michael J; Herrero-Vanrell, Rocío

2011-11-30

The present experimental work describes the use of a novel protein encapsulation method to achieve protection of the biological factor during the microencapsulation procedure. With this aim, the protein is included in poly(lactic-co-glycolic acid) (PLGA) microspheres without any preliminary manipulation, in contrast to the traditional S/O/W (solid-in-oil-in-water) method where the bioactive substance is first dissolved and then freeze-dried in the presence of lyoprotectors. Furthermore, the presented technique involves the use of an oily additive, vitamin E (Vit E), useful from a technological point of view, by promoting additional protein protection and also from a pharmacological point of view, because of its antioxidant and antiproliferative properties. Application of this microencapsulation technique has been performed for GDNF (glial cell line-derived neurotrophic factor) designed for the treatment of optic nerve degenerative diseases, such as glaucoma, the second leading cause of blindness in the western world. The protein was released in vitro in its bioactive form for more than three months, demonstrated by the survival of their potential target cells (photoreceptors and retinal ganglion cells (RGC)). Moreover, the intravitreal injection of GDNF/Vit E PLGA microspheres in an experimental animal model of glaucoma significantly increased RGC survival compared with GDNF, Vit E or blank microspheres (p<0.01). This effect was present for at least eleven weeks, which suggests that the formulation prepared may be clinically useful as a neuroprotective tool in the treatment of glaucomatous optic neuropathy. Copyright © 2011 Elsevier B.V. All rights reserved.

Neurotrophic factors and receptors in the immature and adult spinal cord after mechanical injury or kainic acid.

PubMed

Widenfalk, J; Lundströmer, K; Jubran, M; Brene, S; Olson, L

2001-05-15

Delivery of neurotrophic factors to the injured spinal cord has been shown to stimulate neuronal survival and regeneration. This indicates that a lack of sufficient trophic support is one factor contributing to the absence of spontaneous regeneration in the mammalian spinal cord. Regulation of the expression of neurotrophic factors and receptors after spinal cord injury has not been studied in detail. We investigated levels of mRNA-encoding neurotrophins, glial cell line-derived neurotrophic factor (GDNF) family members and related receptors, ciliary neurotrophic factor (CNTF), and c-fos in normal and injured spinal cord. Injuries in adult rats included weight-drop, transection, and excitotoxic kainic acid delivery; in newborn rats, partial transection was performed. The regulation of expression patterns in the adult spinal cord was compared with that in the PNS and the neonate spinal cord. After mechanical injury of the adult rat spinal cord, upregulations of NGF and GDNF mRNA occurred in meningeal cells adjacent to the lesion. BDNF and p75 mRNA increased in neurons, GDNF mRNA increased in astrocytes close to the lesion, and GFRalpha-1 and truncated TrkB mRNA increased in astrocytes of degenerating white matter. The relatively limited upregulation of neurotrophic factors in the spinal cord contrasted with the response of affected nerve roots, in which marked increases of NGF and GDNF mRNA levels were observed in Schwann cells. The difference between the ability of the PNS and CNS to provide trophic support correlates with their different abilities to regenerate. Kainic acid delivery led to only weak upregulations of BDNF and CNTF mRNA. Compared with several brain regions, the overall response of the spinal cord tissue to kainic acid was weak. The relative sparseness of upregulations of endogenous neurotrophic factors after injury strengthens the hypothesis that lack of regeneration in the spinal cord is attributable at least partly to lack of trophic support.

Glial cell line-derived neurotrophic factor mediates the desirable actions of the anti-addiction drug ibogaine against alcohol consumption.

PubMed

He, Dao-Yao; McGough, Nancy N H; Ravindranathan, Ajay; Jeanblanc, Jerome; Logrip, Marian L; Phamluong, Khanhky; Janak, Patricia H; Ron, Dorit

2005-01-19

Alcohol addiction manifests as uncontrolled drinking despite negative consequences. Few medications are available to treat the disorder. Anecdotal reports suggest that ibogaine, a natural alkaloid, reverses behaviors associated with addiction including alcoholism; however, because of side effects, ibogaine is not used clinically. In this study, we first characterized the actions of ibogaine on ethanol self-administration in rodents. Ibogaine decreased ethanol intake by rats in two-bottle choice and operant self-administration paradigms. Ibogaine also reduced operant self-administration of ethanol in a relapse model. Next, we identified a molecular mechanism that mediates the desirable activities of ibogaine on ethanol intake. Microinjection of ibogaine into the ventral tegmental area (VTA), but not the substantia nigra, reduced self-administration of ethanol, and systemic administration of ibogaine increased the expression of glial cell line-derived neurotrophic factor (GDNF) in a midbrain region that includes the VTA. In dopaminergic neuron-like SHSY5Y cells, ibogaine treatment upregulated the GDNF pathway as indicated by increases in phosphorylation of the GDNF receptor, Ret, and the downstream kinase, ERK1 (extracellular signal-regulated kinase 1). Finally, the ibogaine-mediated decrease in ethanol self-administration was mimicked by intra-VTA microinjection of GDNF and was reduced by intra-VTA delivery of anti-GDNF neutralizing antibodies. Together, these results suggest that GDNF in the VTA mediates the action of ibogaine on ethanol consumption. These findings highlight the importance of GDNF as a new target for drug development for alcoholism that may mimic the effect of ibogaine against alcohol consumption but avoid the negative side effects.

Glial Cell Line-Derived Neurotrophic Factor Mediates the Desirable Actions of the Anti-Addiction Drug Ibogaine against Alcohol Consumption

PubMed Central

He, Dao-Yao; McGough, Nancy N. H.; Ravindranathan, Ajay; Jeanblanc, Jerome; Logrip, Marian L.; Phamluong, Khanhky; Janak, Patricia H.; Ron, Dorit

2005-01-01

Alcohol addiction manifests as uncontrolled drinking despite negative consequences. Few medications are available to treat the disorder. Anecdotal reports suggest that ibogaine, a natural alkaloid, reverses behaviors associated with addiction including alcoholism; however, because of side effects, ibogaine is not used clinically. In this study, we first characterized the actions of ibogaine on ethanol self-administration in rodents. Ibogaine decreased ethanol intake by rats in two-bottle choice and operant self-administration paradigms. Ibogaine also reduced operant self-administration of ethanol in a relapse model. Next, we identified a molecular mechanism that mediates the desirable activities of ibogaine on ethanol intake. Microinjection of ibogaine into the ventral tegmental area (VTA), but not the substantia nigra, reduced self-administration of ethanol, and systemic administration of ibogaine increased the expression of glial cell line-derived neurotrophic factor (GDNF) in a midbrain region that includes the VTA. In dopaminergic neuron-like SHSY5Y cells, ibogaine treatment upregulated the GDNF pathway as indicated by increases in phosphorylation of the GDNF receptor, Ret, and the downstream kinase, ERK1 (extracellular signal-regulated kinase 1). Finally, the ibogaine-mediated decrease in ethanol self-administration was mimicked by intra-VTA microinjection of GDNF and was reduced by intra-VTA delivery of anti-GDNF neutralizing antibodies. Together, these results suggest that GDNF in the VTA mediates the action of ibogaine on ethanol consumption. These findings highlight the importance of GDNF as a new target for drug development for alcoholism that may mimic the effect of ibogaine against alcohol consumption but avoid the negative side effects. PMID:15659598

Glial cell line-derived neurotrophic factor promotes the development of adrenergic neurons in mouse neural crest cultures

PubMed Central

Maxwell, Gerald D.; Reid, Kate; Elefanty, Andrew; Bartlett, Perry F.; Murphy, Mark

1996-01-01

Growth of mouse neural crest cultures in the presence of glial cell line-derived neurotrophic factor (GDNF) resulted in a dramatic dose-dependent increase in the number of tyrosine hydroxylase (TH)-positive cells that developed when 5% chicken embryo extract was present in the medium. In contrast, growth in the presence of bone morphogenetic protein (BMP)-2, BMP-4, BMP-6, transforming growth factor (TGF) β1, TGF-β2, and TGF-β3 elicited no increase in the number of TH-positive cells. The TH-positive cells that developed in the presence of GDNF had neuronal morphology and contained the middle and low molecular weight neurofilament proteins. Numerous TH-negative cells with the morphology of neurons also were observed in GDNF-treated cultures. Analysis revealed that the period from 6 to 12 days in vitro was the critical time for exposure to GDNF to generate the increase in TH-positive cell number. The growth factors neurotrophin-3 and fibroblast growth factor-2 elicited increases in the number of TH-positive cells similar to that seen in response to GDNF. In contrast, nerve growth factor was unable to substitute for GDNF. These findings extend the previously reported biological activities of GDNF by showing that it can act on mouse neural crest cultures to promote the development of neurons. PMID:8917581

Protection of dopaminergic neurons by electroconvulsive shock in an animal model of Parkinson's disease.

PubMed

Anastasia, Agustín; de Erausquin, Gabriel A; Wojnacki, José; Mascó, Daniel H

2007-11-01

Electroconvulsive shock (ECS) improves motor function in Parkinson's disease. In rats, ECS stimulates the expression of various factors some of which have been proposed to exert neuroprotective actions. We have investigated the effects of ECS on 6-hydroxydopamine (6-OHDA)-injected rats. Three weeks after a unilateral administration of 6-OHDA, 85-95% nigral dopaminergic neurons are lost. Chronic ECS prevented this cell loss, protect the nigrostriatal pathway (assessed by FloroGold retrograde labeling) and reduce motor impairment in 6-OHDA-treated animals. Injection of 6-OHDA caused loss of expression of glial cell-line derived neurotrophic factor (GDNF) in the substantia nigra. Chronic ECS completely prevented this loss of GDNF expression in 6-OHDA-treated animals. We also found that protected dopaminergic neurons co-express GDNF receptor proteins. These results strongly suggest that endogenous changes in GDNF expression may participate in the neuroprotective mechanism of ECS against 6-OHDA induced toxicity.

Use of GDNF Releasing Nanofiber Nerve Guide Conduits for the Repair of Conus Medullaris/Cauda Equina Injury in the Nonhuman Primate

DTIC Science & Technology

2015-02-01

repair of conus medullaris/cauda equina injury in rhesus macaques using a biodegradable bridging graft that releasing the trophic factor, GDNF. All...and biodegradable nerve guidance channels as bridging grafts. The follow section describes the methods and protocols for laminectomy, ventral root...in saline until being grafted, 8. For use of a bridging biodegradable NGC segment, the NGCs were fabricated and comprised of electrospun

Glial cell line-derived neurotrophic factor promotes barrier maturation and wound healing in intestinal epithelial cells in vitro.

PubMed

Meir, Michael; Flemming, Sven; Burkard, Natalie; Bergauer, Lisa; Metzger, Marco; Germer, Christoph-Thomas; Schlegel, Nicolas

2015-10-15

Recent data suggest that neurotrophic factors from the enteric nervous system are involved in intestinal epithelial barrier regulation. In this context the glial cell line-derived neurotrophic factor (GDNF) was shown to affect gut barrier properties in vivo directly or indirectly by largely undefined processes in a model of inflammatory bowel disease (IBD). We further investigated the potential role and mechanisms of GDNF in the regulation of intestinal barrier functions. Immunostaining of human gut specimen showed positive GDNF staining in enteric neuronal plexus and in enterocytes. In Western blots of the intestinal epithelial cell lines Caco2 and HT29B6, significant amounts of GDNF were detected, suggesting that enterocytes represent an additional source of GDNF. Application of recombinant GDNF on Caco2 and HT29B6 cells for 24 h resulted in significant epithelial barrier stabilization in monolayers with immature barrier functions. Wound-healing assays showed a significantly faster closure of the wounded areas after GDNF application. GDNF augmented cAMP levels and led to significant inactivation of p38 MAPK in immature cells. Activation of p38 MAPK signaling by SB-202190 mimicked GDNF-induced barrier maturation, whereas the p38 MAPK activator anisomycin blocked GDNF-induced effects. Increasing cAMP levels had adverse effects on barrier maturation, as revealed by permeability measurements. However, increased cAMP augmented the proliferation rate in Caco2 cells, and GDNF-induced proliferation of epithelial cells was abrogated by the PKA inhibitor H89. Our data show that enterocytes represent an additional source of GDNF synthesis. GDNF contributes to wound healing in a cAMP/PKA-dependent manner and promotes barrier maturation in immature enterocytes cells by inactivation of p38 MAPK signaling. Copyright © 2015 the American Physiological Society.

Low-Intensity Extracorporeal Shock Wave Therapy Enhances Brain-Derived Neurotrophic Factor Expression through PERK/ATF4 Signaling Pathway.

PubMed

Wang, Bohan; Ning, Hongxiu; Reed-Maldonado, Amanda B; Zhou, Jun; Ruan, Yajun; Zhou, Tie; Wang, Hsun Shuan; Oh, Byung Seok; Banie, Lia; Lin, Guiting; Lue, Tom F

2017-02-16

Low-intensity extracorporeal shock wave therapy (Li-ESWT) is used in the treatment of erectile dysfunction, but its mechanisms are not well understood. Previously, we found that Li-ESWT increased the expression of brain-derived neurotrophic factor (BDNF). Here we assessed the underlying signaling pathways in Schwann cells in vitro and in penis tissue in vivo after nerve injury. The result indicated that BDNF were significantly increased by the Li-ESWT after nerve injury, as well as the expression of BDNF in Schwann cells (SCs, RT4-D6P2T) in vitro. Li-ESWT activated the protein kinase RNA-like endoplasmic reticulum (ER) kinase (PERK) pathway by increasing the phosphorylation levels of PERK and eukaryotic initiation factor 2a (eIF2α), and enhanced activating transcription factor 4 (ATF4) in an energy-dependent manner. In addition, GSK2656157-an inhibitor of PERK-effectively inhibited the effect of Li-ESWT on the phosphorylation of PERK, eIF2α, and the expression of ATF4. Furthermore, silencing ATF4 dramatically attenuated the effect of Li-ESWT on the expression of BDNF, but had no effect on hypoxia-inducible factor (HIF)1α or glial cell-derived neurotrophic factor (GDNF) in Schwann cells. In conclusion, our findings shed new light on the underlying mechanisms by which Li-ESWT may stimulate the expression of BDNF through activation of PERK/ATF4 signaling pathway. This information may help to refine the use of Li-ESWT to further improve its clinical efficacy.

Low-Intensity Extracorporeal Shock Wave Therapy Enhances Brain-Derived Neurotrophic Factor Expression through PERK/ATF4 Signaling Pathway

PubMed Central

Wang, Bohan; Ning, Hongxiu; Reed-Maldonado, Amanda B.; Zhou, Jun; Ruan, Yajun; Zhou, Tie; Wang, Hsun Shuan; Oh, Byung Seok; Banie, Lia; Lin, Guiting; Lue, Tom F.

2017-01-01

Low-intensity extracorporeal shock wave therapy (Li-ESWT) is used in the treatment of erectile dysfunction, but its mechanisms are not well understood. Previously, we found that Li-ESWT increased the expression of brain-derived neurotrophic factor (BDNF). Here we assessed the underlying signaling pathways in Schwann cells in vitro and in penis tissue in vivo after nerve injury. The result indicated that BDNF were significantly increased by the Li-ESWT after nerve injury, as well as the expression of BDNF in Schwann cells (SCs, RT4-D6P2T) in vitro. Li-ESWT activated the protein kinase RNA-like endoplasmic reticulum (ER) kinase (PERK) pathway by increasing the phosphorylation levels of PERK and eukaryotic initiation factor 2a (eIF2α), and enhanced activating transcription factor 4 (ATF4) in an energy-dependent manner. In addition, GSK2656157—an inhibitor of PERK—effectively inhibited the effect of Li-ESWT on the phosphorylation of PERK, eIF2α, and the expression of ATF4. Furthermore, silencing ATF4 dramatically attenuated the effect of Li-ESWT on the expression of BDNF, but had no effect on hypoxia-inducible factor (HIF)1α or glial cell-derived neurotrophic factor (GDNF) in Schwann cells. In conclusion, our findings shed new light on the underlying mechanisms by which Li-ESWT may stimulate the expression of BDNF through activation of PERK/ATF4 signaling pathway. This information may help to refine the use of Li-ESWT to further improve its clinical efficacy. PMID:28212323

Plasma glial cell line-derived neurotrophic factor in patients with major depressive disorder: a preliminary study.

PubMed

Lee, Bun-Hee; Hong, Jin-Pyo; Hwang, Jung-A; Na, Kyoung-Sae; Kim, Won-Joong; Trigo, Jose; Kim, Yong-Ku

2016-02-01

Some clinical studies have reported reduced peripheral glial cell line-derived neurotrophic factor (GDNF) level in elderly patients with major depressive disorder (MDD). We verified whether a reduction in plasma GDNF level was associated with MDD. Plasma GDNF level was measured in 23 healthy control subjects and 23 MDD patients before and after 6 weeks of treatment. Plasma GDNF level in MDD patients at baseline did not differ from that in healthy controls. Plasma GDNF in MDD patients did not differ significantly from baseline to the end of treatment. GDNF level was significantly lower in recurrent-episode MDD patients than in first-episode patients before and after treatment. Our findings revealed significantly lower plasma GDNF level in recurrent-episode MDD patients, although plasma GDNF levels in MDD patients and healthy controls did not differ significantly. The discrepancy between our study and previous studies might arise from differences in the recurrence of depression or the ages of the MDD patients.

Fibrin matrices with affinity-based delivery systems and neurotrophic factors promote functional nerve regeneration.

PubMed

Wood, Matthew D; MacEwan, Matthew R; French, Alexander R; Moore, Amy M; Hunter, Daniel A; Mackinnon, Susan E; Moran, Daniel W; Borschel, Gregory H; Sakiyama-Elbert, Shelly E

2010-08-15

Glial-derived neurotrophic factor (GDNF) and nerve growth factor (NGF) have both been shown to enhance peripheral nerve regeneration following injury and target different neuronal populations. The delivery of either growth factor at the site of injury may, therefore, result in quantitative differences in motor nerve regeneration and functional recovery. In this study we evaluated the effect of affinity-based delivery of GDNF or NGF from fibrin-filled nerve guidance conduits (NGCs) on motor nerve regeneration and functional recovery in a 13 mm rat sciatic nerve defect. Seven experimental groups were evaluated consisting of GDNF or NGF and the affinity-based delivery system (DS) within NGCs, control groups excluding the DS and/or growth factor, and nerve isografts. Groups with growth factor in the conduit demonstrated equivalent or superior performance in behavioral tests and relative muscle mass measurements compared to isografts at 12 weeks. Additionally, groups with GDNF demonstrated greater specific twitch and tetanic force production in extensor digitorum longus (EDL) muscle than the isograft control, while groups with NGF produced demonstrated similar force production compared to the isograft control. Assessment of motor axon regeneration by retrograde labeling further revealed that the number of ventral horn neurons regenerating across NGCs containing GDNF and NGF DS was similar to the isograft group and these counts were greater than the groups without growth factor. Overall, the GDNF DS group demonstrated superior functional recovery and equivalent motor nerve regeneration compared to the isograft control, suggesting it has potential as a treatment for motor nerve injury.

Human neural progenitor cells over-expressing IGF-1 protect dopamine neurons and restore function in a rat model of Parkinson's disease.

PubMed

Ebert, Allison D; Beres, Amy J; Barber, Amelia E; Svendsen, Clive N

2008-01-01

Growth factors such as glial cell line-derived neurotrophic factor (GDNF) have been shown to prevent neurodegeneration and promote regeneration in many animal models of Parkinson's disease (PD). Insulin-like growth factor 1 (IGF-1) is also known to have neuroprotective effects in a number of disease models but has not been extensively studied in models of PD. We produced human neural progenitor cells (hNPC) releasing either GDNF or IGF-1 and transplanted them into a rat model of PD. hNPC secreting either GDNF or IGF-1 were shown to significantly reduce amphetamine-induced rotational asymmetry and dopamine neuron loss when transplanted 7 days after a 6-hydroxydopamine (6-OHDA) lesion. Neither untransduced hNPC nor a sham transplant had this effect suggesting GDNF and IGF-1 release was required. Interestingly, GDNF, but not IGF-1, was able to protect or regenerate tyrosine hydroxylase-positive fibers in the striatum. In contrast, IGF-1, but not GDNF, significantly increased the overall survival of hNPC both in vitro and following transplantation. This suggests a dual role of IGF-1 to both increase hNPC survival after transplantation and exert trophic effects on degenerating dopamine neurons in this rat model of PD.

Glial cell-derived neurotrophic factor gene polymorpisms affect severity and functionality of bipolar disorder.

PubMed

Safari, Roghaiyeh; Tunca, Zeliha; Özerdem, Ayşegül; Ceylan, Deniz; Yalçın, Yaprak; Sakizli, Meral

2017-01-01

Glial cell-derived neurotrophic factor and other neurotrophins have important role in the development of mental disorders. Here, we aimed to assess the effects of Single nucleotide polymorphisms at potentially regulated regions of GDNF on severity and functionality of bipolar disorder and GDNF serum levels in bipolar disorder patients and healthy volunteers. Severity and functionality of bipolar disorder were evaluated using the Clinical Global Impression and Global Assessment of Functioning scales in sixty-six bipolar disorder patients. The GDNF serum levels obtained from bipolar disorder patients and healthy volunteers who had been already reported SNPs information by our group. GAF scales were lower and GDNF serum levels were higher in Bipolar disorder patients with T/A genotype at 5:37812784 and 5:37812782 compared to patients with T/T genotype. There were significant difference in severity and functionality scores, but not in GDNF serum levels, between patients with G/G and G/A genotype of rs62360370 G > A SNP.rs2075680 C > A and rs79669773 T > C SNPs had no effect on bipolar disorder severity and functionality scores and GDNF serum levels. The results suggest that some SNPs of GDNF have potential association with severity and functionality of bipolar disorder. In addition, except two SNPs, none of GDNF SNPs had association with GDNF serum levels.

Performance on the Wisconsin card-sorting test and serum levels of glial cell line-derived neurotrophic factor in patients with major depressive disorder.

PubMed

Zhang, Xiaobin; Ru, Bu; Sha, Weiwei; Xin, Wang; Zhou, Honghui; Zhang, Yumei

2014-09-01

Some evidence suggests that neurotrophic growth factor systems might be involved in the etiology of major depressive disorder (MDD). Glial cell line-derived neurotrophic factor (GDNF) is a neurotrophic factor from the transforming growth factor-β family that plays a role in the development and function of the brain. This study aimed to test whether GDNF in serum was abnormal in MDD, and whether it was related to the cognitive impairment of MDD. Serum GDNF levels in MDD patients (n = 32) and normal controls (n = 32) were measured with the enzyme-linked immunosorbent assay method. All subjects were assessed for performance on the Wisconsin card-sorting test (WCST). Performance on the WCST in MDD patients was significantly poorer than that in controls. Serum GDNF levels in MDD patients were significantly decreased compared to that of the control subjects (P < 0.001). Furthermore, the decrease in the serum GDNF levels positively correlated with performance in the WCST-% CONC and negatively with performance in the WCST-P in MDD patients. The findings suggest that MDD patients have extensive impairments of executive functioning, and lower serum GDNF might be involved in the pathogenesis of MDD, which may be associated with the cognitive dysfunction in MDD patients. © 2014 Wiley Publishing Asia Pty Ltd.

Targeting the Neural Microenvironment in Prostate Cancer

DTIC Science & Technology

2016-10-01

neurotrophic factor (GDNF), which is expressed by peripheral nerves. GDNF binds to RET, a receptor tyrosine kinase, in conjunction with its co-receptor...kinase, in conjunction with its co- receptor GFRα1 and activates cellular signaling. Studies in pancreatic cancer strongly implicate RET signaling as

Regulators of gene expression in Enteric Neural Crest Cells are putative Hirschsprung disease genes.

PubMed

Schriemer, Duco; Sribudiani, Yunia; IJpma, Arne; Natarajan, Dipa; MacKenzie, Katherine C; Metzger, Marco; Binder, Ellen; Burns, Alan J; Thapar, Nikhil; Hofstra, Robert M W; Eggen, Bart J L

2016-08-01

The enteric nervous system (ENS) is required for peristalsis of the gut and is derived from Enteric Neural Crest Cells (ENCCs). During ENS development, the RET receptor tyrosine kinase plays a critical role in the proliferation and survival of ENCCs, their migration along the developing gut, and differentiation into enteric neurons. Mutations in RET and its ligand GDNF cause Hirschsprung disease (HSCR), a complex genetic disorder in which ENCCs fail to colonize variable lengths of the distal bowel. To identify key regulators of ENCCs and the pathways underlying RET signaling, gene expression profiles of untreated and GDNF-treated ENCCs from E14.5 mouse embryos were generated. ENCCs express genes that are involved in both early and late neuronal development, whereas GDNF treatment induced neuronal maturation. Predicted regulators of gene expression in ENCCs include the known HSCR genes Ret and Sox10, as well as Bdnf, App and Mapk10. The regulatory overlap and functional interactions between these genes were used to construct a regulatory network that is underlying ENS development and connects to known HSCR genes. In addition, the adenosine receptor A2a (Adora2a) and neuropeptide Y receptor Y2 (Npy2r) were identified as possible regulators of terminal neuronal differentiation in GDNF-treated ENCCs. The human orthologue of Npy2r maps to the HSCR susceptibility locus 4q31.3-q32.3, suggesting a role for NPY2R both in ENS development and in HSCR. Copyright © 2016 Elsevier Inc. All rights reserved.

Augmentation of the ascending component of the peristaltic reflex and substance P release by glial cell line-derived neurotrophic factor.

PubMed

Grider, J R; Heuckeroth, R O; Kuemmerle, J F; Murthy, K S

2010-07-01

Glial cell line-derived neurotrophic factor (GDNF) is present in adult gut although its role in the mature enteric nervous system is not well defined. The aim of the present study was to examine the role of GDNF as neuromodulator of the ascending phase of the peristaltic reflex. Colonic segments were prepared as flat sheets and placed in compartmented chambers so as to separate the sensory and motor limbs of the reflex. Ascending contraction was measured in the orad compartment and mucosal stroking stimuli (two to eight strokes) were applied in the caudad compartment. GDNF and substance P (SP) release were measured and the effects of GDNF and GDNF antibody on contraction and release were determined. Mice with reduced levels of GDNF (Gdnf(+/-)) and wild type littermates were also examined. GDNF was released in a stimulus-dependent manner into the orad motor but not caudad sensory compartment. Addition of GDNF to the orad motor but not caudad sensory compartment augmented ascending contraction and SP release. Conversely, addition of GDNF antibody to the orad motor but not caudad sensory compartment reduced ascending contraction and SP release. Similarly, the ascending contraction and SP release into the orad motor compartment was reduced in Gdnf(+/-) mice as compared to wild type littermates. The results suggest that endogenous GDNF is released during the ascending contraction component of the peristaltic reflex where it acts as a neuromodulator to augment SP release from motor neurons thereby augmenting contraction of circular muscle orad to the site of stimulation.

c-RET Molecule in Malignant Melanoma from Oncogenic RET-Carrying Transgenic Mice and Human Cell Lines

PubMed Central

Takeda, Kozue; Iida, Machiko; Kumasaka, Mayuko; Matsumoto, Yoshinari; Kato, Masashi

2010-01-01

Malignant melanoma is one of the most aggressive cancers and its incidence worldwide has been increasing at a greater rate than that of any other cancer. We previously reported that constitutively activated RFP-RET-carrying transgenic mice (RET-mice) spontaneously develop malignant melanoma. In this study, we showed that expression levels of intrinsic c-Ret, glial cell line-derived neurotrophic factor (Gdnf) and Gdnf receptor alpha 1 (Gfra1) transcripts in malignant melanomas from RET-transgenic mice were significantly upregulated compared with those in benign melanocytic tumors. These results suggest that not only introduced oncogenic RET but also intrinsic c-Ret/Gdnf are involved in murine melanomagenesis in RET-mice. We then showed that c-RET and GDNF transcript expression levels in human malignant melanoma cell lines (HM3KO and MNT-1) were higher than those in primary cultured normal human epithelial melanocytes (NHEM), while GFRa1 transcript expression levels were comparable among NHEM, HM3KO and MNT-1. We next showed c-RET and GFRa1 protein expression in HM3KO cells and GDNF-mediated increased levels of their phosphorylated c-RET tyrosine kinase and signal transduction molecules (ERK and AKT) sited potentially downstream of c-RET. Taken together with the finding of augmented proliferation of HM3KO cells after GDNF stimulation, our results suggest that GDNF-mediated c-RET kinase activation is associated with the pathogenesis of malignant melanoma. PMID:20422010

c-RET molecule in malignant melanoma from oncogenic RET-carrying transgenic mice and human cell lines.

PubMed

Ohshima, Yuichiro; Yajima, Ichiro; Takeda, Kozue; Iida, Machiko; Kumasaka, Mayuko; Matsumoto, Yoshinari; Kato, Masashi

2010-04-21

Malignant melanoma is one of the most aggressive cancers and its incidence worldwide has been increasing at a greater rate than that of any other cancer. We previously reported that constitutively activated RFP-RET-carrying transgenic mice (RET-mice) spontaneously develop malignant melanoma. In this study, we showed that expression levels of intrinsic c-Ret, glial cell line-derived neurotrophic factor (Gdnf) and Gdnf receptor alpha 1 (Gfra1) transcripts in malignant melanomas from RET-transgenic mice were significantly upregulated compared with those in benign melanocytic tumors. These results suggest that not only introduced oncogenic RET but also intrinsic c-Ret/Gdnf are involved in murine melanomagenesis in RET-mice. We then showed that c-RET and GDNF transcript expression levels in human malignant melanoma cell lines (HM3KO and MNT-1) were higher than those in primary cultured normal human epithelial melanocytes (NHEM), while GFRa1 transcript expression levels were comparable among NHEM, HM3KO and MNT-1. We next showed c-RET and GFRa1 protein expression in HM3KO cells and GDNF-mediated increased levels of their phosphorylated c-RET tyrosine kinase and signal transduction molecules (ERK and AKT) sited potentially downstream of c-RET. Taken together with the finding of augmented proliferation of HM3KO cells after GDNF stimulation, our results suggest that GDNF-mediated c-RET kinase activation is associated with the pathogenesis of malignant melanoma.

Evaluation of the effect of follicular stimulating hormone on the in vitro bovine spermatogonial stem cells self-renewal: An experimental study

PubMed Central

Jabarpour, Masoome; Tajik, Parviz

2017-01-01

Background: Spermatogonial stem cells (SSCs) are undifferentiated cells which are highly reproducible and expandable. Several studies have been conducted to reproduce these cells in culture. They used growth factors, hormones and different feeder cells to improve survival and proliferation of SSCs. Objective: This study was conducted to evaluate the effects of follicular stimulating hormone (FSH) on gene expression of fibroblast growth factor (FGF2) and glial cell-derived neurotrophic factor (GDNF) in Sertoli cells. Materials and Methods: Sertoli cells and SSCs were isolated from 3-5 month-old calves. Bovine testicular cells were cultured for 15 days with or without FSH. Identification of these cells was confirmed by immunocytochemistry analysis. Colony formation of SSCs was evaluated using an inverted microscope. The gene expression of FGF2 and GDNF and the gene markers bcl6b, thy-1, and C-kit were evaluated using the quantitative RT-PCR technique. Results: The results indicated that FSH increased colonization of SSCs. the expression of GDNF, FGF2, and markers of undifferentiated spermatogonia was increased following culture in control and FSH groups (p<0.05), this increase was more in FSH group. Conversely, the expression of C-kit was decreased in both groups (p<0.05). Conclusion: The results showed that FSH can increase the self-renewal of SSCs in vitro via upregulation of GDNF and FGF2 expression in Sertoli cells. PMID:29492477

Inhibitory Effect of Memantine on Streptozotocin-Induced Insulin Receptor Dysfunction, Neuroinflammation, Amyloidogenesis, and Neurotrophic Factor Decline in Astrocytes.

PubMed

Rajasekar, N; Nath, Chandishwar; Hanif, Kashif; Shukla, Rakesh

2016-12-01

Our earlier studies showed that insulin receptor (IR) dysfunction along with neuroinflammation and amyloidogenesis played a major role in streptozotocin (STZ)-induced toxicity in astrocytes. N-methyl-D-aspartate (NMDA) receptor antagonist-memantine shows beneficial effects in Alzheimer's disease (AD) pathology. However, the protective molecular and cellular mechanism of memantine in astrocytes is not properly understood. Therefore, the present study was undertaken to investigate the effect of memantine on insulin receptors, neurotrophic factors, neuroinflammation, and amyloidogenesis in STZ-treated astrocytes. STZ (100 μM) treatment for 24 h in astrocytes resulted significant decrease in brain-derived neurotrophic factor (BDNF), glial cell line-derived neurotrophic factor (GDNF), and insulin-degrading enzyme (IDE) expression in astrocytes. Treatment with memantine (1-10 μM) improved STZ-induced neurotrophic factor decline (BDNF, GDNF) along with IR dysfunction as evidenced by a significant increase in IR protein expression, phosphorylation of IRS-1, Akt, and GSK-3 α/β in astrocytes. Further, memantine attenuated STZ-induced amyloid precursor protein (APP), β-site APP-cleaving enzyme-1 and amyloid-β 1-42 expression and restored IDE expression in astrocytes. In addition, memantine also displays protective effects against STZ-induced astrocyte activation showed by reduction of inflammatory markers, nuclear factor kappa-B translocation, glial fibrillary acidic protein, cyclooxygenase-2, tumor necrosis factor-α level, and oxidative-nitrostative stress. The results suggest that besides the NMDA receptor antagonisic activity, effect on astroglial IR and neurotrophic factor may also be an important factor in the beneficial effect of memantine in AD pathology. Graphical Abstract Novel neuroprotective mechanisms of memenatine in streptozotocin-induced toxicity in astrocytes.

The importance of neuronal growth factors in the ovary.

PubMed

Streiter, S; Fisch, B; Sabbah, B; Ao, A; Abir, R

2016-01-01

The neurotrophin family consists of nerve growth factor (NGF), neurotrophin 3 (NT3) and neurotrophin 4/5 (NT4/5), in addition to brain-derived neurotrophic factor (BDNF) and the neuronal growth factors, glial cell line-derived neurotrophic factor (GDNF) and vasointestinal peptide (VIP). Although there are a few literature reviews, mainly of animal studies, on the importance of neurotrophins in the ovary, we aimed to provide a complete review of neurotrophins as well as neuronal growth factors and their important roles in normal and pathological processes in the ovary. Follicular assembly is probably stimulated by complementary effects of NGF, NT4/5 and BDNF and their receptors. The neurotrophins, GDNF and VIP and their receptors have all been identified in preantral and antral follicles of mammalian species, including humans. Transgenic mice with mutations in the genes encoding for Ngf, Nt4/5 and Bdnf and their tropomyosin-related kinase β receptor showed a reduction in preantral follicles and an abnormal ovarian morphology, whereas NGF, NT3, GDNF and VIP increased the in vitro activation of primordial follicles in rats and goats. Additionally, NGF, NT3 and GDNF promoted follicular cell proliferation; NGF, BDNF and VIP were shown to be involved in ovulation; VIP inhibited follicular apoptosis; NT4/5, BDNF and GDNF promoted oocyte maturation and NGF, NT3 and VIP stimulated steroidogenesis. NGF may also exert a stimulatory effect in ovarian cancer and polycystic ovarian syndrome (PCOS). Low levels of NGF and BDNF in follicular fluid may be associated with diminished ovarian reserve and high levels with endometriosis. More knowledge of the roles of neuronal growth factors in the ovary has important implications for the development of new therapeutic drugs (such as anti-NGF agents) for ovarian cancer and PCOS as well as various infertility problems, warranting further research. © The Author 2015. Published by Oxford University Press on behalf of the European Society

Local delivery of glial cell line-derived neurotrophic factor improves facial nerve regeneration after late repair.

PubMed

Barras, Florian M; Kuntzer, Thierry; Zurn, Anne D; Pasche, Philippe

2009-05-01

Facial nerve regeneration is limited in some clinical situations: in long grafts, by aged patients, and when the delay between nerve lesion and repair is prolonged. This deficient regeneration is due to the limited number of regenerating nerve fibers, their immaturity and the unresponsiveness of Schwann cells after a long period of denervation. This study proposes to apply glial cell line-derived neurotrophic factor (GDNF) on facial nerve grafts via nerve guidance channels to improve the regeneration. Two situations were evaluated: immediate and delayed grafts (repair 7 months after the lesion). Each group contained three subgroups: a) graft without channel, b) graft with a channel without neurotrophic factor; and c) graft with a GDNF-releasing channel. A functional analysis was performed with clinical observation of facial nerve function, and nerve conduction study at 6 weeks. Histological analysis was performed with the count of number of myelinated fibers within the graft, and distally to the graft. Central evaluation was assessed with Fluoro-Ruby retrograde labeling and Nissl staining. This study showed that GDNF allowed an increase in the number and the maturation of nerve fibers, as well as the number of retrogradely labeled neurons in delayed anastomoses. On the contrary, after immediate repair, the regenerated nerves in the presence of GDNF showed inferior results compared to the other groups. GDNF is a potent neurotrophic factor to improve facial nerve regeneration in grafts performed several months after the nerve lesion. However, GDNF should not be used for immediate repair, as it possibly inhibits the nerve regeneration.

Lesion-induced increase in survival and migration of human neural progenitor cells releasing GDNF

PubMed Central

Behrstock, Soshana; Ebert, Allison D.; Klein, Sandra; Schmitt, Melanie; Moore, Jeannette M.; Svendsen, Clive N.

2009-01-01

The use of human neural progenitor cells (hNPC) has been proposed to provide neuronal replacement or astrocytes delivering growth factors for brain disorders such as Parkinson’s and Huntington’s disease. Success in such studies likely requires migration from the site of transplantation and integration into host tissue in the face of ongoing damage. In the current study, hNPC modified to release glial cell line derived neurotrophic factor (hNPCGDNF) were transplanted into either intact or lesioned animals. GDNF release itself had no effect on the survival, migration or differentiation of the cells. The most robust migration and survival was found using a direct lesion of striatum (Huntington’s model) with indirect lesions of the dopamine system (Parkinson’s model) or intact animals showing successively less migration and survival. No lesion affected differentiation patterns. We conclude that the type of brain injury dictates migration and integration of hNPC which has important consequences when considering transplantation of these cells as a therapy for neurodegenerative diseases. PMID:19044202

Visual detection of glial cell line-derived neurotrophic factor based on a molecular translator and isothermal strand-displacement polymerization reaction.

PubMed

Zhang, Li-Yong; Xing, Tao; Du, Li-Xin; Li, Qing-Min; Liu, Wei-Dong; Wang, Ji-Yue; Cai, Jing

2015-01-01

Glial cell line-derived neurotrophic factor (GDNF) is a small protein that potently promotes the survival of many types of neurons. Detection of GDNF is vital to monitoring the survival of sympathetic and sensory neurons. However, the specific method for GDNF detection is also un-discovered. The purpose of this study is to explore the method for protein detection of GDNF. A novel visual detection method based on a molecular translator and isothermal strand-displacement polymerization reaction (ISDPR) has been proposed for the detection of GDNF. In this study, a molecular translator was employed to convert the input protein to output deoxyribonucleic acid signal, which was further amplified by ISDPR. The product of ISDPR was detected by a lateral flow biosensor within 30 minutes. This novel visual detection method based on a molecular translator and ISDPR has very high sensitivity and selectivity, with a dynamic response ranging from 1 pg/mL to 10 ng/mL, and the detection limit was 1 pg/mL of GDNF. This novel visual detection method exhibits high sensitivity and selectivity, which is very simple and universal for GDNF detection to help disease therapy in clinical practice.

Visual detection of glial cell line-derived neurotrophic factor based on a molecular translator and isothermal strand-displacement polymerization reaction

PubMed Central

Zhang, Li-Yong; Xing, Tao; Du, Li-Xin; Li, Qing-Min; Liu, Wei-Dong; Wang, Ji-Yue; Cai, Jing

2015-01-01

Background Glial cell line-derived neurotrophic factor (GDNF) is a small protein that potently promotes the survival of many types of neurons. Detection of GDNF is vital to monitoring the survival of sympathetic and sensory neurons. However, the specific method for GDNF detection is also un-discovered. The purpose of this study is to explore the method for protein detection of GDNF. Methods A novel visual detection method based on a molecular translator and isothermal strand-displacement polymerization reaction (ISDPR) has been proposed for the detection of GDNF. In this study, a molecular translator was employed to convert the input protein to output deoxyribonucleic acid signal, which was further amplified by ISDPR. The product of ISDPR was detected by a lateral flow biosensor within 30 minutes. Results This novel visual detection method based on a molecular translator and ISDPR has very high sensitivity and selectivity, with a dynamic response ranging from 1 pg/mL to 10 ng/mL, and the detection limit was 1 pg/mL of GDNF. Conclusion This novel visual detection method exhibits high sensitivity and selectivity, which is very simple and universal for GDNF detection to help disease therapy in clinical practice. PMID:25848224

Serum brain-derived neurotrophic factor, glial-derived neurotrophic factor, nerve growth factor, and neurotrophin-3 levels in children with attention-deficit/hyperactivity disorder.

PubMed

Bilgiç, Ayhan; Toker, Aysun; Işık, Ümit; Kılınç, İbrahim

2017-03-01

It has been suggested that neurotrophins are involved in the etiopathogenesis of attention-deficit/hyperactivity disorder (ADHD). This study aimed to investigate whether there are differences in serum brain-derived neurotrophic factor (BDNF), glial-derived neurotrophic factor (GDNF), nerve growth factor (NGF), and neurotrophin-3 (NTF3) levels between children with ADHD and healthy controls. A total of 110 treatment-naive children with the combined presentation of ADHD and 44 healthy controls aged 8-18 years were enrolled in this study. The severity of ADHD symptoms was determined by scores on the Conners' Parent Rating Scale-Revised Short and Conners' Teacher Rating Scale-Revised Short. The severity of depression and anxiety symptoms of the children were evaluated by the self-report inventories. Serum levels of neurotrophins were measured using commercial enzyme-linked immunosorbent assay kits. The multivariate analysis of covariance (MANCOVA) revealed a significant main effect of groups in the levels of serum neurotrophins, an effect that was independent of age, sex, and the severity of the depression and anxiety. The analysis of covariance (ANCOVA) indicated that the mean serum GDNF and NTF3 levels of ADHD patients were significantly higher than that of controls. However, serum BDNF and NGF levels did not show any significant differences between groups. No correlations between the levels of serum neurotrophins and the severity of ADHD were observed. These results suggest that elevated serum GDNF and NTF3 levels may be related to ADHD in children.

CD24 expression does not affect dopamine neuronal survival in a mouse model of Parkinson's disease.

PubMed

Stott, Simon R W; Hayat, Shaista; Carnwath, Tom; Garas, Shaady; Sleeman, Jonathan P; Barker, Roger A

2017-01-01

Parkinson's disease (PD) is a progressive neurodegenerative condition that is characterised by the loss of specific populations of neurons in the brain. The mechanisms underlying this selective cell death are unknown but by using laser capture microdissection, the glycoprotein, CD24 has been identified as a potential marker of the populations of cells that are affected in PD. Using in situ hybridization and immunohistochemistry on sections of mouse brain, we confirmed that CD24 is robustly expressed by many of these subsets of cells. To determine if CD24 may have a functional role in PD, we modelled the dopamine cell loss of PD in Cd24 mutant mice using striatal delivery of the neurotoxin 6-OHDA. We found that Cd24 mutant mice have an anatomically normal dopamine system and that this glycoprotein does not modulate the lesion effects of 6-OHDA delivered into the striatum. We then undertook in situ hybridization studies on sections of human brain and found-as in the mouse brain-that CD24 is expressed by many of the subsets of the cells that are vulnerable in PD, but not those of the midbrain dopamine system. Finally, we sought to determine if CD24 is required for the neuroprotective effect of Glial cell-derived neurotrophic factor (GDNF) on the dopaminergic nigrostriatal pathway. Our results indicate that in the absence of CD24, there is a reduction in the protective effects of GDNF on the dopaminergic fibres in the striatum, but no difference in the survival of the cell bodies in the midbrain. While we found no obvious role for CD24 in the normal development and maintenance of the dopaminergic nigrostriatal system in mice, it may have a role in mediating the neuroprotective aspects of GDNF in this system.

Zirconium oxide ceramic foam: a promising supporting biomaterial for massive production of glial cell line-derived neurotrophic factor*

PubMed Central

Liu, Zhong-wei; Li, Wen-qiang; Wang, Jun-kui; Ma, Xian-cang; Liang, Chen; Liu, Peng; Chu, Zheng; Dang, Yong-hui

2014-01-01

This study investigated the potential application of a zirconium oxide (ZrO2) ceramic foam culturing system to the production of glial cell line-derived neurotrophic factor (GDNF). Three sets of ZrO2 ceramic foams with different pore densities of 10, 20, and 30 pores per linear inch (PPI) were prepared to support a 3D culturing system. After primary astrocytes were cultured in these systems, production yields of GDNF were evaluated. The biomaterial biocompatibility, cell proliferation and activation of cellular signaling pathways in GDNF synthesis and secretion in the culturing systems were also assessed and compared with a conventional culturing system. In this study, we found that the ZrO2 ceramic foam culturing system was biocompatible, using which the GDNF yields were elevated and sustained by stimulated cell proliferation and activation of signaling pathways in astrocytes cultured in the system. In conclusion, the ZrO2 ceramic foam is promising for the development of a GDNF mass production device for Parkinson’s disease treatment. PMID:25471830

Targeting the Neural Microenvironment in Prostate Cancer

DTIC Science & Technology

2017-10-01

exogenous GDNF, implying they are the only PCa cell line that expresses GFRα1. Indeed, as seen in Fig 2A, by western blotting and ELISA for GDNF, all PCa...lysates with anti-GDNF antibody. Positive control is recombinant GDNF. Actin is a loading control. B. GDNF quantitation by ELISA of PCa cell line

Short-term ethanol exposure causes imbalanced neurotrophic factor allocation in the basal forebrain cholinergic system: a novel insight into understanding the initial processes of alcohol addiction.

PubMed

Miki, Takanori; Kusaka, Takashi; Yokoyama, Toshifumi; Ohta, Ken-ichi; Suzuki, Shingo; Warita, Katsuhiko; Jamal, Mostofa; Wang, Zhi-Yu; Ueki, Masaaki; Liu, Jun-Qian; Yakura, Tomiko; Tamai, Motoki; Sumitani, Kazunori; Hosomi, Naohisa; Takeuchi, Yoshiki

2014-02-01

Alcohol ingestion affects both motor and cognitive functions. One brain system that is influenced by ethanol is the basal forebrain (BF) cholinergic projection system, which projects to diverse neocortical and limbic areas. The BF is associated with memory and cognitive function. Our primary interest is the examination of how regions that receive BF cholinergic projections are influenced by short-term ethanol exposure through alterations in the mRNA levels of neurotrophic factors [nerve growth factor/TrkA, brain-derived neurotrophic factor/TrkB, and glial-derived neurotrophic factor (GDNF)/GDNF family receptor α1]. Male BALB/C mice were fed a liquid diet containing 5 % (v/v) ethanol. Pair-fed control mice were maintained on an identical liquid diet, except that the ethanol was isocalorically substituted with sucrose. Mice exhibiting signs of ethanol intoxication (stages 1-2) were used for real-time reverse transcription-polymerase chain reaction analyses. Among the BF cholinergic projection regions, decreased levels of GDNF mRNA and increased levels of TrkB mRNA were observed in the basal nucleus, and increased levels of TrkB mRNA were observed in the cerebral cortex. There were no significant alterations in the levels of expression of relevant neurotrophic factors in the septal nucleus and hippocampus. Given that neurotrophic factors function in retrograde/anterograde or autocrine/paracrine mechanisms and that BF cholinergic projection regions are neuroanatomically connected, these findings suggested that an imbalanced allocation of neurotrophic factor ligands and receptors is an initial phenomenon in alcohol addiction. The exact mechanisms underlying this phenomenon in the BF cholinergic system are unknown. However, our results provide a novel notion for the understanding of the initial processes in alcohol addiction.

Glial cell line-derived neurotrophic factor and endothelial cells promote self-renewal of rabbit germ cells with spermatogonial stem cell properties.

PubMed

Kubota, Hiroshi; Wu, Xin; Goodyear, Shaun M; Avarbock, Mary R; Brinster, Ralph L

2011-08-01

Previous studies suggest that exogenous factors crucial for spermatogonial stem cell (SSC) self-renewal are conserved among several mammalian species. Since glial cell line-derived neurotrophic factor (GDNF) and fibroblast growth factor 2 (FGF2) are critical for rodent SSC self-renewal, we hypothesized that they might promote self-renewal of nonrodent SSCs. Therefore, we cultured testicular germ cells from prepubertal rabbits in the presence of GDNF and FGF2 and found they proliferated indefinitely as cellular clumps that displayed characteristics previously identified for rodent SSCs. The rabbit germ cells could not be maintained on mouse embryonic fibroblast (STO) feeders that support rodent SSC self-renewal in vitro but were rather supported on mouse yolk sac-derived endothelial cell (C166) feeder layers. Proliferation of rabbit germ cells was dependent on GDNF. Of critical importance was that clump-forming rabbit germ cells colonized seminiferous tubules of immunodeficient mice, proliferated for at least 6 mo, while retaining an SSC phenotype in the testes of recipient mice, indicating that they were rabbit SSCs. This study demonstrates that GDNF is a mitogenic factor promoting self-renewal that is conserved between rodent and rabbit SSCs; with an evolutionary separation of ∼ 60 million years. These findings provide a foundation to study the mechanisms governing SSC self-renewal in nonrodent species.

NEUROTROPHIN SELECTIVITY IN ORGANIZING TOPOGRAPHIC REGENERATION OF NOCICEPTIVE AFFERENTS

PubMed Central

Kelamangalath, Lakshmi; Tang, Xiaoqing; Bezik, Kathleen; Sterling, Noelle; Son, Young-Jin; Smith, George M.

2015-01-01

Neurotrophins represent some of the best candidates to enhance regeneration. In the current study, we investigated the effects of artemin, a member of the glial derived neurotrophic factor (GDNF) family, on sensory axon regeneration following a lumbar dorsal root injury and compared these effects with that observed after either NGF or GDNF expression in the rat spinal cord. Unlike previously published data, artemin failed to induce regeneration of large-diameter myelinated sensory afferents when expressed within either the spinal cord or DRG. However, artemin or NGF induced regeneration of calcitonin gene related peptide positive (CGRP+) axons only when expressed within the spinal cord. Accordingly, artemin or NGF enhanced recovery of only nociceptive behavior and showed a cFos distribution similar to the topography of regenerating axons. Artemin and GDNF signaling requires binding to different co-receptors (GFRα3 or GFRα1, respectively) prior to binding to the signaling receptor, cRet. Approximately 70% of DRG neurons express cRet, but only 35% express either co-receptor. To enhance artemin-induced regeneration, we co-expressed artemin with either GFRα3 or GDNF. Co-expression of artemin and GFRα3 only slightly enhanced regeneration of IB4+ non-peptidergic nociceptive axons, but not myelinated axons. Interestingly, this co-expression also disrupted the ability of artemin to produce topographic targeting and lead to significant increases in cFos immunoreactivity within the deep dorsal laminae. This study failed to demonstrate artemin-induced regeneration of myelinated axons, even with co-expression of GFR-α3, which only promoted mistargeted regeneration. PMID:26054884

Neurotrophin selectivity in organizing topographic regeneration of nociceptive afferents.

PubMed

Kelamangalath, Lakshmi; Tang, Xiaoqing; Bezik, Kathleen; Sterling, Noelle; Son, Young-Jin; Smith, George M

2015-09-01

Neurotrophins represent some of the best candidates to enhance regeneration. In the current study, we investigated the effects of artemin, a member of the glial derived neurotrophic factor (GDNF) family, on sensory axon regeneration following a lumbar dorsal root injury and compared these effects with that observed after either NGF or GDNF expression in the rat spinal cord. Unlike previously published data, artemin failed to induce regeneration of large-diameter myelinated sensory afferents when expressed within either the spinal cord or DRG. However, artemin or NGF induced regeneration of calcitonin gene related peptide positive (CGRP(+)) axons only when expressed within the spinal cord. Accordingly, artemin or NGF enhanced recovery of only nociceptive behavior and showed a cFos distribution similar to the topography of regenerating axons. Artemin and GDNF signaling requires binding to different co-receptors (GFRα3 or GFRα1, respectively) prior to binding to the signaling receptor, cRet. Approximately 70% of DRG neurons express cRet, but only 35% express either co-receptor. To enhance artemin-induced regeneration, we co-expressed artemin with either GFRα3 or GDNF. Co-expression of artemin and GFRα3 only slightly enhanced regeneration of IB4(+) non-peptidergic nociceptive axons, but not myelinated axons. Interestingly, this co-expression also disrupted the ability of artemin to produce topographic targeting and lead to significant increases in cFos immunoreactivity within the deep dorsal laminae. This study failed to demonstrate artemin-induced regeneration of myelinated axons, even with co-expression of GFRα3, which only promoted mistargeted regeneration. Copyright © 2015 Elsevier Inc. All rights reserved.

Expression and functional studies of the GDNF family receptor-alpha3 (GFRα3) in the pancreas

PubMed Central

Nivlet, Laure; Herrmann, Joel; Martin, Delia Esteban; Meunier, Aline; Orvain, Christophe; Gradwohl, Gérard

2018-01-01

The generation of therapeutic β-cells from human pluripotent stem cells relies on the identification of growth factors that faithfully mimic pancreatic β-cell development in vitro. In this context, the aim of the study was to determine the expression and function of the Glial cell line derived neurotrophic factor receptor α 3 (GFRα3) and its ligand Artemin in islet cell development and function. GFRα3 and Artn expression were characterized by in situ hybridization, immunochemistry and qRT-PCR. We used GFRα3-deficient mice to study GFRα3 function and generated a transgenic mice overexpressing Artn in the embryonic pancreas to study Artn function. We found that GFRα3 is expressed at the surface of a subset of Ngn3-positive endocrine progenitors as well as of embryonic α- and β-cells, while Artn is found in the pancreatic mesenchyme. Adult β-cells lack GFRα3 but α-cells express the receptor. GFRα3 was also found in parasympathetic and sympathetic intra islets neurons as well as in glial cells in the embryonic and adult pancreas. The loss of GFRα3 or overexpression of Artn has no impact on Ngn3- and islet- cells formation and maintenance in the embryo. Islet organisation and innervation as well as glucose homeostasis is normal in GFRα3-deficient mice suggesting functional redundancy. PMID:26576643

Retinal expression of Fgf2 in RCS rats with subretinal microphotodiode array.

PubMed

Ciavatta, Vincent T; Kim, Moon; Wong, Paul; Nickerson, John M; Shuler, R Keith; McLean, George Y; Pardue, Machelle T

2009-10-01

To test the hypothesis that subretinal electrical stimulation from a microphotodiode array (MPA) exerts a neuroprotective effect in Royal College of Surgeons (RCS) rats through the induction of growth factors. At postnatal day 21, RCS rats were divided into four groups in which one eye per rat received treatment: (A) active MPA, (M) minimally active MPA, (S) sham surgery, or (C) no surgery and the opposite eye was unoperated. Dark- and light-adapted ERGs were recorded 1 week after surgery. A second set of A-, M-, and C-treated RCS rats had weekly ERG recordings for 4 weeks. Real-time RT-PCR was used to measure relative expression of mRNAs (Bdnf, Fgf2, Fgf1, Cntf, Gdnf, and Igf1) in retina samples collected 2 days after the final ERG. One week after surgery, there was a slight difference in dark-adapted ERG b-wave at the brightest flash intensity. Mean retinal Fgf2 expression in the treated eye relative to the opposite eye was greater for the A group (4.67 +/- 0.72) than for the M group (2.80 +/- 0.45; P = 0.0501), S group (2.03 +/- 0.45; P < 0.01), and C group (1.30 +/- 0.22; P < 0.001). No significant change was detected for Bdnf, Cntf, Fgf1, Gdnf, and Igf1. Four weeks after surgery, the A group had significantly larger dark- and light-adapted ERG b-waves than for the M and C groups (P < 0.01). Simultaneously, mean relative Fgf2 expression was again greater for the A group (3.28 +/- 0.61) than for the M (1.28 +/- 0.32; P < 0.05) and C (1.05 +/- 0.04; P < 0.05) groups. The results show subretinal implantation of an MPA induces selective expression of Fgf2 above that expected from a retina-piercing injury. Preservation of ERG b-wave amplitude 4 weeks after implantation is accompanied by elevated Fgf2 expression. These results suggest that Fgf2 may play a role in the neuroprotection provided by subretinal electrical stimulation.

Peritubular Myoid Cells Participate in Male Mouse Spermatogonial Stem Cell Maintenance

PubMed Central

Chen, Liang-Yu; Brown, Paula R.; Willis, William B.

2014-01-01

Peritubular myoid (PM) cells surround the seminiferous tubule and together with Sertoli cells form the cellular boundary of the spermatogonial stem cell (SSC) niche. However, it remains unclear what role PM cells have in determining the microenvironment in the niche required for maintenance of the ability of SSCs to undergo self-renewal and differentiation into spermatogonia. Mice with a targeted disruption of the androgen receptor gene (Ar) in PM cells experienced a progressive loss of spermatogonia, suggesting that PM cells require testosterone (T) action to produce factors influencing SSC maintenance in the niche. Other studies showed that glial cell line-derived neurotrophic factor (GDNF) is required for SSC self-renewal and differentiation of SSCs in vitro and in vivo. This led us to hypothesize that T-regulated GDNF expression by PM cells contributes to the maintenance of SSCs. This hypothesis was tested using an adult mouse PM cell primary culture system and germ cell transplantation. We found that T induced GDNF expression at the mRNA and protein levels in PM cells. Furthermore, when thymus cell antigen 1-positive spermatogonia isolated from neonatal mice were cocultured with PM cells with or without T and transplanted to the testes of germ cell-depleted mice, the number and length of transplant-derived colonies was increased considerably by in vitro T treatment. These results support the novel hypothesis that T-dependent regulation of GDNF expression in PM cells has a significant influence on the microenvironment of the niche and SSC maintenance. PMID:25181385

Update of Neurotrophic Factors in Neurobiology of Addiction and Future Directions

PubMed Central

Koskela, Maryna; Bäck, Susanne; Võikar, Vootele; Richie, Christopher T.; Domanskyi, Andrii; Harvey, Brandon K.; Airavaara, Mikko

2016-01-01

Drug addiction is a chronic brain disease and drugs of abuse cause long lasting neuroadaptations. Addiction is characterized by the loss of control over drug use despite harmful consequences, and high rates of relapse even after long periods of abstinence. Neurotrophic factors (NTFs) are well known for their actions on neuronal survival in the peripheral nervous system. Moreover, NTFs have been shown to be involved in synaptic plasticity in the brain. Brain-derived neurotrophic factor (BDNF) and glial cell line-derived neurotrophic factor (GDNF) are two of the most studied NTFs and both of them have been reported to increase craving when administered into the mesocorticolimbic dopaminergic system after drug self-administration. Here we review recent data on BDNF and GDNF functions in addiction-related behavior and discuss them in relation to previous findings. Finally, we give an insight into how new technologies could aid in further elucidating the role of these factors in drug addiction. PMID:27189755

Control of Aβ release from human neurons by differentiation status and RET signaling.

PubMed

Scholz, Diana; Chernyshova, Yana; Leist, Marcel

2013-01-01

Few studies have compared the processing of endogenous human amyloid precursor protein (APP) in younger and older neurons. Here, we characterized LUHMES cells as a human model to study Alzheimer's disease-related processes during neuronal maturation and aging. Differentiated LUHMES expressed and spontaneously processed APP via the secretase pathways, and they secreted amyloid β (Aβ) peptide. This was inhibited by cholesterol depletion or secretase inhibition, but not by block of tau phosphorylation. In vitro aged cells increased Aβ secretion without upregulation of APP or secretases. We identified the medium constituent glial cell line-derived neurotrophic factor (GDNF) as responsible for this effect. GDNF-triggered Aβ release was associated with rapid upregulation of the GDNF coreceptor "rearranged during transfection" (RET). Other direct (neurturin) or indirect (nerve growth factor) RET activators also increased Aβ, whereas different neurotrophins were ineffective. Downstream of RET, we found activation of protein kinase B (AKT) to be involved. Accordingly, inhibitors of the AKT regulator phosphatidylinositol-3-kinase completely blocked GDNF-triggered AKT phosphorylation and Aβ increase. This suggests that RET signaling affects Aβ release from aging neurons. Copyright © 2013 Elsevier Inc. All rights reserved.

Temporal profiles of age-dependent changes in cytokine mRNA expression and glial cell activation after status epilepticus in postnatal rat hippocampus.

PubMed

Järvelä, Juha T; Lopez-Picon, Francisco R; Plysjuk, Anna; Ruohonen, Saku; Holopainen, Irma E

2011-04-08

Status epilepticus (SE) is proposed to lead to an age-dependent acute activation of a repertoire of inflammatory processes, which may contribute to neuronal damage in the hippocampus. The extent and temporal profiles of activation of these processes are well known in the adult brain, but less so in the developing brain. We have now further elucidated to what extent inflammation is activated by SE by investigating the acute expression of several cytokines and subacute glial reactivity in the postnatal rat hippocampus. SE was induced by an intraperitoneal (i.p.) injection of kainic acid (KA) in 9- and 21-day-old (P9 and P21) rats. The mRNA expression of interleukin-1 beta (IL-1β), tumor necrosis factor-alpha (TNF-α), interleukin-10 (IL-10), matrix metalloproteinase-9 (MMP-9), glial-derived neurotrophic factor (GDNF), interferon gamma (IFN-γ), and transforming growth factor-beta 1 (TGF-β1) were measured from 4 h up to 3 days after KA injection with real-time quantitative PCR (qPCR). IL-1β protein expression was studied with ELISA, GFAP expression with western blotting, and microglial and astrocyte morphology with immunohistochemistry 3 days after SE. SE increased mRNA expression of IL-1β, TNF-α and IL-10 mRNA in hippocampus of both P9 and P21 rats, their induction being more rapid and pronounced in P21 than in P9 rats. MMP-9 expression was augmented similarly in both age groups and GDNF expression augmented only in P21 rats, whereas neither IFN-γ nor TGF-β1 expression was induced in either age group. Microglia and astrocytes exhibited activated morphology in the hippocampus of P21 rats, but not in P9 rats 3 d after SE. Microglial activation was most pronounced in the CA1 region and also detected in the basomedial amygdala. Our results suggest that SE provokes an age-specific cytokine expression in the acute phase, and age-specific glial cell activation in the subacute phase as verified now in the postnatal rat hippocampus. In the juvenile hippocampus

Use of GDNF-Releasing Nanofiber Nerve Guide Conduits for the Repair of Conus medullaris/Cauda Equina Injury in the Non-Human Primate

DTIC Science & Technology

2011-10-01

Cauda equina, non-human primate, ventral root. neural repair, electromyography , magnetic resonance imaging 16. SECURITY CLASSIFICATION OF: 17...of a guidance channel without GDNF release and a peripheral nerve graft to bridge the tissue gap. A comprehensive set of electrodiagnostic, imaging ... Electromyography (EMG) recordings of the external anal sphincter are obtained pre-operatively as baseline records. The external anal sphincter muscle

Evaluating the behavior of cultured sertoli cells in the presence and absence of spermatogonial stem cell

PubMed Central

Jabarpour, Masoome

2018-01-01

Background The complex process of spermatogenesis is regulated by various factors. Several studies have been conducted to proliferate cells involved in the spermatogenesis process, in culture by used growth factors, different hormones and feeder cells. This study was conducted to evaluate the role of Sertoli cells on gene expression of fibroblast growth factor (FGF2) and glial cell derived neurotrophic factor (GDNF) after removal of spermatogonial stem cells (SSCs) from the culture medium. Methods Following isolation, bovine SSCs were co-cultured with Sertoli cells and follicular stimulating hormone (FSH) for 12 days. In the treatment group, SSCs were removed from the culture medium; in the control group no intervention was done in the culture. Colony formation of SSCs was evaluated by using an inverted microscope. Then, the expression of factors genes were assessed by quantitative RT-PCR. Data was analyzed by using paired-samples t-test. Results The results showed that removal of SSCs led to the increase in expression of GDNF and FGF2. These findings suggest that loss of SSCs population or decline in its population leads to changing in behavior of somatic cells which forming niche and consequently stimulates self-renewal and inhibits differentiation of SSCs. Conclusions The present study showed that removal of SSCs from the culture medium could be a model for damage to SSCs; the results revealed that niche cells respond to SSCs removal by upregulation of FGF2 and GDNF to stimulate self-renewal of SSCs and abrogation of differentiation. PMID:29430457

GPER: A new tool to protect dopaminergic neurons?

PubMed

Bessa, Agustina; Campos, Filipa Lopes; Videira, Rita Alexandra; Mendes-Oliveira, Julieta; Bessa-Neto, Diogo; Baltazar, Graça

2015-10-01

Parkinson's disease (PD) is characterized by a selective degeneration of nigrostriatal dopaminergic pathway. Epidemiological studies revealed a male predominance of the disease that has been attributed to the female steroid hormones, mainly the estrogen. Estrogen neuroprotective effects have been shown in several studies, however the mechanisms responsible by these effects are still unclear. Previous data from our group revealed that glial cell line-derived neurotrophic factor (GDNF) is crucial to the dopaminergic protection provided by 17β-estradiol, and also suggest that the intracellular estrogen receptors (ERs) are not required for that neuroprotective effects. The present study aimed to investigate the contribution of the G protein-coupled ER (GPER) activation in estrogen-mediated dopaminergic neuroprotection against an insult induced by 1-methyl-4-phenylpyridinium (MPP(+)), and whether GPER neuroprotective effects involve the regulation of GDNF expression. Using primary mesencephalic cultures, we found that GPER activation protects dopaminergic neurons from MPP(+) toxicity in an extent similar to the promoted by a 17β-estradiol. Moreover, GPER activation promotes an increase in GDNF levels. Both, GDNF antibody neutralization or RNA interference-mediated GDNF knockdown prevented the GPER-mediated dopaminergic protection verified in mesencephalic cultures challenged with MPP(+). Overall, these results revealed that G1, a selective agonist of GPER, is able to protect dopaminergic neurons and that GDNF overexpression is a key feature to GPER induced the neuroprotective effects. Copyright © 2015 Elsevier B.V. All rights reserved.

Artemin growth factor increases nicotinic cholinergic receptor subunit expression and activity in nociceptive sensory neurons.

PubMed

Albers, Kathryn M; Zhang, Xiu Lin; Diges, Charlotte M; Schwartz, Erica S; Yang, Charles I; Davis, Brian M; Gold, Michael S

2014-05-22

Artemin (Artn), a member of the glial cell line-derived growth factor (GDNF) family, supports the development and function of a subpopulation of peptidergic, TRPV1-positive sensory neurons. Artn (enovin, neublastin) is elevated in inflamed tissue and its injection in skin causes transient thermal hyperalgesia. A genome wide expression analysis of trigeminal ganglia of mice that overexpress Artn in the skin (ART-OE mice) showed elevation in nicotinic acetylcholine receptor (nAChR) subunits, suggesting these ion channels contribute to Artn-induced sensitivity. Here we have used gene expression, immunolabeling, patch clamp electrophysiology and behavioral testing assays to investigate the link between Artn, nicotinic subunit expression and thermal hypersensitivity. Reverse transcriptase-PCR validation showed increased levels of mRNAs encoding the nAChR subunits α3 (13.3-fold), β3 (4-fold) and β4 (7.7-fold) in trigeminal ganglia and α3 (4-fold) and β4 (2.8-fold) in dorsal root ganglia (DRG) of ART-OE mice. Sensory ganglia of ART-OE mice had increased immunoreactivity for nAChRα3 and exhibited increased overlap in labeling with GFRα3-positive neurons. Patch clamp analysis of back-labeled cutaneous afferents showed that while the majority of nicotine-evoked currents in DRG neurons had biophysical and pharmacological properties of α7-subunit containing nAChRs, the Artn-induced increase in α3 and β4 subunits resulted in functional channels. Behavioral analysis of ART-OE and wildtype mice showed that Artn-induced thermal hyperalgesia can be blocked by mecamylamine or hexamethonium. Complete Freund's adjuvant (CFA) inflammation of paw skin, which causes an increase in Artn in the skin, also increased the level of nAChR mRNAs in DRG. Finally, the increase in nAChRs transcription was not dependent on the Artn-induced increase in TRPV1 or TRPA1 in ART-OE mice since nAChRs were elevated in ganglia of TRPV1/TRPA1 double knockout mice. These findings suggest that Artn

A novel GLP-1/GIP dual agonist is more effective than liraglutide in reducing inflammation and enhancing GDNF release in the MPTP mouse model of Parkinson's disease.

PubMed

Yuan, Ziyue; Li, Dongfang; Feng, Peng; Xue, Guofang; Ji, Chenhui; Li, Guanglai; Hölscher, Christian

2017-10-05

Type 2 diabetes mellitus (T2DM) is one of the risk factors for Parkinson's disease (PD). Insulin desensitisation has been observed in the brains of patients, which may promote neurodegeneration. Incretins are a family of growth factors that can re-sensitise insulin signalling. We have previously shown that mimetics of glucagon-like peptide-1 (GLP-1) and glucose-dependent insulinotropic polypeptide (GIP) have neuroprotective effects in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropypridine (MPTP) mouse model of PD. Recently, dual GLP-1/GIP receptor agonists have been developed. We therefore tested the novel dual agonist DA3-CH in comparison with the best GLP-1 analogue currently on the market, liraglutide (both drugs 25nmol/kg ip once-daily for 7 days) in the MPTP mouse model of PD (25mg/kg ip once-daily for 7 days). In the Rotarod and grip strength assessment, DA3-CH was superior to liraglutide in reversing the MPTP-induced motor impairment. Dopamine synthesis as indicated by levels of tyrosine hydroxylase was much reduced by MPTP in the substantia nigra and striatum, and DA3-CH reversed this while liragutide only partially reversed this. The chronic inflammation response as shown in increased levels of activated microglia and astrocytes was reduced by both drugs. Importantly, expression levels of the neuroprotective growth factor Glial Derived Neurotrophic Factor (GDNF) was much enhanced by both DA3-CH and liragutide. The results demonstrate that the combination of GLP-1 and GIP receptor activation is superior to single GLP-1 receptor activation alone. Therefore, new dual agonists may be a promising treatment for PD. The GLP-1 receptor agonist exendin-4 has already shown disease modifying effects in clinical trials in PD patients. Copyright © 2017 Elsevier B.V. All rights reserved.

Optimizing neurotrophic factor combinations for neurite outgrowth

NASA Astrophysics Data System (ADS)

Deister, C.; Schmidt, C. E.

2006-06-01

Most neurotrophic factors are members of one of three families: the neurotrophins, the glial cell-line derived neurotrophic factor family ligands (GFLs) and the neuropoietic cytokines. Each family activates distinct but overlapping cellular pathways. Several studies have shown additive or synergistic interactions between neurotrophic factors from different families, though generally only a single combination has been studied. Because of possible interactions between the neurotrophic factors, the optimum concentration of a factor in a mixture may differ from the optimum when applied individually. Additionally, the effect of combinations of neurotrophic factors from each of the three families on neurite extension is unclear. This study examines the effects of several combinations of the neurotrophin nerve growth factor (NGF), the GFL glial cell-line derived neurotrophic factor (GDNF) and the neuropoietic cytokine ciliary neurotrophic factor (CNTF) on neurite outgrowth from young rat dorsal root ganglion (DRG) explants. The combination of 50 ng ml-1 NGF and 10 ng ml-1 of each GDNF and CNTF induced the highest level of neurite outgrowth at a 752 ± 53% increase over untreated DRGs and increased the longest neurite length to 2031 ± 97 µm compared to 916 ± 64 µm for untreated DRGs. The optimum concentrations of the three factors applied in combination corresponded to the optimum concentration of each factor when applied individually. These results indicate that the efficacy of future therapies for nerve repair would be enhanced by the controlled release of a combination of neurotrophins, GFLs and neuropoietic cytokines at higher concentrations than used in previous conduit designs.

Rapid Modulation of Protein Expression in the Rat Hippocampus Following Deep Brain Stimulation of the Fornix.

PubMed

Gondard, Elise; Chau, Hien N; Mann, Amandeep; Tierney, Travis S; Hamani, Clement; Kalia, Suneil K; Lozano, Andres M

2015-01-01

The forniceal area is currently being evaluated as a target for deep brain stimulation (DBS) to improve cognitive function in patients with Alzheimer's disease. The molecular changes at downstream targets within the stimulated circuit are unknown. To analyze the modulation of hippocampal protein expression following 1 h of fornix DBS in the rat. Animals underwent bilateral forniceal DBS for 1 h and sacrificed at different time-points after the initiation of the stimulation (1 h, 2.5 h, 5 h, 25 h). Bilateral hippocampi were isolated for western blot analyses. Forniceal DBS led to a dramatic elevation of cFos post-stimulation, suggesting that forniceal DBS activates the hippocampus. There was also a significant increase in candidate proteins including several trophic factors, such as brain derived neurotrophic factor (BDNF) and vascular endothelial growth factor (VEGF) but not glial cell-derived neurotrophic factor (GDNF). There was in addition, increased expression of the synaptic markers growth associated protein 43 (GAP-43), synaptophysin and α-synuclein. No changes were observed at the studied time-points in Alzheimer's-related proteins including amyloid precursor protein (APP), tau, phosphorylated tau (ptau), or selected chaperone proteins (HSP40, HSP70 and CHIP). Forniceal DBS triggers hippocampal activity and rapidly modulate the expression of neurotrophic factors and markers of synaptic plasticity known to play key roles in memory processing. The clinical effects of DBS of the fornix may, in part, be mediated by producing changes in the expression of these proteins. Copyright © 2015 Elsevier Inc. All rights reserved.

The hormone prolactin is a novel, endogenous trophic factor able to regulate reactive glia and to limit retinal degeneration.

PubMed

Arnold, Edith; Thebault, Stéphanie; Baeza-Cruz, German; Arredondo Zamarripa, David; Adán, Norma; Quintanar-Stéphano, Andrés; Condés-Lara, Miguel; Rojas-Piloni, Gerardo; Binart, Nadine; Martínez de la Escalera, Gonzalo; Clapp, Carmen

2014-01-29

Retinal degeneration is characterized by the progressive destruction of retinal cells, causing the deterioration and eventual loss of vision. We explored whether the hormone prolactin provides trophic support to retinal cells, thus protecting the retina from degenerative pressure. Inducing hyperprolactinemia limited photoreceptor apoptosis, gliosis, and changes in neurotrophin expression, and it preserved the photoresponse in the phototoxicity model of retinal degeneration, in which continuous exposure of rats to bright light leads to retinal cell death and retinal dysfunction. In this model, the expression levels of prolactin receptors in the retina were upregulated. Moreover, retinas from prolactin receptor-deficient mice exhibited photoresponsive dysfunction and gliosis that correlated with decreased levels of retinal bFGF, GDNF, and BDNF. Collectively, these data unveiled prolactin as a retinal trophic factor that may regulate glial-neuronal cell interactions and is a potential therapeutic molecule against retinal degeneration.

Effects of postnatal ethanol exposure at different developmental phases on neurotrophic factors and phosphorylated proteins on signal transductions in rat brain.

PubMed

Tsuji, Ryozo; Fattori, Vittorio; Abe, Shin-ichi; Costa, Lucio G; Kobayashi, Kumiko

2008-01-01

Exposure to ethanol during development induces severe brain damage resulting in a number of CNS dysfunctions including microencephaly and mental retardation in humans and in laboratory animals. The most vulnerable period to ethanol neurotoxicity coincides with the peak of brain growth spurt. Recently, neurotrophic factors and/or their signal transduction pathways have been reported as a potential relevant target for the developmental neurotoxicity of ethanol. The present studies were designed to investigate the effects of ethanol given in various developmental phases during the brain growth spurt in rats. Rat pups were assigned to the three treatment groups and treated with 5 g/kg of ethanol for three days, on postnatal days (PND) 2-4, 6-8 or 13-15. Whole brain weights were reduced only in the PND 6-8 group concurrently with the reduction of GDNF mRNA in cortex in this group. BDNF mRNA expression was reduced in both the PND 6-8 and 13-15 groups, while mRNA expressions of NT-3 and NGF were unchanged in all three groups. Phospho-Akt level was mostly reduced in the PND 6-8 group. Both phospho-MAPK and p-70S6 kinase levels were decreased in all groups whereas no changes were observed in either phospho-PKCzeta or CREB level. The phosphorylation of Akt was immediately inhibited after single administration of ethanol, and its inhibition was correlated with variations in blood ethanol concentration. These findings suggest that GDNF and the phosphorylation of Akt play a possible key role in the ethanol-induced developmental neurotoxicity.

Agavins Increase Neurotrophic Factors and Decrease Oxidative Stress in the Brains of High-Fat Diet-Induced Obese Mice.

PubMed

Franco-Robles, Elena; López, Mercedes G

2016-08-02

Fructans obtained from agave, called agavins, have recently shown significant benefits for human health including obesity. Therefore, we evaluated the potential of agavins as neuroprotectors and antioxidants by determining their effect on brain-derived neurotrophic factor (BDNF) and glial-derived neurotrophic factor (GDNF) as well as oxidative brain damage in of obese mice. Male C57BL/6J mice were fed a high-fat diet (HFD) and treated daily with 5% (HFD/A5) or 10% (HFD/A10) of agavins or a standard diet (SD) for 10 weeks. The levels of BDNF and GDNF were evaluated by ELISA. The oxidative stress was evaluated by lipid peroxidation (TBARS) and carbonyls. SCFAs were also measured with GC-FID. Differences between groups were assessed using ANOVA and by Tukey's test considering p < 0.05. The body weight gain and food intake of mice HFD/A10 group were significantly lower than those in the HFD group. Agavins restored BDNF levels in HFD/A5 group and GDNF levels of HFD/A5 and HFD/A10 groups in cerebellum. Interestingly, agavins decreased TBARS levels in HFD/A5 and HFD/A10 groups in the hippocampus, frontal cortex and cerebellum. Carbonyl levels were also lower in HFD/A5 and HFD/A10 for only the hippocampus and cerebellum. It was also found that agavins enhanced SCFAs production in feces. Agavins may act as bioactive ingredients with antioxidant and protective roles in the brain.

Glial cell line-derived neurotrophic factor in combination with insulin-like growth factor 1 and basic fibroblast growth factor promote in vitro culture of goat spermatogonial stem cells.

PubMed

Bahadorani, M; Hosseini, S M; Abedi, P; Abbasi, H; Nasr-Esfahani, M H

2015-01-01

Growth factors are increasingly considered as important regulators of spermatogonial stem cells (SSCs). This study investigated the effects of various growth factors (GDNF, IGF1, bFGF, EGF and GFRalpha-1) on purification and colonization of undifferentiated goat SSCs under in vitro and in vivo conditions. Irrespective of the culture condition used, the first signs of developing colonies were observed from day 4 of culture onwards. The number of colonies developed in GDNF + IGF1 + bFGF culture condition was significantly higher than the other groups (p < 0.05). In contrast, the size of colonies developed in GDNF + EGF + LIF culture condition was significantly higher than the other groups (p < 0.05). Immunocytochemical stationing for specific biomarkers of somatic cells (vimentin, alpha-inhibin and α-SMA) and spermatogonial cells (PLZF, THY 1, VASA, alpha-1 integrin, bet-1 integrin and DBA) revealed that both cell types existed in developing colonies, irrespective of the culture condition used. Even though, the relative abundance of VASA, FGFR3, OCT4, PLZF, BCL6B and THY1 transcription factors in GDNF + IGF1 + bFGF treatment group was significantly higher than the other groups (p < 0.05). Additionally, goat SSCs developed in the latter culture condition could colonize within the seminiferous tubules of the germ-cell depleted recipient mice following xenotransplantation. Obtained results demonstrated that combination of GDNF with IGF1 and bFGF promote in vitro culture of goat SSCs while precludes uncontrolled proliferation of somatic cells.

The lack of CD131 and the inhibition of Neuro-2a growth by carbamylated erythropoietin.

PubMed

Ding, Jing; Li, Qin-Ying; Yu, Jie-Zhong; Wang, Xin; Lu, Chuan-Zhen; Ma, Cun-Gen; Xiao, Bao-Guo

2015-02-01

Recombinant human erythropoietin (EPO), a glycohormone, is one of the leading biopharmaceutical products, while carbamylated erythropoietin (CEPO), an EPO derivative, is attracting widespread interest due to its neuroprotective effects without erythropoiesis in several cells and animal models. However, exogenous EPO promotes an angiogenic response from tumor cells and is associated with tumor growth, but knowledge of CEPO on tumor growth is lacking. Here we show that CEPO, but not EPO, inhibited Neuro-2a growth and viability. As expected, CEPO--unlike EPO--did not activate JAK-2 either in primary neurons or in Neuro-2a cells. Interestingly, CEPO did not induce GDNF expression and subsequent AKT activation in Neuro-2a cells. Before CEPO/EPO treatment, glial cell line-derived neurotrophic factor (GDNF) neutralization and GFR receptor blocking decreased the viability of EPO-treated Neuro-2a cells but did not influence CEPO-treated Neuro-2a cells. As compared to primary neurons, the expression of CD131, as a receptor complex binding to CEPO, is almost lacking in Neuro-2a cells. In BABL/C-nu mice, CEPO did not promote the growth of Neuro-2a cells nor extended the survival time compared to mice treated with EPO. The results indicate that CEPO did not promote tumor growth because of lower expression of CD131 and subsequent dysfunction of CD131/GDNF/AKT pathway in Neuro-2a cells, revealing its therapeutic potential in future clinical application.

Neurotrophic factors as a therapeutic target for Parkinson's disease.

PubMed

Evans, Jonathan R; Barker, Roger A

2008-04-01

The search for therapeutic agents that might alter the disease course in Parkinson's disease (PD) is ongoing. One area of particular interest involves neurotrophic factors (NTFs), with those of the glial cell line-derived neurotrophic factor (GDNF) family showing greatest promise. The safety and efficacy of these therapies has recently come into question. Furthermore, many of the key questions pertaining to such therapies, such as the optimal method of delivery, timing of treatment and selection of patients most likely to benefit, remain unanswered. In this review we sought to evaluate the therapeutic potential of NTFs in the treatment of PD. We appraised the evidence provided by both in vitro and in vivo work before proceeding to a critical assessment of the relevant clinical trial data. Relevant literature was identified using a PubMed search of articles published up to October 2007. Search terms included: 'Parkinson's disease', 'Neurotrophic factors', 'BDNF' (Brain-derived neurotrophic factor), 'GDNF' and 'Neurturin'. Original articles were reviewed, and relevant citations from these articles were also appraised. NTF therapy has potential in the treatment of nigrostriatal dysfunction in PD but numerous methodological and safety issues will need to be addressed before this approach can be widely adopted. Furthermore PD is now recognized as being more than a pure motor disorder, and one in which neuronal loss is not just confined to the dopaminergic nigrostriatal system. Non-motor symptomatology in PD is unlikely to benefit from therapies that target only the nigrostriatal system, and this must inform our thinking as to the maximal achievable benefit that NTFs are ever likely to provide.

Role of Wnt5a-Ror2 Signaling in Morphogenesis of the Metanephric Mesenchyme during Ureteric Budding

PubMed Central

Qiao, Sen; Miyamoto, Mari; Okinaka, Yuka; Yamada, Makiko; Hashimoto, Ryuju; Iijima, Kazumoto; Otani, Hiroki; Hartmann, Christine; Nishinakamura, Ryuichi

2014-01-01

Development of the metanephric kidney begins with the induction of a single ureteric bud (UB) on the caudal Wolffian duct (WD) in response to GDNF (glial cell line-derived neurotrophic factor) produced by the adjacent metanephric mesenchyme (MM). Mutual interaction between the UB and MM maintains expression of GDNF in the MM, thereby supporting further outgrowth and branching morphogenesis of the UB, while the MM also grows and aggregates around the branched tips of the UB. Ror2, a member of the Ror family of receptor tyrosine kinases, has been shown to act as a receptor for Wnt5a to mediate noncanonical Wnt signaling. We show that Ror2 is predominantly expressed in the MM during UB induction and that Ror2- and Wnt5a-deficient mice exhibit duplicated ureters and kidneys due to ectopic UB induction. During initial UB formation, these mutant embryos show dysregulated positioning of the MM, resulting in spatiotemporally aberrant interaction between the MM and WD, which provides the WD with inappropriate GDNF signaling. Furthermore, the numbers of proliferating cells in the mutant MM are markedly reduced compared to the wild-type MM. These results indicate an important role of Wnt5a-Ror2 signaling in morphogenesis of the MM to ensure proper epithelial tubular formation of the UB required for kidney development. PMID:24891614

Effects of brain-derived and glial cell line-derived neurotrophic factors on startle response and disrupted prepulse inhibition in mice of DBA/2J inbred strain.

PubMed

Naumenko, Vladimir S; Bazovkina, Daria V; Morozova, Maryana V; Popova, Nina K

2013-08-29

Prepulse inhibition (PPI), the reduction in acoustic startle reflex when it is preceded by weak prepulse stimuli, is a measure of critical to normal brain functioning sensorimotor gating. PPI deficit was shown in a variety of psychiatric disorders including schizophrenia, and in DBA/2J mouse strain. In the current study, we examined the effects of brain-derived (BDNF) and glial cell line-derived (GDNF) neurotrophic factors on acoustic startle response and PPI in DBA/2J mice. It was found that BDNF (300 ng, i.c.v.) significantly increased amplitude of startle response and restored disrupted PPI in 7 days after acute administration. GDNF (800 ng, i.c.v.) did not produce significant alteration neither in amplitude of startle response nor in PPI in DBA/2J mice. The reversal effect of BDNF on PPI deficit was unusually long-lasting: significant increase in PPI was found 1.5 months after single acute BDNF administration. Long-term ameliorative effect BDNF on disrupted PPI suggested the implication of epigenetic mechanism in BDNF action on neurogenesis. BDNF rather than GDNF could be a perspective drug for the treatment of sensorimotor gating impairments. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

The regulation of trefoil factor 2 expression by the transcription factor Sp3.

PubMed

Liu, Jingjing; Wang, Xu; Cai, Yiling; Zhou, Jingping; Guleng, Bayasi; Shi, Huaxiu; Ren, Jianlin

2012-10-19

Trefoil factor family 2 (TFF2) participates in mucus stabilization and repair, apoptosis, and inflammatory responses. Previously published reports have indicated that several growth factors and basal transcription factors are associated with the expression of TFF2. However, the detailed mechanisms that regulate TFF2 expression are not fully understood. The present study was designed to assess the essential role of the transcription factor SP3 with respect to TFF2 expression. We first demonstrated that there was a negative correlation between the expression levels of SP3 and TFF2. Thus, in the examined cells, the overexpression of SP3 decreased the expression level of TFF2, whereas the inhibition of SP3 increased the expression level of TFF2. Moreover, we discovered two GC boxes in the TFF2 promoter and confirmed the specific binding of SP3 to this promoter. On the whole, this study indicated that Sp3 was a major regulator of TFF2 expression. This knowledge should contribute to our understanding of the role that is played by SP3 in the regulation of TFF2 expression. Copyright © 2012 Elsevier Inc. All rights reserved.

Expression of virulence factors by Staphylococcus aureus grown in serum.

PubMed

Oogai, Yuichi; Matsuo, Miki; Hashimoto, Masahito; Kato, Fuminori; Sugai, Motoyuki; Komatsuzawa, Hitoshi

2011-11-01

Staphylococcus aureus produces many virulence factors, including toxins, immune-modulatory factors, and exoenzymes. Previous studies involving the analysis of virulence expression were mainly performed by in vitro experiments using bacterial medium. However, when S. aureus infects a host, the bacterial growth conditions are quite different from those in a medium, which may be related to the different expression of virulence factors in the host. In this study, we investigated the expression of virulence factors in S. aureus grown in calf serum. The expression of many virulence factors, including hemolysins, enterotoxins, proteases, and iron acquisition factors, was significantly increased compared with that in bacterial medium. In addition, the expression of RNA III, a global regulon for virulence expression, was significantly increased. This effect was partially restored by the addition of 300 μM FeCl₃ into serum, suggesting that iron depletion is associated with the increased expression of virulence factors in serum. In chemically defined medium without iron, a similar effect was observed. In a mutant with agr inactivated grown in serum, the expression of RNA III, psm, and sec4 was not increased, while other factors were still induced in the mutant, suggesting that another regulatory factor(s) is involved. In addition, we found that serum albumin is a major factor for the capture of free iron to prevent the supply of iron to bacteria grown in serum. These results indicate that S. aureus expresses virulence factors in adaptation to the host environment.

The histone code reader SPIN1 controls RET signaling in liposarcoma

PubMed Central

Franz, Henriette; Greschik, Holger; Willmann, Dominica; Ozretić, Luka; Jilg, Cordula Annette; Wardelmann, Eva; Jung, Manfred; Buettner, Reinhard; Schüle, Roland

2015-01-01

The histone code reader Spindlin1 (SPIN1) has been implicated in tumorigenesis and tumor growth, but the underlying molecular mechanisms remain poorly understood. Here, we show that reducing SPIN1 levels strongly impairs proliferation and increases apoptosis of liposarcoma cells in vitro and in xenograft mouse models. Combining signaling pathway, genome-wide chromatin binding, and transcriptome analyses, we found that SPIN1 directly enhances expression of GDNF, an activator of the RET signaling pathway, in cooperation with the transcription factor MAZ. Accordingly, knockdown of SPIN1 or MAZ results in reduced levels of GDNF and activated RET explaining diminished liposarcoma cell proliferation and survival. In line with these observations, levels of SPIN1, GDNF, activated RET, and MAZ are increased in human liposarcoma compared to normal adipose tissue or lipoma. Importantly, a mutation of SPIN1 within the reader domain interfering with chromatin binding reduces liposarcoma cell proliferation and survival. Together, our data describe a molecular mechanism for SPIN1 function in liposarcoma and suggest that targeting SPIN1 chromatin association with small molecule inhibitors may represent a novel therapeutic strategy. PMID:25749382

Comparative study of the distribution of the alpha-subunits of voltage-gated sodium channels in normal and axotomized rat dorsal root ganglion neurons.

PubMed

Fukuoka, Tetsuo; Kobayashi, Kimiko; Yamanaka, Hiroki; Obata, Koichi; Dai, Yi; Noguchi, Koichi

2008-09-10

We compared the distribution of the alpha-subunit mRNAs of voltage-gated sodium channels Nav1.1-1.3 and Nav1.6-1.9 and a related channel, Nax, in histochemically identified neuronal subpopulations of the rat dorsal root ganglia (DRG). In the naïve DRG, the expression of Nav1.1 and Nav1.6 was restricted to A-fiber neurons, and they were preferentially expressed by TrkC neurons, suggesting that proprioceptive neurons possess these channels. Nav1.7, -1.8, and -1.9 mRNAs were more abundant in C-fiber neurons compared with A-fiber ones. Nax was evenly expressed in both populations. Although Nav1.8 and -1.9 were preferentially expressed by TrkA neurons, other alpha-subunits were expressed independently of TrkA expression. Actually, all IB4(+) neurons expressed both Nav1.8 and -1.9, and relatively limited subpopulations of IB4(+) neurons (3% and 12%, respectively) expressed Nav1.1 and/or Nav1.6. These findings provide useful information in interpreting the electrophysiological characteristics of some neuronal subpopulations of naïve DRG. After L5 spinal nerve ligation, Nav1.3 mRNA was up-regulated mainly in A-fiber neurons in the ipsilateral L5 DRG. Although previous studies demonstrated that nerve growth factor (NGF) and glial cell-derived neurotrophic factor (GDNF) reversed this up-regulation, the Nav1.3 induction was independent of either TrkA or GFRalpha1 expression, suggesting that the induction of Nav1.3 may be one of the common responses of axotomized DRG neurons without a direct relationship to NGF/GDNF supply. (c) 2008 Wiley-Liss, Inc.

The effect of long-term hindlimb unloading on the expression of risk neurogenes encoding elements of serotonin-, dopaminergic systems and apoptosis; comparison with the effect of actual spaceflight on mouse brain.

PubMed

Kulikova, E A; Kulikov, V A; Sinyakova, N A; Kulikov, A V; Popova, N K

2017-02-15

The study of spaceflight effects on the brain is technically complex concern; complicated by the problem of applying an adequate ground model. The most-widely used experimental model to study the effect of microgravity is the tail-suspension hindlimb unloading model; however, its compliance with the effect of actual spaceflight on the brain is still unclear. We evaluated the effect of one month hindlimb unloading on the expression of genes related to the brain neuroplasticity-brain neutotrophic factors (Gdnf, Cdnf), apoptotic factors (Bcl-xl, Bax), serotonin- and dopaminergic systems (5-HT 2A , Maoa, Maob, Th, D1r, Comt), and compared the results with the data obtained on mice that spent one month in spaceflight on Russian biosatellite Bion-M1. No effect of hindlimb unloading was observed on the expression of most genes, which were considered as risk neurogenes for long-term actual spaceflight. The opposite effect of hindlimb unloading and spaceflight was found on the level of mRNA of D1 dopamine receptor and catechol-O-methyltransferase in the striatum. At the same time, the expression of Maob in the midbrain decreased, and the expression of Bcl-xl genes increased in the hippocampus, which corresponds to the effect of spaceflight. However, the hindlimb unloading model failed to reproduce the majority of effects of long-term spaceflight on serotonin-, dopaminergic systems and some apoptotic factors. Copyright © 2017 Elsevier B.V. All rights reserved.

Hox11 paralogous genes are essential for metanephric kidney induction

PubMed Central

Wellik, Deneen M.; Hawkes, Patrick J.; Capecchi, Mario R.

2002-01-01

The mammalian Hox complex is divided into four linkage groups containing 13 sets of paralogous genes. These paralogous genes have retained functional redundancy during evolution. For this reason, loss of only one or two Hox genes within a paralogous group often results in incompletely penetrant phenotypes which are difficult to interpret by molecular analysis. For example, mice individually mutant for Hoxa11 or Hoxd11 show no discernible kidney abnormalities. Hoxa11/Hoxd11 double mutants, however, demonstrate hypoplasia of the kidneys. As described in this study, removal of the last Hox11 paralogous member, Hoxc11, results in the complete loss of metanephric kidney induction. In these triple mutants, the metanephric blastema condenses, and expression of early patterning genes, Pax2 and Wt1, is unperturbed. Eya1 expression is also intact. Six2 expression, however, is absent, as is expression of the inducing growth factor, Gdnf. In the absence of Gdnf, ureteric bud formation is not initiated. Molecular analysis of this phenotype demonstrates that Hox11 control of early metanephric induction is accomplished by the interaction of Hox11 genes with the pax-eya-six regulatory cascade, a pathway that may be used by Hox genes more generally for the induction of multiple structures along the anteroposterior axis. PMID:12050119

Hox11 paralogous genes are essential for metanephric kidney induction.

PubMed

Wellik, Deneen M; Hawkes, Patrick J; Capecchi, Mario R

2002-06-01

The mammalian Hox complex is divided into four linkage groups containing 13 sets of paralogous genes. These paralogous genes have retained functional redundancy during evolution. For this reason, loss of only one or two Hox genes within a paralogous group often results in incompletely penetrant phenotypes which are difficult to interpret by molecular analysis. For example, mice individually mutant for Hoxa11 or Hoxd11 show no discernible kidney abnormalities. Hoxa11/Hoxd11 double mutants, however, demonstrate hypoplasia of the kidneys. As described in this study, removal of the last Hox11 paralogous member, Hoxc11, results in the complete loss of metanephric kidney induction. In these triple mutants, the metanephric blastema condenses, and expression of early patterning genes, Pax2 and Wt1, is unperturbed. Eya1 expression is also intact. Six2 expression, however, is absent, as is expression of the inducing growth factor, Gdnf. In the absence of Gdnf, ureteric bud formation is not initiated. Molecular analysis of this phenotype demonstrates that Hox11 control of early metanephric induction is accomplished by the interaction of Hox11 genes with the pax-eya-six regulatory cascade, a pathway that may be used by Hox genes more generally for the induction of multiple structures along the anteroposterior axis.

RNA Expression Profiling Reveals Differentially Regulated Growth Factor and Receptor Expression in Redirected Cancer Cells.

PubMed

Schmucker, Hannah S; Park, Jang Pyo; Coissieux, Marie-May; Bentires-Alj, Mohamed; Feltus, F Alex; Booth, Brian W

2017-05-01

Tumorigenic cells can be redirected to adopt a normal phenotype when transplanted into cleared mammary fat pads of juvenile female mice in specific ratios with normal epithelial cells. The redirected tumorigenic cells enter stem cell niches and provide progeny that differentiate into all mammary epithelial subtypes. We have developed an in vitro model that mimics the in vivo phenomenon. The shift in phenotype to redirection should be accomplished through a return to a normal gene expression state. To measure this shift, we interrogated the transcriptome of various in vitro model states in search for casual genes. For this study, expression of growth factors, cytokines, and their associated receptors was examined. In all, we queried 251 growth factor and cytokine-related genes. We found numerous growth factor and cytokine genes whose expression levels switched from expression levels seen in cancer cells to expression levels observed in normal cells. The comparisons of gene expression between normal mammary epithelial cells, tumor-derived cells, and redirected cancer cells have revealed insight into active and inactive growth factors and cytokines in cancer cell redirection.

Absence of Ret Signaling in Mice Causes Progressive and Late Degeneration of the Nigrostriatal System

PubMed Central

Kramer, Edgar R; Aron, Liviu; Ramakers, Geert M. J; Seitz, Sabine; Zhuang, Xiaoxi; Beyer, Klaus; Smidt, Marten P; Klein, Rüdiger

2007-01-01

Support of ageing neurons by endogenous neurotrophic factors such as glial cell line–derived neurotrophic factor (GDNF) and brain-derived neurotrophic factor (BDNF) may determine whether the neurons resist or succumb to neurodegeneration. GDNF has been tested in clinical trials for the treatment of Parkinson disease (PD), a common neurodegenerative disorder characterized by the loss of midbrain dopaminergic (DA) neurons. BDNF modulates nigrostriatal functions and rescues DA neurons in PD animal models. The physiological roles of GDNF and BDNF signaling in the adult nigrostriatal DA system are unknown. We generated mice with regionally selective ablations of the genes encoding the receptors for GDNF (Ret) and BDNF (TrkB). We find that Ret, but not TrkB, ablation causes progressive and adult-onset loss of DA neurons specifically in the substantia nigra pars compacta, degeneration of DA nerve terminals in striatum, and pronounced glial activation. These findings establish Ret as a critical regulator of long-term maintenance of the nigrostriatal DA system and suggest conditional Ret mutants as useful tools for gaining insights into the molecular mechanisms involved in the development of PD. PMID:17298183

MicroRNA-202 maintains spermatogonial stem cells by inhibiting cell cycle regulators and RNA binding proteins

PubMed Central

Chen, Jian; Cai, Tanxi; Zheng, Chunwei; Lin, Xiwen; Wang, Guojun; Liao, Shangying; Wang, Xiuxia; Gan, Haiyun; Zhang, Daoqin; Hu, Xiangjing; Wang, Si; Li, Zhen; Feng, Yanmin

2017-01-01

Abstract miRNAs play important roles during mammalian spermatogenesis. However, the function of most miRNAs in spermatogenesis and the underlying mechanisms remain unknown. Here, we report that miR-202 is highly expressed in mouse spermatogonial stem cells (SSCs), and is oppositely regulated by Glial cell-Derived Neurotrophic Factor (GDNF) and retinoic acid (RA), two key factors for SSC self-renewal and differentiation. We used inducible CRISPR-Cas9 to knockout miR-202 in cultured SSCs, and found that the knockout SSCs initiated premature differentiation accompanied by reduced stem cell activity and increased mitosis and apoptosis. Target genes were identified with iTRAQ-based proteomic analysis and RNA sequencing, and are enriched with cell cycle regulators and RNA-binding proteins. Rbfox2 and Cpeb1 were found to be direct targets of miR-202 and Rbfox2 but not Cpeb1, is essential for the differentiation of SSCs into meiotic cells. Accordingly, an SSC fate-regulatory network composed of signaling molecules of GDNF and RA, miR-202 and diverse downstream effectors has been identified. PMID:27998933

[Effect of cryotherapy over the expression of vascular endothelial growth factor and pigment epithelium-derived factor].

PubMed

Toscano-Garibay, Julia Dolores; Quiroz-Mercado, Hugo; Espitia-Pinzón, Clara; Gil-Carrasco, Félix; Flores-Estrada, José Javier

2014-01-01

Cryotherapy is a no invasive technique that uses intense cold to freeze and destroy cancer tissues. There are no descriptions of its effects over the expression of vascular endothelial growth factor and pigment epithelium-derived factor. Experimental study in cryogenic spot were applied in the right sclera of twelve pigs for ten minutes. Other 3 pigs were used as normal controls. Animals were sacrificed at 7, 14 and 21 and the tissues of choriodes and retina were dissected in areas of approximately 1 cm2 surrounding cryogenic spots. Expression levels of vascular endothelial growth factor and pigment epithelium-derived factor were determined analyzed using polymerase chain reaction coupled to reverse-transcription. Vascular endothelial growth factor was significantly downregulated (24%, p< 0.05) seven days post-treatment meanwhile pigment epithelium-derived factor levels increased 44.8% (p< 0.05) as compared to normal controls (untreated). Both vascular endothelial growth factor and pigment epithelium-derived factor levels remain the same until day 14 but returned to basal expression at day 21. This work expose the relation of cryotherapy with the expression of two factors related to angiogenesis. RESULTS showed significant changes on the expression of vascular endothelial growth factor and pigment epithelium-derived factor illustrating that both proteins are regulated in response to cryogenic treatment in relatively short periods (21 days).

Specific induction of neuronal cells from bone marrow stromal cells and application for autologous transplantation

PubMed Central

Dezawa, Mari; Kanno, Hiroshi; Hoshino, Mikio; Cho, Hirotomi; Matsumoto, Naoya; Itokazu, Yutaka; Tajima, Nobuyoshi; Yamada, Hitoshi; Sawada, Hajime; Ishikawa, Hiroto; Mimura, Toshirou; Kitada, Masaaki; Suzuki, Yoshihisa; Ide, Chizuka

2004-01-01

Bone marrow stromal cells (MSCs) have the capability under specific conditions of differentiating into various cell types such as osteocytes, chondrocytes, and adipocytes. Here we demonstrate a highly efficient and specific induction of cells with neuronal characteristics, without glial differentiation, from both rat and human MSCs using gene transfection with Notch intracellular domain (NICD) and subsequent treatment with bFGF, forskolin, and ciliary neurotrophic factor. MSCs expressed markers related to neural stem cells after transfection with NICD, and subsequent trophic factor administration induced neuronal cells. Some of them showed voltage-gated fast sodium and delayed rectifier potassium currents and action potentials compatible with characteristics of functional neurons. Further treatment of the induced neuronal cells with glial cell line–derived neurotrophic factor (GDNF) increased the proportion of tyrosine hydroxylase–positive and dopamine-producing cells. Transplantation of these GDNF-treated cells showed improvement in apomorphine-induced rotational behavior and adjusting step and paw-reaching tests following intrastriatal implantation in a 6-hydroxy dopamine rat model of Parkinson disease. This study shows that a population of neuronal cells can be specifically generated from MSCs and that induced cells may allow for a neuroreconstructive approach. PMID:15199405

[Partial dorsal root rhizotomy increases the anterograde transportation of neunotrophic factors in primary sensory neuron].

PubMed

Long, Shuang-lian; Li, Yong-mei; Yuan, Yuan; Wang, Ting-hua; Wu, Lin-yan

2005-05-01

To investigate whether partial dorsal root rhizotomy promotes the anterograde Five adult cats were transportation of BDNF, NT-3 and GDNF in the primary sensory neuron. Subjected to unilateral spared root rhizotomy (the DRGs of L1-L5 and L7-S2 were removed, but L6 DRG was spared) and bilateral dorsal roots of L6 were ligated at the same time. Three days after operation, dorsal roots were taken out and made into frozen sections 20 microm in thickness. The sections were stained using specific BDNF, NT-3, GDNF antibody (1:1500) by ABC method. The immunoreactive density was observed in a site near DRG and a site near spinal cord. In the control group (with spared L6 DRG), there were no marked differences in NT-3 and GDNF immunoreactivity between the site near DRG and the site near spinal cord, while BDNF immunoreactivity was more intense in the site near DRG than that in the site near spinal cord. In the operation group, the immunoreactivity of each neurotrophin in the site near DRG was stronger than that in the site near spinal cord, and the immunoreactivities of BDNF, NT-3, GDNF in the site near DRG of the operation were stronger than those of the control group respectively. The increasing of immunoreactivities of neurotrophins near DRG following partial dorsal root rhizotomy suggests that partial dorsal root rhizotomy can promote their anterograde transportation from spared DRG to the spinal cord.

Neuronal expression of fibroblast growth factor receptors in zebrafish.

PubMed

Rohs, Patricia; Ebert, Alicia M; Zuba, Ania; McFarlane, Sarah

2013-12-01

Fibroblast growth factor (FGF) signaling is important for a host of developmental processes such as proliferation, differentiation, tissue patterning, and morphogenesis. In vertebrates, FGFs signal through a family of four fibroblast growth factor receptors (FGFR 1-4), one of which is duplicated in zebrafish (FGFR1). Here we report the mRNA expression of the five known zebrafish fibroblast growth factor receptors at five developmental time points (24, 36, 48, 60, and 72h postfertilization), focusing on expression within the central nervous system. We show that the receptors have distinct and dynamic expression in the developing zebrafish brain, eye, inner ear, lateral line, and pharynx. In many cases, the expression patterns are similar to those of homologous FGFRs in mouse, chicken, amphibians, and other teleosts. Copyright © 2013 Elsevier B.V. All rights reserved.

Cerebrolysin Attenuates Heat Shock Protein (HSP 72 KD) Expression in the Rat Spinal Cord Following Morphine Dependence and Withdrawal: Possible New Therapy for Pain Management

PubMed Central

Sharma, Hari S; Ali, Syed F; Patnaik, Ranjana; Zimmermann-Meinzingen, Sibilla; Sharma, Aruna; Muresanu, Dafin F

2011-01-01

The possibility that pain perception and processing in the CNS results in cellular stress and may influence heat shock protein (HSP) expression was examined in a rat model of morphine dependence and withdrawal. Since activation of pain pathways result in exhaustion of growth factors, we examined the influence of cerebrolysin, a mixture of potent growth factors (BDNF, GDNF, NGF, CNTF etc,) on morphine induced HSP expression. Rats were administered morphine (10 mg/kg, s.c. /day) for 12 days and the spontaneous withdrawal symptoms were developed by cessation of the drug administration on day 13th that were prominent on day 14th and continued up to day 15th (24 to 72 h periods). In a separate group of rats, cerebrolysin was infused intravenously (5 ml/kg) once daily from day one until day 15th. In these animals, morphine dependence and withdrawal along with HSP immunoreactivity was examined using standard protocol. In untreated group mild HSP immunoreaction was observed during morphine tolerance, whereas massive upregulation of HSP was seen in CNS during withdrawal phase that correlated well with the withdrawal symptoms and neuronal damage. Pretreatment with cerebrolysin did not affect morphine tolerance but reduced the HSP expression during this phase. Furthermore, cerebrolysin reduced the withdrawal symptoms on day 14th to 15th. Taken together these observations suggest that cellular stress plays an important role in morphine induced pain pathology and exogenous supplement of growth factors, i.e. cerebrolysin attenuates HSP expression in the CNS and induce neuroprotection. This indicates a new therapeutic role of cerebrolysin in the pathophysiology of drugs of abuse, not reported earlier. PMID:21886595

Improvement in the neural stem cell proliferation in rats treated with modified "Shengyu" decoction may contribute to the neurorestoration.

PubMed

Chen, Miao-Miao; Zhao, Guang-Wei; He, Peng; Jiang, Zheng-Lin; Xi, Xin; Xu, Shi-Hui; Ma, Dong-Ming; Wang, Yong; Li, Yong-Cai; Wang, Guo-Hua

2015-05-13

"Shengyu" decoction, a traditional Chinese medicine, has been used to treat diseases with deficit in "qi" and "blood". The modified "Shengyu" decoction (MSD) used in the present study was designed to treat traumatic brain injury (TBI) on the basis of the "Shengyu" decoction, in which additional four herbs were added. Many ingredients in these herbs have been demonstrated to be effective for the treatment of brain injury. The present study was performed to evaluate the neurorestorative effect and the underlying mechanisms of MSD on the rat brain after a TBI. TBI was induced in the right cerebral cortex of adult rats using Feeney's weight-drop method. Intragastrical administration of MSD (1.0 ml/200 g) was begun 6h after TBI. The neurological functions and neuronal loss in the cortex and hippocampus were determined. The levels of nerve growth-related factors GDNF, NGF, NCAM, TN-C, and Nogo-A and the number of GFAP(+)/GDNF(+), BrdU(+)/nestin(+), BrdU(+)/NeuN(+) immunoreactive cells in the brain ipsilateral to TBI were also measured. Moreover, the influences of MSD on these variables were observed at the same time. We found that treatment with MSD in TBI rats ameliorated the neurological functions and alleviated neuronal loss. MSD treatment elevated the expression of GDNF, NGF, NCAM, and TN-C, and inhibited the expression of Nogo-A. Moreover, MSD treatment increased the number of GFAP(+)/GDNF(+), BrdU(+)/nestin(+), and BrdU(+)/NeuN(+) immunoreactive cells in the cortex and hippocampus. The present results suggest that MSD treatment in TBI rats could improve the proliferation of neural stem/progenitor cells and differentiation into neurons, which may facilitate neural regeneration and tissue repair and thus contribute to the recovery of neurological functions. These effects of modified "Shengyu" decoction may provide a foundation for the use of MSD as a prescription of medicinal herbs in the traditional medicine to treat brain injuries in order to improve the

Dose-Dependent Differential Effect of Neurotrophic Factors on In Vitro and In Vivo Regeneration of Motor and Sensory Neurons

PubMed Central

Santos, Daniel; Gonzalez-Perez, Francisco; Navarro, Xavier

2016-01-01

Although peripheral axons can regenerate after nerve transection and repair, functional recovery is usually poor due to inaccurate reinnervation. Neurotrophic factors promote directional guidance to regenerating axons and their selective application may help to improve functional recovery. Hence, we have characterized in organotypic cultures of spinal cord and dorsal root ganglia the effect of GDNF, FGF-2, NGF, NT-3, and BDNF at different concentrations on motor and sensory neurite outgrowth. In vitro results show that GDNF and FGF-2 enhanced both motor and sensory neurite outgrowth, NGF and NT-3 were the most selective to enhance sensory neurite outgrowth, and high doses of BDNF selectively enhanced motor neurite outgrowth. Then, NGF, NT-3, and BDNF (as the most selective factors) were delivered in a collagen matrix within a silicone tube to repair the severed sciatic nerve of rats. Quantification of Fluorogold retrolabeled neurons showed that NGF and NT-3 did not show preferential effect on sensory regeneration whereas BDNF preferentially promoted motor axons regeneration. Therefore, the selective effects of NGF and NT-3 shown in vitro are lost when they are applied in vivo, but a high dose of BDNF is able to selectively enhance motor neuron regeneration both in vitro and in vivo. PMID:27867665

Rapid transient isoform-specific neuregulin1 transcription in motor neurons is regulated by neurotrophic factors and axon-target interactions.

PubMed

Wang, Jiajing; Hmadcha, Abdelkrim; Zakarian, Vaagn; Song, Fei; Loeb, Jeffrey A

2015-09-01

The neuregulins (NRGs) are a family of alternatively spliced factors that play important roles in nervous system development and disease. In motor neurons, NRG1 expression is regulated by activity and neurotrophic factors, however, little is known about what controls isoform-specific transcription. Here we show that NRG1 expression in the chick embryo increases in motor neurons that have extended their axons and that limb bud ablation before motor axon outgrowth prevents this induction, suggesting a trophic role from the developing limb. Consistently, NRG1 induction after limb bud ablation can be rescued by adding back the neurotrophic factors BDNF and GDNF. Mechanistically, BDNF induces a rapid and transient increase in type I and type III NRG1 mRNAs that peak at 4h in rat embryonic ventral spinal cord cultures. Blocking MAPK or PI3K signaling or blocking transcription with Actinomycin D blocks BDNF induced NRG1 gene induction. BDNF had no effect on mRNA degradation, suggesting that transcriptional activation rather than message stability is important. Furthermore, BDNF activates a reporter construct that includes 700bp upstream of the type I NRG1 start site. Protein synthesis is also required for type I NRG1 mRNA transcription as cycloheximide produced a super-induction of type I, but not type III NRG1 mRNA, possibly through a mechanism involving sustained activation of MAPK and PI3K. These results reveal the existence of highly responsive, transient transcriptional regulatory mechanisms that differentially modulate NRG1 isoform expression as a function of extracellular and intracellular signaling cascades and mediated by neurotrophic factors and axon-target interactions. Copyright © 2015 Elsevier Inc. All rights reserved.

GPR30 Activation Contributes to the Puerarin-Mediated Neuroprotection in MPP+-Induced SH-SY5Y Cell Death.

PubMed

Cheng, Yue-Fa; Zhu, Guoqi; Wu, Qing-Wen; Xie, Yue-Sheng; Jiang, Yan; Guo, Lan; Guan, Ya-Li; Liu, Ying-Shuo; Zhang, Jun

2017-02-01

The neuroprotective action of puerarin in Parkinson's disease (PD) models has been well investigated. However, the mechanisms involved in protection have not been completely understood. G protein-coupled receptor 30 (GPR30) is a G protein-coupled estrogen receptor and considered a potential target in the neuroprotection against PD. In this study, we investigated whether puerarin prevented against 1-methyl-4-phenylpyridinium (MPP + )-induced cell death via GPR30. Our results showed that the GPR30 agonist, G1, exhibited puerarin-mediated neuroprotection against MPP + -induced cell death of SH-SY5Y cells. This protective action was reversed by the GPR30 antagonist. Moreover, a time- and concentration-dependent effect of puerarin on GPR30 expression was verified at the protein level but not at the mRNA level. Further, we showed that an mTor-dependent new GPR30 synthesis contributed to the protection conferred by puerarin. Finally, glial cell line-derived neurotrophic factor (GDNF) levels were enhanced by puerarin and G1 in both control and MPP + -lesioned cells via GPR30. Taken together, our data strongly suggest that puerarin prevents MPP + -induced cell death via facilitating GPR30 expression and GDNF release.

Temporal expression of growth factors triggered by epiregulin regulates inflammation development.

PubMed

Harada, Masaya; Kamimura, Daisuke; Arima, Yasunobu; Kohsaka, Hitoshi; Nakatsuji, Yuji; Nishida, Makoto; Atsumi, Toru; Meng, Jie; Bando, Hidenori; Singh, Rajeev; Sabharwal, Lavannya; Jiang, Jing-Jing; Kumai, Noriko; Miyasaka, Nobuyuki; Sakoda, Saburo; Yamauchi-Takihara, Keiko; Ogura, Hideki; Hirano, Toshio; Murakami, Masaaki

2015-02-01

In this study, we investigated the relationship between several growth factors and inflammation development. Serum concentrations of epiregulin, amphiregulin, betacellulin, TGF-α, fibroblast growth factor 2, placental growth factor (PLGF), and tenascin C were increased in rheumatoid arthritis patients. Furthermore, local blockades of these growth factors suppressed the development of cytokine-induced arthritis in mice by inhibiting chemokine and IL-6 expressions. We found that epiregulin expression was early and followed by the induction of other growth factors at different sites of the joints. The same growth factors then regulated the expression of epiregulin at later time points of the arthritis. These growth factors were increased in patients suffering from multiple sclerosis (MS) and also played a role in the development of an MS model, experimental autoimmune encephalomyelitis. The results suggest that the temporal expression of growth factors is involved in the inflammation development seen in several diseases, including rheumatoid arthritis and MS. Therefore, various growth factor pathways might be good therapeutic targets for various inflammatory diseases. Copyright © 2015 by The American Association of Immunologists, Inc.

Hemoglobin enhances tissue factor expression on human malignant cells.

PubMed

Siddiqui, F A; Amirkhosravi, A; Amaya, M; Meyer, T; Biggerstaff, J; Desai, H; Francis, J L

2001-04-01

Tissue Factor (TF) is a transmembrane glycoprotein that complexes with factor VII/activated factor VII to initiate blood coagulation. TF may be expressed on the surface of various cells including monocytes and endothelial cells. Over-expression of TF in human tumor cell lines promotes metastasis. We recently showed that hemoglobin (Hb) forms a specific complex with TF purified from human malignant melanoma cells and enhances its procoagulant activity (PCA). To further study this interaction, we examined the effect of Hb on the expression of TF on human malignant (TF+) cells and KG1 myeloid leukemia (TF-) cells. Human melanoma A375 and J82 bladder carcinoma cells, which express TF at moderate and relatively high levels, respectively, were incubated with varying concentrations (0-1.5 mg/ml) of Hb. After washing, cells were analyzed for Hb binding and TF expression using flow cytometry and confocal microscopy. Hb bound to the cells in a concentration-dependent manner, and increased both TF expression and PCA. The human A375 malignant melanoma cells incubated with Hb (1 mg/ml) expressed up to six times more TF antigen than cells without Hb. This increase in TF expression and PCA of intact cells incubated with Hb was significantly inhibited by cycloheximide at a concentration of 10 microg/ml (P < 0.01). An increase in total cellular TF antigen content was demonstrated by specific immunoassay. In contrast, Hb (5 mg/ml) did not induce TF expression and PCA on KG1 cells as determined by flow cytometry and TF (FXAA) activity. We conclude that Hb specifically binds to TF-bearing malignant cells and increases their PCA. This effect seems to be at least partly due to de novo synthesis of TF and increased surface expression. However, the exact mechanism by which Hb binds and upregulates TF expression remains to be determined.

Encapsulated cell device approach for combined electrical stimulation and neurotrophic treatment of the deaf cochlea.

PubMed

Konerding, W S; Janssen, H; Hubka, P; Tornøe, J; Mistrik, P; Wahlberg, L; Lenarz, T; Kral, A; Scheper, V

2017-07-01

Profound hearing impairment can be overcome by electrical stimulation (ES) of spiral ganglion neurons (SGNs) via a cochlear implant (CI). Thus, SGN survival is critical for CI efficacy. Application of glial cell line-derived neurotrophic factor (GDNF) has been shown to reduce SGN degeneration following deafness. We tested a novel method for local, continuous GDNF-delivery in combination with ES via a CI. The encapsulated cell (EC) device contained a human ARPE-19 cell-line, genetically engineered for secretion of GDNF. In vitro, GDNF delivery was stable during ES delivered via a CI. In the chronic in vivo part, cats were systemically deafened and unilaterally implanted into the scala tympani with a CI and an EC device, which they wore for six months. The implantation of control devices (same cell-line not producing GDNF) had no negative effect on SGN survival. GDNF application without ES led to an unexpected reduction in SGN survival, however, the combination of GDNF with initial, short-term ES resulted in a significant protection of SGNs. A tight fibrous tissue formation in the scala tympani of the GDNF-only group is thought to be responsible for the increased SGN degeneration, due to mechanisms related to an aggravated foreign body response. Furthermore, the fibrotic encapsulation of the EC device led to cell death or cessation of GDNF release within the EC device during the six months in vivo. In both in vitro and in vivo, fibrosis was reduced by CI stimulation, enabling the neuroprotective effect of the combined treatment. Thus, fibrous tissue growth limits treatment possibilities with an EC device. For a stable and successful long-term neurotrophic treatment of the SGN via EC devices in human CI users, it would be necessary to make changes in the treatment approach (provision of anti-inflammatories), the EC device surface (reduced cell adhesion) and the ES (initiation prior to fibrosis formation). Copyright © 2017 Elsevier B.V. All rights reserved.

Extended Duration of Transgene Expression from Pegylated POD Nanoparticles Enables Attenuation of Photoreceptor Degeneration

PubMed Central

Binder, Christina; Cashman, Siobhan M.; Kumar-Singh, Rajendra

2013-01-01

Retinitis pigmentosa (RP) is the most genetically heterogeneous disorder known to cause blindness, involving over 50 different genes. Previously, we have described nanoparticles (NPs) 150 nm in size, comprised of a 3.5 kD peptide (POD) complexed to PEG and DNA (PEGPOD DNA). These NPs expressing GDNF enabled rescue of photoreceptor degeneration in mice up to 11 days post injection. In the current study we examine use of scaffold/ matrix attachment regions (S/MARs), CpG depletion and titration of DNA content of PEGPOD DNA NPs to extend the duration of transgene expression. S/MARs and CpGs did not significantly influence the duration of transgene expression, but did influence its stability. These parameters enabled us to extend transgene expression from 48 hours to 10 weeks. At 77 days post injection, we observed a 76% rescue of the thickness of the retinal outer nuclear layer (ONL) and at 37 days post injection we observed 53% and 55% rescue of the A and B wave ERG amplitudes respectively and 60% rescue of the ONL. Our studies suggest that PEGPOD DNA NPs have potential as gene delivery vectors for the retina. PMID:24278479

Reconstruction of Mouse Testicular Cellular Microenvironments in Long-Term Seminiferous Tubule Culture

PubMed Central

Mäkelä, Juho-Antti; Toppari, Jorma; Rivero-Müller, Adolfo; Ventelä, Sami

2014-01-01

Research on spermatogonia is hampered by complex architecture of the seminiferous tubule, poor viability of testicular tissue ex vivo and lack of physiologically relevant long-term culture systems. Therefore there is a need for an in vitro model that would enable long term survival and propagation of spermatogonia. We aimed at the most simplified approach to enable all different cell types within the seminiferous tubules to contribute to the creation of a niche for spermatogonia. In the present study we describe the establishment of a co-culture of mouse testicular cells that is based on proliferative and migratory activity of seminiferous tubule cells and does not involve separation, purification or differential plating of individual cell populations. The co-culture is composed of the constituents of testicular stem cell niche: Sertoli cells [identified by expression of Wilm's tumour antigen 1 (WT1) and secretion of glial cell line-derived neurotrophic factor, GDNF], peritubular myoid cells (expressing alpha smooth muscle actin, αSMA) and spermatogonia [expressing MAGE-B4, PLZF (promyelocytic leukaemia zinc finger), LIN28, Gpr125 (G protein-coupled receptor 125), CD9, c-Kit and Nanog], and can be maintained for at least five weeks. GDNF was found in the medium at a sufficient concentration to support proliferating spermatogonial stem cells (SSCs) that were able to start spermatogenic differentiation after transplantation to an experimentally sterile recipient testis. Gdnf mRNA levels were elevated by follicle-stimulating hormone (FSH) which shows that the Sertoli cells in the co-culture respond to physiological stimuli. After approximately 2–4 weeks of culture a spontaneous formation of cord-like structures was monitored. These structures can be more than 10 mm in length and branch. They are formed by peritubular myoid cells, Sertoli cells, fibroblasts and spermatogonia as assessed by gene expression profiling. In conclusion, we have managed to establish in

Nerve Growth Factor Expression Is Not Associated with Perineural Invasion in Extrahepatic Cholangiocarcinoma.

PubMed

Urabe, Kazuhide; Murakami, Yoshiaki; Kondo, Naru; Uemura, Kenichiro; Hashimoto, Yasushi; Nakagawa, Naoya; Sasaki, Hayato; Hiyama, Eiso; Takahashi, Shinya; Sueda, Taijiro

2016-03-01

Although the presence of perineural invasion has been recognized as a poor prognostic factor in extrahepatic cholangiocarcinoma, the molecular mechanisms of perineural invasion in extrahepatic cholangiocarcinoma remain unclear. Nerve growth factor has been reported to be a candidate predictive biomarker of perineural invasion in some cancers. To investigate the impact of intratumoral nerve growth factor expression in resected extrahepatic cholangiocarcinoma on survival. Intratumoral nerve growth factor expression was investigated immunohistochemically in 112 patients with resected extrahepatic cholangiocarcinoma. Associations between nerve growth factor expression and clinicopathological factors were statistically evaluated, and risk factors for poor survival were analyzed using univariate and multivariate analyses. High and low nerve growth factor expression was observed in 62 (55%) and 50 (45%) patients, respectively. For all 112 patients, no significant correlation was found between nerve growth factor expression and presence of perineural invasion (P = 0.942). Moreover, nerve growth factor expression was not associated with recurrence-free survival (P = 0.861) and overall survival (P = 0.973). In multivariate analysis, lymph node metastasis (P = 0.004) was identified as an independent risk factor for early recurrence and the presence of perineural invasion (P = 0.002) and lymph node metastasis (P < 0.001) was identified as independent risk factors for poor survival. Intratumoral nerve growth factor expression is not associated with perineural invasion or recurrence-free and overall survival in patients with resected extrahepatic cholangiocarcinoma.

Developmental expression patterns of candidate co-factors for vertebrate Six family transcription factors

PubMed Central

Neilson, Karen M.; Pignoni, Francesca; Yan, Bo; Moody, Sally A.

2010-01-01

Six family transcription factors play important roles in craniofacial development. Their transcriptional activity can be modified by co-factor proteins. Two Six genes and one co-factor gene (Eya1) are involved in the human Branchio-otic (BO) and Branchio-otic-renal (BOR) syndromes. However, mutations in Six and Eya genes only account for about half of these patients. To discover potential new causative genes, we searched the Xenopus genome for orthologues of Drosophila co-factor proteins that interact with the fly Six-related factor, SO. We identified 33 Xenopus genes with high sequence identity to 20 of the 25 fly SO-interacting proteins. We provide the developmental expression patterns of the Xenopus orthologues for 11 of the fly genes, and demonstrate that all are expressed in developing craniofacial tissues with at least partial overlap with Six1/Six2. We speculate that these genes may function as Six-interacting partners with important roles in vertebrate craniofacial development and perhaps congenital syndromes. PMID:21089078

Low-intensity pulsed ultrasound promotes Schwann cell viability and proliferation via the GSK-3β/β-catenin signaling pathway

PubMed Central

Ren, Cong; Chen, Xiaohui; Du, Ning; Geng, Shuo; Hu, Yingying; Liu, Xin; Wu, Xianxian; Lin, Yuan; Bai, Xue; Yin, Wenzhe; Cheng, Shi; Yang, Lei; Zhang, Yong

2018-01-01

Background: It has been reported that ultrasound enhances peripheral nerve regeneration, but the mechanism remains elusive. Low-intensity pulsed ultrasound (LIPUS) has been reported to enhance proliferation and alter protein production in various types of cells. In this study, we detected the effects of LIPUS on Schwann cells. Material and methods: Schwann cells were separated from new natal Sprague-Dawley rat sciatic nerves and were cultured and purified. The Schwann cells were treated by LIPUS for 10 minutes every day, with an intensity of 27.37 mW/cm2. After treatment for 5 days, MTT, EdU staining, and flow cytometry were performed to examine cell viability and proliferation. Neurotrophic factors, including FGF, NGF, BDNF, and GDNF, were measured by western blot and real-time PCR. GSK-3β, p-GSK-3β, β-catenin and Cyclin D1 protein levels were detected using a western blot analysis. The expression of Cyclin D1 was also detected by immunofluorescence. Results: MTT and EdU staining showed that LIPUS increased the Schwann cells viability and proliferation. Compared to the control group, LIPUS increased the expression of growth factors and neurotrophic factors, including FGF, NGF, BDNF, GDNF, and Cyclin D1. Meanwhile, GSK-3β activity was inhibited in the LIPUS group as demonstrated by the increased level of p-GSK-3β and the ratio of the p-GSK-3β/GSK-3β level. The mRNA and protein expressions of β-catenin were increased in the LIPUS group. However, SB216763, a GSK-3β inhibitor, reversed the effects of LIPUS on Schwann cells. Conclusion: LIPUS promotes Schwann cell viability and proliferation by increasing Cyclin D1 expression via enhancing the GSK-3β/β-catenin signaling pathway.

Biological basis of neuroprotection and neurotherapeutic effects of Whole Body Periodic Acceleration (pGz).

PubMed

Adams, Jose A; Uryash, Arkady; Bassuk, Jorge; Sackner, Marvin A; Kurlansky, Paul

2014-06-01

Exercise is a well known neuroprotective and neurotherapeutic strategy in animal models and humans with brain injury and cognitive dysfunction. In part, exercise induced beneficial effects relate to endothelial derived nitric oxide (eNO) production and induction of the neurotrophins; Brain Derived Neurotrophic Factor (BDNF) and Glial Derived Neurotrophic Factor (GDNF). Whole Body Periodic Acceleration (WBPA (pGz), is the motion of the supine body headward to footward in a sinusoidal fashion, at frequencies of 100-160 cycles/min, inducing pulsatile shear stress to the vascular endothelium. WBPA (pGz) increases eNO in the cardiovascular system in animal models and humans. We hypothesized that WBPA (pGz) has neuroprotective and neurotherapeutic effects due to enhancement of biological pathways that include eNOS, BDNF and GDNF. We discuss protein expression analysis of these in brain of rodents. Animal and observational human data affirm a neuroprotective and neurotherapeutic role for WBPA (pGz). These findings suggest that WBPA (pGz) in addition to its well known beneficial cardiovascular effects can be a simple non-invasive neuroprotective and neurotherapeutic strategy with far reaching health benefits. Copyright © 2014 Elsevier Ltd. All rights reserved.

Tissue factor expression by endothelial cells in sickle cell anemia.

PubMed

Solovey, A; Gui, L; Key, N S; Hebbel, R P

1998-05-01

The role of the vascular endothelium in activation of the coagulation system, a fundamental homeostatic mechanism of mammalian biology, is uncertain because there is little evidence indicating that endothelial cells in vivo express tissue factor (TF), the system's triggering mechanism. As a surrogate for vessel wall endothelium, we examined circulating endothelial cells (CEC) from normals and patients with sickle cell anemia, a disease associated with activation of coagulation. We find that sickle CEC abnormally express TF antigen (expressed as percent CEC that are TF-positive), with 66+/-13% positive in sickle patients in steady-state, 83+/-19% positive in sickle patients presenting with acute vasoocclusive episodes, and only 10+/-13% positive in normal controls. Repeated samplings confirmed this impression that TF expression is greater when sickle patients develop acute vasoocclusive episodes. Sickle CEC are also positive for TF mRNA, with excellent concurrence between antigen and mRNA expression. The TF expressed on the antigen-positive CEC is functional, as demonstrated by a binding assay for Factor VIIa and a chromogenic assay sensitive to generation of Factor Xa. By establishing that endothelial cells in vivo can express TF, these data imply that the vast endothelial surface area does provide an important pathophysiologic trigger for coagulation activation.

Evaluation of procoagulant tissue factor expression in canine hemangiosarcoma cell lines.

PubMed

Witter, Lauren E; Gruber, Erika J; Lean, Fabian Z X; Stokol, Tracy

2017-01-01

OBJECTIVE To evaluate expression of procoagulant tissue factor (TF) by canine hemangiosarcoma cells in vitro. SAMPLES 4 canine hemangiosarcoma cell lines (SB-HSA [mouse-passaged cutaneous tumor], Emma [primary metastatic brain tumor], and Frog and Dal-1 [primary splenic tumors]) and 1 nonneoplastic canine endothelial cell line (CnAoEC). PROCEDURES TF mRNA and TF antigen expression were evaluated by quantitative real-time PCR assay and flow cytometry, respectively. Thrombin generation was measured in canine plasma and in coagulation factor-replete or specific coagulation factor-deficient human plasma by calibrated automated thrombography. Corn trypsin inhibitor and annexin V were used to examine contributions of contact activation and membrane-bound phosphatidylserine, respectively, to thrombin generation. RESULTS All cell lines expressed TF mRNA and antigen, with significantly greater expression of both products in SB-HSA and Emma cells than in CnAoEC. A greater percentage of SB-HSA cells expressed TF antigen, compared with other hemangiosarcoma cell lines. All hemangiosarcoma cell lines generated significantly more thrombin than did CnAoEC in canine or factor-replete human plasma. Thrombin generation induced by SB-HSA cells was significantly lower in factor VII-deficient plasma than in factor-replete plasma and was abolished in factor X-deficient plasma; residual thrombin generation in factor VII-deficient plasma was abolished by incubation of cells with annexin V. Thrombin generation by SB-HSA cells was unaffected by the addition of corn trypsin inhibitor. CONCLUSIONS AND CLINICAL RELEVANCE Hemangiosarcoma cell lines expressed procoagulant TF in vitro. Further research is needed to determine whether TF can be used as a biomarker for hemostatic dysfunction in dogs with hemangiosarcoma.

Alternative Sigma Factor Over-Expression Enables Heterologous Expression of a Type II Polyketide Biosynthetic Pathway in Escherichia coli

PubMed Central

Stevens, David Cole; Conway, Kyle R.; Pearce, Nelson; Villegas-Peñaranda, Luis Roberto; Garza, Anthony G.; Boddy, Christopher N.

2013-01-01

Background Heterologous expression of bacterial biosynthetic gene clusters is currently an indispensable tool for characterizing biosynthetic pathways. Development of an effective, general heterologous expression system that can be applied to bioprospecting from metagenomic DNA will enable the discovery of a wealth of new natural products. Methodology We have developed a new Escherichia coli-based heterologous expression system for polyketide biosynthetic gene clusters. We have demonstrated the over-expression of the alternative sigma factor σ54 directly and positively regulates heterologous expression of the oxytetracycline biosynthetic gene cluster in E. coli. Bioinformatics analysis indicates that σ54 promoters are present in nearly 70% of polyketide and non-ribosomal peptide biosynthetic pathways. Conclusions We have demonstrated a new mechanism for heterologous expression of the oxytetracycline polyketide biosynthetic pathway, where high-level pleiotropic sigma factors from the heterologous host directly and positively regulate transcription of the non-native biosynthetic gene cluster. Our bioinformatics analysis is consistent with the hypothesis that heterologous expression mediated by the alternative sigma factor σ54 may be a viable method for the production of additional polyketide products. PMID:23724102

Kruppel-like factor 2 inhibits hypoxia-inducible factor 1alpha expression and function in the endothelium.

PubMed

Kawanami, Daiji; Mahabeleshwar, Ganapati H; Lin, Zhiyong; Atkins, G Brandon; Hamik, Anne; Haldar, Saptarsi M; Maemura, Koji; Lamanna, Joseph C; Jain, Mukesh K

2009-07-31

Hypoxia-inducible factor 1 (HIF-1) is a central regulator of the hypoxic response in many cell types. In endothelial cells, HIF-1 induces the expression of key proangiogenic factors to promote angiogenesis. Recent studies have identified Kruppel-like factor 2 (KLF2) as a potent inhibitor of angiogenesis. However, the role of KLF2 in regulating HIF-1 expression and function has not been evaluated. KLF2 expression was induced acutely by hypoxia in endothelial cells. Adenoviral overexpression of KLF2 inhibited hypoxia-induced expression of HIF-1alpha and its target genes such as interleukin 8, angiopoietin-2, and vascular endothelial growth factor in endothelial cells. Conversely, knockdown of KLF2 increased expression of HIF-1alpha and its targets. Furthermore, KLF2 inhibited hypoxia-induced endothelial tube formation, whereas endothelial cells from mice with haploinsufficiency of KLF2 showed increased tube formation in response to hypoxia. Consistent with this ex vivo observation, KLF2 heterozygous mice showed increased microvessel density in the brain. Mechanistically, KLF2 promoted HIF-1alpha degradation in a von Hippel-Lindau protein-independent but proteasome-dependent manner. Finally, KLF2 disrupted the interaction between HIF-1alpha and its chaperone Hsp90, suggesting that KLF2 promotes degradation of HIF-1alpha by affecting its folding and maturation. These observations identify KLF2 as a novel inhibitor of HIF-1alpha expression and function. Therefore, KLF2 may be a target for modulating the angiogenic response in disease states.

Recombinant expression of extracellular domain of mutant Epidermal Growth Factor Receptor in prokaryotic and baculovirus expression systems.

PubMed

Vettath, Sunitha Kodengil; Shivashankar, Gaganashree; Menon, Krishnakumar N; Vijayachandran, Lakshmi S

2018-04-15

Epidermal Growth Factor Receptor variant III (EGFRvIII) is a tumor specific antigen detected in various tumors including gliomas, breast cancer, lung cancer, head and neck squamous cell carcinoma (HNSCC). Screening of EGFRvIII targeting drug molecules can be accelerated by developing drug screening platforms using recombinantly expressed protein. Choice of expression system is one of the major factors deciding the success of recombinant expression of a protein. In our study, we have tried to express and purify the extracellular domain (ECD) of this highly unstable protein using bacterial and baculovirus expression systems to select the expression system suited for our purpose. Even though the protein was successfully expressed in prokaryotic system, purification could be done only under denaturing conditions. But in the baculovirus expression system, the protein was expressed in soluble form and could be purified under native conditions, with single step of purification. Based on our results, we conclude that insect cells are better choice over E. coli cells for expressing EGFRvIII ECD in soluble form. This study provides insights for other researchers involved in expression of similar unstable membrane proteins, on selecting the best expression system and challenges involved. Copyright © 2018 Elsevier B.V. All rights reserved.

Endocrine gland-derived vascular endothelial growth factor (EG-VEGF) expression in colorectal cancer.

PubMed

Nagano, Hideki; Goi, Takanori; Koneri, Kenji; Hirono, Yasuo; Katayama, Kanji; Yamaguchi, Akio

2007-12-01

Vascular endothelial growth factor (VEGF) is known as an important factor in the growth and metastasis of cancer cells. In 2001, a novel angiogenesis factor, endocrine gland-derived vascular endothelial growth factor (EG-VEGF), was cloned. In this study, we investigated the expression of EG-VEGF in colorectal cancer, the relationship between its expression and clinicopathological factors, and the in vitro activity of EG-VEGF transfectants. We determined expression levels of EG-VEGF in 113 advanced colorectal cancers resected in our hospital by quantitative PCR, and compared the expression levels and clinicopathological findings by multivariate analyses. The expression of EG-VEGF mRNA was positive in 31 cancers and negative in 82 cancers. We found that compared with the negative expression of the EG-VEGF gene, its positive expression was more frequently associated with hematogenous metastasis, and was associated with a poorer survival rate. In addition, EG-VEGF transfectants showed a higher degree of in vitro tubular formation than control cells. We speculate that, in colorectal cancers, the EG-VEGF gene functions as an important factor in angiogenesis in primary and metastatic lesions, and consider that it is useful as a novel prognostic factor. EG-VEGF molecule-targeted therapy has the potential for improving survival rates.

Clinical validation of nuclear factor kappa B expression in invasive breast cancer.

PubMed

Agrawal, Anil Kumar; Pielka, Ewa; Lipinski, Artur; Jelen, Michal; Kielan, Wojciech; Agrawal, Siddarth

2018-01-01

Breast cancer is the most commonly diagnosed cancer in Polish women. The expression of transcription nuclear factor kappa B, a key inducer of inflammatory response promoting carcinogenesis and cancer progression in breast cancer, is not well-established. We assessed the nuclear factor kappa B expression in a total of 119 invasive breast carcinomas and 25 healthy control samples and correlated this expression pattern with several clinical and pathologic parameters including histologic type and grade, tumor size, lymph node status, estrogen receptor status, and progesterone receptor status. The data used for the analysis were derived from medical records. An immunohistochemical analysis of nuclear factor kappa B, estrogen receptor, and progesterone receptor was carried out and evaluation of stainings was performed. The expression of nuclear factor kappa B was significantly higher than that in the corresponding healthy control samples. No statistical difference was demonstrated in nuclear factor kappa B expression in relation to age, menopausal status, lymph node status, tumor size and location, grade and histologic type of tumor, and hormonal status (estrogen receptor and progesterone receptor). Nuclear factor kappa B is significantly overexpressed in invasive breast cancer tissues. Although nuclear factor kappa B status does not correlate with clinicopathological findings, it might provide important additional information on prognosis and become a promising object for targeted therapy.

Autocrine expression of the epidermal growth factor receptor ligand heparin-binding EGF-like growth factor in cervical cancer.

PubMed

Schrevel, Marlies; Osse, E Michelle; Prins, Frans A; Trimbos, J Baptist M Z; Fleuren, Gert Jan; Gorter, Arko; Jordanova, Ekaterina S

2017-06-01

In cervical cancer, the epidermal growth factor receptor (EGFR) is overexpressed in 70-90% of the cases and has been associated with poor prognosis. EGFR-based therapy is currently being explored in cervical cancer. We investigated which EGFR ligand is primarily expressed in cervical cancer and which cell type functions as the major source of this ligand. We hypothesized that macrophages are the main source of EGFR ligands and that a paracrine loop between tumor cells and macrophages is responsible for ligand expression. mRNA expression analysis was performed on 32 cervical cancer cases to determine the expression of the EGFR ligands amphiregulin, β-cellulin, epidermal growth factor (EGF), epiregulin, heparin-binding EGF-like growth factor (HB‑EGF) and transforming growth factor α (TGFα). Subsequently, protein expression was determined immunohistochemically on 36 additional cases. To assess whether macrophages are the major source of EGFR ligands, immunohistochemical double staining was performed on four representative tissue slides. Expression of the chemokines granulocyte-macrophage colony-stimulating factor (GM-CSF) and C-C motif ligand 2 (CCL2) was determined by mRNA in situ hybridization. Of the known EGFR ligands, HB‑EGF had the highest mRNA expression and HB‑EGF and EGFR protein expression were highly correlated. Tumor specimens with high EGFR expression showed higher numbers of macrophages, and higher expression of GM-CSF and CCL2, but only a small subset (9%) of macrophages was found to be HB‑EGF-positive. Strikingly, 78% of cervical cancer specimens were found to express HB‑EGF. Standardized assessment of staining intensity, using spectral imaging analysis, showed that HB‑EGF expression was higher in the tumor compartment than in the stromal compartment. These results suggest that HB‑EGF is an important EGFR ligand in cervical cancer and that cervical cancer cells are the predominant source of HB‑EGF. Therefore, we propose an autocrine

Factor XIIIa is expressed by fibroblasts in fibrovascular tumors.

PubMed

Nemeth, A J; Penneys, N S

1989-10-01

Factor XIIIa (FXIIIa), a blood and intracellularly produced coagulation factor, has been found in a variety of cell types including fibroblast-like mesenchymal cells, and has been shown to stimulate the proliferation of fibroblasts and some neoplastic cells in vitro. We have already shown that the dendritic fibroblasts composing the fibrous papule contain this factor. We hypothesized that histopathologically similar fibrovascular tumors may also express FXIIIa and, in this report, show that the large stellate fibroblasts found in acquired digital fibrokeratomas, angiofibromas (adenoma sebaceum of Pringle), and oral fibroma (oral fibrous hyperplasia) also express FXIIIa. We postulate that FXIIIa, possibly acting as a growth factor, may be a common denominator in the pathogenesis of these tumors. Another possibility is that these tumors may be the consequence of a local overproduction of FXIIIa in response to an, as yet, unidentified stimulus.

E74-like factor 2 regulates valosin-containing protein expression

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

Zhang, Binglin; Tomita, Yasuhiko; Qiu, Ying

2007-05-11

Enhanced expression of valosin-containing protein (VCP) correlates with invasion and metastasis of cancers. To clarify the transcription mechanism of VCP, human and mouse genomic sequence was compared, revealing a 260 bp DNA sequence in the 5'-flanking region of VCP gene to be highly conserved between the two, in which binding motif of E74-like factor 2/new Ets-related factor (ELF2/NERF) was identified. Chromatin immunoprecipitation assay showed binding of ELF2/NERF to the 5'-flanking region of VCP gene. Knock-down of ELF2/NERF by siRNA decreased expression level of VCP. Viability of cells under tumor necrosis factor-alpha treatment significantly reduced in ELF2/NERF-knock-down breast cancer cell line.more » Immunohistochemical analysis on clinical breast cancer specimens showed a correlation of nuclear ELF2/NERF expression with VCP expression and proliferative activity of cells shown by Ki-67 immunohistochemistry. These findings indicate that ELF2/NERF promotes VCP transcription and that ELF2/NERF-VCP pathway might be important for cell survival and proliferation under cytokine stress.« less

The protective effects of resveratrol on Schwann cells with toxicity induced by ethanol in vitro.

PubMed

Yuan, Hongtu; Zhang, Jingfen; Liu, Huaxiang; Li, Zhenzhong

2013-09-01

Schwann cells (SCs) are the myelin forming cells in the peripheral nervous system, they play a key role in the pathology of various polyneuropathies and provide trophic support to axons via expression of various neurotrophic factors, such as nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF) and glial cell line-derived neurotrophic factor (GDNF). Ethanol (EtOH) adversely affected both SCs proliferation and myelin formation in culture. Resveratrol (Res) has been shown to regulate many cellular processes and to display multiple protective and therapeutic effects. Whether Res has protective effects on SCs with EtOH-induced toxicity is still unclear. The protective efficacy of Res on EtOH-treated SCs in vitro was investigated in the present study. Res improved cell viability of the EtOH-treated SCs. Hoechst 33342 staining and terminal deoxynucleotidyl transferase (TdT)-mediated deoxyuridine triphosphate nick-end labeling analysis showed that the EtOH-induced apoptosis was inhibited by Res. The effects of Res were blocked by the 5'-adenosine monophosphate-activated protein kinase inhibitor Compound C and the silencing information regulator T1 inhibitor nicotinamide. Res could increase the mRNA and protein levels of BDNF and GDNF in the EtOH-treated SCs. However, the EtOH-induced increase of NGF in the SCs is inhibited by Res. The data from the present study indicate that Res protects SCs from EtOH-induced cell death and regulates the expression of neurotrophicfactors. Res and its derivative may be effective for the treatment of neuropathic diseases induced by EtOH. Copyright © 2013 Elsevier Ltd. All rights reserved.

Krüppel Like Factors Family Expression in Cervical Cancer Cells.

PubMed

Marrero-Rodríguez, Daniel; la Cruz, Hugo Arreola-De; Taniguchi-Ponciano, Keiko; Gomez-Virgilio, Laura; Huerta-Padilla, Victor; Ponce-Navarrete, Gustavo; Andonegui-Elguera, Sergio; Jimenez-Vega, Florinda; Romero-Morelos, Pablo; Rodriguez-Esquivel, Miriam; Meraz-Rios, Marco; Figueroa-Corona, Ma Del Pilar; Monroy, Alberto; Pérez-González, Oscar; Salcedo, Mauricio

2017-05-01

Krüppel Like Factors (KLF) refers to a family of seventeen members of transcription factors. Involved in several cellular processes. As other cancer types, Cervical Cancer (CC) presents molecular deregulations in transcription factors, but especially Human Papilloma Virus (HPV) sequences. Here in this work we analyzed the mRNA expression of all KLF family members in CC-derived cell lines and CC tissues. The cell lines used were HeLa, INBL, RoVa, C4-I, Ms751, ViPa, CaLo, SiHa, CaSki, C33a and ViBo and the non-tumorigenic HaCaT. mRNA expression was analyzed by means of expression microarray and RT-PCR, and KLF5 protein by immunofluorescence. The cell lines were grouped according to HPV genotype as HPV16, HPV18 positive or HPV negative cells. Heterogeneous expression was observed among the cell lines. Despite the heterogeneous expression profile, KLF3, -5, -12, -15 and -16 transcripts were present in all cell lines, KLF4 and -10 which were not expressed in CaSki; KLF11 and 13 were not expressed by Vipa and C4-I, and KLF7 was not expressed by C4-I and Rova. The CC tissue analysis shows expression of most of the KLF members, such as KLF5. KLF5 immunosignal was positive in the three cell lines analyzed. We suggest that KLF expression could not be related to HPV presence/genotype, at least at transcriptional level, and the expression of KLF family members may be necessary in the biology of the CC cells. Copyright © 2017 IMSS. Published by Elsevier Inc. All rights reserved.

Evaluation of procoagulant tissue factor expression in canine hemangiosarcoma cell lines

PubMed Central

Witter, Lauren E.; Gruber, Erika J.; Lean, Fabian Z. X.; Stokol, Tracy

2017-01-01

OBJECTIVE To evaluate expression of procoagulant tissue factor (TF) by canine hemangiosarcoma cells in vitro. SAMPLES 4 canine hemangiosarcoma cell lines (SB-HSA [mouse-passaged cutaneous tumor], Emma [primary metastatic brain tumor], and Frog and Dal-1 [primary splenic tumors]) and 1 nonneoplastic canine endothelial cell line (CnAoEC). PROCEDURES TF mRNA and TF antigen expression were evaluated by quantitative real-time PCR assay and flow cytometry, respectively. Thrombin generation was measured in canine plasma and in coagulation factor–replete or specific coagulation factor–deficient human plasma by calibrated automated thrombography. Corn trypsin inhibitor and annexin V were used to examine contributions of contact activation and membrane-bound phosphatidylserine, respectively, to thrombin generation. RESULTS All cell lines expressed TF mRNA and antigen, with significantly greater expression of both products in SB-HSA and Emma cells than in CnAoEC. A greater percentage of SB-HSA cells expressed TF antigen, compared with other hemangiosarcoma cell lines. All hemangiosarcoma cell lines generated significantly more thrombin than did CnAoEC in canine or factor-replete human plasma. Thrombin generation induced by SB-HSA cells was significantly lower in factor VII-deficient plasma than in factor-replete plasma and was abolished in factor X–deficient plasma; residual thrombin generation in FVII-deficient plasma was abolished by incubation of cells with annexin V. Thrombin generation by SB-HSA cells was unaffected by the addition of corn trypsin inhibitor. CONCLUSIONS AND CLINICAL RELEVANCE Hemangiosarcoma cell lines expressed procoagulant TF in vitro. Further research is needed to determine whether TF can be used as a biomarker for hemostatic dysfunction in dogs with hemangiosarcoma. PMID:28029283

Accurate expressions for solar cell fill factors including series and shunt resistances

NASA Astrophysics Data System (ADS)

Green, Martin A.

2016-02-01

Together with open-circuit voltage and short-circuit current, fill factor is a key solar cell parameter. In their classic paper on limiting efficiency, Shockley and Queisser first investigated this factor's analytical properties showing, for ideal cells, it could be expressed implicitly in terms of the maximum power point voltage. Subsequently, fill factors usually have been calculated iteratively from such implicit expressions or from analytical approximations. In the absence of detrimental series and shunt resistances, analytical fill factor expressions have recently been published in terms of the Lambert W function available in most mathematical computing software. Using a recently identified perturbative relationship, exact expressions in terms of this function are derived in technically interesting cases when both series and shunt resistances are present but have limited impact, allowing a better understanding of their effect individually and in combination. Approximate expressions for arbitrary shunt and series resistances are then deduced, which are significantly more accurate than any previously published. A method based on the insights developed is also reported for deducing one-diode fits to experimental data.

Factors Expressed in an Animal Model of Anteroinferior Glenohumeral Instability

PubMed Central

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

2015-01-01

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

Insulin-like growth factor and fibroblast growth factor expression profiles in growth-restricted fetal sheep pancreas.

PubMed

Chen, Xiaochuan; Rozance, Paul J; Hay, William W; Limesand, Sean W

2012-05-01

Placental insufficiency results in intrauterine growth restriction (IUGR), impaired fetal insulin secretion and less fetal pancreatic β-cell mass, partly due to lower β-cell proliferation rates. Insulin-like growth factors (IGFs) and fibroblast growth factors (FGFs) regulate fetal β-cell proliferation and pancreas development, along with transcription factors, such as pancreatic and duodenal homeobox 1 (PDX-1). We determined expression levels for these growth factors, their receptors and IGF binding proteins in ovine fetal pancreas and isolated islets. In the IUGR pancreas, relative mRNA expression levels of IGF-I, PDX-1, FGF7 and FGFR2IIIb were 64% (P < 0.01), 76% (P < 0.05), 76% (P < 0.05) and 52% (P < 0.01) lower, respectively, compared with control fetuses. Conversely, insulin-like growth factor binding protein 2 (IGFBP-2) mRNA and protein concentrations were 2.25- and 1.2-fold greater (P < 0.05) in the IUGR pancreas compared with controls. In isolated islets from IUGR fetuses, IGF-II and IGFBP-2 mRNA concentrations were 1.5- and 3.7-fold greater (P < 0.05), and insulin mRNA was 56% less (P < 0.05) than control islets. The growth factor expression profiles for IGF and FGF signaling pathways indicate that declines in β-cell mass are due to decreased growth factor signals for both pancreatic progenitor epithelial cell and mature β-cell replication.

Epidermal Growth Factor Increases LRF/Pokemon Expression in Human Prostate Cancer Cells

PubMed Central

Aggarwal, Himanshu; Aggarwal, Anshu; Agrawal, Devendra K.

2011-01-01

Leukemia/lymphoma related factor/POK erythroid myeloid ontogenic factor (LRF/Pokemon) is a member of the POK family of proteins that promotes oncogenesis in several forms of cancer. Recently, we found higher LRF expression in human breast and prostate carcinomas compared to the corresponding normal tissues. The aim of this study was to examine the regulation of LRF expression in human prostate cells. Epidermal growth factor (EGF) and its receptors mediate several tumorigenic cascades that regulate cell differentiation, proliferation, migration and survival of prostate cancer cells. There was significantly higher level of LRF expression in the nucleus of LNCaP and PC-3 cells than RWPE-1 cells. A significant increase in LRF expression was observed with increasing doses of EGF in more aggressive and androgen-sensitive prostate cancer cells suggesting that EGF signaling pathway is critical in upregulating the expression of LRF/Pokemon to promote oncogenesis. PMID:21640721

Tissue factor expression as a possible determinant of thromboembolism in ovarian cancer

PubMed Central

Uno, K; Homma, S; Satoh, T; Nakanishi, K; Abe, D; Matsumoto, K; Oki, A; Tsunoda, H; Yamaguchi, I; Nagasawa, T; Yoshikawa, H; Aonuma, K

2007-01-01

Ovarian cancer, and clear cell carcinoma in particular, reportedly increases the risk of venous thromboembolism (VTE). However, the mechanisms remain unclear. Tissue factor (TF) supposedly represents a major factor in the procoagulant activities of cancer cells. The present study examined the involvement of TF expression in VTE for patients with ovarian cancer. Subjects comprised 32 consecutive patients (mean age 49.8 years) with histologically confirmed ovarian cancer. Presence of VTE was examined using a combination of clinical features, D-dimer levels and venous ultrasonography. Immunohistochemical analysis was used to evaluate TF expression into 4 degrees. Venous thromboembolism was identified in 10 of the 32 patients (31%), including five of the 11 patients with clear cell carcinoma. Tissue factor expression was detected in cancer tissues from 24 patients and displayed significant correlations with VTE development (P=0.0003), D-dimer concentration (P=0.003) and clear cell carcinoma (P<0.05). Multivariate analysis identified TF expression as an independent predictive factor of VTE development (P<0.05). Tissue factor (TF) expression is a possible determinant of VTE development in ovarian cancer. In particular, clear cell carcinoma may produce excessive levels of TF and is more likely to develop VTE. PMID:17211468

Reactivation of Latent HIV-1 Expression by Engineered TALE Transcription Factors.

PubMed

Perdigão, Pedro; Gaj, Thomas; Santa-Marta, Mariana; Barbas, Carlos F; Goncalves, Joao

2016-01-01

The presence of replication-competent HIV-1 -which resides mainly in resting CD4+ T cells--is a major hurdle to its eradication. While pharmacological approaches have been useful for inducing the expression of this latent population of virus, they have been unable to purge HIV-1 from all its reservoirs. Additionally, many of these strategies have been associated with adverse effects, underscoring the need for alternative approaches capable of reactivating viral expression. Here we show that engineered transcriptional modulators based on customizable transcription activator-like effector (TALE) proteins can induce gene expression from the HIV-1 long terminal repeat promoter, and that combinations of TALE transcription factors can synergistically reactivate latent viral expression in cell line models of HIV-1 latency. We further show that complementing TALE transcription factors with Vorinostat, a histone deacetylase inhibitor, enhances HIV-1 expression in latency models. Collectively, these findings demonstrate that TALE transcription factors are a potentially effective alternative to current pharmacological routes for reactivating latent virus and that combining synthetic transcriptional activators with histone deacetylase inhibitors could lead to the development of improved therapies for latent HIV-1 infection.

Reactivation of Latent HIV-1 Expression by Engineered TALE Transcription Factors

PubMed Central

Perdigão, Pedro; Gaj, Thomas; Santa-Marta, Mariana; Goncalves, Joao

2016-01-01

The presence of replication-competent HIV-1 –which resides mainly in resting CD4+ T cells–is a major hurdle to its eradication. While pharmacological approaches have been useful for inducing the expression of this latent population of virus, they have been unable to purge HIV-1 from all its reservoirs. Additionally, many of these strategies have been associated with adverse effects, underscoring the need for alternative approaches capable of reactivating viral expression. Here we show that engineered transcriptional modulators based on customizable transcription activator-like effector (TALE) proteins can induce gene expression from the HIV-1 long terminal repeat promoter, and that combinations of TALE transcription factors can synergistically reactivate latent viral expression in cell line models of HIV-1 latency. We further show that complementing TALE transcription factors with Vorinostat, a histone deacetylase inhibitor, enhances HIV-1 expression in latency models. Collectively, these findings demonstrate that TALE transcription factors are a potentially effective alternative to current pharmacological routes for reactivating latent virus and that combining synthetic transcriptional activators with histone deacetylase inhibitors could lead to the development of improved therapies for latent HIV-1 infection. PMID:26933881

Keratinocyte growth factor expression in human gingival fibroblasts and stimulation of in vitro gene expression by retinoic acid.

PubMed

Mackenzie, I C; Gao, Z

2001-04-01

Keratinocyte growth factor (KGF) is a stromally derived growth factor of the fibroblast growth factor (FGF) family with paracrine effects targeted to influence the growth and differentiation of epithelia. Regional and temporal changes in KGF expression play important roles in the development and maintenance of epithelial structures and in epithelial wound healing. Differing patterns of expression of KGF by fibroblasts in the gingival region could therefore be related to the observed regional variation in the differentiation and behavior of gingival epithelia. The in vitro and in vivo patterns of expression of KGF mRNA in human gingival and periodontal fibroblasts were examined using reverse transcription polymerase chain reactions (RT-PCR) and in situ hybridization with digoxigenin-labeled riboprobes. The patterns observed for human gingiva were compared with those for human skin and for murine tissues. Gingival and periodontal fibroblasts showed expression of KGF transcripts in vitro, and the degree of expression was markedly influenced by the presence of retinoic acid, an agent known to influence patterns of epithelial differentiation. Sections of human and murine gingiva and skin showed regionally variable expression of transcripts with the cells expressing KGF in the subepithelial, rather than the deeper, connective tissues and periodontium. The results point to a role of KGF in the maintenance of normal growth and differentiation of gingival epithelia. A lack of KGF expression by periodontal fibroblasts in vivo is expected to hinder apical epithelial migration and thus stabilize the epithelial attachment. The effects of retinoic acid (RA) on KGF expression in vitro provide an indirect mechanism by which RA may regulate the growth and differentiation of gingival epithelia.

Epidermal growth factor increases LRF/Pokemon expression in human prostate cancer cells.

PubMed

Aggarwal, Himanshu; Aggarwal, Anshu; Agrawal, Devendra K

2011-10-01

Leukemia/lymphoma related factor/POK erythroid myeloid ontogenic factor (LRF/Pokemon) is a member of the POK family of proteins that promotes oncogenesis in several forms of cancer. Recently, we found higher LRF expression in human breast and prostate carcinomas compared to the corresponding normal tissues. The aim of this study was to examine the regulation of LRF expression in human prostate cells. Epidermal growth factor (EGF) and its receptors mediate several tumorigenic cascades that regulate cell differentiation, proliferation, migration and survival of prostate cancer cells. There was significantly higher level of LRF expression in the nucleus of LNCaP and PC-3 cells than RWPE-1 cells. A significant increase in LRF expression was observed with increasing doses of EGF in more aggressive and androgen-sensitive prostate cancer cells suggesting that EGF signaling pathway is critical in upregulating the expression of LRF/Pokemon to promote oncogenesis. Copyright © 2011 Elsevier Inc. All rights reserved.

Environmental signals modulate ToxT-dependent virulence factor expression in Vibrio cholerae.

PubMed

Schuhmacher, D A; Klose, K E

1999-03-01

The regulatory protein ToxT directly activates the transcription of virulence factors in Vibrio cholerae, including cholera toxin (CT) and the toxin-coregulated pilus (TCP). Specific environmental signals stimulate virulence factor expression by inducing the transcription of toxT. We demonstrate that transcriptional activation by the ToxT protein is also modulated by environmental signals. ToxT expressed from an inducible promoter activated high-level expression of CT and TCP in V. cholerae at 30 degrees C, but expression of CT and TCP was significantly decreased or abolished by the addition of 0.4% bile to the medium and/or an increase of the temperature to 37 degrees C. Also, expression of six ToxT-dependent TnphoA fusions was modulated by temperature and bile. Measurement of ToxT-dependent transcription of genes encoding CT and TCP by ctxAp- and tcpAp-luciferase fusions confirmed that negative regulation by 37 degrees C or bile occurs at the transcriptional level in V. cholerae. Interestingly, ToxT-dependent transcription of these same promoters in Salmonella typhimurium was relatively insensitive to regulation by temperature or bile. These data are consistent with ToxT transcriptional activity being modulated by environmental signals in V. cholerae and demonstrate an additional level of complexity governing the expression of virulence factors in this pathogen. We propose that negative regulation of ToxT-dependent transcription by environmental signals prevents the incorrect temporal and spatial expression of virulence factors during cholera pathogenesis.

Expression of vascular endothelial growth factor in Juvenile Angiofibroma.

PubMed

Hota, Ashutosh; Sarkar, Chitra; Gupta, Siddhartha Datta; Kumar, Rakesh; Bhalla, Ashu Seith; Thakar, Alok

2015-06-01

To examine Juvenile Angiofibroma (JA) tissue for expression of vascular endothelial growth factor (VEGF), and to explore its relationship with puberty status, stage, recurrence and the intraoperative blood loss. Retrospective cohort study of 36 histologically proven cases of JA. Minimum follow up period was 3 years. VEGF expression on tumor cells assessed by immunohistochemistry and graded on two criteria--percentage of cells expressing positivity and the intensity of positivity. These two parameters assessed for impact on puberty status, stage, recurrence, and blood loss. VEGF expression noted on the tumor endothelial cells in 36/36, and on the tumor stromal cells in 34/36. The percentage of cells expressing VEGF and the intensity of expression were not significantly related to puberty status, tumor stage, recurrence, or intra-operative blood loss (p values 0.3-1.0). VEGF expression is near universal in JA. Such expression is independent of puberty status and stage, and does not impact on intra operative blood loss and recurrence. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Accuracy of expressions for the fill factor of a solar cell in terms of open-circuit voltage and ideality factor

NASA Astrophysics Data System (ADS)

Leilaeioun, Mehdi; Holman, Zachary C.

2016-09-01

An approximate expression proposed by Green predicts the maximum obtainable fill factor (FF) of a solar cell from its open-circuit voltage (Voc). The expression was originally suggested for silicon solar cells that behave according to a single-diode model and, in addition to Voc, it requires an ideality factor as input. It is now commonly applied to silicon cells by assuming a unity ideality factor—even when the cells are not in low injection—as well as to non-silicon cells. Here, we evaluate the accuracy of the expression in several cases. In particular, we calculate the recombination-limited FF and Voc of hypothetical silicon solar cells from simulated lifetime curves, and compare the exact FF to that obtained with the approximate expression using assumed ideality factors. Considering cells with a variety of recombination mechanisms, wafer doping densities, and photogenerated current densities reveals the range of conditions under which the approximate expression can safely be used. We find that the expression is unable to predict FF generally: For a typical silicon solar cell under one-sun illumination, the error is approximately 6% absolute with an assumed ideality factor of 1. Use of the expression should thus be restricted to cells under very low or very high injection.

Liver-enriched transcription factors uncoupled from expression of hepatic functions in hepatoma cell lines.

PubMed Central

Chaya, D; Fougère-Deschatrette, C; Weiss, M C

1997-01-01

Among the liver-enriched transcription factors identified to date, only expression of hepatocyte nuclear factor 4 (HNF4) and hepatocyte nuclear factor 1 (HNF1) is in strict correlation with hepatic differentiation in cultured rat hepatoma cells. Indeed, differentiated hepatoma cells that stably express an extensive set of adult hepatic functions express liver-enriched transcription factors, while dedifferentiated cells that have lost expression of all these hepatic functions no longer express HNF4 and HNF1. We describe a new heritable phenotype, designated as uncoupled, in which there is a spontaneous dissociation between the expression of these transcription factors and that of the hepatic functions. Cells presenting this phenotype, isolated from differentiated hepatoma cells, cease to accumulate all transcripts coding for hepatic functions but nevertheless maintain expression of HNF4 and HNF1. Transitory transfection experiments indicate that these two factors present in these cells have transcriptional activity similar to that of differentiated hepatoma cells. Characterization of the appropriate intertypic cell hybrids demonstrates that this new phenotype is recessive to the dedifferentiated state and fails to be complemented by differentiated cells. These results indicate the existence of mechanisms that inhibit transcription of genes coding for hepatocyte functions in spite of the presence of functional HNF4 and HNF1. Cells of the uncoupled phenotype present certain properties of oval cells described for pathological states of the liver. PMID:9343392

Liver-enriched transcription factors uncoupled from expression of hepatic functions in hepatoma cell lines.

PubMed

Chaya, D; Fougère-Deschatrette, C; Weiss, M C

1997-11-01

Among the liver-enriched transcription factors identified to date, only expression of hepatocyte nuclear factor 4 (HNF4) and hepatocyte nuclear factor 1 (HNF1) is in strict correlation with hepatic differentiation in cultured rat hepatoma cells. Indeed, differentiated hepatoma cells that stably express an extensive set of adult hepatic functions express liver-enriched transcription factors, while dedifferentiated cells that have lost expression of all these hepatic functions no longer express HNF4 and HNF1. We describe a new heritable phenotype, designated as uncoupled, in which there is a spontaneous dissociation between the expression of these transcription factors and that of the hepatic functions. Cells presenting this phenotype, isolated from differentiated hepatoma cells, cease to accumulate all transcripts coding for hepatic functions but nevertheless maintain expression of HNF4 and HNF1. Transitory transfection experiments indicate that these two factors present in these cells have transcriptional activity similar to that of differentiated hepatoma cells. Characterization of the appropriate intertypic cell hybrids demonstrates that this new phenotype is recessive to the dedifferentiated state and fails to be complemented by differentiated cells. These results indicate the existence of mechanisms that inhibit transcription of genes coding for hepatocyte functions in spite of the presence of functional HNF4 and HNF1. Cells of the uncoupled phenotype present certain properties of oval cells described for pathological states of the liver.

Thymidylate synthase (TS) protein expression as a prognostic factor in advanced colorectal cancer: a comparison with TS mRNA expression.

PubMed

Nakagawa, Tateo; Shimada, Mitsuo; Kurita, Nobuhiro; Iwata, Takashi; Nishioka, Masanori; Yoshikawa, Kozo; Higashijima, Jun; Utsunomiya, Tohru

2012-06-01

The role of intratumoral thymidylate synthase (TS) mRNA or protein expression is still controversial and little has been reported regarding relation of them in colorectal cancer. Forty-six patients with advanced colorectal cancer who underwent surgical resection were included. TS mRNA expression was determined by the Danenberg tumor profile method based on laser-captured micro-dissection of the tumor cells. TS protein expression was evaluated using immunohistochemical staining. TS mRNA expression tended to relate TS protein expression. Statistical significance was not found in overall survival between the TS mRNA high group and low group regardless of performing adjuvant chemotherapy. The overall survival in the TS protein negative group was significantly higher than that in positive group in all and the patients without adjuvant chemotherapy. Multivariate analysis showed TS protein expression was as an independent prognostic factor. TS protein expression tends to be related TS mRNA expression and is an independent prognostic factor in advanced colorectal cancer.

Targeting the receptor tyrosine kinase RET in combination with aromatase inhibitors in ER positive breast cancer xenografts.

PubMed

Andreucci, Elena; Francica, Paola; Fearns, Antony; Martin, Lesley-Ann; Chiarugi, Paola; Isacke, Clare M; Morandi, Andrea

2016-12-06

The majority of breast cancers are estrogen receptor positive (ER+). Blockade of estrogen biosynthesis by aromatase inhibitors (AIs) is the first-line endocrine therapy for post-menopausal women with ER+ breast cancers. However, AI resistance remains a major challenge. We have demonstrated previously that increased GDNF/RET signaling in ER+ breast cancers promotes AI resistance. Here we investigated the efficacy of different small molecule RET kinase inhibitors, sunitinib, cabozantinib, NVP-BBT594 and NVP-AST487, and the potential of combining a RET inhibitor with the AI letrozole in ER+ breast cancers. The most effective inhibitor identified, NVP-AST487, suppressed GDNF-stimulated RET downstream signaling and 3D tumor spheroid growth. Ovariectomized mice were inoculated with ER+ aromatase-overexpressing MCF7-AROM1 cells and treated with letrozole, NVP-AST487 or the two drugs in combination. Surprisingly, the three treatment regimens showed similar efficacy in impairing MCF7-AROM1 tumor growth in vivo. However in vitro, NVP-AST487 was superior to letrozole in inhibiting the GDNF-induced motility and tumor spheroid growth of MCF7-AROM1 cells and required in combination with letrozole to inhibit GDNF-induced motility in BT474-AROM3 aromatase expressing cells. These data indicate that inhibiting RET is as effective as the current therapeutic regimen of AI therapy but that a combination treatment may delay cancer cell dissemination and metastasis.

Effects of Two Training Programs on Transcriptional Levels of Neurotrophins and Glial Cells Population in Hippocampus of Experimental Multiple Sclerosis.

PubMed

Naghibzadeh, Maryam; Ranjbar, Rouhollah; Tabandeh, Mohammad Reza; Habibi, Abdolhamid

2018-05-18

The aim of the present study was to investigate the effects of high-intensity interval training (HIIT) versus low-intensity continuous training (LICT) on transcriptional levels of neurotrophic factors and oligodendrocyte/microglia cell loss in a cuprizone (CP) induced animal model of demyelination. Male C57BL/6 mice were assigned to six groups: control (C), cuprizone-induced demyelination (CP), interval training (IT), continuous training (CT), IT plus CP (ITCP), and CT plus CP (CTCP). Training programs on the treadmill were performed for four weeks, and then demyelination was induced by feeding mice a diet containing 0.2% cuprizone for five weeks. Animals that received cuprizone showed poorer motor function, lower expression of BDNF, GDNF, NGF, and fewer oligodendrocytes in the hippocampus compared to the control animals. The numbers of oligodendrocyte and microglia cells increased in the ITCP group compared to the CTCP group (P<0.05). Both training programs increased the mRNA levels of BDNF, GDNF and NGF, and the HIIT program was more effective than the LICT program (P<0.05). Both exercise programs prevented the abnormal neurological movements induced by cuprizone. Our results indicated that HIIT versus LICT had a greater neuroprotective effect against multiple sclerosis by improving gene expression for abnormal neurotrophins and cellular loss in the hippocampus. © Georg Thieme Verlag KG Stuttgart · New York.

Endometrial Expression of Steroidogenic Factor 1 Promotes Cystic Glandular Morphogenesis

PubMed Central

Vasquez, Yasmin M.; Wu, San-Pin; Anderson, Matthew L.; Hawkins, Shannon M.; Creighton, Chad J.; Ray, Madhumita; Tsai, Sophia Y.; Tsai, Ming-Jer; Lydon, John P.

2016-01-01

Epigenetic silencing of steroidogenic factor 1 (SF1) is lost in endometriosis, potentially contributing to de novo local steroidogenesis favoring inflammation and growth of ectopic endometrial tissue. In this study, we examine the impact of SF1 expression in the eutopic uterus by a novel mouse model that conditionally expresses SF1 in endometrium. In vivo SF1 expression promoted the development of enlarged endometrial glands and attenuated estrogen and progesterone responsiveness. Endometriosis induction by autotransplantation of uterine tissue to the mesenteric membrane resulted in the increase in size of ectopic lesions from SF1-expressing mice. By integrating the SF1-dependent transcriptome with the whole genome binding profile of SF1, we identified uterine-specific SF1-regulated genes involved in Wingless and Progesterone receptor-Hedgehog-Chicken ovalbumin upstream promoter transcription factor II signaling for gland development and epithelium-stroma interaction, respectively. The present results indicate that SF1 directly contributes to the abnormal uterine gland morphogenesis, an inhibition of steroid hormone signaling and activation of an immune response, in addition to previously postulated estrogen production. PMID:27018534

Analysis of DOK-6 function in downstream signaling of RET in human neuroblastoma cells.

PubMed

Kurotsuchi, Ai; Murakumo, Yoshiki; Jijiwa, Mayumi; Kurokawa, Kei; Itoh, Yasutomo; Kodama, Yoshinori; Kato, Takuya; Enomoto, Atsushi; Asai, Naoya; Terasaki, Hiroko; Takahashi, Masahide

2010-05-01

Point mutations and structural alterations of the RET tyrosine kinase gene cause multiple endocrine neoplasia type 2 (MEN 2) and papillary thyroid carcinoma, respectively. RET activation by glial cell line-derived neurotrophic factor (GDNF) is essential for the development of the enteric nervous system and the kidney. The signal through RET tyrosine kinase requires several adaptor proteins including the DOK (downstream of kinase) family of proteins. Of the seven members of the DOK protein family, DOK-1, -4, -5, and -6 have been reported to play roles in the GDNF-RET signaling pathway. Although DOK-6 has been shown to bind to RET and promote GDNF-induced neurite outgrowth in mouse Neuro2A cells, DOK-6 function in human cells remains unclear. In the present study, we investigated the role of DOK-6 in GDNF-RET signaling in human cells including neuroblastoma cells. DOK-6 was constitutively localized to the plasma membrane via its pleckstrin homology (PH) domain, and was phosphorylated following RET activation via a MEN2A mutation or GDNF stimulation. However, DOK-6 could not significantly affect downstream signaling and neurite outgrowth in human neuroblastoma cells. The binding affinity of the DOK-6 phosphotyrosine-binding (PTB) domain to RET was much lower than that of the DOK-1, DOK-4, and SHC PTB domains to RET. These findings indicate that DOK-6 is involved in RET signaling with less influence when compared with DOK-1, DOK-4, and SHC.

p27{sup Kip1} inhibits tissue factor expression

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

Breitenstein, Alexander, E-mail: alexander.breitenstein@usz.ch; Cardiovascular Research, Physiology Institute, University of Zurich; Center for Integrative Human Physiology

2013-10-04

Highlights: •p27{sup Kip1}regulates the expression of tissue factor at the transcriptional level. •This inhibitory effect of p27{sup Kip1} is independently of its cell regulatory action. •The current study provides new insights into a pleiotrophic function of p27{sup Kip1}. -- Abstract: Background: The cyclin-dependent kinase inhibitor (CDKI) p27{sup Kip1} regulates cell proliferation and thus inhibits atherosclerosis and vascular remodeling. Expression of tissue factor (TF), the key initator of the coagulation cascade, is associated with atherosclerosis. Yet, it has not been studied whether p27{sup Kip1} influences the expression of TF. Methods and results: p27{sup Kip1} overexpression in human aortic endothelial cells wasmore » achieved by adenoviral transfection. Cells were rendered quiescent for 24 h in 0.5% fetal-calf serum. After stimulation with TNF-α (5 ng/ml), TF protein expression and activity was significantly reduced (n = 4; P < 0.001) in cells transfected with p27{sup Kip1}. In line with this, p27{sup Kip1} overexpression reduced cytokine-induced TF mRNA expression (n = 4; P < 0.01) and TF promotor activity (n = 4; P < 0.05). In contrast, activation of the MAP kinases p38, ERK and JNK was not affected by p27{sup Kip1} overexpression. Conclusion: This in vitro study suggests that p27{sup Kip1} inhibits TF expression at the transcriptional level. These data indicate an interaction between p27{sup Kip1} and TF in important pathological alterations such as atherosclerosis and vascular remodeling.« less

Fibroblast growth factor 8 is expressed at higher levels in lactating human breast and in breast cancer.

PubMed

Zammit, C; Coope, R; Gomm, J J; Shousha, S; Johnston, C L; Coombes, R C

2002-04-08

Fibroblast growth factor 8 can transform NIH3T3 cells and its expression has been found to be associated with breast and prostate cancer. Following our finding that fibroblast growth factor 8 mRNA expression is increased in breast cancer, we have undertaken an immunohistochemistry study of fibroblast growth factor 8 expression in a series of human breast tissues and other normal tissues. Our findings confirm increased expression of fibroblast growth factor 8 in malignant breast tissue but also show significant fibroblast growth factor 8 expression in non-malignant breast epithelial cells. No significant difference in fibroblast growth factor 8 expression was found between different grades of ductal carcinoma, lobular carcinoma and ductal carcinoma in-situ or cancer of different oestrogen receptor, progesterone receptor or nodal status. The highest levels of fibroblast growth factor 8 expression were found in lactating breast tissues and fibroblast growth factor 8 was also detected in human milk. A survey of other normal tissues showed that fibroblast growth factor 8 is expressed in the proliferative cells of the dermis and epithelial cells in colon, ovary fallopian tube and uterus. Fibroblast growth factor 8 appears to be expressed in several organs in man and appears to have an importance in lactation.

The strategy of fusion genes construction determines efficient expression of introduced transcription factors.

PubMed

Adamus, Tomasz; Konieczny, Paweł; Sekuła, Małgorzata; Sułkowski, Maciej; Majka, Marcin

2014-01-01

The main goal in gene therapy and biomedical research is an efficient transcription factors (TFs) delivery system. SNAIL, a zinc finger transcription factor, is strongly involved in tumor, what makes its signaling pathways an interesting research subject. The necessity of tracking activation of intracellular pathways has prompted fluorescent proteins usage as localization markers. Advanced molecular cloning techniques allow to generate fusion proteins from fluorescent markers and transcription factors. Depending on fusion strategy, the protein expression levels and nuclear transport ability are significantly different. The P2A self-cleavage motif through its cleavage ability allows two single proteins to be simultaneously expressed. The aim of this study was to compare two strategies for introducing a pair of genes using expression vector system. We have examined GFP and SNAI1 gene fusions by comprising common nucleotide polylinker (multiple cloning site) or P2A motif in between them, resulting in one fusion or two independent protein expressions respectively. In each case transgene expression levels and translation efficiency as well as nuclear localization of expressed protein have been analyzed. Our data showed that usage of P2A motif provides more effective nuclear transport of SNAIL transcription factor than conventional genes linker. At the same time the fluorescent marker spreads evenly in subcellular space.

Tumor necrosis factor receptor-associated factor 6 (TRAF6) participates in anti-lipopolysaccharide factors (ALFs) gene expression in mud crab.

PubMed

Sun, Wan-Wei; Zhang, Xin-Xu; Wan, Wei-Song; Wang, Shu-Qi; Wen, Xiao-Bo; Zheng, Huai-Ping; Zhang, Yue-Ling; Li, Sheng-Kang

2017-02-01

Tumor necrosis factor receptor-associated factor 6 (TRAF6) is a key cytoplasm signal adaptor that mediates signals activated by tumor necrosis factor receptor (TNFR) superfamily and the Interleukin-1 receptor/Toll-like receptor (IL-1/TLR) superfamily. The full-length 2492 bp TRAF6 (Sp-TRAF6) from Scylla paramamosain contains 1800 bp of open reading frame (ORF) encoding 598 amino acids, including an N-terminal RING-type zinc finger, two TRAF-type zinc fingers and a conserved C-terminal meprin and TRAF homology (MATH) domain. Multiple alignment analysis shows that the putative amino acid sequence of Sp-TRAf6 has highest identity of 88% with Pt-TRAF6 from Portunus trituberculatus, while the similarity of Sp-TRAF6 with other crustacean sequences was 54-55%. RT-PCR analysis indicated that Sp-TRAF6 transcripts were predominantly expressed in the hepatopancreas and stomach, whereas it was barely detected in the heart and hemocytes in our study. Moreover, Sp-TRAF6 transcripts were significantly up-regulated after Vibrio parahemolyticus and LPS challenges. RNA interference assay was carried out used by siRNA to investigate the genes expression patterns regulated by Sp-TRAF6. The qRT-PCR results showed that silencing Sp-TRAF6 gene could inhibit SpALF1, SpALF2, SpALF5 and SpALF6 expression in hemocytes, while inhibit SpALF1, SpALF3, SpALF4, SpALF5 and SpALF6 expression in hepatopancreas. Taken together, the acute-phase response to immune challenges and the inhibition of SpALFs gene expression indicate that Sp-TRAF6 plays an important role in host defense against pathogen invasions via regulation of ALF gene expression in S. paramamosain. Copyright © 2016. Published by Elsevier Ltd.

Expression of a transmembrane phosphotyrosine phosphatase inhibits cellular response to platelet-derived growth factor and insulin-like growth factor-1.

PubMed

Mooney, R A; Freund, G G; Way, B A; Bordwell, K L

1992-11-25

Tyrosine phosphorylation is a mechanism of signal transduction shared by many growth factor receptors and oncogene products. Phosphotyrosine phosphatases (PTPases) potentially modulate or counter-regulate these signaling pathways. To test this hypothesis, the transmembrane PTPase CD45 (leukocyte common antigen) was expressed in the murine cell line C127. Hormone-dependent autophosphorylation of the platelet-derived growth factor (PDGF) and insulin-like growth factor-1 (IGF-1) receptors was markedly reduced in cells expressing the transmembrane PTPase. Tyrosine phosphorylation of other PDGF-dependent phosphoproteins (160, 140, and 55 kDa) and IGF-1-dependent phosphoproteins (145 kDa) was similarly decreased. Interestingly, the pattern of growth factor-independent tyrosine phosphorylations was comparable in cells expressing the PTPase and control cells. This suggests a selectivity or accessibility of the PTPase limited to a subset of cellular phosphotyrosyl proteins. The maximum mitogenic response to PDGF and IGF-1 in cells expressing the PTPase was decreased by 67 and 71%, respectively. These results demonstrate that a transmembrane PTPase can both affect the tyrosine phosphorylation state of growth factor receptors and modulate proximal and distal cellular responses to the growth factors.

Localization of cholinergic innervation and neurturin receptors in adult mouse heart and expression of the neurturin gene.

PubMed

Mabe, Abigail M; Hoard, Jennifer L; Duffourc, Michelle M; Hoover, Donald B

2006-10-01

Neurturin (NRTN) is a neurotrophic factor required during development for normal cholinergic innervation of the heart, but whether NRTN continues to function in the adult heart is unknown. We have therefore evaluated NRTN expression in adult mouse heart and the association of NRTN receptors with intracardiac cholinergic neurons and nerve fibers. Mapping the regional distribution and density of cholinergic nerves in mouse heart was an integral part of this goal. Analysis of RNA from adult C57BL/6 mouse hearts demonstrated NRTN expression in atrial and ventricular tissue. Virtually all neurons in the cardiac parasympathetic ganglia exhibited the cholinergic phenotype, and over 90% of these cells contained both components of the NRTN receptor, Ret tyrosine kinase and GDNF family receptor alpha2 (GFRalpha2). Cholinergic nerve fibers, identified by labeling for the high affinity choline transporter, were abundant in the sinus and atrioventricular nodes, ventricular conducting system, interatrial septum, and much of the right atrium, but less abundant in the left atrium. The right ventricular myocardium contained a low density of cholinergic nerves, which were sparse in other regions of the working ventricular myocardium. Some cholinergic nerves were also associated with coronary vessels. GFRalpha2 was present in most cholinergic nerve fibers and in Schwann cells and their processes throughout the heart. Some cholinergic nerve fibers, such as those in the sinus node, also exhibited Ret immunoreactivity. These findings provide the first detailed mapping of cholinergic nerves in mouse heart and suggest that the neurotrophic influence of NRTN on cardiac cholinergic innervation continues in mature animals.

Quantitative gene expression deregulation in mantle-cell lymphoma: correlation with clinical and biologic factors.

PubMed

Kienle, Dirk; Katzenberger, Tiemo; Ott, German; Saupe, Doreen; Benner, Axel; Kohlhammer, Holger; Barth, Thomas F E; Höller, Sylvia; Kalla, Jörg; Rosenwald, Andreas; Müller-Hermelink, Hans Konrad; Möller, Peter; Lichter, Peter; Döhner, Hartmut; Stilgenbauer, Stephan

2007-07-01

There is evidence for a direct role of quantitative gene expression deregulation in mantle-cell lymphoma (MCL) pathogenesis. Our aim was to investigate gene expression associations with other pathogenic factors and the significance of gene expression in a multivariate survival analysis. Quantitative expression of 20 genes of potential relevance for MCL prognosis and pathogenesis were analyzed using real-time reverse transcriptase polymerase chain reaction and correlated with clinical and genetic factors, tumor morphology, and Ki-67 index in 65 MCL samples. Genomic losses at the loci of TP53, RB1, and P16 were associated with reduced transcript levels of the respective genes, indicating a gene-dosage effect as the pathomechanism. Analysis of gene expression correlations between the candidate genes revealed a separation into two clusters, one dominated by proliferation activators, another by proliferation inhibitors and regulators of apoptosis. Whereas only weak associations were identified between gene expression and clinical parameters or blastoid morphology, several genes were correlated closely with the Ki-67 index, including the short CCND1 variant (positive correlation) and RB1, ATM, P27, and BMI (negative correlation). In multivariate survival analysis, expression levels of MYC, MDM2, EZH2, and CCND1 were the strongest prognostic factors independently of tumor proliferation and clinical factors. These results indicate a pathogenic contribution of several gene transcript levels to the biology and clinical course of MCL. Genes can be differentiated into factors contributing to proliferation deregulation, either by enhancement or loss of inhibition, and proliferation-independent factors potentially contributing to MCL pathogenesis by apoptosis impairment.

Expression of connective tissue growth factor is a prognostic marker for patients with intrahepatic cholangiocarcinoma.

PubMed

Gardini, A; Corti, B; Fiorentino, M; Altimari, A; Ercolani, G; Grazi, G L; Pinna, A D; Grigioni, W F; D'Errico Grigioni, A

2005-04-01

Connective tissue growth factor is a member of the 'CCN' protein family. Consistent with its profibrotic properties, it is over-expressed in several human epithelial malignancies. We have retrospectively evaluated by immunohistochemistry the presence of connective tissue growth factor in archival tissues from 55 resected intrahepatic cholangiocarcinomas and compared its expression to the main pathological parameters, disease free and overall survival. Tumours were scored as high and low/absent expressers (> or =50%, 0-50% cells, respectively). Thirty-three of 55 cholangiocarcinomas (60%) were high and 22 (40%) low expressers. No significant correlation was found between connective tissue growth factor and tumour grade, tumour location, vascular and perineural invasion. Eighteen of 22 (82%) low/absent expressers and 12/33 (36%) high expressers had recurrence of disease (P=0.001). Low/absent expressers showed a poor disease free and overall survival compared with the higher expressers (P<0.001). Vascular invasion was related to tumour recurrence (P=0.025) and to decreased disease free survival (P<0.05). During proportional hazard regression analysis, only connective tissue growth factor was found to influence disease free survival (P=0.01). Expression of connective tissue growth factor is an independent prognostic indicator of both tumour recurrence and overall survival for intrahepatic cholangiocarcinoma patients regardless of tumour location, tumour grade, vascular and perineural invasion.

Several fibroblast growth factors are expressed during pre-attachment bovine conceptus development and regulate interferon-tau expression from trophectoderm.

PubMed

Cooke, Flavia N T; Pennington, Kathleen A; Yang, Qien; Ealy, Alan D

2009-02-01

The trophectoderm-derived factor interferon tau (IFNT) maintains the uterus in a pregnancy-receptive state in cattle and sheep. Fibroblast growth factors (FGFs) are implicated in regulating IFNT expression and potentially other critical events associated with early conceptus development in cattle. The overall objectives of this work were to identify the various FGFs and FGF receptors (FGFRs) expressed in elongating pre-attachment bovine conceptuses and determine if these FGFs regulate conceptus development and/or mediate IFNT production. In vitro-derived bovine blastocysts and in vivo-derived elongated conceptuses collected at day 17 of pregnancy express at least four FGFR subtypes (R1c, R2b, R3c, R4). In addition, transcripts for FGF1, 2, and 10 but not FGF7 are present in elongated bovine conceptuses. The expression pattern of FGF10 most closely resembled that of IFNT, with both transcripts remaining low in day 8 and day 11 conceptuses and increasing substantially in day 14 and day 17 conceptuses. Supplementation with recombinant FGF1, 2 or 10 increased IFNT mRNA levels in bovine trophectoderm cells and bovine blastocysts and increased IFNT protein concentrations in trophectoderm-conditioned medium. Blastocyst development was not affected by any of the FGFs. In summary, at least four FGFRs reside in pre- and peri-attachment bovine conceptuses. Moreover, conceptuses express at least three candidate FGFs during elongation, the time of peak IFNT expression. These findings provide new insight for how conceptus-derived factors such as FGF1, 2, and 10 may control IFNT expression during early pregnancy in cattle.

Hypoxia-inducible factor regulates alphavbeta3 integrin cell surface expression.

PubMed

Cowden Dahl, Karen D; Robertson, Sarah E; Weaver, Valerie M; Simon, M Celeste

2005-04-01

Hypoxia-inducible factor (HIF)-deficient placentas exhibit a number of defects, including changes in cell fate adoption, lack of fetal angiogenesis, hypocellularity, and poor invasion into maternal tissue. HIF is a heterodimeric transcription factor consisting of alpha and beta aryl hydrocarbon receptor nuclear translocator or ARNT) subunits. We used undifferentiated trophoblast stem (TS) cells to characterize HIF-dependent adhesion, migration, and invasion. Arnt(-/-) and Hifalpha(-/-) TS cells exhibit reduced adhesion and migration toward vitronectin compared with wild-type cells. Furthermore, this defect is associated with decreased cell surface expression of integrin alphavbeta3 and significantly decreased expression of this integrin in focal adhesions. Because of the importance of adhesion and migration in tumor progression (in addition to placental development), we examined the affect of culturing B16F0 melanoma cells in 1.5% oxygen (O(2)). Culturing B16F0 melanoma cells at 1.5% O(2) resulted in increased alphavbeta3 integrin surface expression and increased adhesion to and migration toward vitronectin. Together, these data suggest that HIF and O(2) tension influence placental invasion and tumor migration by increasing cell surface expression of alphavbeta3 integrin.

Transcription Factor Binding Site Enrichment Analysis in Co-Expression Modules in Celiac Disease

PubMed Central

Romero-Garmendia, Irati; Jauregi-Miguel, Amaia; Plaza-Izurieta, Leticia; Cros, Marie-Pierre; Legarda, Maria; Irastorza, Iñaki; Herceg, Zdenko; Fernandez-Jimenez, Nora

2018-01-01

The aim of this study was to construct celiac co-expression patterns at a whole genome level and to identify transcription factors (TFs) that could drive the gliadin-related changes in coordination of gene expression observed in celiac disease (CD). Differential co-expression modules were identified in the acute and chronic responses to gliadin using expression data from a previous microarray study in duodenal biopsies. Transcription factor binding site (TFBS) and Gene Ontology (GO) annotation enrichment analyses were performed in differentially co-expressed genes (DCGs) and selection of candidate regulators was performed. Expression of candidates was measured in clinical samples and the activation of the TFs was further characterized in C2BBe1 cells upon gliadin challenge. Enrichment analyses of the DCGs identified 10 TFs and five were selected for further investigation. Expression changes related to active CD were detected in four TFs, as well as in several of their in silico predicted targets. The activation of TFs was further characterized in C2BBe1 cells upon gliadin challenge, and an increase in nuclear translocation of CAMP Responsive Element Binding Protein 1 (CREB1) and IFN regulatory factor-1 (IRF1) in response to gliadin was observed. Using transcriptome-wide co-expression analyses we are able to propose novel genes involved in CD pathogenesis that respond upon gliadin stimulation, also in non-celiac models. PMID:29748492

Transcription Factor Binding Site Enrichment Analysis in Co-Expression Modules in Celiac Disease.

PubMed

Romero-Garmendia, Irati; Garcia-Etxebarria, Koldo; Hernandez-Vargas, Hector; Santin, Izortze; Jauregi-Miguel, Amaia; Plaza-Izurieta, Leticia; Cros, Marie-Pierre; Legarda, Maria; Irastorza, Iñaki; Herceg, Zdenko; Fernandez-Jimenez, Nora; Bilbao, Jose Ramon

2018-05-10

The aim of this study was to construct celiac co-expression patterns at a whole genome level and to identify transcription factors (TFs) that could drive the gliadin-related changes in coordination of gene expression observed in celiac disease (CD). Differential co-expression modules were identified in the acute and chronic responses to gliadin using expression data from a previous microarray study in duodenal biopsies. Transcription factor binding site (TFBS) and Gene Ontology (GO) annotation enrichment analyses were performed in differentially co-expressed genes (DCGs) and selection of candidate regulators was performed. Expression of candidates was measured in clinical samples and the activation of the TFs was further characterized in C2BBe1 cells upon gliadin challenge. Enrichment analyses of the DCGs identified 10 TFs and five were selected for further investigation. Expression changes related to active CD were detected in four TFs, as well as in several of their in silico predicted targets. The activation of TFs was further characterized in C2BBe1 cells upon gliadin challenge, and an increase in nuclear translocation of CAMP Responsive Element Binding Protein 1 (CREB1) and IFN regulatory factor-1 (IRF1) in response to gliadin was observed. Using transcriptome-wide co-expression analyses we are able to propose novel genes involved in CD pathogenesis that respond upon gliadin stimulation, also in non-celiac models.

Growth Factors and COX2 Expression in Canine Perivascular Wall Tumors.

PubMed

Avallone, G; Stefanello, D; Boracchi, P; Ferrari, R; Gelain, M E; Turin, L; Tresoldi, E; Roccabianca, P

2015-11-01

Canine perivascular wall tumors (PWTs) are a group of subcutaneous soft tissue sarcomas developing from vascular mural cells. Mural cells are involved in angiogenesis through a complex crosstalk with endothelial cells mediated by several growth factors and their receptors. The evaluation of their expression may have relevance since they may represent a therapeutic target in the control of canine PWTs. The expression of vascular endothelial growth factor (VEGF) and receptors VEGFR-I/II, basic fibroblast growth factor (bFGF) and receptor Flg, platelet-derived growth factor B (PDGFB) and receptor PDGFRβ, transforming growth factor β1 (TGFβ1) and receptors TGFβR-I/II, and cyclooxygenase 2 (COX2) was evaluated on frozen sections of 40 PWTs by immunohistochemistry and semiquantitatively scored to identify their potential role in PWT development. Statistical analysis was performed to analyze possible correlations between Ki67 labeling index and the expression of each molecule. Proteins of the VEGF-, PDGFB-, and bFGF-mediated pathways were highly expressed in 27 (67.5%), 30 (75%), and 19 (47.5%) of 40 PWTs, respectively. Proteins of the TGFβ1- and COX2-mediated pathways were highly expressed in 4 (10%) and 14 (35%) of 40 cases. Statistical analysis identified an association between VEGF and VEGFR-I/II (P = .015 and .003, respectively), bFGF and Flg (P = .038), bFGF and PDGFRβ (P = .003), and between TGFβ1 and COX2 (P = .006). These findings were consistent with the mechanisms that have been reported to play a role in angiogenesis and in tumor development. No association with Ki67 labeling index was found. VEGF-, PDGFB-, and bFGF-mediated pathways seem to have a key role in PWT development and growth. Blockade of tyrosine kinase receptors after surgery could represent a promising therapy with the aim to reduce the PWT relapse rate and prolong the time to relapse. © The Author(s) 2015.

Trophic and neurotrophic factors in human pituitary adenomas (Review).

PubMed

Spoletini, Marialuisa; Taurone, Samanta; Tombolini, Mario; Minni, Antonio; Altissimi, Giancarlo; Wierzbicki, Venceslao; Giangaspero, Felice; Parnigotto, Pier Paolo; Artico, Marco; Bardella, Lia; Agostinelli, Enzo; Pastore, Francesco Saverio

2017-10-01

The pituitary gland is an organ that functionally connects the hypothalamus with the peripheral organs. The pituitary gland is an important regulator of body homeostasis during development, stress, and other processes. Pituitary adenomas are a group of tumors arising from the pituitary gland: they may be subdivided in functional or non-functional, depending on their hormonal activity. Some trophic and neurotrophic factors seem to play a key role in the development and maintenance of the pituitary function and in the regulation of hypothalamo-pituitary-adrenocortical axis activity. Several lines of evidence suggest that trophic and neurotrophic factors may be involved in pituitary function, thus suggesting a possible role of the trophic and neurotrophic factors in the normal development of pituitary gland and in the progression of pituitary adenomas. Additional studies might be necessary to better explain the biological role of these molecules in the development and progression of this type of tumor. In this review, in light of the available literature, data on the following neurotrophic factors are discussed: ciliary neurotrophic factor (CNTF), transforming growth factors β (TGF‑β), glial cell line-derived neurotrophic factor (GDNF), nerve growth factor (NGF), vascular endothelial growth factor (VEGF), vascular endothelial growth inhibitor (VEGI), fibroblast growth factors (FGFs) and epidermal growth factor (EGF) which influence the proliferation and growth of pituitary adenomas.

Enhancing Peripheral Nerve Regeneration with a Novel Drug Delivering Nerve Conduit

DTIC Science & Technology

2017-12-01

control group ) or a conduit that released GDNF. The main outcome measures were muscle atrophy, electrophysiology, motor endplate reinnervation...prepared NGF+GDNF ( Control groups ). 8 Gastrocnemius Atrophy The gastrocnemius muscle weight of the GDNF treated group was ~ 60% of the non...experimental side at 10 weeks. GDNF conduit group (49.4±1.4 %) had statistically less muscle atrophy than the control group (65.1±5.1 %) (pɘ.05) at 10

Expression of the Maize Dof1 Transcription Factor in Wheat and Sorghum

PubMed Central

Peña, Pamela A.; Quach, Truyen; Sato, Shirley; Ge, Zhengxiang; Nersesian, Natalya; Changa, Taity; Dweikat, Ismail; Soundararajan, Madhavan; Clemente, Tom E.

2017-01-01

Nitrogen is essential for plant growth and development. Improving the ability of plants to acquire and assimilate nitrogen more efficiently is a key agronomic parameter that will augment sustainability in agriculture. A transcription factor approach was pursued to address improvement of nitrogen use efficiency in two major commodity crops. To this end, the Zea mays Dof1 (ZmDof1) transcription factor was expressed in both wheat (Triticum aestivum) and sorghum (Sorghum bicolor) either constitutively, UBI4 promoter from sugarcane, or in a tissue specific fashion via the maize rbcS1 promoter. The primary transcription activation target of ZmDof1, phosphoenolpyruvate carboxylase (PEPC), is observed in transgenic wheat events. Expression ZmDof1 under control of the rbcs1 promoter translates to increase in biomass and yield components in wheat. However, constitutive expression of ZmDof1 led to the down-regulation of genes involved in photosynthesis and the functional apparatus of chloroplasts, and an outcome that negatively impacts photosynthesis, height, and biomass in wheat. Similar patterns were also observed in sorghum transgenic events harboring the constitutive expression cassette of ZmDof1. These results indicate that transcription factor strategies to boost agronomic phenotypic outcomes in crops need to consider expression patterns of the genetic elements to be introduced. PMID:28424717

Hepatocyte growth factor/scatter factor enhances the invasion of mesothelioma cell lines and the expression of matrix metalloproteinases

PubMed Central

Harvey, P; Clark, I M; Jaurand, M-C; Warn, R M; Edwards, D R

2000-01-01

Hepatocyte growth factor/scatter factor (HGF/SF) is a multifunctional factor involved both in development and tissue repair, as well as pathological processes such as cancer and metastasis. It has been identified in vivo in many types of tumours together with its tyrosine kinase receptor, Met. We show here that exogenous HGF/SF acts as a strong chemoattractant for human mesothelioma cell lines. The factor also enhanced cell adhesion to and invasion into Matrigel. The mesothelioma cell lines synthesized a panel of matrix metalloproteinases critical for tumour progression such as MMP-1, 2, 3, 9 and membrane-bound MT1-MMP. HGF/SF stimulated the expression of MMP-1, 9 and MT1-MMP and had a slight effect on expression of the MMP inhibitor TIMP-1 but not TIMP-2. However, there was no simple correlation between the levels of MMPs and TIMPs of the cell lines and their different invasion properties or between HGF/SF stimulatory effects on MMP expression and invasion. In addition, effects of protease inhibitors on invasion suggested that serine proteases were also expressed in human mesothelioma cell lines and were involved in HGF/SF-induced invasion. The results show a predominant role for HGF/SF in mesothelioma cell invasion, stimulating simultaneously adhesion, motility, invasion and regulation of MMP and TIMP levels. © 2000 Cancer Research Campaign PMID:11027427

Targeting tissue factor-expressing tumor angiogenesis and tumors with EF24 conjugated to factor VIIa.

PubMed

Shoji, Mamoru; Sun, Aiming; Kisiel, Walter; Lu, Yang J; Shim, Hyunsuk; McCarey, Bernard E; Nichols, Christopher; Parker, Ernest T; Pohl, Jan; Mosley, Cara A; Alizadeh, Aaron R; Liotta, Dennis C; Snyder, James P

2008-04-01

Tissue factor (TF) is aberrantly expressed on tumor vascular endothelial cells (VECs) and on cancer cells in many malignant tumors, but not on normal VECs, making it a promising target for cancer therapy. As a transmembrane receptor for coagulation factor VIIa (fVIIa), TF forms a high-affinity complex with its cognate ligand, which is subsequently internalized through receptor-mediated endocytosis. Accordingly, we developed a method for selectively delivering EF24, a potent synthetic curcumin analog, to TF-expressing tumor vasculature and tumors using fVIIa as a drug carrier. EF24 was chemically conjugated to fVIIa through a tripeptide-chloromethyl ketone. After binding to TF-expressing targets by fVIIa, EF24 will be endocytosed along with the drug carrier and will exert its cytotoxicity. Our results showed that the conjugate inhibits vascular endothelial growth factor-induced angiogenesis in a rabbit cornea model and in a Matrigel model in athymic nude mice. The conjugate-induced apoptosis in tumor cells and significantly reduced tumor size in human breast cancer xenografts in athymic nude mice as compared with the unconjugated EF24. By conjugating potent drugs to fVIIa, this targeted drug delivery system has the potential to enhance therapeutic efficacy, while reducing toxic side effects. It may also prove to be useful for treating drug-resistant tumors and micro-metastases in addition to primary tumors.

Systematic identification and comparison of expressed profiles of lncRNAs and circRNAs with associated co-expression and ceRNA networks in mouse germline stem cells

PubMed Central

Wu, Ji

2017-01-01

Accumulating evidence indicates that long noncoding RNAs (lncRNAs) and circular RNAs (circRNAs) involve in germ cell development. However, little is known about the functions and mechanisms of lncRNAs and circRNAs in self-renewal and differentiation of germline stem cells. Therefore, we explored the expression profiles of mRNAs, lncRNAs, and circRNAs in male and female mouse germline stem cells by high-throughput sequencing. We identified 18573 novel lncRNAs and 18822 circRNAs in the germline stem cells and further confirmed the existence of these lncRNAs and circRNAs by RT-PCR. The results showed that male and female germline stem cells had similar GDNF signaling mechanism. Subsequently, 8115 mRNAs, 3996 lncRNAs, and 921 circRNAs exhibited sex-biased expression that may be associated with germline stem cell acquisition of the sex-specific properties required for differentiation into gametes. Gene Ontology (GO) and KEGG pathway enrichment analyses revealed different functions for these sex-biased lncRNAs and circRNAs. We further constructed correlated expression networks including coding–noncoding co-expression and competing endogenous RNAs with bioinformatics. Co-expression analysis showed hundreds of lncRNAs were correlated with sex differences in mouse germline stem cells, including lncRNA Gm11851, lncRNA Gm12840, lncRNA 4930405O22Rik, and lncRNA Atp10d. CeRNA network inferred that lncRNA Meg3 and cirRNA Igf1r could bind competitively with miRNA-15a-5p increasing target gene Inha, Acsl3, Kif21b, and Igfbp2 expressions. These findings provide novel perspectives on lncRNAs and circRNAs and lay a foundation for future research into the regulating mechanisms of lncRNAs and circRNAs in germline stem cells. PMID:28404936

In vitro efficacy of a gene-activated nerve guidance conduit incorporating non-viral PEI-pDNA nanoparticles carrying genes encoding for NGF, GDNF and c-Jun.

PubMed

Lackington, William A; Raftery, Rosanne M; O'Brien, Fergal J

2018-06-07

Despite the success of tissue engineered nerve guidance conduits (NGCs) for the treatment of small peripheral nerve injuries, autografts remain the clinical gold standard for larger injuries. The delivery of neurotrophic factors from conduits might enhance repair for more effective treatment of larger injuries but the efficacy of such systems is dependent on a safe, effective platform for controlled and localised therapeutic delivery. Gene therapy might offer an innovative approach to control the timing, release and level of neurotrophic factor production by directing cells to transiently sustain therapeutic protein production in situ. In this study, a gene-activated NGC was developed by incorporating non-viral polyethyleneimine-plasmid DNA (PEI-pDNA) nanoparticles (N/P 7 ratio, 2μg dose) with the pDNA encoding for nerve growth factor (NGF), glial derived neurotrophic factor (GDNF) or the transcription factor c-Jun. The physicochemical properties of PEI-pDNA nanoparticles, morphology, size and charge, were shown to be suitable for gene delivery and demonstrated high Schwann cell transfection efficiency (60±13%) in vitro. While all three genes showed therapeutic potential in terms of enhancing neurotrophic cytokine production while promoting neurite outgrowth, delivery of the gene encoding for c-Jun showed the greatest capacity to enhance regenerative cellular processes in vitro. Ultimately, this gene-activated NGC construct was shown to be capable of transfecting both Schwann cells (S42 cells) and neuronal cells (PC12 and dorsal root ganglia) in vitro, demonstrating potential for future therapeutic applications in vivo. The basic requirements of biomaterial-based nerve guidance conduits have now been well established and include being able to bridge a nerve injury to support macroscopic guidance between nerve stumps, while being strong enough to withstand longitudinal tension and circumferential compression, in addition to being mechanically sound to facilitate

Understanding Transcription Factor Regulation by Integrating Gene Expression and DNase I Hypersensitive Sites.

PubMed

Wang, Guohua; Wang, Fang; Huang, Qian; Li, Yu; Liu, Yunlong; Wang, Yadong

2015-01-01

Transcription factors are proteins that bind to DNA sequences to regulate gene transcription. The transcription factor binding sites are short DNA sequences (5-20 bp long) specifically bound by one or more transcription factors. The identification of transcription factor binding sites and prediction of their function continue to be challenging problems in computational biology. In this study, by integrating the DNase I hypersensitive sites with known position weight matrices in the TRANSFAC database, the transcription factor binding sites in gene regulatory region are identified. Based on the global gene expression patterns in cervical cancer HeLaS3 cell and HelaS3-ifnα4h cell (interferon treatment on HeLaS3 cell for 4 hours), we present a model-based computational approach to predict a set of transcription factors that potentially cause such differential gene expression. Significantly, 6 out 10 predicted functional factors, including IRF, IRF-2, IRF-9, IRF-1 and IRF-3, ICSBP, belong to interferon regulatory factor family and upregulate the gene expression levels responding to the interferon treatment. Another factor, ISGF-3, is also a transcriptional activator induced by interferon alpha. Using the different transcription factor binding sites selected criteria, the prediction result of our model is consistent. Our model demonstrated the potential to computationally identify the functional transcription factors in gene regulation.

A sigma factor toolbox for orthogonal gene expression in Escherichia coli

PubMed Central

Van Brempt, Maarten; Van Nerom, Katleen; Van Hove, Bob; Maertens, Jo; De Mey, Marjan; Charlier, Daniel

2018-01-01

Abstract Synthetic genetic sensors and circuits enable programmable control over timing and conditions of gene expression and, as a result, are increasingly incorporated into the control of complex and multi-gene pathways. Size and complexity of genetic circuits are growing, but stay limited by a shortage of regulatory parts that can be used without interference. Therefore, orthogonal expression and regulation systems are needed to minimize undesired crosstalk and allow for dynamic control of separate modules. This work presents a set of orthogonal expression systems for use in Escherichia coli based on heterologous sigma factors from Bacillus subtilis that recognize specific promoter sequences. Up to four of the analyzed sigma factors can be combined to function orthogonally between each other and toward the host. Additionally, the toolbox is expanded by creating promoter libraries for three sigma factors without loss of their orthogonal nature. As this set covers a wide range of transcription initiation frequencies, it enables tuning of multiple outputs of the circuit in response to different sensory signals in an orthogonal manner. This sigma factor toolbox constitutes an interesting expansion of the synthetic biology toolbox and may contribute to the assembly of more complex synthetic genetic systems in the future. PMID:29361130

[Effect of human oviductal embryotrophic factors on gene expression of mouse preimplantation embryos].

PubMed

Yao, Yuan-Qing; Lee, Kai-Fai; Xu, Jia-Seng; Ho, Pak-Chung; Yeung, Shu-Biu

2007-09-01

To investigate the effect of embryotrophic factors (ETF) from human oviductal cells on gene expression of mouse early developmental embryos and discuss the role of fallopian tube in early development of embryos. ETF was isolated from conditioned medium of human oviductal cell line by sequential liquid chromatographic systems. Mouse embryos were treated by ETF in vitro. Using differential display RT-PCR, the gene expression of embryos treated by ETF was compared with embryos without ETF treatment. The differentially expressed genes were separated, re-amplified, cloned and sequenced. Gene expression profiles of embryos with ETF treatment was different from embryos without this treatment. Eight differentially expressed genes were cloned and sequenced. These genes functioned in RNA degradation, synthesis, splicing, protein trafficking, cellular differentiation and embryo development. Embryotrophic factors from human oviductal cells affect gene expression of early developmental embryos. The human oviductal cells play wide roles in early developmental stages of embryos.

Role of Estrogens in the Size of Neuronal Somata of Paravaginal Ganglia in Ovariectomized Rabbits

PubMed Central

Hernández-Aragón, Laura G.; García-Villamar, Verónica; Carrasco-Ruiz, María de los Ángeles; Nicolás-Toledo, Leticia; Ortega, Arturo; Cuevas-Romero, Estela; Martínez-Gómez, Margarita

2017-01-01

We aimed to determine the role of estrogens in modulating the size of neuronal somata of paravaginal ganglia. Rabbits were allocated into control (C), ovariectomized (OVX), and OVX treated with estradiol benzoate (OVX + EB) groups to evaluate the neuronal soma area; total serum estradiol (E2) and testosterone (T) levels; the percentage of immunoreactive (ir) neurons anti-aromatase, anti-estrogen receptor (ERα, ERβ) and anti-androgen receptor (AR); the intensity of the immunostaining anti-glial cell line-derived neurotrophic factor (GDNF) and the GDNF family receptor alpha type 1 (GFRα1); and the number of satellite glial cells (SGCs) per neuron. There was a decrease in the neuronal soma size for the OVX group, which was associated with low T, high percentages of aromatase-ir and neuritic AR-ir neurons, and a strong immunostaining anti-GDNF and anti-GFRα1. The decrease in the neuronal soma size was prevented by the EB treatment that increased the E2 without affecting the T levels. Moreover, there was a high percentage of neuritic AR-ir neurons, a strong GDNF immunostaining in the SGC, and an increase in the SGCs per neuron. Present findings show that estrogens modulate the soma size of neurons of the paravaginal ganglia, likely involving the participation of the SGC. PMID:28316975

Expression of connective tissue growth factor in the livers of non-viral hepatocellular carcinoma patients with metabolic risk factors.

PubMed

Akahoshi, Keiichi; Tanaka, Shinji; Mogushi, Kaoru; Shimada, Shu; Matsumura, Satoshi; Akiyama, Yoshimitsu; Aihara, Arihiro; Mitsunori, Yusuke; Ban, Daisuke; Ochiai, Takanori; Kudo, Atsushi; Arii, Shigeki; Tanabe, Minoru

2016-09-01

The incidence of hepatocellular carcinoma (HCC) associated with metabolic risk factors, such as diabetes and obesity, has been increasing. However, the underlying mechanism that links these diseases remains unclear. We performed genome-wide expression analysis of human liver tissues of non-viral HCC patients with or without metabolic risk factors. The upregulated genes that associated with diabetes and obesity were investigated by in vitro and in vivo experiments, and immunohistochemistry of human liver tissues was performed. Among the upregulated genes, connective tissue growth factor (CTGF) expression was induced to a greater extent by combined glucose and insulin administration to human hepatoma cells. Genome-wide expression analysis revealed upregulation of a chemokine network in CTGF-overexpressing hepatoma cells, which displayed an increased ability to induce in vitro activation of macrophages, and in vivo infiltration of liver macrophages. Immunohistochemistry of human liver tissues validated the correlations between CTGF expression and diabetes or obesity as well as activation of liver macrophages in patients with non-viral HCC. Recurrence-free survival was significantly poorer in the CTGF-positive patients compared with the CTGF-negative patients (p = 0.002). Multivariate analysis determined that CTGF expression (HR 2.361; 95 % CI 1.195-4.665; p = 0.013) and vascular invasion (HR 2.367; 95 % CI 1.270-4.410; p = 0.007) were independent prognostic factors for recurrence of non-viral HCC. Our data suggest that CTGF could be involved in oncogenic pathways promoting non-viral HCC associated with metabolic risk factors via induction of liver inflammation and is expected to be a novel HCC risk biomarker and potential therapeutic target.

Advanced Glycation End-Products affect transcription factors regulating insulin gene expression

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

Puddu, A., E-mail: alep100@hotmail.com; Storace, D.; Odetti, P.

2010-04-23

Advanced Glycation End-Products (AGEs) are generated by the covalent interaction of reducing sugars with proteins, lipids or nucleic acids. AGEs are implicated in diabetic complications and pancreatic {beta}-cell dysfunction. We previously demonstrated that exposure of the pancreatic islet cell line HIT-T15 to high concentrations of AGEs leads to a significant decrease of insulin secretion and content. Insulin gene transcription is positively regulated by the beta cell specific transcription factor PDX-1 (Pancreatic and Duodenal Homeobox-1). On the contrary, the forkhead transcription factor FoxO1 inhibits PDX-1 gene transcription. Activity of FoxO1 is regulated by post-translational modifications: phosphorylation deactivates FoxO1, and acetylation preventsmore » FoxO1 ubiquitination. In this work we investigated whether AGEs affect expression and subcellular localization of PDX-1 and FoxO1. HIT-T15 cells were cultured for 5 days in presence of AGEs. Cells were then lysed and processed for subcellular fractionation. We determined intracellular insulin content, then we assessed the expression and subcellular localization of PDX-1, FoxO1, phosphoFoxO1 and acetylFoxO1. As expected intracellular insulin content was lower in HIT-T15 cells cultured with AGEs. The results showed that AGEs decreased expression and nuclear localization of PDX-1, reduced phosphorylation of FoxO1, and increased expression and acetylation of FoxO1. These results suggest that AGEs decrease insulin content unbalancing transcription factors regulating insulin gene expression.« less

Muscle and Liver Carbohydrates: Response to Military Task Performance by Women and Men

DTIC Science & Technology

2000-10-01

rapidly synthesize glycogen from three-carbon compounds generated by muscle metabolism and taken up by the liver ( gluconeogenesis ). 20 UNPUBLISHED DATA...49008 Glial cell line-derived neturotrophic factor (GDNF) is a recently discovered nerrotrophic factor that afflets peripheral motor neurons . Increased

Placental growth factor expression is reversed by antivascular endothelial growth factor therapy under hypoxic conditions.

PubMed

Zhou, Ai-Yi; Bai, Yu-Jing; Zhao, Min; Yu, Wen-Zhen; Huang, Lv-Zhen; Li, Xiao-Xin

2014-08-01

Clinical trials have revealed that the antivascular endothelial growth factor (VEGF) therapies are effective in retinopathy of prematurity (ROP). But the low level of VEGF was necessary as a survival signal in healthy conditions, and endogenous placental growth factor (PIGF) is redundant for development. The purpose of this study was to elucidate the PIGF expression under hypoxia as well as the influence of anti-VEGF therapy on PIGF. CoCl2-induced hypoxic human umbilical vein endothelial cells (HUVECs) were used for an in vitro study, and oxygen-induced retinopathy (OIR) mice models were used for an in vivo study. The expression patterns of PIGF under hypoxic conditions and the influence of anti-VEGF therapy on PIGF were evaluated by quantitative reverse transcription-polymerase chain reaction (RTPCR). The retinal avascular areas and neovascularization (NV) areas of anti-VEGF, anti-PIGF and combination treatments were calculated. Retina PIGF concentration was evaluated by ELISA after treatment. The vasoactive effects of exogenous PIGF on HUVECs were investigated by proliferation and migration studies. PIGF mRNA expression was reduced by hypoxia in OIR mice, in HUVECs under hypoxia and anti-VEGF treatment. However, PIGF expression was reversed by anti-VEGF therapy in the OIR model and in HUVECs under hypoxia. Exogenous PIGF significantly inhibited HUVECs proliferation and migration under normal conditions, but it stimulated cell proliferation and migration under hypoxia. Anti-PIGF treatment was effective for neovascular tufts in OIR mice (P<0.05). The finding that PIGF expression is iatrogenically up-regulated by anti-VEGF therapy provides a consideration to combine it with anti-PIGF therapy.

Nuclear factor I-A represses expression of the cell adhesion molecule L1

PubMed Central

2009-01-01

Background The neural cell adhesion molecule L1 plays a crucial role in development and plasticity of the nervous system. Neural cells thus require precise control of L1 expression. Results We identified a full binding site for nuclear factor I (NFI) transcription factors in the regulatory region of the mouse L1 gene. Electrophoretic mobility shift assay (EMSA) showed binding of nuclear factor I-A (NFI-A) to this site. Moreover, for a brain-specific isoform of NFI-A (NFI-A bs), we confirmed the interaction in vivo using chromatin immunoprecipitation (ChIP). Reporter gene assays showed that in neuroblastoma cells, overexpression of NFI-A bs repressed L1 expression threefold. Conclusion Our findings suggest that NFI-A, in particular its brain-specific isoform, represses L1 gene expression, and might act as a second silencer of L1 in addition to the neural restrictive silencer factor (NRSF). PMID:20003413

Expression of transcription factors during sodium phenylacetate induced erythroid differentiation in K562 cells.

PubMed

Rath, A V; Schmahl, G E; Niemeyer, C M

1997-01-01

During 15 days of treatment of K562 cells with sodium phenylacetate, we observed an increase in the cellular hemoglobin concentration with a similar increase in the expression of gamma-globin mRNA. Morphological studies demonstrated characteristic features of erythroid differentiation and maturation. At the same time there was no change in the level of expression of the cell surface antigenes CD33, CD34, CD45, CD71 and glycophorin A. Likewise, the level of expression of the erythroid transcription factors GATA-1, GATA-2, NF-E2, SCL and RBTN2, all expressed in untreated K562 cells, did not increase during sodium phenylacetate induced erythroid differentiation. The expression of the nuclear factors Evi-1 and c-myb, known to inhibit erythroid differentiation, did not decrease. We conclude that sodium phenylacetate treatment of K562 cells increases gamma-globin mRNA and induces cell maturation as judged by morphology without affecting the expression of the erythroid transcription factors, some of which are known to be involved in the regulation of beta-like globin genes.

Extracellular matrix metalloproteinase inducer (EMMPRIN) expression correlates positively with active angiogenesis and negatively with basic fibroblast growth factor expression in epithelial ovarian cancer.

PubMed

Szubert, Sebastian; Szpurek, Dariusz; Moszynski, Rafal; Nowicki, Michal; Frankowski, Andrzej; Sajdak, Stefan; Michalak, Slawomir

2014-03-01

The primary aim of this paper was to evaluate the expression of extracellular matrix metalloproteinase inducer (EMMPRIN) and its relationship with proangiogenic factors and microvessel density (MVD) in ovarian cancer. The study group included 58 epithelial ovarian cancers (EOCs), 35 benign ovarian tumors, and 21 normal ovaries. The expression of EMMPRIN, vascular endothelial growth factor (VEGF), and basic fibroblast growth factor (bFGF) was assessed by ELISA of tissue homogenates. Antibodies against CD105, CD31, and CD34 were used to immunohistochemically assess MVD. We have found significantly higher EMMPRIN expression in EOC than in benign ovarian tumors and normal ovaries. Similarly, the VEGF expression was higher in EOC than in benign ovarian tumors and normal ovaries. By contrast, bFGF expression was lower in EOC than in benign ovarian tumors and ovary samples. EMMPRIN expression in EOC was directly correlated with VEGF expression and CD105-MVD, but inversely correlated with bFGF expression. Grade 2/3 ovarian cancers had increased expression of EMMPRIN and VEGF, increased CD105-MVD, and lowered expression of bFGF compared to grade 1 ovarian cancers. Moreover, EMMPRIN expression was higher in advanced (FIGO III and IV) ovarian cancer. The upregulation of EMMPRIN and VEGF expression is correlated with increased CD105-MVD and silenced bFGF, which suggests early and/or reactivated angiogenesis in ovarian cancer. Aggressive EOC is characterized by the following: high expression of EMMPRIN and VEGF, high CD105-MVD, and low expression of bFGF.

Fibroblast growth factor-2 promotes keratan sulfate proteoglycan expression by keratocytes in vitro

NASA Technical Reports Server (NTRS)

Long, C. J.; Roth, M. R.; Tasheva, E. S.; Funderburgh, M.; Smit, R.; Conrad, G. W.; Funderburgh, J. L.

2000-01-01

Keratocytes of the corneal stroma produce a specialized extracellular matrix responsible for corneal transparency. Corneal keratan sulfate proteoglycans (KSPG) are unique products of keratocytes that are down-regulated in corneal wounds and in vitro. This study used cultures of primary bovine keratocytes to define factors affecting KSPG expression in vitro. KSPG metabolically labeled with [(35)S]sulfate decreased during the initial 2-4 days of culture in quiescent cultures with low serum concentrations (0.1%). Addition of fetal bovine serum, fibroblast growth factor-2 (FGF-2), transforming growth factor beta, or platelet derived growth factor all stimulated cell division, but only FGF-2 stimulated KSPG secretion. Combined with serum, FGF-2 also prevented serum-induced KSPG down-regulation. KSPG secretion was lost during serial subculture with or without FGF-2. Expression of KSPG core proteins (lumican, mimecan, and keratocan) was stimulated by FGF-2, and steady state mRNA pools for these proteins, particularly keratocan, were significantly increased by FGF-2 treatment. KSPG expression therefore is supported by exogenous FGF-2 and eliminated by subculture of the cells in presence of serum. FGF-2 stimulates KSPG core protein expression primarily through an increase in mRNA pools.

Transforming growth factor-beta inhibits the expression of clock genes.

PubMed

Gast, Heidemarie; Gordic, Sonja; Petrzilka, Saskia; Lopez, Martin; Müller, Andreas; Gietl, Anton; Hock, Christoph; Birchler, Thomas; Fontana, Adriano

2012-07-01

Disturbances of sleep-wake rhythms are an important problem in Alzheimer's disease (AD). Circadian rhythms are regulated by clock genes. Transforming growth factor-beta (TGF-β) is overexpressed in neurons in AD and is the only cytokine that is increased in cerebrospinal fluid (CSF). Our data show that TGF-β2 inhibits the expression of the clock genes Period (Per)1, Per2, and Rev-erbα, and of the clock-controlled genes D-site albumin promoter binding protein (Dbp) and thyrotroph embryonic factor (Tef). However, our results showed that TGF-β2 did not alter the expression of brain and muscle Arnt-like protein-1 (Bmal1). The concentrations of TGF-β2 in the CSF of 2 of 16 AD patients and of 1 of 7 patients with mild cognitive impairment were in the dose range required to suppress the expression of clock genes. TGF-β2-induced dysregulation of clock genes may alter neuronal pathways, which may be causally related to abnormal sleep-wake rhythms in AD patients. © 2012 New York Academy of Sciences.

Obesity-induced endoplasmic reticulum stress suppresses nuclear factor-Y expression.

PubMed

Liu, Yulan; Zhang, Yuwei; Zhang, Yanjie; Zhang, Jinlong; Liu, Yin; Feng, Peiqun; Su, Zhiguang

2017-02-01

Nuclear transcription factor Y (NF-Y) is an evolutionarily conserved transcription factor composed of three subunits, NF-YA, NF-YB, and NF-YC. NF-Y plays crucial roles in pre-adipocyte maintenance and/or commitment to adipogenesis. NF-YA dysfunction in adipocyte resulted in an age-dependent progressive loss of adipose tissue associated with metabolic complications. Endoplasmic reticulum (ER) stress has emerged as an important mediator in the pathogenesis of obesity. However, it is not known if NF-YA is involved in the ER stress-mediated pathogenesis of obesity. We first examined the effects of ER stress on the NF-YA expression in cultured 3T3-L1 adipocytes; then in ob/ob genetic obesity mice, we tested the effect of chemical chaperones alleviating ER stress on the expression levels of NF-YA. Subsequently, we inhibited the new mRNA synthesis using actinomycin D in 3T3-L1 cells to explore the mechanism modulating NF-YA expression. Finally, we evaluated the involvement of PPARg in the regulation of NF-YA expression by ER stress. We demonstrated that both obesity- and chemical chaperone -induced ER stress suppressed NF-YA expression and alleviation of ER stress by chemical chaperone could recover NF-YA expression in ob/ob mice. Moreover, we showed that ER stress suppressed NF-YA mRNA transcription through the involvement of peroxisome proliferator-activated receptor gamma (PPARg). Activation of PPARg ameliorates the ER stress-induced NF-YA suppression. Our findings may point to a possible role of NF-YA in stress conditions that occur in chronic obesity, ER stress might be involved in the pathogenesis of obesity through NF-YA depletion.

High-Dimensional Sparse Factor Modeling: Applications in Gene Expression Genomics

PubMed Central

Carvalho, Carlos M.; Chang, Jeffrey; Lucas, Joseph E.; Nevins, Joseph R.; Wang, Quanli; West, Mike

2010-01-01

We describe studies in molecular profiling and biological pathway analysis that use sparse latent factor and regression models for microarray gene expression data. We discuss breast cancer applications and key aspects of the modeling and computational methodology. Our case studies aim to investigate and characterize heterogeneity of structure related to specific oncogenic pathways, as well as links between aggregate patterns in gene expression profiles and clinical biomarkers. Based on the metaphor of statistically derived “factors” as representing biological “subpathway” structure, we explore the decomposition of fitted sparse factor models into pathway subcomponents and investigate how these components overlay multiple aspects of known biological activity. Our methodology is based on sparsity modeling of multivariate regression, ANOVA, and latent factor models, as well as a class of models that combines all components. Hierarchical sparsity priors address questions of dimension reduction and multiple comparisons, as well as scalability of the methodology. The models include practically relevant non-Gaussian/nonparametric components for latent structure, underlying often quite complex non-Gaussianity in multivariate expression patterns. Model search and fitting are addressed through stochastic simulation and evolutionary stochastic search methods that are exemplified in the oncogenic pathway studies. Supplementary supporting material provides more details of the applications, as well as examples of the use of freely available software tools for implementing the methodology. PMID:21218139

Expression of transforming growth factor alpha and epidermal growth factor receptor messenger RNA in neoplastic and nonneoplastic human kidney tissue.

PubMed

Mydlo, J H; Michaeli, J; Cordon-Cardo, C; Goldenberg, A S; Heston, W D; Fair, W R

1989-06-15

Using Northern blot analysis, we have demonstrated that mRNA for transforming growth factor alpha (TGF-alpha) was expressed in five malignant kidney tissue specimens but was not detected in their autologous nonneoplastic homologues. In addition, the expression of epidermal growth factor (EGF) receptor mRNA in these malignant tissues was 2- to 3-fold greater than in nontransformed tissues. In two cases examined using immunohistochemistry, we were able to correlate the increased expression of the mRNA with an increase in protein expression. Since TGF-alpha is known to bind to the EGF receptor, the finding of an increased expression of both TGF-alpha and EGF receptor mRNA in kidney tumor tissue suggests that interaction between TGF-alpha and the EGF receptor may play a role in promoting transformation and/or proliferation of kidney neoplasms, perhaps by an autocrine mechanism.

Connective tissue growth factor immunohistochemical expression is associated with gallbladder cancer progression.

PubMed

Garcia, Patricia; Leal, Pamela; Alvarez, Hector; Brebi, Priscilla; Ili, Carmen; Tapia, Oscar; Roa, Juan C

2013-02-01

Gallbladder cancer (GBC) is an aggressive neoplasia associated with late diagnosis, unsatisfactory treatment, and poor prognosis. Molecular mechanisms involved in GBC pathogenesis remain poorly understood. Connective tissue growth factor (CTGF) is thought to play a role in the pathologic processes and is overexpressed in several human cancers, including GBC. No information is available about CTGF expression in early stages of gallbladder carcinogenesis. Objective.- To evaluate the expression level of CTGF in benign and malignant lesions of gallbladder and its correlation with clinicopathologic features and GBC prognosis. Connective tissue growth factor protein was examined by immunohistochemistry on tissue microarrays containing tissue samples of chronic cholecystitis (n = 51), dysplasia (n = 15), and GBC (n = 169). The samples were scored according to intensity of staining as low/absent and high CTGF expressers. Statistical analysis was performed using the χ(2) test or Fisher exact probability test with a significance level of P < .05. Survival analysis was assessed by the Kaplan-Meier method and the log-rank test. Connective tissue growth factor expression showed a progressive increase from chronic cholecystitis to dysplasia and then to early and advanced carcinoma. Immunohistochemical expression (score ≥2) was significantly higher in advanced tumors, in comparison with chronic cholecystitis (P < .001) and dysplasia (P = .03). High levels of CTGF expression correlated with better survival (P = .04). Our results suggest a role for CTGF in GBC progression and a positive association with better prognosis. In addition, they underscore the importance of considering the involvement of inflammation on GBC development.

Neural progenitor cell implants modulate vascular endothelial growth factor and brain-derived neurotrophic factor expression in rat axotomized neurons.

PubMed

Talaverón, Rocío; Matarredona, Esperanza R; de la Cruz, Rosa R; Pastor, Angel M

2013-01-01

Axotomy of central neurons leads to functional and structural alterations which largely revert when neural progenitor cells (NPCs) are implanted in the lesion site. The new microenvironment created by NPCs in the host tissue might modulate in the damaged neurons the expression of a high variety of molecules with relevant roles in the repair mechanisms, including neurotrophic factors. In the present work, we aimed to analyze changes in neurotrophic factor expression in axotomized neurons induced by NPC implants. For this purpose, we performed immunofluorescence followed by confocal microscopy analysis for the detection of vascular endothelial growth factor (VEGF), brain-derived neurotrophic factor (BDNF), neurotrophin-3 (NT-3) and nerve growth factor (NGF) on brainstem sections from rats with axotomy of abducens internuclear neurons that received NPC implants (implanted group) or vehicle injections (axotomized group) in the lesion site. Control abducens internuclear neurons were strongly immunoreactive to VEGF and BDNF but showed a weak staining for NT-3 and NGF. Comparisons between groups revealed that lesioned neurons from animals that received NPC implants showed a significant increase in VEGF content with respect to animals receiving vehicle injections. However, the immunoreactivity for BDNF, which was increased in the axotomized group as compared to control, was not modified in the implanted group. The modifications induced by NPC implants on VEGF and BDNF content were specific for the population of axotomized abducens internuclear neurons since the neighboring abducens motoneurons were not affected. Similar levels of NT-3 and NGF immunolabeling were obtained in injured neurons from axotomized and implanted animals. Among all the analyzed neurotrophic factors, only VEGF was expressed by the implanted cells in the lesion site. Our results point to a role of NPC implants in the modulation of neurotrophic factor expression by lesioned central neurons, which might

Expression of LIM-homeodomain transcription factors in the developing and mature mouse retina

PubMed Central

Balasubramanian, Revathi; Bui, Andrew; Ding, Qian; Gan, Lin

2014-01-01

LIM-homeodomain (LIM-HD) transcription factors have been extensively studied for their role in the development of the central nervous system. Their function is key to several developmental events like cell proliferation, differentiation and subtype specification. However, their roles in retinal neurogenesis remain largely unknown. Here we report a detailed expression study of LIM-HD transcription factors LHX9 and LHX2, LHX3 and LHX4, and LHX6 in the developing and mature mouse retina using immunohistochemistry and in situ hybridization techniques. We show that LHX9 is expressed during the early stages of development in the retinal ganglion cell layer and the inner nuclear layer. We also show that LHX9 is expressed in a subset of amacrine cells in the adult retina. LHX2 is known to be expressed in retinal progenitor cells during development and in Müller glial cells and a subset of amacrine cells in the adult retina. We found that the LHX2 subset of amacrine cells is not cholinergic and that a very few of LHX2 amacrine cells express calretinin. LHX3 and LHX4 are expressed in a subset of bipolar cells in the adult retina. LHX6 is expressed in cells in the ganglion cell layer and the neuroblast layer starting at embryonic stage 13.5 (E13.5) and continues to be expressed in cells in the ganglion cell layer and inner nuclear layer, postnatally, suggesting its likely expression in amacrine cells or a subset thereof. Taken together, our comprehensive assay of expression patterns of LIM-HD transcription factors during mouse retinal development will help further studies elucidating their biological functions in the differentiation of retinal cell subtypes. PMID:24333658

Clinically significant association of elevated expression of nuclear factor E2-related factor 2 expression with higher glucose uptake and progression of upper urinary tract cancer.

PubMed

Nukui, Akinori; Narimatsu, Takahiro; Kambara, Tsunehito; Abe, Hideyuki; Sakamoto, Setsu; Yoshida, Ken-Ichiro; Kamai, Takao

2018-05-02

There is growing evidence that the transcription factor nuclear factor E2-related factor 2 (Nrf2) is the major participant in regulating antioxidants and pathways for detoxifying reactive oxygen species (ROS), as well as having a vital role in tumor proliferation, invasion, and chemoresistance. It was also recently reported that Nrf2 supports cell proliferation by promoting metabolic activity. Thus, Nrf2 is involved in progression of cancer. Upper urinary tract urothelial carcinoma (UTUC) is a biologically aggressive tumor with high rates of recurrence and progression, resulting in a poor prognosis. However, the role of Nrf2 in UTUC is largely unknown. In order to study the role of Nrf2 in UTUC from the metabolic perspective, we retrospectively assessed Nrf2 expression in the surgical specimen and the preoperative maximum standard glucose uptake (SUVmax) on [ 18 F]fluorodeoxy-glucose positron emission tomography ( 18 F-FDG-PET) of 107 patients with UTUC who underwent radical nephroureterectomy. Increased expression of Nrf2 in the primary lesion was correlated with less differentiated histology, local invasion, and lymph node metastasis, and was also an independent indicator of shorter overall survival according to multivariate analysis. Furthermore, increased expression of Nrf2 was associated with higher preoperative SUVmax by the primary tumor on 18 F-FDG-PET, while Nrf2 expression and SUVmax were also significantly correlated in the metastatic lymph nodes. Among the 18 patients with lymph node metastasis at nephroureterectomy who underwent retroperitoneal lymph node dissection and received adjuvant chemotherapy, the patients with higher Nrf2 expression in the primary tumor had worse recurrence-free survival. These results suggest that constitutive activation of Nrf2 might be linked with tumor aerobic glycolysis and progression of UTUC, indicating that Nrf2 signaling in the tumor microenvironment promotes progression of UTUC.

Regulation of endogenous human gene expression by ligand-inducible TALE transcription factors.

PubMed

Mercer, Andrew C; Gaj, Thomas; Sirk, Shannon J; Lamb, Brian M; Barbas, Carlos F

2014-10-17

The construction of increasingly sophisticated synthetic biological circuits is dependent on the development of extensible tools capable of providing specific control of gene expression in eukaryotic cells. Here, we describe a new class of synthetic transcription factors that activate gene expression in response to extracellular chemical stimuli. These inducible activators consist of customizable transcription activator-like effector (TALE) proteins combined with steroid hormone receptor ligand-binding domains. We demonstrate that these ligand-responsive TALE transcription factors allow for tunable and conditional control of gene activation and can be used to regulate the expression of endogenous genes in human cells. Since TALEs can be designed to recognize any contiguous DNA sequence, the conditional gene regulatory system described herein will enable the design of advanced synthetic gene networks.

The phosphodiesterase inhibitor, ibudilast, attenuates neuroinflammation in the MPTP model of Parkinson’s disease

PubMed Central

Joniec-Maciejak, Ilona; Cudna, Agnieszka; Mirowska-Guzel, Dagmara; Kurkowska-Jastrzębska, Iwona

2017-01-01

Background/Aims Since the degeneration of the nigrostriatal dopaminergic pathway in Parkinson’s disease (PD) is associated with the inflammation process and decreased levels of cyclic nucleotides, inhibition of up-regulated cyclic nucleotide phosphodiesterases (PDEs) appears to be a promising therapeutic strategy. We used ibudilast (IBD), a non-selective PDE3,4,10,11 inhibitor, due to the abundant PDE 4 and 10 expression in the striatum. The present study for the first time examined the efficacy of IBD in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) mouse model of PD. Methods IBD [0, 20, 30, 40, or 50 mg/kg] was injected b.i.d. subcutaneously for nine days to three-month-old male C57Bl/10Tar mice, beginning two days prior to MPTP (60 mg/kg) intoxication. High-pressure liquid chromatography, Western blot analysis, and real time RT-PCR methods were applied. Results Our study demonstrated that chronic administration of IBD attenuated astroglial reactivity and increased glial cell-derived neurotrophic factor (GDNF) production in the striatum. Moreover, IBD reduced TNF-α, IL-6, and IL-1β expression. Conclusion IBD had a well-defined effect on astroglial activation in the mouse model of PD; however, there was no protective effect in the acute phase of injury. Diminished inflammation and an increased level of GDNF may provide a better outcome in the later stages of neurodegeneration. PMID:28753652

Identification of trans-acting factors regulating SamDC expression in Oryza sativa

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

Basu, Supratim, E-mail: supratim_genetics@yahoo.co.in; Division of Plant Biology, Bose Institute, Kolkata; Roychoudhury, Aryadeep

2014-03-07

Highlights: • Identification of cis elements responsible for SamDC expression by in silico analysis. • qPCR analysis of SamDC expression to abiotic and biotic stress treatments. • Detection of SamDC regulators using identified cis-elements as probe by EMSA. • Southwestern Blot analysis to predict the size of the trans-acting factors. - Abstract: Abiotic stress affects the growth and productivity of crop plants; to cope with the adverse environmental conditions, plants have developed efficient defense machinery comprising of antioxidants like phenolics and flavonoids, and osmolytes like polyamines. SamDC is a key enzyme in the polyamine biosynthesis pathway in plants. In ourmore » present communication we have done in silico analysis of the promoter region of SamDC to look for the presence of different cis-regulatory elements contributing to its expression. Based on the presence of different cis-regulatory elements we completed comparative analysis of SamDC gene expression in rice lamina of IR-29 and Nonabokra by qPCR in response to the abiotic stress treatments of salinity, drought, cold and the biotic stress treatments of ABA and light. Additionally, to explore the role of the cis-regulatory elements in regulating the expression of SamDC gene in plants we comparatively analyzed the binding of rice nuclear proteins prepared from IR-29 and Nonabokra undergoing various stress treatments. The intensity of the complex formed was low and inducible in IR-29 in contrast to Nonabokra. Southwestern blot analysis helped in predicting the size of the trans-acting factors binding to these cis-elements. To our knowledge this is the first report on the comprehensive analysis of SamDC gene expression in rice and identification of the trans-acting factors regulating its expression.« less

Vitamin D regulates tyrosine hydroxylase expression: N-cadherin a possible mediator.

PubMed

Cui, X; Pertile, R; Liu, P; Eyles, D W

2015-09-24

Vitamin D is a neuroactive steroid. Its genomic actions are mediated via the active form of vitamin D, 1,25(OH)2D3, binding to the vitamin D receptor (VDR). The VDR emerges in the rat mesencephalon at embryonic day 12, representing the peak period of dopaminergic cell birth. Our prior studies reveal that developmental vitamin D (DVD)-deficiency alters the ontogeny of dopaminergic neurons in the developing mesencephalon. There is also consistent evidence from others that 1,25(OH)2D3 promotes the survival of dopaminergic neurons in models of dopaminergic toxicity. In both developmental and toxicological studies it has been proposed that 1,25(OH)2D3 may modulate the differentiation and maturation of dopaminergic neurons; however, to date there is lack of direct evidence. The aim of the current study is to investigate this both in vitro using a human SH-SY5Y cell line transfected with rodent VDR and in vivo using a DVD-deficient model. Here we show that in VDR-expressing SH-SY5Y cells, 1,25(OH)2D3 significantly increased production of tyrosine hydroxylase (TH), the rate-limiting enzyme in dopamine synthesis. This effect was dose- and time-dependent, but was not due to an increase in TH-positive cell number, nor was it due to the production of trophic survival factors for dopamine neurons such as glial-derived neurotrophic factor (GDNF). In accordance with 1,25(OH)2D3's anti-proliferative actions in the brain, 1,25(OH)2D3 reduced the percentage of dividing cells from approximately 15-10%. Given the recently reported role of N-cadherin in the direct differentiation of dopaminergic neurons, we examined here whether it may be elevated by 1,25(OH)2D3. We confirmed this in vitro and more importantly, we showed DVD-deficiency decreases N-cadherin expression in the embryonic mesencephalon. In summary, in our in vitro model we have shown 1,25(OH)2D3 increases TH expression, decreases proliferation and elevates N-cadherin, a potential factor that mediates these processes

Cancer Associated Fibroblasts express pro-inflammatory factors in human breast and ovarian tumors

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

Erez, Neta, E-mail: netaerez@post.tau.ac.il; Glanz, Sarah; Raz, Yael

Highlights: •CAFs in human breast and ovarian tumors express pro-inflammatory factors. •Expression of pro-inflammatory factors correlates with tumor invasiveness. •Expression of pro-inflammatory factors is associated with NF-κb activation in CAFs. -- Abstract: Inflammation has been established in recent years as a hallmark of cancer. Cancer Associated Fibroblasts (CAFs) support tumorigenesis by stimulating angiogenesis, cancer cell proliferation and invasion. We previously demonstrated that CAFs also mediate tumor-enhancing inflammation in a mouse model of skin carcinoma. Breast and ovarian carcinomas are amongst the leading causes of cancer-related mortality in women and cancer-related inflammation is linked with both these tumor types. However, themore » role of CAFs in mediating inflammation in these malignancies remains obscure. Here we show that CAFs in human breast and ovarian tumors express high levels of the pro-inflammatory factors IL-6, COX-2 and CXCL1, previously identified to be part of a CAF pro-inflammatory gene signature. Moreover, we show that both pro-inflammatory signaling by CAFs and leukocyte infiltration of tumors are enhanced in invasive ductal carcinoma as compared with ductal carcinoma in situ. The pro-inflammatory genes expressed by CAFs are known NF-κB targets and we show that NF-κB is up-regulated in breast and ovarian CAFs. Our data imply that CAFs mediate tumor-promoting inflammation in human breast and ovarian tumors and thus may be an attractive target for stromal-directed therapeutics.« less

Gene therapy for Parkinson's disease: Disease modification by GDNF family of ligands.

PubMed

Kirik, Deniz; Cederfjäll, Erik; Halliday, Glenda; Petersén, Åsa

2017-01-01

Gene transfer is a promising drug delivery method of advanced therapeutic entities for Parkinson's disease. One advantage over conventional therapies, such as peripheral delivery of the dopamine pre-cursor l-DOPA, is site-specific expression of proteins with regenerative, disease-modifying and potentially neuroprotective capacity. Several clinical trials have been performed to test the capacity of glial-cell line derived neurotrophic factor and neurturin to rescue degenerating dopaminergic neurons in the substantia nigra and their axon terminals in the striatum by delivery of these neurotrophic factors either as purified protein or by means of viral vector mediated gene delivery to the brain. Although gene therapy approaches tested so far have been shown to be safe, none met their primary endpoints in phase II clinical trials designed and powered to test the efficacy of the intervention. Within the scope of this review we aim to describe the state-of-the-art in the field, how different technical parameters were translated from pre-clinical studies in non-human primates to clinical trials, and what these trials taught us regarding important factors that may pave the way to the success of gene therapy for the treatment of Parkinson's disease. Copyright © 2016 Elsevier Inc. All rights reserved.

[Expression of connective tissue growth factor in colorectal cancer and its association with prognosis].

PubMed

Sun, Zheng; Yang, Ping; Liang, Li-yuan; Zhang, Tong; Zhang, Wei-jian; Cao, Jie

2012-11-01

To investigate the expression of connective tissue growth factor (CTGF) in colorectal cancer(CRC) and its association with clinicopathologic parameters and overall survival rate. Fresh tumor tissues and matched distal normal colon tissues were collected from 92 patients diagnosed as CRC by surgical operation. The expression level of CTGF mRNA was quantified by quantitative reverse transcription PCR. Thirty out of 92 pairs of tissue specimens were selected randomly to detect CTGF protein by immunohistochemistry. All the cases were followed up to identify prognostic factors for survival. CTGF mRNA expression was up-regulated in CRC. The positive rate of CTGF protein expression tissues (73.3%) was significantly higher than that in the corresponding normal tissues (23.3%, P<0.01). CTGF expression was lower in patients with lymphatic metastasis or stage III/IIII disease (all P<0.05). A negative association was also observed between the CTGF protein positive rate and tumor infiltration depth (P<0.05). The relative expression of CTGF mRNA in tumor tissues was classified into high and low expression groups. The 5-year cumulative survival rate was lower in patients with low CTGF expression (29.3%) as compared to those with high CTGF expressions (68.3%) (P<0.01). Cox regression analysis revealed that the relative expression level of CTGF was independent factor of overall survival (RR=2.960, 95%CI:1.491-1.587, P<0.01). ROC curve analysis showed that sensitivity and specificity of CTGF mRNA expression for prediction of 5-year survival were 64.9% and 74.5%, respectively. The aberrant expression of CTGF is associated with the malignant biological behaviors of CRC. Low expression of CTGF is associated with worse prognosis of CRC.

Immunohistochemical expression of vascular endothelial growth factor in canine oral squamous cell carcinomas.

PubMed

Martano, Manuela; Restucci, Brunella; Ceccarelli, Dora Maria; Lo Muzio, Lorenzo; Maiolino, Paola

2016-01-01

Angiogenesis is crucial for the growth and metastasis of malignant tumours, and various proangiogenic factors promote this process. One of these factors is vascular endothelial growth factor (VEGF), which appears to play a key role in tumour angiogenesis. The aim of the present study was to assess whether VEGF expression is associated with angiogenesis, disease progression and neoplastic proliferation in canine oral squamous cell carcinoma (OSCC) tissue. VEGF immunoreactivity was quantified by immunohistochemistry in 30 specimens, including normal oral mucosa and OSCC tissues graded as well, moderately or poorly differentiated. VEGF expression was correlated with tumour cell proliferation, as assessed using the proliferating cell nuclear antigen (PCNA) marker and microvessel density (data already published). The present results revealed that VEGF and PCNA expression increased significantly between normal oral tissue and neoplastic tissue, and between well and moderately/poorly differentiated tumours. In addition, VEGF expression was strongly correlated with PCNA expression and microvessel density. It was concluded that VEGF may promote angiogenesis through a paracrine pathway, stimulating endothelial cell proliferation and, similarly, may induce tumour cell proliferation through an autocrine pathway. The present results suggest that the evaluation of VEGF may be a useful additional criterion for estimating malignancy and growth potential in canine OSCCs.

A previously unrecognized role of C3a in proteinuric progressive nephropathy

PubMed Central

Morigi, Marina; Locatelli, Monica; Rota, Cinzia; Buelli, Simona; Corna, Daniela; Rizzo, Paola; Abbate, Mauro; Conti, Debora; Perico, Luca; Longaretti, Lorena; Benigni, Ariela; Zoja, Carlamaria; Remuzzi, Giuseppe

2016-01-01

Podocyte loss is the initial event in the development of glomerulosclerosis, the structural hallmark of progressive proteinuric nephropathies. Understanding mechanisms underlying glomerular injury is the key challenge for identifying novel therapeutic targets. In mice with protein-overload induced by bovine serum albumin (BSA), we evaluated whether the alternative pathway (AP) of complement mediated podocyte depletion and podocyte-dependent parietal epithelial cell (PEC) activation causing glomerulosclerosis. Factor H (Cfh−/−) or factor B-deficient mice were studied in comparison with wild-type (WT) littermates. WT+BSA mice showed podocyte depletion accompanied by glomerular complement C3 and C3a deposits, PEC migration to capillary tuft, proliferation, and glomerulosclerosis. These changes were more prominent in Cfh−/− +BSA mice. The pathogenic role of AP was documented by data that factor B deficiency preserved glomerular integrity. In protein-overload mice, PEC dysregulation was associated with upregulation of CXCR4 and GDNF/c-Ret axis. In vitro studies provided additional evidence of a direct action of C3a on proliferation and CXCR4-related migration of PECs. These effects were enhanced by podocyte-derived GDNF. In patients with proteinuric nephropathy, glomerular C3/C3a paralleled PEC activation, CXCR4 and GDNF upregulation. These results indicate that mechanistically uncontrolled AP complement activation is not dispensable for podocyte-dependent PEC activation resulting in glomerulosclerosis. PMID:27345360

Phenotypic differences between oral and skin fibroblasts in wound contraction and growth factor expression.

PubMed

Shannon, Diane B; McKeown, Scott T W; Lundy, Fionnuala T; Irwin, Chris R

2006-01-01

Wounds of the oral mucosa heal in an accelerated fashion with reduced scarring compared with cutaneous wounds. The differences in healing outcome between oral mucosa and skin could be because of phenotypic differences between the respective fibroblast populations. This study compared paired mucosal and dermal fibroblasts in terms of collagen gel contraction, alpha-smooth muscle actin expression (alpha-SMA), and production of the epithelial growth factors: keratinocyte growth factor (KGF) and hepatocyte growth factor/scatter factor (HGF). The effects of transforming growth factor -beta1 and -beta3 on each parameter were also determined. Gel contraction in floating collagen lattices was determined over a 7-day period. alpha-SMA expression by fibroblasts was determined by Western blotting. KGF and HGF expression were determined by an enzyme-linked immunosorbent assay. Oral fibroblasts induced accelerated collagen gel contraction, yet surprisingly expressed lower levels of alpha-SMA. Oral cells also produced significantly greater levels of both KGF and HGF than their dermal counterparts. Transforming growth factor-beta1 and -beta3, over the concentration range of 0.1-10 ng/mL, had similar effects on cell function, stimulating both gel contraction and alpha-SMA production, but inhibiting KGF and HGF production by both cell types. These data indicate phenotypic differences between oral and dermal fibroblasts that may well contribute to the differences in healing outcome between these two tissues.

Dynamic expression of transcription factor Brn3b during mouse cranial nerve development

PubMed Central

Sajgo, Szilard; Ali, Seid; Popescu, Octavian; Badea, Tudor Constantin

2015-01-01

During development transcription factor combinatorial codes define a large variety of morphologically and physiologically distinct neurons. Such a combinatorial code has been proposed for the differentiation of projection neurons of the somatic and visceral components of cranial nerves. It is possible that individual neuronal cell types are not specified by unique transcription factors, but rather emerge through the intersection of their expression domains. Brn3a, Brn3b and Brn3c, in combination with each other and/or transcription factors of other families, can define subgroups of Retinal Ganglion Cells (RGC), Spiral and Vestibular Ganglia, inner ear and vestibular hair cell neurons in the vestibuloacoustic system, and groups of somatosensory neurons in the Dorsal Root Ganglia (DRG). In the present study we investigated the expression and potential role of the Brn3b transcription factor in cranial nerves and associated nuclei of the brainstem. We report the dynamic expression of Brn3b in the somatosensory component of cranial nerves II, V, VII and VIII and visceromotor nuclei of nerves VII, IX, X, as well as other brainstem nuclei during different stages of development into adult stage. We find that genetically identified Brn3bKO RGC axons show correct but delayed pathfinding during the early stages of embryonic development. However loss of Brn3b does not affect the anatomy of the other cranial nerves normally expressing this transcription factor. PMID:26356988

Increased expression of placental growth factor in high-grade endometrial carcinoma.

PubMed

Coenegrachts, Lieve; Schrauwen, Stefanie; Van Bree, Rita; Despierre, Evelyn; Luyten, Catherine; Jonckx, Bart; Stassen, Jean Marie; Vergote, Ignace; Amant, Frédéric

2013-02-01

Placental growth factor (PlGF), a homolog of vascular endothelial growth factor (VEGF), exerts pleiotropic functions in cancer by affecting tumor cells as well as endothelial and inflammatory cells. Moreover, PlGF expression correlates with tumor stage, recurrence, metastasis and patient outcome in different types of cancer. Recently, administration of anti-PlGF therapy reduced tumor growth and metastasis in preclinical tumor models. In the present study, we evaluated the diagnostic and prognostic value of systemic and local expression of PlGF in primary endometrial carcinomas. PlGF levels in tumor lysates (n=128) and serum (n=88) of patients with primary endometrial cancer were determined using ELISA. PlGF mRNA expression in endometrial carcinoma tissues was quantified by quantitative qRT-PCR. Results were compared to endometrial cancer stage and grade. Systemic PlGF levels were not altered in patients with endometrial cancer (FIGO stage I-II-III) as compared to healthy controls. Only in FIGO stage IV patients, serum PlGF levels were slightly increased. Local PlGF mRNA and protein expression in endometrial tumors progressively increased with tumor grade. In endometrioid carcinomas, PlGF mRNA expression was significantly increased in endometrioid grade 3 tumors as compared to normal endometrial tissue. PlGF protein expression was significantly increased in endometrioid grade 2 and 3 carcinomas and in serous carcinomas as compared to normal endometrial tissue. Our study showed that systemic/serum PlGF levels cannot be used as a diagnostic or prognostic marker in endometrial cancer. However, the increased local expression of PlGF, primarily in high-grade carcinomas, underscores the possibility for preclinical assessment of anti-PlGF therapy in endometrial cancer.

Correlation between spontaneous apoptosis and the expression of angiogenic factors in advanced gastric adenocarcinoma.

PubMed

Ikeguchi, M; Cai, J; Fukuda, K; Oka, S; Katano, K; Tsujitani, S; Maeta, M; Kaibara, N

2001-06-01

The aim of this study was to investigate whether angiogenic factors influence the occurrence of spontaneous apoptosis in advanced gastric cancer. The apoptotic indices (AIs) of 97 tumors from 97 patients with advanced gastric cancer (pT3, pN0, pM0, Stage II) were analyzed by the terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate biotin nick end labeling (TUNEL) method. Intratumoral microvessel densities (IMVDs) of tumors stained with anti-CD34 monoclonal antibody were quantified under x 200 magnification using computer-assisted image analysis. The expressions of angiogenic factors, such as vascular endothelial growth factor (VEGF), thymidine phosphorylase (dThdPase), transforming growth factor-alpha (TGF-alpha), and p53 were analyzed immunohistochemically and compared with IMVDs and AIs. The mean IMVD of the 97 tumors was 365/mm2 (range 147-990/mm2). The mean AI of tumors was 2.1% (range 0-11.3%). A significant inverse correlation between the AIs and the IMVDs was shown (p = -0.278, P = 0.0064). The mean IMVDs of tumors with high expressions of dThdPase, TGF-alpha, or p53 were significantly higher than those of tumors with low expressions of these factors. The mean AI of tumors with high expressions of dThdPase was significantly lower than that of tumors with low expressions of dThdPase (P = 0.023). However, no significant correlations were detected between AIs and the expression levels of VEGF, TGF-alpha, or p53. In gastric cancer, dThdPase may play an important role in tumor progression by increasing microvessels and by suppressing apoptosis of cancer cells.

CD2-associated Protein (CD2AP) Enhances Casitas B Lineage Lymphoma-3/c (Cbl-3/c)-mediated Ret Isoform-specific Ubiquitination and Degradation via Its Amino-terminal Src Homology 3 Domains*

PubMed Central

Calco, Gina N.; Stephens, Olivia R.; Donahue, Laura M.; Tsui, Cynthia C.; Pierchala, Brian A.

2014-01-01

Ret is the receptor tyrosine kinase for the glial cell line-derived neurotrophic factor (GDNF) family of neuronal growth factors. Upon activation by GDNF, Ret is rapidly polyubiquitinated and degraded. This degradation process is isoform-selective, with the longer Ret51 isoform exhibiting different degradation kinetics than the shorter isoform, Ret9. In sympathetic neurons, Ret degradation is induced, at least in part, by a complex consisting of the adaptor protein CD2AP and the E3-ligase Cbl-3/c. Knockdown of Cbl-3/c using siRNA reduced the GDNF-induced ubiquitination and degradation of Ret51 in neurons and podocytes, suggesting that Cbl-3/c was a predominant E3 ligase for Ret. Coexpression of CD2AP with Cbl-3/c augmented the ubiquitination of Ret51 as compared with the expression of Cbl-3/c alone. Ret51 ubiquitination by the CD2AP·Cbl-3/c complex required a functional ring finger and TKB domain in Cbl-3/c. The SH3 domains of CD2AP were sufficient to drive the Cbl-3/c-dependent ubiquitination of Ret51, whereas the carboxyl-terminal coiled-coil domain of CD2AP was dispensable. Interestingly, activated Ret induced the degradation of CD2AP, but not Cbl-3/c, suggesting a potential inhibitory feedback mechanism. There were only two major ubiquitination sites in Ret51, Lys1060 and Lys1107, and the combined mutation of these lysines almost completely eliminated both the ubiquitination and degradation of Ret51. Ret9 was not ubiquitinated by the CD2AP·Cbl-3/c complex, suggesting that Ret9 was down-regulated by a fundamentally different mechanism. Taken together, these results suggest that only the SH3 domains of CD2AP were necessary to enhance the E3 ligase activity of Cbl-3/c toward Ret51. PMID:24425877

Industrial production of clotting factors: Challenges of expression, and choice of host cells.

PubMed

Kumar, Sampath R

2015-07-01

The development of recombinant forms of blood coagulation factors as safer alternatives to plasma derived factors marked a major advance in the treatment of common coagulation disorders. These are complex proteins, mostly enzymes or co-enzymes, involving multiple post-translational modifications, and therefore are difficult to express. This article reviews the nature of the expression challenges for the industrial production of these factors, vis-à-vis the translational and post-translational bottlenecks, as well as the choice of host cell lines for high-fidelity production. For achieving high productivities of vitamin K dependent proteins, which include factors II (prothrombin), VII, IX and X, and protein C, host cell limitation of γ-glutamyl carboxylation is a major bottleneck. Despite progress in addressing this, involvement of yet unidentified protein(s) impedes a complete cell engineering solution. Human factor VIII expresses at very low levels due to limitations at several steps in the protein secretion pathway. Protein and cell engineering, vector improvement and alternate host cells promise improvement in the productivity. Production of Von Willebrand factor is constrained by its large size, complex structure, and the need for extensive glycosylation and disulfide-bonded oligomerization. All the licensed therapeutic factors are produced in CHO, BHK or HEK293 cells. While HEK293 is a recent adoption, BHK cells appear to be disfavored. Copyright © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

G =  MAT: linking transcription factor expression and DNA binding data.

PubMed

Tretyakov, Konstantin; Laur, Sven; Vilo, Jaak

2011-01-31

Transcription factors are proteins that bind to motifs on the DNA and thus affect gene expression regulation. The qualitative description of the corresponding processes is therefore important for a better understanding of essential biological mechanisms. However, wet lab experiments targeted at the discovery of the regulatory interplay between transcription factors and binding sites are expensive. We propose a new, purely computational method for finding putative associations between transcription factors and motifs. This method is based on a linear model that combines sequence information with expression data. We present various methods for model parameter estimation and show, via experiments on simulated data, that these methods are reliable. Finally, we examine the performance of this model on biological data and conclude that it can indeed be used to discover meaningful associations. The developed software is available as a web tool and Scilab source code at http://biit.cs.ut.ee/gmat/.

G = MAT: Linking Transcription Factor Expression and DNA Binding Data

PubMed Central

Tretyakov, Konstantin; Laur, Sven; Vilo, Jaak

2011-01-01

Transcription factors are proteins that bind to motifs on the DNA and thus affect gene expression regulation. The qualitative description of the corresponding processes is therefore important for a better understanding of essential biological mechanisms. However, wet lab experiments targeted at the discovery of the regulatory interplay between transcription factors and binding sites are expensive. We propose a new, purely computational method for finding putative associations between transcription factors and motifs. This method is based on a linear model that combines sequence information with expression data. We present various methods for model parameter estimation and show, via experiments on simulated data, that these methods are reliable. Finally, we examine the performance of this model on biological data and conclude that it can indeed be used to discover meaningful associations. The developed software is available as a web tool and Scilab source code at http://biit.cs.ut.ee/gmat/. PMID:21297945

The expression of pigment epithelium-derived factor in bladder transitional cell carcinoma.

PubMed

Jang, Tae Jung; Kim, Sung Woo; Lee, Kyung Seop

2012-06-01

Pigment epithelium-derived factor (PEDF) is an anti-angiogenic factor. The purpose of this study is to examine the involvement of PEDF in the angiogenesis and biological behavior of bladder transitional cell carcinoma (TCC). We examined the expression of PEDF in 99 bladder TCCs and ten non-neoplastic tissues, and evaluated microvessel density (MVD). The positive immunoreactivity for PEDF was seen in normal urothelium in 60% (6/10) and TCC in 13% (13/99). The PEDF expression had a significant correlation with MVD, i.e., a low MVD in 42% (5/12), a middle MVD in 11% (8/76) and a high MVD 0% (0/11) of tumors. The PEDF expression was not significantly correlated with the differentiation and invasion of TCC, but the degree of MVD was significantly higher in both high grade TCC and the pT2 tumors. The degree of PEDF expression is significantly higher in normal bladder urothelium than bladder TCC; it is inversely correlated with the angiogenesis; and it is not related to the differentiation and progression of TCC. It can therefore be concluded that bladder TCC would initially occur if there is a lack of the PEDF expression.

Pathway-based factor analysis of gene expression data produces highly heritable phenotypes that associate with age.

PubMed

Anand Brown, Andrew; Ding, Zhihao; Viñuela, Ana; Glass, Dan; Parts, Leopold; Spector, Tim; Winn, John; Durbin, Richard

2015-03-09

Statistical factor analysis methods have previously been used to remove noise components from high-dimensional data prior to genetic association mapping and, in a guided fashion, to summarize biologically relevant sources of variation. Here, we show how the derived factors summarizing pathway expression can be used to analyze the relationships between expression, heritability, and aging. We used skin gene expression data from 647 twins from the MuTHER Consortium and applied factor analysis to concisely summarize patterns of gene expression to remove broad confounding influences and to produce concise pathway-level phenotypes. We derived 930 "pathway phenotypes" that summarized patterns of variation across 186 KEGG pathways (five phenotypes per pathway). We identified 69 significant associations of age with phenotype from 57 distinct KEGG pathways at a stringent Bonferroni threshold ([Formula: see text]). These phenotypes are more heritable ([Formula: see text]) than gene expression levels. On average, expression levels of 16% of genes within these pathways are associated with age. Several significant pathways relate to metabolizing sugars and fatty acids; others relate to insulin signaling. We have demonstrated that factor analysis methods combined with biological knowledge can produce more reliable phenotypes with less stochastic noise than the individual gene expression levels, which increases our power to discover biologically relevant associations. These phenotypes could also be applied to discover associations with other environmental factors. Copyright © 2015 Brown et al.

Pathway-Based Factor Analysis of Gene Expression Data Produces Highly Heritable Phenotypes That Associate with Age

PubMed Central

Anand Brown, Andrew; Ding, Zhihao; Viñuela, Ana; Glass, Dan; Parts, Leopold; Spector, Tim; Winn, John; Durbin, Richard

2015-01-01

Statistical factor analysis methods have previously been used to remove noise components from high-dimensional data prior to genetic association mapping and, in a guided fashion, to summarize biologically relevant sources of variation. Here, we show how the derived factors summarizing pathway expression can be used to analyze the relationships between expression, heritability, and aging. We used skin gene expression data from 647 twins from the MuTHER Consortium and applied factor analysis to concisely summarize patterns of gene expression to remove broad confounding influences and to produce concise pathway-level phenotypes. We derived 930 “pathway phenotypes” that summarized patterns of variation across 186 KEGG pathways (five phenotypes per pathway). We identified 69 significant associations of age with phenotype from 57 distinct KEGG pathways at a stringent Bonferroni threshold (P<5.38×10−5). These phenotypes are more heritable (h2=0.32) than gene expression levels. On average, expression levels of 16% of genes within these pathways are associated with age. Several significant pathways relate to metabolizing sugars and fatty acids; others relate to insulin signaling. We have demonstrated that factor analysis methods combined with biological knowledge can produce more reliable phenotypes with less stochastic noise than the individual gene expression levels, which increases our power to discover biologically relevant associations. These phenotypes could also be applied to discover associations with other environmental factors. PMID:25758824

Expression of the Eukaryotic Translation Initiation Factors 4E and 2α in Non-Hodgkin’s Lymphomas

PubMed Central

Wang, Songtao; Rosenwald, Igor B.; Hutzler, Michael J.; Pihan, German A.; Savas, Lou; Chen, Jane-Jane; Woda, Bruce A.

1999-01-01

Transition of cells from quiescence to proliferation requires an increase in the rate of protein synthesis, which is regulated in part by two key translation initiation factors, 4E and 2α. The expression and activity of both factors are increased transiently when normal resting cells are stimulated to proliferate. They are constitutively elevated in oncogene transformed cultured cells, and overexpression of either initiation factor in rodent cells makes them tumorigenic. In this study we investigate an association between the expression of translation initiation factors and lymphomagenesis. We have analyzed the expression of the protein synthesis initiation factors 4E and 2α by immunohistochemistry in reactive lymph nodes and several types of non-Hodgkin’s lymphoma representing a wide range of clinical behaviors based on the Revised European-American Lymphoma behavioral classification. The study included 7 benign lymph nodes with follicular hyperplasia, 26 indolent lymphomas (6 marginal zone lymphomas, 7 small lymphocytic lymphomas, and 13 follicular lymphomas, grades 1 and 2), 16 moderately aggressive lymphomas (8 mantle cell lymphomas and 8 follicular lymphomas, grade 3), 24 aggressive lymphomas (14 large-B-cell lymphomas and 10 anaplastic large-cell lymphomas), and 15 highly aggressive lymphomas (7 lymphoblastic lymphomas and 8 Burkitt’s lymphomas). Strong expression of initiation factors 4E and 2α was demonstrated in the germinal centers of reactive follicles. Minimal or no expression was seen in the mantle zones and surrounding paracortices, indicating that high expression of initiation factors 4E and 2α is associated with the active proliferation of lymphocytes. Most cases of aggressive and highly aggressive lymphomas showed strong expression of initiation factors 4E and 2α, in contrast to the cases of indolent and moderately aggressive lymphoma, in which their expression was intermediate between the germinal centers and the mantles of reactive

Expression of pigment epithelium-derived factor and tumor necrosis factor-α is correlated in bladder tumor and is related to tumor angiogenesis.

PubMed

Feng, Chen-Chen; Wang, Pao-Hsun; Ding, Qiang; Guan, Ming; Zhang, Yuan-Fang; Jiang, Hao-Wen; Wen, Hui; Wu, Zhong

2013-02-01

Angiogenesis is a pivotal process on which solid tumor growth is substantially dependent. Pigment epithelium-derived factor (PEDF) is the most potent natural anti-angiogenic factor, which has seldom been studied in bladder tumor, and whose functioning pathway remains unclear. We have thus investigated PEDF expression in relation to tumor necrosis factor-α (TNF-α) and microvessel density (MVD) with immunohistochemistry. Antibodies of PEDF and TNF-α were examined by Western blotting before immunohistochemistry. Sixty-four urothelial tumor sections and 23 normal controls were stained and expression of PEDF, TNF-α, and MVD were studied. Decreased PEDF expression and increased TNF-α expression was noticed in tumorous tissue compared with healthy urothelium. Lower PEDF expression was related to higher tumor grade but stage. Increased TNF-α expression was noticed in recurrent, larger tumors as well as in tumors with progression in grade and stage. Expression of PEDF and TNF-α was correlated in bladder tumor. PEDF or TNF-α was correlated with MVD negatively or positively, respectively, in cancerous tissue and tumorous grouping without correlation in papillary urothelial neoplasm of low malignant potential. Expressional change of PEDF and TNF-α is in relation to angiogenesis of bladder tumor, especially in bladder cancer development. Copyright © 2013 Elsevier Inc. All rights reserved.

Increased expression of placental growth factor in high-grade endometrial carcinoma

PubMed Central

COENEGRACHTS, LIEVE; SCHRAUWEN, STEFANIE; VAN BREE, RITA; DESPIERRE, EVELYN; LUYTEN, CATHERINE; JONCKX, BART; STASSEN, JEAN MARIE; VERGOTE, IGNACE; AMANT, FRÉDÉRIC

2013-01-01

Placental growth factor (PlGF), a homolog of vascular endothelial growth factor (VEGF), exerts pleiotropic functions in cancer by affecting tumor cells as well as endothelial and inflammatory cells. Moreover, PlGF expression correlates with tumor stage, recurrence, metastasis and patient outcome in different types of cancer. Recently, administration of anti-PlGF therapy reduced tumor growth and metastasis in preclinical tumor models. In the present study, we evaluated the diagnostic and prognostic value of systemic and local expression of PlGF in primary endometrial carcinomas. PlGF levels in tumor lysates (n=128) and serum (n=88) of patients with primary endometrial cancer were determined using ELISA. PlGF mRNA expression in endometrial carcinoma tissues was quantified by quantitative qRT-PCR. Results were compared to endometrial cancer stage and grade. Systemic PlGF levels were not altered in patients with endometrial cancer (FIGO stage I-II-III) as compared to healthy controls. Only in FIGO stage IV patients, serum PlGF levels were slightly increased. Local PlGF mRNA and protein expression in endometrial tumors progressively increased with tumor grade. In endometrioid carcinomas, PlGF mRNA expression was significantly increased in endometrioid grade 3 tumors as compared to normal endometrial tissue. PlGF protein expression was significantly increased in endometrioid grade 2 and 3 carcinomas and in serous carcinomas as compared to normal endometrial tissue. Our study showed that systemic/serum PlGF levels cannot be used as a diagnostic or prognostic marker in endometrial cancer. However, the increased local expression of PlGF, primarily in high-grade carcinomas, underscores the possibility for preclinical assessment of anti-PlGF therapy in endometrial cancer. PMID:23232836

Fluoxetine Ameliorates Behavioral and Neuropathological Deficits in a Transgenic Model Mouse of α-synucleinopathy

PubMed Central

Ubhi, Kiren; Inglis, Chandra; Mante, Michael; Patrick, Christina; Adame, Anthony; Spencer, Brian; Rockenstein, Edward; May, Verena; Winkler, Juergen; Masliah, Eliezer

2013-01-01

The term α-synucleinopathies refers to a group of age-related neurological disorders including Parkinson’s disease (PD), Dementia with Lewy Bodies (DLB) and Multiple System Atrophy (MSA) that display an abnormal accumulation of alpha-synuclein (α-syn). In contrast to the neuronal α-syn accumulation observed in PD and DLB, MSA is characterized by a widespread oligodendrocytic α-syn accumulation. Transgenic mice expressing human α-syn under the oligodendrocyte-specific myelin basic protein promoter (MBP1-hαsyn tg mice) model many of the behavioral and neuropathological alterations observed in MSA. Fluoxetine, a selective serotonin reuptake inhibitor, has been shown to be protective in toxin-induced models of PD, however its effects in an in vivo transgenic model of α-synucleinopathy remain unclear. In this context, this study examined the effect of fluoxetine in the MBP1-hαsyn tg mice, a model of MSA. Fluoxetine adminstration ameliorated motor deficits in the MBP1-hαsyn tg mice, with a concomitant decrease in neurodegenerative pathology in the basal ganglia, neocortex and hippocampus. Fluoxetine adminstration also increased levels of the neurotrophic factors, GDNF (glial-derived neurotrophic factor) and BDNF (brain-derived neurotrophic factor) in the MBP1-hαsyn tg mice compared to vehicle-treated tg mice. This fluoxetine-induced increase in GDNF and BDNF protein levels was accompanied by activation of the ERK signaling pathway. The effects of fluoxetine adminstration on myelin and serotonin markers were also examined. Collectively these results indicate that fluoxetine may represent a novel therapeutic intervention for MSA and other neurodegenerative disorders. PMID:22281106

Aberrant RNA splicing in cancer; expression changes and driver mutations of splicing factor genes.

PubMed

Sveen, A; Kilpinen, S; Ruusulehto, A; Lothe, R A; Skotheim, R I

2016-05-12

Alternative splicing is a widespread process contributing to structural transcript variation and proteome diversity. In cancer, the splicing process is commonly disrupted, resulting in both functional and non-functional end-products. Cancer-specific splicing events are known to contribute to disease progression; however, the dysregulated splicing patterns found on a genome-wide scale have until recently been less well-studied. In this review, we provide an overview of aberrant RNA splicing and its regulation in cancer. We then focus on the executors of the splicing process. Based on a comprehensive catalog of splicing factor encoding genes and analyses of available gene expression and somatic mutation data, we identify cancer-associated patterns of dysregulation. Splicing factor genes are shown to be significantly differentially expressed between cancer and corresponding normal samples, and to have reduced inter-individual expression variation in cancer. Furthermore, we identify enrichment of predicted cancer-critical genes among the splicing factors. In addition to previously described oncogenic splicing factor genes, we propose 24 novel cancer-critical splicing factors predicted from somatic mutations.

Dynamic Modulation of Expression of Lentiviral Restriction Factors in Primary CD4+ T Cells following Simian Immunodeficiency Virus Infection.

PubMed

Rahmberg, Andrew R; Rajakumar, Premeela A; Billingsley, James M; Johnson, R Paul

2017-04-01

Although multiple restriction factors have been shown to inhibit HIV/SIV replication, little is known about their expression in vivo Expression of 45 confirmed and putative HIV/SIV restriction factors was analyzed in CD4 + T cells from peripheral blood and the jejunum in rhesus macaques, revealing distinct expression patterns in naive and memory subsets. In both peripheral blood and the jejunum, memory CD4 + T cells expressed higher levels of multiple restriction factors compared to naive cells. However, relative to their expression in peripheral blood CD4 + T cells, jejunal CCR5 + CD4 + T cells exhibited significantly lower expression of multiple restriction factors, including APOBEC3G , MX2 , and TRIM25 , which may contribute to the exquisite susceptibility of these cells to SIV infection. In vitro stimulation with anti-CD3/CD28 antibodies or type I interferon resulted in upregulation of distinct subsets of multiple restriction factors. After infection of rhesus macaques with SIVmac239, the expression of most confirmed and putative restriction factors substantially increased in all CD4 + T cell memory subsets at the peak of acute infection. Jejunal CCR5 + CD4 + T cells exhibited the highest levels of SIV RNA, corresponding to the lower restriction factor expression in this subset relative to peripheral blood prior to infection. These results illustrate the dynamic modulation of confirmed and putative restriction factor expression by memory differentiation, stimulation, tissue microenvironment and SIV infection and suggest that differential expression of restriction factors may play a key role in modulating the susceptibility of different populations of CD4 + T cells to lentiviral infection. IMPORTANCE Restriction factors are genes that have evolved to provide intrinsic defense against viruses. HIV and simian immunodeficiency virus (SIV) target CD4 + T cells. The baseline level of expression in vivo and degree to which expression of restriction factors is

Factors affecting expression of the recF gene of Escherichia coli K-12.

PubMed

Sandler, S J; Clark, A J

1990-01-31

This report describes four factors which affect expression of the recF gene from strong upstream lambda promoters under temperature-sensitive cIAt2-encoded repressor control. The first factor was the long mRNA leader sequence consisting of the Escherichia coli dnaN gene and 95% of the dnaA gene and lambda bet, N (double amber) and 40% of the exo gene. When most of this DNA was deleted, RecF became detectable in maxicells. The second factor was the vector, pBEU28, a runaway replication plasmid. When we substituted pUC118 for pBEU28, RecF became detectable in whole cells by the Coomassie blue staining technique. The third factor was the efficiency of initiation of translation. We used site-directed mutagenesis to change the mRNA leader, ribosome-binding site and the 3 bp before and after the translational start codon. Monitoring the effect of these mutational changes by translational fusion to lacZ, we discovered that the efficiency of initiation of translation was increased 30-fold. Only an estimated two- or threefold increase in accumulated levels of RecF occurred, however. This led us to discover the fourth factor, namely sequences in the recF gene itself. These sequences reduce expression of the recF-lacZ fusion genes 100-fold. The sequences responsible for this decrease in expression occur in four regions in the N-terminal half of recF. Expression is reduced by some sequences at the transcriptional level and by others at the translational level.

Transforming growth factor β-activated kinase 1 negatively regulates interleukin-1α-induced stromal-derived factor-1 expression in vascular smooth muscle cells

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

Yang, Bin; Li, Wei; Zheng, Qichang

Stromal-derived Factor-1 (SDF-1) derived from vascular smooth muscle cells (VSMCs) contributes to vascular repair and remodeling in various vascular diseases. In this study, the mechanism underlying regulation of SDF-1 expression by interleukin-1α (IL-1α) was investigated in primary rat VSMCs. We found IL-1α promotes SDF-1 expression by up-regulating CCAAT-enhancer-binding protein β (C/EBPβ) in an IκB kinase β (IKKβ) signaling-dependent manner. Moreover, IL-1α-induced expression of C/EBPβ and SDF-1 was significantly potentiated by knockdown of transforming growth factor β-activated kinase 1 (TAK1), an upstream activator of IKKβ signaling. In addition, we also demonstrated that TAK1/p38 mitogen-activated protein kinase (p38 MAPK) signaling exerted negativemore » effect on IL-1α-induced expression of C/EBPβ and SDF-1 through counteracting ROS-dependent up-regulation of nuclear factor erythroid 2-related factor 2 (NRF2). In conclusion, TAK1 acts as an important regulator of IL-1α-induced SDF-1 expression in VSMCs, and modulating activity of TAK1 may serve as a potential strategy for modulating vascular repair and remodeling. - Highlights: • IL-1α induces IKKβ signaling-dependent SDF-1 expression by up-regulating C/EBPβ. • Activation of TAK1 by IL-1α negatively regulates C/EBPβ-dependent SDF-1 expression. • IL-1α-induced TAK1/p38 MAPK signaling counteracts ROS-dependent SDF-1 expression. • TAK1 counteracts IL-1α-induced SDF-1 expression by attenuating NRF2 up-regulation.« less

Hypoxia-inducible factors regulate pluripotency factor expression by ZNF217- and ALKBH5-mediated modulation of RNA methylation in breast cancer cells.

PubMed

Zhang, Chuanzhao; Zhi, Wanqing Iris; Lu, Haiquan; Samanta, Debangshu; Chen, Ivan; Gabrielson, Edward; Semenza, Gregg L

2016-10-04

Exposure of breast cancer cells to hypoxia increases the percentage of breast cancer stem cells (BCSCs), which are required for tumor initiation and metastasis, and this response is dependent on the activity of hypoxia-inducible factors (HIFs). We previously reported that exposure of breast cancer cells to hypoxia induces the ALKBH5-mediated demethylation of N6-methyladenosine (m6A) in NANOG mRNA leading to increased expression of NANOG, which is a pluripotency factor that promotes BCSC specification. Here we report that exposure of breast cancer cells to hypoxia also induces ZNF217-dependent inhibition of m6A methylation of mRNAs encoding NANOG and KLF4, which is another pluripotency factor that mediates BCSC specification. Although hypoxia induced the BCSC phenotype in all breast-cancer cell lines analyzed, it did so through variable induction of pluripotency factors and ALKBH5 or ZNF217. However, in every breast cancer line, the hypoxic induction of pluripotency factor and ALKBH5 or ZNF217 expression was HIF-dependent. Immunohistochemistry revealed that expression of HIF-1α and ALKBH5 was concordant in all human breast cancer biopsies analyzed. ALKBH5 knockdown in MDA-MB-231 breast cancer cells significantly decreased metastasis from breast to lungs in immunodeficient mice. Thus, HIFs stimulate pluripotency factor expression and BCSC specification by negative regulation of RNA methylation.

EBF factors drive expression of multiple classes of target genes governing neuronal development.

PubMed

Green, Yangsook S; Vetter, Monica L

2011-04-30

Early B cell factor (EBF) family members are transcription factors known to have important roles in several aspects of vertebrate neurogenesis, including commitment, migration and differentiation. Knowledge of how EBF family members contribute to neurogenesis is limited by a lack of detailed understanding of genes that are transcriptionally regulated by these factors. We performed a microarray screen in Xenopus animal caps to search for targets of EBF transcriptional activity, and identified candidate targets with multiple roles, including transcription factors of several classes. We determined that, among the most upregulated candidate genes with expected neuronal functions, most require EBF activity for some or all of their expression, and most have overlapping expression with ebf genes. We also found that the candidate target genes that had the most strongly overlapping expression patterns with ebf genes were predicted to be direct transcriptional targets of EBF transcriptional activity. The identification of candidate targets that are transcription factor genes, including nscl-1, emx1 and aml1, improves our understanding of how EBF proteins participate in the hierarchy of transcription control during neuronal development, and suggests novel mechanisms by which EBF activity promotes migration and differentiation. Other candidate targets, including pcdh8 and kcnk5, expand our knowledge of the types of terminal differentiated neuronal functions that EBF proteins regulate.

Pattern identification in time-course gene expression data with the CoGAPS matrix factorization.

PubMed

Fertig, Elana J; Stein-O'Brien, Genevieve; Jaffe, Andrew; Colantuoni, Carlo

2014-01-01

Patterns in time-course gene expression data can represent the biological processes that are active over the measured time period. However, the orthogonality constraint in standard pattern-finding algorithms, including notably principal components analysis (PCA), confounds expression changes resulting from simultaneous, non-orthogonal biological processes. Previously, we have shown that Markov chain Monte Carlo nonnegative matrix factorization algorithms are particularly adept at distinguishing such concurrent patterns. One such matrix factorization is implemented in the software package CoGAPS. We describe the application of this software and several technical considerations for identification of age-related patterns in a public, prefrontal cortex gene expression dataset.

Transcription factor CREB is involved in CaSR-mediated cytoskeleton gene expression.

PubMed

Huang, Shuaishuai; Ren, Yu; Wang, Ping; Li, Yanyuan; Wang, Xue; Zhuang, Haihui; Fang, Rong; Wang, Yuduo; Liu, Ningsheng; Hehir, Michael; Zhou, Jeff X

2015-03-01

Our previous studies illustrated that a steady increase of intracellular calcium concentration ([Ca2+]i) was important for maintaining microtubules (MTs) rearrangement in apoptotic cells. However, little is known about the effect of calcium sensing receptor (CaSR)-mediated increase in [Ca2+]i on cytoskeleton gene expression. We examined the impact of taxol or CaSR agonist/antagonist on the regulation of [Ca2+]i concentration, cytoskeleton arrangement, phosphorylated CREB and cytoskeleton gene expressions in HeLa cells with dominant negative plasmid of CREB (PM). This study demonstrated that Gdcl3 (a specific CaSR agonist) evoked a rapid increase of [Ca2+]i, formed a rigid bundle of MTs which surrounded the nucleus and decreased the cytoskeleton gene expressions in HeLa cells. These effects were rescued by addition of NPS2390 (a specific CaSR antagonist). Moreover, CaSR activity affected cytoskeleton gene expression through transcription factor CREB. Histoscores of pCREB immunoreactivity in tissues of cervical adenocarcinoma, renal clear cell carcinoma, and diffuse large B-cell lymphoma were markedly increased compared with non malignant tissue. These data demonstrate, for the first time, that CaSR-mediated increase in [Ca2+]i probably modulate cytoskeleton organization and gene expression via transcription factor. © 2014 Wiley Periodicals, Inc.

The Expression of Pigment Epithelium-Derived Factor in Bladder Transitional Cell Carcinoma

PubMed Central

Kim, Sung Woo; Lee, Kyung Seop

2012-01-01

Background Pigment epithelium-derived factor (PEDF) is an anti-angiogenic factor. The purpose of this study is to examine the involvement of PEDF in the angiogenesis and biological behavior of bladder transitional cell carcinoma (TCC). Methods We examined the expression of PEDF in 99 bladder TCCs and ten non-neoplastic tissues, and evaluated microvessel density (MVD). Results The positive immunoreactivity for PEDF was seen in normal urothelium in 60% (6/10) and TCC in 13% (13/99). The PEDF expression had a significant correlation with MVD, i.e., a low MVD in 42% (5/12), a middle MVD in 11% (8/76) and a high MVD 0% (0/11) of tumors. The PEDF expression was not significantly correlated with the differentiation and invasion of TCC, but the degree of MVD was significantly higher in both high grade TCC and the pT2 tumors. Conclusions The degree of PEDF expression is significantly higher in normal bladder urothelium than bladder TCC; it is inversely correlated with the angiogenesis; and it is not related to the differentiation and progression of TCC. It can therefore be concluded that bladder TCC would initially occur if there is a lack of the PEDF expression. PMID:23110012

Expressions of apoptosis-regulating factors in bovine retained placenta.

PubMed

Kamemori, Y; Wakamiya, K; Nishimura, R; Hosaka, Y; Ohtani, S; Okuda, K

2011-01-01

The aim of the present study was to evaluate the relationship between the retention of fetal membranes (RFM) and apoptosis of the cells in fetal membranes. The present study investigated mRNA and protein expressions of apoptosis-regulating factors: FAS, cellular FLICE-like inhibiting protein (cFLIP), BAX, BCL2, caspase-8 (CASP8), and CASP3 in fetal membranes. Placentomes were manually collected from the uterus immediately after parturition and classified into two groups (RFM; n = 8 and non-RFM; n = 8) according to whether placental membranes were expelled or not within 12 h after delivery. FAS mRNA expression in maternal placental tissue was less in RFM cows than in non-RFM cows (P < 0.05). cFLIP mRNA expression in maternal and fetal placental tissue was greater in RFM cows than in non-RFM cows (P < 0.05). CASP3 mRNA expression in maternal placental tissue was greater in RFM cows than in non-RFM cows (P < 0.05). However, the protein expressions of FAS, cFLIP and cleaved CASP3 were not significantly different between the two groups. mRNA and protein expressions of BAX, BCL2 and CASP8 were also not significantly different between the two groups. In the immunohistochemical study, single-stranded DNA, which appears specifically in the apoptotic cells, was mainly found in the maternal placenta of non-RFM cows. Together these results suggest that RFM occurs at least in part due to a dysfunctional apoptotic process caused by the inhibition of FAS expression in the maternal placenta, and the increase of cFLIP expression in the maternal and fetal placenta. Copyright © 2010 Elsevier Ltd. All rights reserved.

TERATOGENIC RESPONSES ARE MODULATED IN MICE LACKING EXPRESSION OF EPIDERMAL GROWTH FACTOR (EGF) AND TRANSFORMING GROWTH FACTOR-ALPHA (TGF)

EPA Science Inventory

TITLE:
TERATOGENIC RESPONSES ARE MODULATED IN MICE LACKING EXPRESSION OF EPIDERMAL GROWTH FACTOR (EGF) AND TRANSFORMING GROWTH FACTOR-ALPHA (TGF). AUTHORS (ALL): Abbott, Barbara D.1; Best, Deborah S.1; Narotsky, Michael G.1. SPONSOR NAME: None INSTITUTIONS (ALL): 1. Repro Tox ...

Sequence and Expression Analyses of Ethylene Response Factors Highly Expressed in Latex Cells from Hevea brasiliensis

PubMed Central

Piyatrakul, Piyanuch; Yang, Meng; Putranto, Riza-Arief; Pirrello, Julien; Dessailly, Florence; Hu, Songnian; Summo, Marilyne; Theeravatanasuk, Kannikar; Leclercq, Julie; Kuswanhadi; Montoro, Pascal

2014-01-01

The AP2/ERF superfamily encodes transcription factors that play a key role in plant development and responses to abiotic and biotic stress. In Hevea brasiliensis, ERF genes have been identified by RNA sequencing. This study set out to validate the number of HbERF genes, and identify ERF genes involved in the regulation of latex cell metabolism. A comprehensive Hevea transcriptome was improved using additional RNA reads from reproductive tissues. Newly assembled contigs were annotated in the Gene Ontology database and were assigned to 3 main categories. The AP2/ERF superfamily is the third most represented compared with other transcription factor families. A comparison with genomic scaffolds led to an estimation of 114 AP2/ERF genes and 1 soloist in Hevea brasiliensis. Based on a phylogenetic analysis, functions were predicted for 26 HbERF genes. A relative transcript abundance analysis was performed by real-time RT-PCR in various tissues. Transcripts of ERFs from group I and VIII were very abundant in all tissues while those of group VII were highly accumulated in latex cells. Seven of the thirty-five ERF expression marker genes were highly expressed in latex. Subcellular localization and transactivation analyses suggested that HbERF-VII candidate genes encoded functional transcription factors. PMID:24971876

Sequence and expression analyses of ethylene response factors highly expressed in latex cells from Hevea brasiliensis.

PubMed

Piyatrakul, Piyanuch; Yang, Meng; Putranto, Riza-Arief; Pirrello, Julien; Dessailly, Florence; Hu, Songnian; Summo, Marilyne; Theeravatanasuk, Kannikar; Leclercq, Julie; Kuswanhadi; Montoro, Pascal

2014-01-01

The AP2/ERF superfamily encodes transcription factors that play a key role in plant development and responses to abiotic and biotic stress. In Hevea brasiliensis, ERF genes have been identified by RNA sequencing. This study set out to validate the number of HbERF genes, and identify ERF genes involved in the regulation of latex cell metabolism. A comprehensive Hevea transcriptome was improved using additional RNA reads from reproductive tissues. Newly assembled contigs were annotated in the Gene Ontology database and were assigned to 3 main categories. The AP2/ERF superfamily is the third most represented compared with other transcription factor families. A comparison with genomic scaffolds led to an estimation of 114 AP2/ERF genes and 1 soloist in Hevea brasiliensis. Based on a phylogenetic analysis, functions were predicted for 26 HbERF genes. A relative transcript abundance analysis was performed by real-time RT-PCR in various tissues. Transcripts of ERFs from group I and VIII were very abundant in all tissues while those of group VII were highly accumulated in latex cells. Seven of the thirty-five ERF expression marker genes were highly expressed in latex. Subcellular localization and transactivation analyses suggested that HbERF-VII candidate genes encoded functional transcription factors.

Glucocorticoid receptor represses brain-derived neurotrophic factor expression in neuron-like cells.

PubMed

Chen, Hui; Lombès, Marc; Le Menuet, Damien

2017-04-12

Brain-derived neurotrophic factor (BDNF) is involved in many functions such as neuronal growth, survival, synaptic plasticity and memorization. Altered expression levels are associated with many pathological situations such as depression, epilepsy, Alzheimer's, Huntington's and Parkinson's diseases. Glucocorticoid receptor (GR) is also crucial for neuron functions, via binding of glucocorticoid hormones (GCs). GR actions largely overlap those of BDNF. It has been proposed that GR could be a regulator of BDNF expression, however the molecular mechanisms involved have not been clearly defined yet. Herein, we analyzed the effect of a GC agonist dexamethasone (DEX) on BDNF expression in mouse neuronal primary cultures and in the newly characterized, mouse hippocampal BZ cell line established by targeted oncogenesis. Mouse Bdnf gene exhibits a complex genomic structure with 8 untranslated exons (I to VIII) splicing onto one common and unique coding exon IX. We found that DEX significantly downregulated total BDNF mRNA expression by around 30%. Expression of the highly expressed exon IV and VI containing transcripts was also reduced by DEX. The GR antagonist RU486 abolished this effect, which is consistent with specific GR-mediated action. Transient transfection assays allowed us to define a short 275 bp region within exon IV promoter responsible for GR-mediated Bdnf repression. Chromatin immunoprecipitation experiments demonstrated GR recruitment onto this fragment, through unidentified transcription factor tethering. Altogether, GR downregulates Bdnf expression through direct binding to Bdnf regulatory sequences. These findings bring new insights into the crosstalk between GR and BDNF signaling pathways both playing a major role in physiology and pathology of the central nervous system.

Orf virus interleukin-10 and vascular endothelial growth factor-E modulate gene expression in cultured equine dermal fibroblasts.

PubMed

Wise, Lyn M; Bodaan, Christa J; Mercer, Andrew A; Riley, Christopher B; Theoret, Christine L

2016-10-01

Wounds in horses often exhibit sustained inflammation and inefficient vascularization, leading to excessive fibrosis and clinical complications such as "proud flesh". Orf virus-derived proteins, vascular endothelial growth factor (VEGF)-E and interleukin (ovIL)-10, enhance angiogenesis and control inflammation and fibrosis in skin wounds of laboratory animals. The study aimed to determine if equine dermal cells respond to VEGF-E and ovIL-10. Equine dermal cells are expected to express VEGF and IL-10 receptors, so viral protein treatment is likely to alter cellular gene expression and behaviour in a manner conducive to healing. Skin samples were harvested from the lateral thoracic wall of two healthy thoroughbred horses. Equine dermal cells were isolated using a skin explant method and their phenotype assessed by immunofluorescence. Cells were treated with recombinant proteins, with or without inflammatory stimuli. Gene expression was examined using standard and quantitative reverse transcriptase PCR. Cell behaviour was evaluated in a scratch assay. Cultured cells were half vimentin(+ve) fibroblasts and half alpha smooth muscle actin(+ve) and vimentin(+ve) myofibroblasts. VEGF-E increased basal expression of IL-10 mRNA, whereas VEGF-A and collagenase-1 mRNA expression was increased by ovIL-10. In cells exposed to inflammatory stimulus, both treatments dampened tumour necrosis factor mRNA expression, and ovIL-10 exacerbated expression of monocyte chemoattractant protein. Neither viral protein influenced cell migration greatly. This study shows that VEGF-E and ovIL-10 are active on equine dermal cells and exert anti-inflammatory and anti-fibrotic effects that may enhance skin wound healing in horses. © 2016 ESVD and ACVD.

Inflammatory monocytes expressing tissue factor drive SIV and HIV coagulopathy

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

Schechter, Melissa E.; Andrade, Bruno B.; He, Tianyu

In HIV infection, persistent inflammation despite effective antiretroviral therapy is linked to increased risk of noninfectious chronic complications such as cardiovascular and thromboembolic disease. Thus, a better understanding of inflammatory and coagulation pathways in HIV infection is needed to optimize clinical care. Markers of monocyte activation and coagulation independently predict morbidity and mortality associated with non-AIDS events. We identified a specific subset of monocytes that express tissue factor (TF), persist after virological suppression, and trigger the coagulation cascade by activating factor X. This subset of monocytes expressing TF had a distinct gene signature with up-regulated innate immune markers and evidencemore » of robust production of multiple proinflammatory cytokines, including interleukin-1β (IL-1β), tumor necrosis factor–α (TNF-α), and IL-6, ex vivo and in vitro upon lipopolysaccharide stimulation. We validated our findings in a nonhuman primate model, showing that TF-expressing inflammatory monocytes were associated with simian immunodeficiency virus (SIV)–related coagulopathy in the progressive [pigtail macaques (PTMs)] but not in the nonpathogenic (African green monkeys) SIV infection model. Last, Ixolaris, an anticoagulant that inhibits the TF pathway, was tested and potently blocked functional TF activity in vitro in HIV and SIV infection without affecting monocyte responses to Toll-like receptor stimulation. Strikingly, in vivo treatment of SIV-infected PTMs with Ixolaris was associated with significant decreases in D-dimer and immune activation. These data suggest that TF-expressing monocytes are at the epicenter of inflammation and coagulation in chronic HIV and SIV infection and may represent a potential therapeutic target.« less

Inflammatory monocytes expressing tissue factor drive SIV and HIV coagulopathy

DOE PAGES

Schechter, Melissa E.; Andrade, Bruno B.; He, Tianyu; ...

2017-08-30

In HIV infection, persistent inflammation despite effective antiretroviral therapy is linked to increased risk of noninfectious chronic complications such as cardiovascular and thromboembolic disease. Thus, a better understanding of inflammatory and coagulation pathways in HIV infection is needed to optimize clinical care. Markers of monocyte activation and coagulation independently predict morbidity and mortality associated with non-AIDS events. We identified a specific subset of monocytes that express tissue factor (TF), persist after virological suppression, and trigger the coagulation cascade by activating factor X. This subset of monocytes expressing TF had a distinct gene signature with up-regulated innate immune markers and evidencemore » of robust production of multiple proinflammatory cytokines, including interleukin-1β (IL-1β), tumor necrosis factor–α (TNF-α), and IL-6, ex vivo and in vitro upon lipopolysaccharide stimulation. We validated our findings in a nonhuman primate model, showing that TF-expressing inflammatory monocytes were associated with simian immunodeficiency virus (SIV)–related coagulopathy in the progressive [pigtail macaques (PTMs)] but not in the nonpathogenic (African green monkeys) SIV infection model. Last, Ixolaris, an anticoagulant that inhibits the TF pathway, was tested and potently blocked functional TF activity in vitro in HIV and SIV infection without affecting monocyte responses to Toll-like receptor stimulation. Strikingly, in vivo treatment of SIV-infected PTMs with Ixolaris was associated with significant decreases in D-dimer and immune activation. These data suggest that TF-expressing monocytes are at the epicenter of inflammation and coagulation in chronic HIV and SIV infection and may represent a potential therapeutic target.« less

Egr-1 and serum response factor are involved in growth factors- and serum-mediated induction of E2-EPF UCP expression that regulates the VHL-HIF pathway.

PubMed

Lim, Jung Hwa; Jung, Cho-Rok; Lee, Chan-Hee; Im, Dong-Soo

2008-11-01

E2-EPF ubiquitin carrier protein (UCP) has been shown to be highly expressed in common human cancers and target von Hippel-Lindau (VHL) for proteosomal degradation in cells, thereby stabilizing hypoxia-inducible factor (HIF)-1alpha. Here, we investigated cellular factors that regulate the expression of UCP gene. Promoter deletion assay identified binding sites for early growth response-1 (Egr-1) and serum response factor (SRF) in the UCP promoter. Hepatocyte or epidermal growth factor (EGF), or phorbol 12-myristate 13-acetate induced UCP expression following early induction of Egr-1 expression in HeLa cells. Serum increased mRNA and protein levels of SRF and UCP in the cell. By electrophoretic mobility shift and chromatin immunoprecipitation assays, sequence-specific DNA-binding of Egr-1 and SRF to the UCP promoter was detected in nuclear extracts from HeLa cells treated with EGF and serum, respectively. Overexpression of Egr-1 or SRF increased UCP expression. RNA interference-mediated depletion of endogenous Egr-1 or SRF impaired EGF- or serum-mediated induction of UCP expression, which was required for cancer cell proliferation. Systemic delivery of EGF into mice also increased UCP expression following early induction of Egr-1 expression in mouse liver. The induced UCP expression by the growth factors or serum increased HIF-1alpha protein level under non-hypoxic conditions, suggesting that the Egr-1/SRF-UCP-VHL pathway is in part responsible for the increased HIF-1alpha protein level in vitro and in vivo. Thus, growth factors and serum induce expression of Egr-1 and SRF, respectively, which in turn induces UCP expression that positively regulates cancer cell growth.

Growth differentiation factor 9 and its spatiotemporal expression and regulation in the zebrafish ovary.

PubMed

Liu, Lin; Ge, Wei

2007-02-01

Growth differentiation factor 9 (GDF9) is a member of the transforming growth factor beta (TGFB) superfamily. As an oocyte-specific growth factor, GDF9 plays critical roles in controlling folliculogenesis in mammals. In the present study, we cloned a 2.1-kb cDNA of the zebrafish GDF9 homolog (Gdf9, gdf9), which shares approximately 60% homology with that of mammals in the mature region. RT-PCR analysis showed that zebrafish gdf9 expression was present only in the gonads and Northern blot analysis revealed a single transcript of about 2.0 kb in the ovary. Real-time RT-PCR analysis revealed that gdf9 expression was highest in primary growth (PG, stage I) follicles and gradually decreased during follicular development, with the lowest level being found in fully grown (FG) follicles. The expression of gdf9 was maintained through fertilization and early embryonic development until gastrulation, at which point the expression level dramatically decreased. Expression was barely detectable after the late gastrula stage. Within the follicle, gdf9 mRNA was localized exclusively in the oocytes, as demonstrated by RT-PCR of denuded oocytes and freshly isolated follicle layers as well as by in situ hybridization. Interestingly, when amplified for high numbers of cycles, the expression of gdf9 was detected in cultured zebrafish follicular cells that were free of oocytes. The expression of gdf9 was downregulated by hCG in both ovarian fragments and isolated follicles in dose- and time-dependent manners, and this inhibition appeared to be stage-dependent, with the strongest inhibition observed for the FG follicles and no effect seen for the PG follicles. This correlates well with the expression profile of the LH receptor (lhcgr) in zebrafish follicles. In conclusion, as an oocyte-derived growth factor, GDF9 is highly conserved across vertebrates. With its biological advantages, zebrafish provides an alternative model for studying gene function and regulation.

TALE factors use two distinct functional modes to control an essential zebrafish gene expression program.

PubMed

Ladam, Franck; Stanney, William; Donaldson, Ian J; Yildiz, Ozge; Bobola, Nicoletta; Sagerström, Charles G

2018-06-18

TALE factors are broadly expressed embryonically and known to function in complexes with transcription factors (TFs) like Hox proteins at gastrula/segmentation stages, but it is unclear if such generally expressed factors act by the same mechanism throughout embryogenesis. We identify a TALE-dependent gene regulatory network (GRN) required for anterior development and detect TALE occupancy associated with this GRN throughout embryogenesis. At blastula stages, we uncover a novel functional mode for TALE factors, where they occupy genomic DECA motifs with nearby NF-Y sites. We demonstrate that TALE and NF-Y form complexes and regulate chromatin state at genes of this GRN. At segmentation stages, GRN-associated TALE occupancy expands to include HEXA motifs near PBX:HOX sites. Hence, TALE factors control a key GRN, but utilize distinct DNA motifs and protein partners at different stages - a strategy that may also explain their oncogenic potential and may be employed by other broadly expressed TFs. © 2018, Ladam et al.

Expression of vascular endothelial growth factor mRNA in non-small-cell lung carcinomas

PubMed Central

Fontanini, G; Boldrini, L; Chinè, S; Pisaturo, F; Basolo, F; Calcinai, A; Lucchi, M; Mussi, A; Angeletti, C A; Bevilacqua, G

1999-01-01

The vascular endothelial growth factor (VEGF) has been shown to be strictly related to vascular permeability and endothelial cell growth under physiological and pathological conditions. In tumour development and progression, VEGF plays a pivotal role in the development of the tumoral vascular network, and useful information in the progression of human cancer can be obtained by analysing the vascular endothelial growth factor expression of the tumours. In this study, we investigated the vascular endothelial growth factor transcript expression in non-small-cell lung carcinomas to evaluate the significance of this factor in a group of cancers in which the vascular pattern has been shown to significantly affect progression. Surgical samples of 42 patients with NSCLC were studied using reverse transcription polymerase chain reaction (PCR) analysis and in situ hybridization. Thirty-three out of 42 cases (78.6%) showed VEGF transcript expression predominantly as transcripts for the secretory forms of VEGF (isoforms 121 and 165). In situ hybridization, performed on 24 out of 42 samples, showed that the VEGF transcript expression was in several cases present in the cytoplasm both of neoplastic and normal cells, even if the VEGF mRNA was less expressed in the corresponding non-tumoral part. The VEGF 121 expression was associated with hilar and/or mediastinal nodal involvement (P = 0.02), and, taken together, the VEGF isoforms were shown to significantly influence overall (P = 0.02) and disease-free survival (P = 0.03). As a regulator of tumour angiogenesis, VEGF may represent a useful indicator of progression and poor prognosis in non-small-cell lung carcinomas. © 1999 Cancer Research Campaign PMID:9888482

Expression of the human blood coagulation protein factor XIIIa in Saccharomyces cerevisiae: dependence of the expression levels from host-vector systems and medium conditions.

PubMed

Bröker, M; Bäuml, O; Göttig, A; Ochs, J; Bodenbenner, M; Amann, E

1991-03-01

The human blood coagulation protein Factor XIIIa (FXIIIa) was expressed in Saccharomyces cerevisiae employing Escherichia coli-yeast shuttle vectors based on a 2-mu plasmid. Several factors affecting high production yield of recombinant FXIIIa were analysed. The use of the regulatable GAL-CYC1 hybrid promoter resulted in higher FXIIIa expression when compared with the constitutive ADCI promoter. Screening for suitable yeast strains for expression of FXIIIa under the transcriptional control of the GAL-CYC1 hybrid promoter revealed a broad spectrum of productivity. No obvious correlation between the expression rate and the genetic markers of the strains could be identified. The medium composition markedly influenced the FXIIIa expression rates. The expression of FXIIIa was strictly regulated by the carbon source. Glucose as the only sugar and energy source repressed the synthesis of FXIIIa, whereas addition of galactose induced FXIIIa expression. Special feeding schemes resulted in a productivity of up to 100 mg FXIIIa/l in shake flasks.

Expression of macrophage migration inhibitory factor in footpad skin lesions with diabetic neuropathy.

PubMed

Up Noh, Sun; Lee, Won-Young; Kim, Won-Serk; Lee, Yong-Taek; Jae Yoon, Kyung

2018-01-01

Background Diabetic neuropathy originating in distal lower extremities is associated with pain early in the disease course, overwhelming in the feet. However, the pathogenesis of diabetic neuropathy remains unclear. Macrophage migration inhibitory factor has been implicated in the onset of neuropathic pain and the development of diabetes. Objective of this study was to observe pain syndromes elicited in the footpad of diabetic neuropathy rat model and to assess the contributory role of migration inhibitory factor in the pathogenesis of diabetic neuropathy. Methods Diabetic neuropathy was made in Sprague Dawley rats by streptozotocin. Pain threshold was evaluated using von Frey monofilaments for 24 weeks. On comparable experiment time after streptozotocin injection, all footpads were prepared for following procedures; glutathione assay, terminal deoxynucleotidyl transferase-mediated biotinylated UTP nick end labeling staining, immunohistochemistry staining, real-time reverse transcription polymerase chain reaction, and Western blot. Additionally, human HaCaT skin keratinocytes were treated with methylglyoxal, transfected with migration inhibitory factor/control small interfering RNA, and prepared for real-time reverse transcription polymerase chain reaction and Western blot. Results As compared to sham group, pain threshold was significantly reduced in diabetic neuropathy group, and glutathione was decreased in footpad skin, simultaneously, cell death was increased. Over-expression of migration inhibitory factor, accompanied by low expression of glyoxalase-I and intraepidermal nerve fibers, was shown on the footpad skin lesions of diabetic neuropathy. But, there was no significance in expression of neurotransmitters and inflammatory mediators such as transient receptor potential vanilloid 1, mas-related G protein coupled receptor D, nuclear factor kappa B, tumor necrosis factor-alpha, and interleukin-6 between diabetic neuropathy group and sham group. Intriguingly

Factors for C-Kit Expression in Cardiac Outgrowth Cells and Human Heart Tissue.

PubMed

Matsushita, Satoshi; Minematsu, Kazuo; Yamamoto, Taira; Inaba, Hirotaka; Kuwaki, Kenji; Shimada, Akie; Yokoyama, Yasutaka; Amano, Atsushi

2017-12-12

We determined the factors associated with the expression of c-kit in the heart and the proliferation of c-kit-positive (c-kit pos ) cardiac stem cells among the outgrowth cells cultured from human cardiac explants.Samples of the right atrium (RA), left atrium (LA), and left ventricle obtained from patients during open-heart surgery were processed for cell culture of outgrowth cells and tissue analysis. The total number of growing cells and the population of c-kit pos cells were measured and compared with c-kit expression in native tissues and characteristics of the patients according to the region of the heart.We analyzed 452 samples from 334 patients. Atrial fibrillation (AF) in the patients reduced the number of outgrowth cells from the RA and LA, and aging was a co-factor for the LA. The c-kit pos population from the RA was associated with serum brain natriuretic peptide (BNP). C-kit expression in native tissue was also associated with BNP expression. However, we observed no relationship in expression between outgrowth cells and native tissue. In addition, the RA tissue provided the highest number of c-kit pos cells, and the left ventricle provided the lowest.C-kit was weakly expressed in response to damage. In addition, no correlation between outgrowth cells and native tissue was found for c-kit expression.

Hypoxia-inducible Factor Regulates αvβ3 Integrin Cell Surface Expression

PubMed Central

Cowden Dahl, Karen D.; Robertson, Sarah E.; Weaver, Valerie M.; Simon, M. Celeste

2005-01-01

Hypoxia-inducible factor (HIF)-deficient placentas exhibit a number of defects, including changes in cell fate adoption, lack of fetal angiogenesis, hypocellularity, and poor invasion into maternal tissue. HIF is a heterodimeric transcription factor consisting of α and β aryl hydrocarbon receptor nuclear translocator or ARNT) subunits. We used undifferentiated trophoblast stem (TS) cells to characterize HIF-dependent adhesion, migration, and invasion. Arnt-/- and Hifα-/- TS cells exhibit reduced adhesion and migration toward vitronectin compared with wild-type cells. Furthermore, this defect is associated with decreased cell surface expression of integrin αvβ3 and significantly decreased expression of this integrin in focal adhesions. Because of the importance of adhesion and migration in tumor progression (in addition to placental development), we examined the affect of culturing B16F0 melanoma cells in 1.5% oxygen (O2). Culturing B16F0 melanoma cells at 1.5% O2 resulted in increased αvβ3 integrin surface expression and increased adhesion to and migration toward vitronectin. Together, these data suggest that HIF and O2 tension influence placental invasion and tumor migration by increasing cell surface expression of αvβ3 integrin. PMID:15689487

Induction of neurite outgrowth in 3D hydrogel-based environments.

PubMed

Assunção-Silva, Rita C; Oliveira, Cátia Costa; Ziv-Polat, Ofra; Gomes, Eduardo D; Sahar, Abraham; Sousa, Nuno; Silva, Nuno A; Salgado, António J

2015-10-20

The ability of peripheral nervous system (PNS) axons to regenerate and re-innervate their targets after an injury has been widely recognized. However, despite the considerable advances made in microsurgical techniques, complete functional recovery is rarely achieved, especially for severe peripheral nerve injuries (PNIs). Therefore, alternative therapies that can successfully repair peripheral nerves are still essential. In recent years the use of biodegradable hydrogels enriched with growth-supporting and guidance cues, cell transplantation, and biomolecular therapies have been explored for the treatment of PNIs. Bearing this in mind, the aim of this study was to assess whether Gly-Arg-Gly-Asp-Ser synthetic peptide (GRGDS)-modified gellan gum (GG) based hydrogels could foster an amenable environment for neurite/axonal growth. Additionally, strategies to further improve the rate of neurite outgrowth were also tested, namely the use of adipose tissue derived stem cells (ASCs), as well as the glial derived neurotrophic factor (GDNF). In order to increase its stability and enhance its bioactivity, the GDNF was conjugated covalently to iron oxide nanoparticles (IONPs). The impact of hydrogel modification as well as the effect of the GDNF-IONPs on ASC behavior was also screened. The results revealed that the GRGDS-GG hydrogel was able to support dorsal root ganglia (DRG)-based neurite outgrowth, which was not observed for non-modified hydrogels. Moreover, the modified hydrogels were also able to support ASCs attachment. In contrast, the presence of the GDNF-IONPs had no positive or negative impact on ASC behavior. Further experiments revealed that the presence of ASCs in the hydrogel improved axonal growth. On the other hand, GDNF-IONPs alone or combined with ASCs significantly increased neurite outgrowth from DRGs, suggesting a beneficial role of the proposed strategy for future applications in PNI regenerative medicine.

The dependence receptor Ret induces apoptosis in somatotrophs through a Pit-1/p53 pathway, preventing tumor growth

PubMed Central

Cañibano, Carmen; Rodriguez, Noela L; Saez, Carmen; Tovar, Sulay; Garcia-Lavandeira, Montse; Borrello, Maria Grazia; Vidal, Anxo; Costantini, Frank; Japon, Miguel; Dieguez, Carlos; Alvarez, Clara V

2007-01-01

Somatotrophs are the only pituitary cells that express Ret, GFRα1 and GDNF. This study investigated the effects of Ret in a somatotroph cell line, in primary pituitary cultures and in Ret KO mice. Ret regulates somatotroph numbers by inducing Pit-1 overexpression, leading to increased p53 expression and apoptosis, both of which can be prevented with Ret or Pit-1 siRNA. The Pit-1 overexpression is mediated by sustained activation of PKCδ, JNK, c/EBPα and CREB induced by a complex of Ret, caspase 3 and PKCδ. In the presence of GDNF, Akt is activated, and the Pit-1 overexpression and resulting apoptosis are blocked. The adenopituitary of Ret KO mice is larger than normal, showing Pit-1 and somatotroph hyperplasia. In normal animals, activation of the Ret/Pit-1/p53 pathway by retroviral introduction of Ret blocked tumor growth in vivo. Thus, somatotrophs have an intrinsic mechanism for controlling Pit-1/GH production through an apoptotic/survival pathway. Ret might be of value for treatment of pituitary adenomas. PMID:17380130

The dependence receptor Ret induces apoptosis in somatotrophs through a Pit-1/p53 pathway, preventing tumor growth.

PubMed

Cañibano, Carmen; Rodriguez, Noela L; Saez, Carmen; Tovar, Sulay; Garcia-Lavandeira, Montse; Borrello, Maria Grazia; Vidal, Anxo; Costantini, Frank; Japon, Miguel; Dieguez, Carlos; Alvarez, Clara V

2007-04-18

Somatotrophs are the only pituitary cells that express Ret, GFRalpha1 and GDNF. This study investigated the effects of Ret in a somatotroph cell line, in primary pituitary cultures and in Ret KO mice. Ret regulates somatotroph numbers by inducing Pit-1 overexpression, leading to increased p53 expression and apoptosis, both of which can be prevented with Ret or Pit-1 siRNA. The Pit-1 overexpression is mediated by sustained activation of PKCdelta, JNK, c/EBPalpha and CREB induced by a complex of Ret, caspase 3 and PKCdelta. In the presence of GDNF, Akt is activated, and the Pit-1 overexpression and resulting apoptosis are blocked. The adenopituitary of Ret KO mice is larger than normal, showing Pit-1 and somatotroph hyperplasia. In normal animals, activation of the Ret/Pit-1/p53 pathway by retroviral introduction of Ret blocked tumor growth in vivo. Thus, somatotrophs have an intrinsic mechanism for controlling Pit-1/GH production through an apoptotic/survival pathway. Ret might be of value for treatment of pituitary adenomas.

Comparative study of angiostatic and anti-invasive gene expressions as prognostic factors in gastric cancer.

PubMed

Lee, J H; Koh, J T; Shin, B A; Ahn, K Y; Roh, J H; Kim, Y J; Kim, K K

2001-02-01

Genes involving angiogenesis and metastasis play an important role in the progression and infiltration of cancer. We examined the expressions of various angiostatic and potential invasion/metastasis suppressor genes through RT-PCR analyses in 32 gastric cancer specimens with or without distant metastasis. The expressions of the invasion/metastasis suppressor, nm23 and E-cadherin increased much more in the cancer tissue (CT) and metastatic lymph node (MLN) than in the extraneoplastic mucosa (EM) and non-metastatic lymph node (NLN), respectively. The expressions of the angiostatic factor, angiopoietin 2 and thrombospondin 2 increased in the CT and MLN as compared with the EM and NLN, respectively. The newly cloned angiostatic factor, brain-specific angiogenesis inhibitor 1 (BAI1) decreased much more in the CT and MLN than the EM and NLN, respectively. However, BAI1 increased in the CT compared with the EM among the patients with poor prognosis and distant metastasis, such as liver or peritoneum. The expressions of the invasive factor, matrix metalloproteinase-2 and its suppressor, tissue inhibitor metalloproteinase-2 (TIMP-2) increased in the CM as compared with the EM, but the increased expression pattern of these genes in the CT became blunted among the patients with good prognosis. Our results indicate that BAI1 and TIMP-2 expressions in the extraneoplastic mucosa and non-metastatic lymph nodes were not suppressed in the patients with good prognosis, but increased expressions of angiopoietin 2, thrombospondin 2, TIMP-2, nm23 and E-cadherin in the tumor tissue did not lead to a long survival after operation. It is suggested that the extent of BAI1 and TIMP-2 expression in the gastric mucosa may be an important prognostic factor for predicting survival in gastric cancer.

The spatial expression and regulation of transcription factors IDEF1 and IDEF2

PubMed Central

Kobayashi, Takanori; Ogo, Yuko; Aung, May Sann; Nozoye, Tomoko; Itai, Reiko Nakanishi; Nakanishi, Hiromi; Yamakawa, Takashi; Nishizawa, Naoko K.

2010-01-01

Background and Aims Under conditions of low iron availability, rice plants induce genes involved in iron uptake and utilization. The iron deficiency-responsive cis-acting element binding factors 1 and 2 (IDEF1 and IDEF2) regulate transcriptional response to iron deficiency in rice roots. Clarification of the functions of IDEF1 and IDEF2 could uncover the gene regulation mechanism. Methods Spatial patterns of IDEF1 and IDEF2 expression were analysed by histochemical staining of IDEF1 and IDEF2 promoter-GUS transgenic rice lines. Expression patterns of the target genes of IDEF1 and IDEF2 were analysed using transformants with induced or repressed expression of IDEF1 or IDEF2 grown in iron-rich or in iron-deficient solutions for 1 d. Key Results IDEF1 and IDEF2 were highly expressed in the basal parts of the lateral roots and vascular bundles. IDEF1 and IDEF2 expression was dominant in leaf mesophyll and vascular cells, respectively. These expression patterns were similar under both iron-deficient and iron-sufficient conditions. IDEF1 was strongly expressed in pollen, ovaries, the aleurone layer and embryo. IDEF2 was expressed in pollen, ovaries and the dorsal vascular region of the endosperm. During seed germination, IDEF1 and IDEF2 were expressed in the endosperm and embryo. Expression of IDEF1 target genes was regulated in iron-rich roots similar to early iron-deficiency stages. In addition, the expression patterns of IDEF2 target genes were similar between iron-rich conditions and early or subsequent iron deficiency. Conclusions IDEF1 and IDEF2 are constitutively expressed during both vegetative and reproductive stages. The spatial expression patterns of IDEF1 and IDEF2 overlap with their target genes in restricted cell types, but not in all cells. The spatial expression patterns and gene regulation of IDEF1 and IDEF2 in roots are generally conserved under conditions of iron sufficiency and deficiency, suggesting complicated interactions with unknown factors for

Angiogenesis and expression of vascular endothelial growth factor, tumour necrosis factor-α and hypoxia inducible factor-1α in canine renal cell carcinoma.

PubMed

Yhee, J Y; Yu, C H; Kim, J H; Im, K S; Kim, N H; Brodersen, B W; Doster, A R; Sur, J-H

2012-01-01

The aim of the present study was to determine the distribution and characteristics of microvessels in various histological types of canine renal cell carcinoma (RCC). The study compared microvessel density (MVD) and distribution of blood vessels according to histological type and evaluated the presence of angiogenesis-related proteins. Nine archival samples of canine RCC were studied. MVD was calculated as the mean number of blood vessels per mm(2). The diameter of blood vessels was calculated by determining either the length of the long axis of blood vessels (diameter(max)) or the mean distance from the centre of each blood vessel to the tunica adventia (diameter(mean)). A significant difference in MVD was evident between RCCs and normal kidneys (46.6 ± 28.0 versus 8.4 ± 2.2 microvessels/mm(2)). Diameter(max) in canine RCCs (34.1 ± 14.7 μm) was also significantly different from normal canine kidney (23.2 ± 3.4 μm). Vascular endothelial growth factor (VEGF) was expressed by tumour cells and vascular endothelial cells and tumour necrosis factor (TNF)-α expression was observed in vascular endothelial cells in both neoplastic and normal kidney. Although VEGF is involved in angiogenesis and correlates with tumour stage of development, no correlation was found between VEGF expression and MVD. Tumour-associated macrophages expressing TNF-α and hypoxia inducible factor 1α were identified in peritumoural tissue and may play an important role in angiogenesis. Copyright © 2011 Elsevier Ltd. All rights reserved.

Identification and expression profiles of the WRKY transcription factor family in Ricinus communis.

PubMed

Li, Hui-Liang; Zhang, Liang-Bo; Guo, Dong; Li, Chang-Zhu; Peng, Shi-Qing

2012-07-25

In plants, WRKY proteins constitute a large family of transcription factors. They are involved in many biological processes, such as plant development, metabolism, and responses to biotic and abiotic stresses. A large number of WRKY transcription factors have been reported from Arabidopsis, rice, and other higher plants. The recent publication of the draft genome sequence of castor bean (Ricinus communis) has allowed a genome-wide search for R. communis WRKY (RcWRKY) transcription factors and the comparison of these positively identified proteins with their homologs in model plants. A total of 47 WRKY genes were identified in the castor bean genome. According to the structural features of the WRKY domain, the RcWRKY are classified into seven main phylogenetic groups. Furthermore, putative orthologs of RcWRKY proteins in Arabidopsis and rice could now be assigned. An analysis of expression profiles of RcWRKY genes indicates that 47 WRKY genes display differential expressions either in their transcript abundance or expression patterns under normal growth conditions. Copyright © 2012 Elsevier B.V. All rights reserved.

Differential expression of growth factors at the cellular level in virus-infected brain

PubMed Central

Prosniak, Mikhail; Zborek, Anna; Scott, Gwen S.; Roy, Anirban; Phares, Timothy W.; Koprowski, Hilary; Hooper, D. Craig

2003-01-01

The contribution of host factors to rabies virus (RV) transcription/replication and axonal/transsynaptic spread is largely unknown. We previously identified several host genes that are up-regulated in the mouse brain during RV infection, including neuroleukin, which is involved in neuronal growth and survival, cell motility, and differentiation, and fibroblast growth factor homologous factor 4 (FHF4), which has been implicated in limb and nervous system development. In this study, we used real-time quantitative RT-PCR to assess the expression of mRNAs specific for neuroleukin, the two isoforms of FHF4 (FHF4-1a and -1b) encoded by the FHF4 gene, and N protein of RV in neurons and astrocytes isolated by laser capture microdissection from mouse brains infected with the laboratory-adapted RV strain CVS-N2c or with a street RV of silver-haired bat origin. Differences in the gene expression patterns suggest that the capacity of RV strains to infect nonneuronal cells and differentially modulate host gene expression may be important in virus replication and spread in the CNS. PMID:12736376

Engineering synthetic TALE and CRISPR/Cas9 transcription factors for regulating gene expression.

PubMed

Kabadi, Ami M; Gersbach, Charles A

2014-09-01

Engineered DNA-binding proteins that can be targeted to specific sites in the genome to manipulate gene expression have enabled many advances in biomedical research. This includes generating tools to study fundamental aspects of gene regulation and the development of a new class of gene therapies that alter the expression of endogenous genes. Designed transcription factors have entered clinical trials for the treatment of human diseases and others are in preclinical development. High-throughput and user-friendly platforms for designing synthetic DNA-binding proteins present innovative methods for deciphering cell biology and designing custom synthetic gene circuits. We review two platforms for designing synthetic transcription factors for manipulating gene expression: Transcription activator-like effectors (TALEs) and the RNA-guided clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 system. We present an overview of each technology and a guide for designing and assembling custom TALE- and CRISPR/Cas9-based transcription factors. We also discuss characteristics of each platform that are best suited for different applications. Copyright © 2014 Elsevier Inc. All rights reserved.

Activated Monocytes Enhance Platelet-Driven Contraction of Blood Clots via Tissue Factor Expression.

PubMed

Peshkova, Alina D; Le Minh, Giang; Tutwiler, Valerie; Andrianova, Izabella A; Weisel, John W; Litvinov, Rustem I

2017-07-11

Platelet-driven reduction in blood clot volume (clot contraction or retraction) has been implicated to play a role in hemostasis and thrombosis. Although these processes are often linked with inflammation, the role of inflammatory cells in contraction of blood clots and thrombi has not been investigated. The aim of this work was to study the influence of activated monocytes on clot contraction. The effects of monocytes were evaluated using a quantitative optical tracking methodology to follow volume changes in a blood clot formed in vitro. When a physiologically relevant number of isolated human monocytes pre-activated with phorbol-12-myristate-13-acetate (PMA) were added back into whole blood, the extent and rate of clot contraction were increased compared to addition of non-activated cells. Inhibition of tissue factor expression or its inactivation on the surface of PMA-treated monocytes reduced the extent and rate of clot contraction back to control levels with non-activated monocytes. On the contrary, addition of tissue factor enhanced clot contraction, mimicking the effects of tissue factor expressed on the activated monocytes. These data suggest that the inflammatory cells through their expression of tissue factor can directly affect hemostasis and thrombosis by modulating the size and density of intra- and extravascular clots and thrombi.

Valproic acid disrupts the oscillatory expression of core circadian rhythm transcription factors.

PubMed

Griggs, Chanel A; Malm, Scott W; Jaime-Frias, Rosa; Smith, Catharine L

2018-01-15

Valproic acid (VPA) is a well-established therapeutic used in treatment of seizure and mood disorders as well as migraines and a known hepatotoxicant. About 50% of VPA users experience metabolic disruptions, including weight gain, hyperlipidemia, and hyperinsulinemia, among others. Several of these metabolic abnormalities are similar to the effects of circadian rhythm disruption. In the current study, we examine the effect of VPA exposure on the expression of core circadian transcription factors that drive the circadian clock via a transcription-translation feedback loop. In cells with an unsynchronized clock, VPA simultaneously upregulated the expression of genes encoding core circadian transcription factors that regulate the positive and negative limbs of the feedback loop. Using low dose glucocorticoid, we synchronized cultured fibroblast cells to a circadian oscillatory pattern. Whether VPA was added at the time of synchronization or 12h later at CT12, we found that VPA disrupted the oscillatory expression of multiple genes encoding essential transcription factors that regulate circadian rhythm. Therefore, we conclude that VPA has a potent effect on the circadian rhythm transcription-translation feedback loop that may be linked to negative VPA side effects in humans. Furthermore, our study suggests potential chronopharmacology implications of VPA usage. Copyright © 2017. Published by Elsevier Inc.

Identity-expression interaction in face perception: sex, visual field, and psychophysical factors.

PubMed

Godard, Ornella; Baudouin, Jean-Yves; Bonnet, Philippe; Fiori, Nicole

2013-01-01

We investigated the psychophysical factors underlying the identity-emotion interaction in face perception. Visual field and sex were also taken into account. Participants had to judge whether a probe face, presented in either the left or the right visual field, and a central target face belonging to same person while emotional expression varied (Experiment 1) or to judge whether probe and target faces expressed the same emotion while identity was manipulated (Experiment 2). For accuracy we replicated the mutual facilitation effect between identity and emotion; no sex or hemispheric differences were found. Processing speed measurements, however, showed a lesser degree of interference in women than in men, especially for matching identity when faces expressed different emotions after a left visual presentation probe face. Psychophysical indices can be used to determine whether these effects are perceptual (A') or instead arise at a post-perceptual decision-making stage (B"). The influence of identity on the processing of facial emotion seems to be due to perceptual factors, whereas the influence of emotion changes on identity processing seems to be related to decisional factors. In addition, men seem to be more "conservative" after a LVF/RH probe-face presentation when processing identity. Women seem to benefit from better abilities to extract facial invariant aspects relative to identity.

The Activity of Differentiation Factors Induces Apoptosis in Polyomavirus Large T-Expressing Myoblasts

PubMed Central

Fimia, Gian Maria; Gottifredi, Vanesa; Bellei, Barbara; Ricciardi, Maria Rosaria; Tafuri, Agostino; Amati, Paolo; Maione, Rossella

1998-01-01

It is commonly accepted that pathways that regulate proliferation/differentiation processes, if altered in their normal interplay, can lead to the induction of programmed cell death. In a previous work we reported that Polyoma virus Large Tumor antigen (PyLT) interferes with in vitro terminal differentiation of skeletal myoblasts by binding and inactivating the retinoblastoma antioncogene product. This inhibition occurs after the activation of some early steps of the myogenic program. In the present work we report that myoblasts expressing wild-type PyLT, when subjected to differentiation stimuli, undergo cell death and that this cell death can be defined as apoptosis. Apoptosis in PyLT-expressing myoblasts starts after growth factors removal, is promoted by cell confluence, and is temporally correlated with the expression of early markers of myogenic differentiation. The block of the initial events of myogenesis by transforming growth factor β or basic fibroblast growth factor prevents PyLT-induced apoptosis, while the acceleration of this process by the overexpression of the muscle-regulatory factor MyoD further increases cell death in this system. MyoD can induce PyLT-expressing myoblasts to accumulate RB, p21, and muscle- specific genes but is unable to induce G00 arrest. Several markers of different phases of the cell cycle, such as cyclin A, cdk-2, and cdc-2, fail to be down-regulated, indicating the occurrence of cell cycle progression. It has been frequently suggested that apoptosis can result from an unbalanced cell cycle progression in the presence of a contrasting signal, such as growth factor deprivation. Our data involve differentiation pathways, as a further contrasting signal, in the generation of this conflict during myoblast cell apoptosis. PMID:9614186

Roles for Arabidopsis CAMTA transcription factors in cold-regulated gene expression and freezing tolerance.

PubMed

Doherty, Colleen J; Van Buskirk, Heather A; Myers, Susan J; Thomashow, Michael F

2009-03-01

The Arabidopsis thaliana CBF cold response pathway plays a central role in cold acclimation. It is characterized by rapid cold induction of genes encoding the CBF1-3 transcription factors, followed by expression of the CBF gene regulon, which imparts freezing tolerance. Our goal was to further the understanding of the cis-acting elements and trans-acting factors involved in expression of CBF2. We identified seven conserved DNA motifs (CM), CM1 to 7, that are present in the promoters of CBF2 and another rapidly cold-induced gene encoding a transcription factor, ZAT12. The results presented indicate that in the CBF2 promoter, CM4 and CM6 have negative regulatory activity and that CM2 has both negative and positive activity. A Myc binding site in the CBF2 promoter was also found to have positive regulatory effects. Moreover, our results indicate that members of the calmodulin binding transcription activator (CAMTA) family of transcription factors bind to the CM2 motif, that CAMTA3 is a positive regulator of CBF2 expression, and that double camta1 camta3 mutant plants are impaired in freezing tolerance. These results establish a role for CAMTA proteins in cold acclimation and provide a possible point of integrating low-temperature calcium and calmodulin signaling with cold-regulated gene expression.

Dimensions of assertiveness: factors underlying the college self-expression scale.

PubMed

Kipper, D A; Jaffe, Y

1978-02-01

A total of 447 Israeli students, both males and females, from four educational institutions were administered the College Self-expression Scale, a measure of assertiveness. The obtained responses were factor analyzed using the principal axis solution and the varimax rotation method. The results showed four main factors which included 43 of the 50 items of the original scale. These factors were identified as the willingness to take risks in interpersonal interactions, the ability to communicate feelings, setting rules and rectifying injustices, and the presence or absence of a tendency to invoke a self-punitive attitude. The findings were interpreted as adding support to the validity of the scale as a measure of assertiveness.

Expression of Metallothionein and Vascular Endothelial Growth Factor Isoforms in Breast Cancer Cells.

PubMed

Wierzowiecka, Barbara; Gomulkiewicz, Agnieszka; Cwynar-Zajac, Lucja; Olbromski, Mateusz; Grzegrzolka, Jedrzej; Kobierzycki, Christopher; Podhorska-Okolow, Marzenna; Dziegiel, Piotr

2016-01-01

Metallothioneins (MTs) are low-molecular-weight and cysteine-rich proteins that bind heavy metal ions and oxygen-free radicals. MTs are commonly expressed in various tissues of mammals and are involved in regulation of cell proliferation and differentiation, and may be engaged in angiogenesis. Expression of MTs has been studied in many cancer types, especially breast cancer. The research results indicate that MTs may play important, although not yet fully known, roles in cancer angiogenesis. The aim of this study was to analyze the level of gene expression of selected MT isoforms induced with zinc ions in correlation with vascular endothelial growth factor (VEGF) isoforms in in vitro models of breast cancer. The studies were carried out in three breast cancer cell lines (MCF-7, SK-BR-3, MDA-MB-231). An epithelial cell line derived from normal breast tissue (Me16c) was used as a control. The levels of expression of selected MT isoforms and selected genes involved in angiogenesis were studied with real-time PCR. Expression of different MT isoforms was induced by zinc ions to differing degrees in individual breast cancer cell lines. An increase in the expression of some MT isoforms was associated with a slight increase in the level of expression of VEGFA. The research results may indicate certain correlation between an increased expression of selected MT isoforms and a pro-angiogenic factor VEGF in specific types of breast cancer cells. Copyright © 2016 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

Coordinating Regulation of Gene Expression in Cardiovascular Disease: Interactions between Chromatin Modifiers and Transcription Factors

PubMed Central

Bauer, Ashley J.; Martin, Kathleen A.

2017-01-01

Cardiovascular disease is a leading cause of death with increasing economic burden. The pathogenesis of cardiovascular diseases is complex, but can arise from genetic and/or environmental risk factors. This can lead to dysregulated gene expression in numerous cell types including cardiomyocytes, endothelial cells, vascular smooth muscle cells, and inflammatory cells. While initial studies addressed transcriptional control of gene expression, epigenetics has been increasingly appreciated to also play an important role in this process through alterations in chromatin structure and gene accessibility. Chromatin-modifying proteins including enzymes that modulate DNA methylation, histone methylation, and histone acetylation can influence gene expression in numerous ways. These chromatin modifiers and their marks can promote or prevent transcription factor recruitment to regulatory regions of genes through modifications to DNA, histones, or the transcription factors themselves. This review will focus on the emerging question of how epigenetic modifiers and transcription factors interact to coordinately regulate gene expression in cardiovascular disease. While most studies have addressed the roles of either epigenetic or transcriptional control, our understanding of the integration of these processes is only just beginning. Interrogating these interactions is challenging, and improved technical approaches will be needed to fully dissect the temporal and spatial relationships between transcription factors, chromatin modifiers, and gene expression in cardiovascular disease. We summarize the current state of the field and provide perspectives on limitations and future directions. Through studies of epigenetic and transcriptional interactions, we can advance our understanding of the basic mechanisms of cardiovascular disease pathogenesis to develop novel therapeutics. PMID:28428957

Erythropoietin protects myocardin-expressing cardiac stem cells against cytotoxicity of tumor necrosis factor-{alpha}

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

Madonna, Rosalinda; Institute of Cardiology, and Center of Excellence on Aging, 'G. d'Annunzio' University, Chieti; Shelat, Harnath

2009-10-15

Cardiac stem cells are vulnerable to inflammation caused by infarction or ischemic injury. The growth factor, erythropoietin (Epo), ameliorates the inflammatory response of the myocardium to ischemic injury. This study was designed to assess the role of Epo in regulation of expression and activation of the cell death-associated intracellular signaling components in cardiac myoblasts stimulated with the proinflammatory cytokine tumor necrosis factor (TNF)-{alpha}. Cardiac myoblasts isolated from canine embryonic hearts characterized by expression of myocardin A, a promyogenic transcription factor for cardiovascular muscle development were pretreated with Epo and then exposed to TNF-{alpha}. Compared to untreated cells, the Epo-treated cardiacmore » myoblasts exhibited better morphology and viability. Immunoblotting revealed lower levels of active caspase-3 and reductions in iNOS expression and NO production in Epo-treated cells. Furthermore, Epo pretreatment reduced nuclear translocation of NF-{kappa}B and inhibited phosphorylation of inhibitor of kappa B (I{kappa}B) in TNF-{alpha}-stimulated cardiac myoblasts. Thus, Epo protects cardiac myocyte progenitors or myoblasts against the cytotoxic effects of TNF-{alpha} by inhibiting NF-{kappa}B-mediated iNOS expression and NO production and by preventing caspase-3 activation.« less

The Forkhead Transcription Factor, Foxd1, Is Necessary for Pituitary Luteinizing Hormone Expression in Mice

PubMed Central

Gumbel, Jason H.; Patterson, Elizabeth M.; Owusu, Sarah A.; Kabat, Brock E.; Jung, Deborah O.; Simmons, Jasmine; Hopkins, Torin; Ellsworth, Buffy S.

2012-01-01

The pituitary gland regulates numerous physiological functions including growth, reproduction, temperature and metabolic homeostasis, lactation, and response to stress. Pituitary organogenesis is dependent on signaling factors that are produced in and around the developing pituitary. The studies described in this report reveal that the forkhead transcription factor, Foxd1, is not expressed in the developing mouse pituitary gland, but rather in the mesenchyme surrounding the pituitary gland, which is an essential source of signaling factors that regulate pituitary organogenesis. Loss of Foxd1 causes a morphological defect in which the anterior lobe of the pituitary gland protrudes through the cartilage plate that is developing ventral to the pituitary at embryonic days (e)14.5, e16.5, and e18.5. The number of proliferating pituitary cells is increased at e14.5 and e16.5. Loss of Foxd1 also results in significantly decreased levels of Lhb expression at e18.5. This decrease in Lhb expression does not appear to be due to a change in the number of gonadotrope cells in the pituitary gland. Previous studies have shown that loss of the LIM homeodomain factor, Lhx3, which is activated by the FGF signaling pathway, results in loss of LH production. Although there is a difference in Lhb expression in Foxd1 null mice, the expression pattern of LHX3 is not altered in Foxd1 null mice. These studies suggest that Foxd1 is indirectly required for normal Lhb expression and cartilage formation. PMID:23284914

Transdifferentiation of brain-derived neurotrophic factor (BDNF)-secreting mesenchymal stem cells significantly enhance BDNF secretion and Schwann cell marker proteins.

PubMed

Bierlein De la Rosa, Metzere; Sharma, Anup D; Mallapragada, Surya K; Sakaguchi, Donald S

2017-11-01

The use of genetically modified mesenchymal stem cells (MSCs) is a rapidly growing area of research targeting delivery of therapeutic factors for neuro-repair. Cells can be programmed to hypersecrete various growth/trophic factors such as brain-derived neurotrophic factor (BDNF), glial cell line-derived neurotrophic factor (GDNF), and nerve growth factor (NGF) to promote regenerative neurite outgrowth. In addition to genetic modifications, MSCs can be subjected to transdifferentiation protocols to generate neural cell types to physically and biologically support nerve regeneration. In this study, we have taken a novel approach by combining these two unique strategies and evaluated the impact of transdifferentiating genetically modified MSCs into a Schwann cell-like phenotype. After 8 days in transdifferentiation media, approximately 30-50% of transdifferentiated BDNF-secreting cells immunolabeled for Schwann cell markers such as S100β, S100, and p75 NTR . An enhancement was observed 20 days after inducing transdifferentiation with minimal decreases in expression levels. BDNF production was quantified by ELISA, and its biological activity tested via the PC12-TrkB cell assay. Importantly, the bioactivity of secreted BDNF was verified by the increased neurite outgrowth of PC12-TrkB cells. These findings demonstrate that not only is BDNF actively secreted by the transdifferentiated BDNF-MSCs, but also that it has the capacity to promote neurite sprouting and regeneration. Given the fact that BDNF production remained stable for over 20 days, we believe that these cells have the capacity to produce sustainable, effective, BDNF concentrations over prolonged time periods and should be tested within an in vivo system for future experiments. Copyright © 2017 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

Evaluation of approaches to monitor Staphylococcus aureus virulence factor expression during human disease.

PubMed

Rozemeijer, Wouter; Fink, Pamela; Rojas, Eduardo; Jones, C Hal; Pavliakova, Danka; Giardina, Peter; Murphy, Ellen; Liberator, Paul; Jiang, Qin; Girgenti, Douglas; Peters, Remco P H; Savelkoul, Paul H M; Jansen, Kathrin U; Anderson, Annaliesa S; Kluytmans, Jan

2015-01-01

Staphylococcus aureus is a versatile pathogen of medical significance, using multiple virulence factors to cause disease. A prophylactic S. aureus 4-antigen (SA4Ag) vaccine comprising capsular polysaccharide (types 5 and 8) conjugates, clumping factor A (ClfA) and manganese transporter C (MntC) is under development. This study was designed to characterize S. aureus isolates recovered from infected patients and also to investigate approaches for examining expression of S. aureus vaccine candidates and the host response during human infection. Confirmation of antigen expression in different disease states is important to support the inclusion of these antigens in a prophylactic vaccine. Hospitalized patients with diagnosed S. aureus wound (27) or bloodstream (24) infections were enrolled. Invasive and nasal carriage S. aureus isolates were recovered and characterized for genotypic diversity. S. aureus antigen expression was evaluated directly by real-time, quantitative, reverse-transcriptase PCR (qRT-PCR) analysis and indirectly by serology using a competitive Luminex immunoassay. Study isolates were genotypically diverse and all had the genes encoding the antigens present in the SA4Ag vaccine. S. aureus nasal carriage was detected in 55% of patients, and in those subjects 64% of the carriage isolates matched the invasive strain. In swab samples with detectable S. aureus triosephosphate isomerase housekeeping gene expression, RNA transcripts encoding the S. aureus virulence factors ClfA, MntC, and capsule polysaccharide were detected by qRT-PCR. Antigen expression was indirectly confirmed by increases in antibody titer during the course of infection from acute to convalescent phase. Demonstration of bacterial transcript expression together with immunological response to the SA4Ag antigens in a clinically relevant patient population provides support for inclusion of these antigens in a prophylactic vaccine.

Low-Concentration Tributyltin Decreases GluR2 Expression via Nuclear Respiratory Factor-1 Inhibition

PubMed Central

Ishida, Keishi; Aoki, Kaori; Takishita, Tomoko; Miyara, Masatsugu; Sakamoto, Shuichiro; Sanoh, Seigo; Kimura, Tomoki; Kanda, Yasunari; Ohta, Shigeru; Kotake, Yaichiro

2017-01-01

Tributyltin (TBT), which has been widely used as an antifouling agent in paints, is a common environmental pollutant. Although the toxicity of high-dose TBT has been extensively reported, the effects of low concentrations of TBT are relatively less well studied. We have previously reported that low-concentration TBT decreases α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA)-type glutamate receptor subunit 2 (GluR2) expression in cortical neurons and enhances neuronal vulnerability to glutamate. However, the mechanism of this TBT-induced GluR2 decrease remains unknown. Therefore, we examined the effects of TBT on the activity of transcription factors that control GluR2 expression. Exposure of primary cortical neurons to 20 nM TBT for 3 h to 9 days resulted in a decrease in GluR2 mRNA expression. Moreover, TBT inhibited the DNA binding activity of nuclear respiratory factor-1 (NRF-1), a transcription factor that positively regulates the GluR2. This result indicates that TBT inhibits the activity of NRF-1 and subsequently decreases GluR2 expression. In addition, 20 nM TBT decreased the expression of genes such as cytochrome c, cytochrome c oxidase (COX) 4, and COX 6c, which are downstream of NRF-1. Our results suggest that NRF-1 inhibition is an important molecular action of the neurotoxicity induced by low-concentration TBT. PMID:28800112

Low-Concentration Tributyltin Decreases GluR2 Expression via Nuclear Respiratory Factor-1 Inhibition.

PubMed

Ishida, Keishi; Aoki, Kaori; Takishita, Tomoko; Miyara, Masatsugu; Sakamoto, Shuichiro; Sanoh, Seigo; Kimura, Tomoki; Kanda, Yasunari; Ohta, Shigeru; Kotake, Yaichiro

2017-08-11

Tributyltin (TBT), which has been widely used as an antifouling agent in paints, is a common environmental pollutant. Although the toxicity of high-dose TBT has been extensively reported, the effects of low concentrations of TBT are relatively less well studied. We have previously reported that low-concentration TBT decreases α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA)-type glutamate receptor subunit 2 ( GluR2 ) expression in cortical neurons and enhances neuronal vulnerability to glutamate. However, the mechanism of this TBT-induced GluR2 decrease remains unknown. Therefore, we examined the effects of TBT on the activity of transcription factors that control GluR2 expression. Exposure of primary cortical neurons to 20 nM TBT for 3 h to 9 days resulted in a decrease in GluR2 mRNA expression. Moreover, TBT inhibited the DNA binding activity of nuclear respiratory factor-1 (NRF-1), a transcription factor that positively regulates the GluR2 . This result indicates that TBT inhibits the activity of NRF-1 and subsequently decreases GluR2 expression. In addition, 20 nM TBT decreased the expression of genes such as cytochrome c, cytochrome c oxidase (COX) 4, and COX 6c, which are downstream of NRF-1. Our results suggest that NRF-1 inhibition is an important molecular action of the neurotoxicity induced by low-concentration TBT.

Quantitative PET Imaging of Tissue Factor Expression Using 18F-Labeled Active Site-Inhibited Factor VII.

PubMed

Nielsen, Carsten H; Erlandsson, Maria; Jeppesen, Troels E; Jensen, Mette M; Kristensen, Lotte K; Madsen, Jacob; Petersen, Lars C; Kjaer, Andreas

2016-01-01

Tissue factor (TF) is upregulated in many solid tumors, and its expression is linked to tumor angiogenesis, invasion, metastasis, and prognosis. A noninvasive assessment of tumor TF expression status is therefore of obvious clinical relevance. Factor VII is the natural ligand to TF. Here we report the development of a new PET tracer for specific imaging of TF using an (18)F-labeled derivative of factor VII. Active site-inhibited factor VIIa (FVIIai) was obtained by inactivation with phenylalanine-phenylalanine-arginine-chloromethyl ketone. FVIIai was radiolabeled with N-succinimidyl 4-(18)F-fluorobenzoate and purified. The corresponding product, (18)F-FVIIai, was injected into nude mice with subcutaneous human pancreatic xenograft tumors (BxPC-3) and investigated using small-animal PET/CT imaging 1, 2, and 4 h after injection. Ex vivo biodistribution was performed after the last imaging session, and tumor tissue was preserved for molecular analysis. A blocking experiment was performed in a second set of mice. The expression pattern of TF in the tumors was visualized by immunohistochemistry and the amount of TF in tumor homogenates was measured by enzyme-linked immunosorbent assay and correlated with the uptake of (18)F-FVIIai in the tumors measured in vivo by PET imaging. The PET images showed high uptake of (18)F-FVIIai in the tumor regions, with a mean uptake of 2.5 ± 0.3 percentage injected dose per gram (%ID/g) (mean ± SEM) 4 h after injection of 7.3-9.3 MBq of (18)F-FVIIai and with an average maximum uptake in the tumors of 7.1 ± 0.7 %ID/g at 4 h. In comparison, the muscle uptake was 0.2 ± 0.01 %ID/g at 4 h. At 4 h, the tumors had the highest uptake of any organ. Blocking with FVIIai significantly reduced the uptake of (18)F-FVIIai from 2.9 ± 0.1 to 1.4 ± 0.1 %ID/g (P < 0.001). The uptake of (18)F-FVIIai measured in vivo by PET imaging correlated (r = 0.72, P < 0.02) with TF protein level measured ex vivo. (18)F-FVIIai is a promising PET tracer for

BFDCA: A Comprehensive Tool of Using Bayes Factor for Differential Co-Expression Analysis.

PubMed

Wang, Duolin; Wang, Juexin; Jiang, Yuexu; Liang, Yanchun; Xu, Dong

2017-02-03

Comparing the gene-expression profiles between biological conditions is useful for understanding gene regulation underlying complex phenotypes. Along this line, analysis of differential co-expression (DC) has gained attention in the recent years, where genes under one condition have different co-expression patterns compared with another. We developed an R package Bayes Factor approach for Differential Co-expression Analysis (BFDCA) for DC analysis. BFDCA is unique in integrating various aspects of DC patterns (including Shift, Cross, and Re-wiring) into one uniform Bayes factor. We tested BFDCA using simulation data and experimental data. Simulation results indicate that BFDCA outperforms existing methods in accuracy and robustness of detecting DC pairs and DC modules. Results of using experimental data suggest that BFDCA can cluster disease-related genes into functional DC subunits and estimate the regulatory impact of disease-related genes well. BFDCA also achieves high accuracy in predicting case-control phenotypes by using significant DC gene pairs as markers. BFDCA is publicly available at http://dx.doi.org/10.17632/jdz4vtvnm3.1. Copyright © 2016 Elsevier Ltd. All rights reserved.

Macrophage Migration Inhibitory Factor Stimulates Angiogenic Factor Expression and Correlates With Differentiation and Lymph Node Status in Patients With Esophageal Squamous Cell Carcinoma

PubMed Central

Ren, Yi; Law, Simon; Huang, Xin; Lee, Ping Yin; Bacher, Michael; Srivastava, Gopesh; Wong, John

2005-01-01

Objective: The objectives of this study were: 1) to examine the expression of macrophage migration inhibitory factor (MIF) in esophageal squamous cell carcinoma (ESCC); 2) to see if a relationship exists between MIF expression, clinicopathologic features, and long-term prognosis; and 3) to ascertain the possible biologic function of MIF in angiogenesis. Summary Background Data: MIF has been linked to fundamental processes such as those controlling cell proliferation, cell survival, angiogenesis, and tumor progression. Its role in ESCC, and the correlation of MIF expression and tumor pathologic features in patients, has not been elucidated. Methods: The expression of MIF in tumor and nontumor tissues was examined by immunohistochemical staining. Concentrations of MIF, vascular endothelial growth factor (VEGF), and interleukin-8 (IL-8) in patients’ sera and in the supernatant of tumor cells culture were examined by ELISA. Correlations with clinicopathologic factors were made. Results: In 72 patients with ESCC, intracellular MIF was overexpressed in esophagectomy specimens. The expression of MIF correlated with both tumor differentiation and lymph node status. The median survival in the low-MIF expression group (<50% positively stained cancer cells on immunohistochemistry) and high expression group (≥50% positively stained cancer cells) was 28.3 months and 15.8 months, respectively (P = 0.03). The 3-year survival rates for the 2 groups were 37.7% and 12.1%, respectively. MIF expression was related to microvessel density; increased MIF serum levels also correlated with higher serum levels of VEGF. In addition, in vitro MIF stimulation of esophageal cancer cell lines induced a dose-dependent increase in VEGF and IL-8 secretion. Conclusions: These results demonstrate, for the first time, that human esophageal carcinomas express and secrete large amounts of MIF. Through its effects on VEGF and IL-8, MIF may serve as an autocrine factor in angiogenesis and thus play an

Individual Differences in the Expression of Conditioned Fear Are Associated with Endogenous Fibroblast Growth Factor 2

ERIC Educational Resources Information Center

Graham, Bronwyn M.; Richardson, Rick

2016-01-01

These experiments examined the relationship between the neurotrophic factor fibroblast growth factor 2 (FGF2) and individual differences in the expression of conditioned fear. Experiments 1 and 2 demonstrated that rats naturally expressing low levels of contextual or cued fear have higher levels of hippocampal FGF2 relative to rats that express…

Expression Profile of Interferon Regulatory Factor 1 in Chronic Hepatitis B Virus-Infected Liver Transplant Patients.

PubMed

Janfeshan, Sahar; Yaghobi, Ramin; Eidi, Akram; Karimi, Mohammad Hossein; Geramizadeh, Bita; Malekhosseini, Seyed Ali; Kafilzadeh, Farshid

2017-12-01

Hepatitis B virus, which mainly affects normal liver function, leads to severe acute and chronic hepatitis, resulting in cirrhosis and hepatocellular carcinoma, but can be safely treated after liver transplant. Evaluation of determinative biomarkers may facilitate more effective treatment of posttransplant rejection. Therefore, we investigated interferon regulatory factor 1 expression in hepatitis B virus-infected liver transplant patients with and without previous rejection compared with controls. Hepatitis B virus-infected liver recipients were divided into those with (20 patients) and without a rejection (26 patients), confirmed by pathologic analyses in those who had a rejection. In addition, a healthy control group composed of 13 individuals was included. Expression levels of interferon regulatory factor 1 were evaluated during 3 follow-ups after transplant using an in-house comparative SYBR green real-time polymerase chain reaction method. Statistical analyses were performed with SPSS software (SPSS: An IBM Company, version 16.0, IBM Corporation, Armonk, NY, USA). Modifications of interferon regulatory factor 1 gene expression levels in patient groups with and without rejection were not significant between days 1, 4, and 7 after liver transplant. Interferon regulatory factor 1 mRNA expression levels were down-regulated in patients without rejection versus patients with rejection, although not significantly at day 1 (P = .234) and day 4 (P = .302) but significantly at day 7 (P = .004) after liver transplant. Down-regulation of interferon regulatory factor 1 gene expression in hepatitis B virus patients without rejection emphasized counteraction between hepatitis B virus replication and interferon regulatory factor 1 production. On the other hand, interferon regulatory factor 1 gene overexpression in patients with rejection may result in inflammatory reactions and ischemic-reperfusion injury. Therefore, a better understanding of the association between

Identification and embryonic expression of a new AP-2 transcription factor, AP-2 epsilon.

PubMed

Wang, Hao-Ven; Vaupel, Kristina; Buettner, Reinhard; Bosserhoff, Anja-Katrin; Moser, Markus

2004-09-01

AP-2 proteins comprise a family of highly related transcription factors, which are expressed during mouse embryogenesis in a variety of ectodermal, neuroectodermal, and mesenchymal tissues. AP-2 transcription factors were shown to be involved in morphogenesis of craniofacial, urogenital, neural crest-derived, and placental tissues. By means of a partial cDNA fragment identified during an expressed sequence tag search for AP-2 genes, we identified a fifth, previously unknown AP-2-related gene, AP-2 epsilon. AP-2 epsilon encodes an open reading frame of 434 amino acids, which reveals the typical modular structure of AP-2 transcription factors with highly conserved C-terminal DNA binding and dimerization domains. Although the N-terminally localized activation domain is less homologous, position and identity of amino acids essential for transcriptional transactivation are conserved. Reverse transcriptase-polymerase chain reaction analyses of murine embryos revealed AP-2 epsilon expression from gestational stage embryonic day 7.5 throughout all later embryonic stages until birth. Whole-mount in situ hybridization using a specific AP-2 epsilon cDNA fragment demonstrated that during embryogenesis, expression of AP-2 epsilon is mainly restricted to neural tissue, especially the midbrain, hindbrain, and olfactory bulb. This expression pattern was confirmed by immunohistochemistry with an AP-2 epsilon-specific antiserum. By using this antiserum, we could further localize AP-2 epsilon expression in a hypothalamic nucleus and the neuroepithelium of the vomeronasal organ, suggesting an important function of AP-2 epsilon for the development of the olfactory system.

Validation of a Theory of Planned Behavior-Based Questionnaire to Examine Factors Associated With Milk Expression.

PubMed

Bai, Yeon K; Dinour, Lauren M

2017-11-01

A proper assessment of multidimensional needs for breastfeeding mothers in various settings is crucial to facilitate and support breastfeeding and its exclusivity. The theory of planned behavior (TPB) has been used frequently to measure factors associated with breastfeeding. Full utility of the TPB requires accurate measurement of theory constructs. Research aim: This study aimed to develop and confirm the psychometric properties of an instrument, Milk Expression on Campus, based on the TPB and to establish the reliability and validity of the instrument. In spring 2015, 218 breastfeeding (current or in the recent past) employees and students at one university campus in northern New Jersey completed the online questionnaire containing demography and theory-based items. Internal consistency (α) and split-half reliability ( r) tests and factor analyses established and confirmed the reliability and construct validity of this instrument. Milk Expression on Campus showed strong and significant reliabilities as a full scale (α = .78, r = .74, p < .001) and theory construct subscales. Validity was confirmed as psychometric properties corresponded to the factors extracted from the scale. Four factors extracted from the direct construct subscales accounted for 79.49% of the total variability. Four distinct factors from the indirect construct subscales accounted for 73.68% of the total variability. Milk Expression on Campus can serve as a model TPB-based instrument to examine factors associated with women's milk expression behavior. The utility of this instrument extends to designing effective promotion programs to foster breastfeeding and milk expression behaviors in diverse settings.

Identifying Stress Transcription Factors Using Gene Expression and TF-Gene Association Data

PubMed Central

Wu, Wei-Sheng; Chen, Bor-Sen

2007-01-01

Unicellular organisms such as yeasts have evolved to survive environmental stresses by rapidly reorganizing the genomic expression program to meet the challenges of harsh environments. The complex adaptation mechanisms to stress remain to be elucidated. In this study, we developed Stress Transcription Factor Identification Algorithm (STFIA), which integrates gene expression and TF-gene association data to identify the stress transcription factors (TFs) of six kinds of stresses. We identified some general stress TFs that are in response to various stresses, and some specific stress TFs that are in response to one specific stress. The biological significance of our findings is validated by the literature. We found that a small number of TFs may be sufficient to control a wide variety of expression patterns in yeast under different stresses. Two implications can be inferred from this observation. First, the adaptation mechanisms to different stresses may have a bow-tie structure. Second, there may exist extensive regulatory cross-talk among different stress responses. In conclusion, this study proposes a network of the regulators of stress responses and their mechanism of action. PMID:20066130

Transcription Factors Expressed in Lateral Organ Boundaries: Identification of Downstream Targets

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

Springer, Patricia S

2010-07-12

The processes of lateral organ initiation and patterning are central to the generation of mature plant form. Characterization of the molecular mechanisms underlying these processes is essential to our understanding of plant development. Communication between the shoot apical meristem and initiating organ primordia is important both for functioning of the meristem and for proper organ patterning, and very little is known about this process. In particular, the boundary between meristem and leaf is emerging as a critical region that is important for SAM maintenance and regulation of organogenesis. The goal of this project was to characterize three boundary-expressed genes thatmore » encode predicted transcription factors. Specifically, we have studied LATERAL ORGAN BOUNDARIES (LOB), LATERAL ORGAN FUSION1 (LOF1), and LATERAL ORGAN FUSION2 (LOF2). LOB encodes the founding member of the LOB-DOMAIN (LBD) plant-specific DNA binding transcription factor family and LOF1 and LOF2 encode paralogous MYB-domain transcription factors. We characterized the genetic relationship between these three genes and other boundary and meristem genes. We also used an ectopic inducible expression system to identify direct targets of LOB.« less

The over-expression of a chrysanthemum WRKY transcription factor enhances aphid resistance.

PubMed

Li, Peiling; Song, Aiping; Gao, Chunyan; Jiang, Jiafu; Chen, Sumei; Fang, Weimin; Zhang, Fei; Chen, Fadi

2015-10-01

Members of the large WRKY transcription factor family are responsible for the regulation of plant growth, development and the stress response. Here, five WRKY members were isolated from chrysanthemum. They each contained a single WRKY domain and a C2H2 zinc finger motif, so were classified into group II. Transient expression experiments demonstrated that all five were expressed in the nucleus, although CmWRKY42 was also expressed in the cytoplasm. When expressed heterologously in yeast, the products of CmWRKY22 and CmWRKY48 exhibited transactivation activity, while those of CmWRKY21, CmWRKY40 and CmWRKY42 did not. The transcription of the five CmWRKY genes was profiled when the plants were challenged with a variety of abiotic and biotic stress agents, as well as being treated with various phytohormones. CmWRKY21 proved to be markedly induced by salinity stress, and suppressed by high temperature exposure; CmWRKY22 was induced by high temperature exposure; CmWRKY40 was highly induced by salinity stress, and treatment with either abscisic acid (ABA) or methyl jasmonate (MeJA); CmWRKY42 was up-regulated by salinity stress, low temperature, ABA and MeJA treatment and aphid infestation; CmWRKY48 was induced by drought stress, ABA and MeJA treatment and aphid infestation. The function of CmWRKY48 was further investigated by over-expressing it transgenically. The constitutive expression of this transcription factor inhibited the aphids' population growth capacity, suggesting that it may represent an important component of the plant's defense machinery against aphids. Copyright © 2015 Elsevier Masson SAS. All rights reserved.

Conservation of transcription factor binding events predicts gene expression across species

PubMed Central

Hemberg, Martin; Kreiman, Gabriel

2011-01-01

Recent technological advances have made it possible to determine the genome-wide binding sites of transcription factors (TFs). Comparisons across species have suggested a relatively low degree of evolutionary conservation of experimentally defined TF binding events (TFBEs). Using binding data for six different TFs in hepatocytes and embryonic stem cells from human and mouse, we demonstrate that evolutionary conservation of TFBEs within orthologous proximal promoters is closely linked to function, defined as expression of the target genes. We show that (i) there is a significantly higher degree of conservation of TFBEs when the target gene is expressed in both species; (ii) there is increased conservation of binding events for groups of TFs compared to individual TFs; and (iii) conserved TFBEs have a greater impact on the expression of their target genes than non-conserved ones. These results link conservation of structural elements (TFBEs) to conservation of function (gene expression) and suggest a higher degree of functional conservation than implied by previous studies. PMID:21622661

Analgesia for neuropathic pain by dorsal root ganglion transplantation of genetically engineered mesenchymal stem cells: initial results.

PubMed

Yu, Hongwei; Fischer, Gregory; Ebert, Allison D; Wu, Hsiang-En; Bai, Xiaowen; Hogan, Quinn H

2015-02-12

Cell-based therapy may hold promise for treatment of chronic pain. Mesenchymal stem cells (MSCs) are readily available and robust, and their secretion of therapeutic peptides can be enhanced by genetically engineering. We explored the analgesic potential of transplanting bone marrow-derived MSCs that have been transduced with lentivectors. To optimize efficacy and safety, primary sensory neurons were targeted by MSC injection into the dorsal root ganglia (DRGs). MSCs were transduced using lentivectors to express enhanced green fluorescent protein (EGFP) or to co-express the analgesic peptide glial cell line-derived neurotrophic factor (GDNF) and EGFP by a viral 2A bicistronic transgene cassette. Engineered MSCs were injected into the 4(th) lumbar (L4) and L5 DRGs of adult allogeneic rats to evaluate survival in the DRGs. MSCs were detected by immunofluorescence staining up to 2-3 weeks after injection, distributed in the extracellular matrix space without disrupting satellite glial cell apposition to sensory neurons, suggesting well-tolerated integration of engrafted MSCs into DRG tissue. To examine their potential for inhibiting development of neuropathic pain, MSCs were injected into the L4 and L5 DRGs ipsilateral to a spinal nerve ligation injury. Animals injected with GDNF-engineered MSCs showed moderate but significant reduction in mechanical allodynia and hyperalgesia compared to controls implanted with MSCs expressing EGFP alone. We also observed diminished long-term survival of allografted MSCs at 3 weeks, and the development of a highly-proliferating population of MSCs in 12% of DRGs after transplantation. These data indicate that genetically modified MSCs secreting analgesic peptides could potentially be developed as a novel DRG-targeted cell therapy for treating neuropathic pain. However, further work is needed to address the challenges of MSC survival and excess proliferation, possibly with trials of autologous MSCs, evaluation of clonally selected

Parental expression of disappointment: should it be a factor in Hoffman's model of parental discipline?

PubMed

Patrick, Renee B; Gibbs, John C

2007-06-01

The authors addressed whether parental expression of disappointment should be included as a distinct factor in M. L. Hoffman's well-established typology of parenting styles (induction, love withdrawal, power assertion). Hoffman's 3-factor model, along with a more inclusive 4-factor model (induction, love withdrawal, power assertion, and expressions of disappointment), were respectively evaluated in exploratory factor analyses. The analysis utilized extant data comprised of responses by children (N = 73) and their mothers (N = 67) to an adaptation of M. L. Hoffman and H. D. Saltzstein's parental discipline measure. The findings supported Hoffman's original model. Disappointment may be reducible to love withdrawal or induction, although disappointment may be a more appropriate induction for adolescents.

Expression of allograft inflammatory factor-1 in inflammatory skin disorders.

PubMed

Orsmark, Christina; Skoog, Tiina; Jeskanen, Leila; Kere, Juha; Saarialho-Kere, Ulpu

2007-01-01

Allograft inflammatory factor-1 (AIF-1) is an evolutionarily conserved, inflammatory protein produced by activated macrophages during chronic transplant rejection and in inflammatory brain lesions. Since T-cell-mediated inflammation is common to various dermatoses and nothing is known about AIF-1 in skin, we studied its protein expression at the tissue level and regulation in monocytic cell lines by various agents. Using immunohistochemistry, we found that AIF-1 is expressed at low levels in normal skin, but is highly upregulated in various inflammatory skin disorders, such as psoriasis, lichen planus, graft-versus-host disease and mycosis fungoides. The main cell types expressing AIF-1 in affected skin are macrophages and Langerhans' cells. We also show by real-time PCR that AIF-1 mRNA levels in monocytic THP-1 and U937 cell lines are significantly upregulated by retinoic acid as well as a number of cytokines. We conclude that AIF-1 may mediate survival and pro-inflammatory properties of macrophages in skin diseases.

Effect of hypoxia on tissue factor pathway inhibitor expression in breast cancer.

PubMed

Cui, X Y; Tinholt, M; Stavik, B; Dahm, A E A; Kanse, S; Jin, Y; Seidl, S; Sahlberg, K K; Iversen, N; Skretting, G; Sandset, P M

2016-02-01

ESSENTIALS: A hypoxic microenvironment is a common feature of tumors that may influence activation of coagulation. MCF-7 and SK-BR-3 breast cancer cells and breast cancer tissue samples were used. The results showed transcriptional repression of tissue factor pathway inhibitor expression in hypoxia. Hypoxia-inducible factor 1α may be a target for the therapy of cancer-related coagulation and thrombosis. Activation of coagulation is a common finding in patients with cancer, and is associated with an increased risk of venous thrombosis. As a hypoxic microenvironment is a common feature of solid tumors, we investigated the role of hypoxia in the regulation of tissue factor (TF) pathway inhibitor (TFPI) expression in breast cancer. To explore the transcriptional regulation of TFPI by hypoxia-inducible factor (HIF)-1α in breast cancer cells and their correlation in breast cancer tissues. MCF-7 and SK-BR-3 breast cancer cells were cultured in 1% oxygen or treated with cobalt chloride (CoCl2 ) to mimic hypoxia. Time-dependent and dose-dependent downregulation of TFPI mRNA (quantitative RT-PCR) and of free TFPI protein (ELISA) were observed in hypoxia. Western blotting showed parallel increases in the levels of HIF-1α protein and TF. HIF-1α inhibitor abolished or attenuated the hypoxia-induced downregulation of TFPI. Luciferase reporter assay showed that both hypoxia and HIF-1α overexpression caused strong repression of TFPI promoter activity. Subsequent chromatin immunoprecipitation and mutagenesis analysis demonstrated a functional hypoxia response element within the TFPI promoter, located at -1065 to -1060 relative to the transcriptional start point. In breast cancer tissue samples, gene expression analyses showed a positive correlation between the mRNA expression of TFPI and that of HIF-1α. This study demonstrates that HIF-1α is involved in the transcriptional regulation of the TFPI gene, and suggests that a hypoxic microenvironment inside a breast tumor may

Fluoxetine ameliorates behavioral and neuropathological deficits in a transgenic model mouse of α-synucleinopathy.

PubMed

Ubhi, Kiren; Inglis, Chandra; Mante, Michael; Patrick, Christina; Adame, Anthony; Spencer, Brian; Rockenstein, Edward; May, Verena; Winkler, Juergen; Masliah, Eliezer

2012-04-01

The term α-synucleinopathies refers to a group of age-related neurological disorders including Parkinson's disease (PD), Dementia with Lewy Bodies (DLB) and Multiple System Atrophy (MSA) that display an abnormal accumulation of alpha-synuclein (α-syn). In contrast to the neuronal α-syn accumulation observed in PD and DLB, MSA is characterized by a widespread oligodendrocytic α-syn accumulation. Transgenic mice expressing human α-syn under the oligodendrocyte-specific myelin basic protein promoter (MBP1-hαsyn tg mice) model many of the behavioral and neuropathological alterations observed in MSA. Fluoxetine, a selective serotonin reuptake inhibitor, has been shown to be protective in toxin-induced models of PD, however its effects in an in vivo transgenic model of α-synucleinopathy remain unclear. In this context, this study examined the effect of fluoxetine in the MBP1-hαsyn tg mice, a model of MSA. Fluoxetine administration ameliorated motor deficits in the MBP1-hαsyn tg mice, with a concomitant decrease in neurodegenerative pathology in the basal ganglia, neocortex and hippocampus. Fluoxetine administration also increased levels of the neurotrophic factors, GDNF (glial-derived neurotrophic factor) and BDNF (brain-derived neurotrophic factor) in the MBP1-hαsyn tg mice compared to vehicle-treated tg mice. This fluoxetine-induced increase in GDNF and BDNF protein levels was accompanied by activation of the ERK signaling pathway. The effects of fluoxetine administration on myelin and serotonin markers were also examined. Collectively these results indicate that fluoxetine may represent a novel therapeutic intervention for MSA and other neurodegenerative disorders. Copyright © 2011 Elsevier Inc. All rights reserved.

Breast Angiosarcoma: Case Series and Expression of Vascular Endothelial Growth Factor

PubMed Central

Brar, Rondeep; West, Robert; Witten, Daniela; Raman, Bhargav; Jacobs, Charlotte; Ganjoo, Kristen

2009-01-01

Purpose Angiosarcoma of the breast is a rare, malignant tumor for which little is known regarding prognostic indicators and optimal therapeutic regimens. To address this issue, we performed a retrospective analysis of breast angiosarcoma cases seen at Stanford University along with immunohistochemical analysis for markers of angiogenesis. Methods Breast angiosarcoma cases seen between 1980 and 2008 were examined. Viable tissue blocks were analyzed for expression of vascular endothelial growth factor and its receptors. Results A total of 16 cases were identified. Data was collected regarding epidemiology, treatment, response rates, disease-free survival, and the use of various imaging modalities. Five tissue blocks remained viable for immunohistochemical analysis. Vascular endothelial growth factor-A was positively expressed in 3 of these samples. Conclusion Angiosarcoma of the breast is an aggressive malignancy with a propensity for both local recurrence and distant metastases. Angiogenesis inhibition may represent a novel therapeutic modality in this rare, vascular malignancy. PMID:20737044

Vascular endothelial growth factor-C (VEGF-C) expression predicts lymph node metastasis of transitional cell carcinoma of the bladder.

PubMed

Suzuki, Kazumi; Morita, Tatsuo; Tokue, Akihiko

2005-02-01

It has been found that expression of vascular endothelial growth factor-C (VEGF-C) in several carcinomas is significantly associated with angiogenesis, lymphangiogenesis and regional lymph node metastasis. However, VEGF-C expression in bladder transitional cell carcinoma (TCC) has not yet been reported. To elucidate the role of VEGF-C in bladder TCC, we examined VEGF-C expression in bladder TCC and pelvic lymph node metastasis specimens obtained from patients who underwent radical cystectomy. Eighty-seven patients who underwent radical cystectomy for clinically organ-confined TCC of the bladder were enrolled in the present study. No neoadjuvant treatments, except transurethral resection of the tumor, were given to these patients. The VEGF-C expressions of 87 bladder tumors and 20 pelvic lymph node metastasis specimens were examined immunohistochemically and the association between VEGF-C expression and clinicopathological factors, including angiogenesis as evaluated by microvessel density (MVD), was also examined. Vascular endothelial growth factor-C expression was found in the cytoplasm of tumor cells, but not in the normal transitional epithelium. Vascular endothelial growth factor-C expression was significantly associated with the pathological T stage (P = 0.0289), pelvic lymph node metastasis (P < 0.0001), lymphatic involvement (P = 0.0008), venous involvement (P = 0.0002) and high MVD (P = 0.0043). The multivariate analysis demonstrated that VEGF-C expression and high MVD in bladder TCC were independent risk factors influencing the pelvic lymph node metastasis. Moreover, the patients with VEGF-C-positive tumors had significantly poorer prognoses than those with the VEGF-C-negative tumors (P = 0.0087) in the univariate analysis. The multivariate analysis based on Cox proportional hazard model showed that the independent prognostic factors were patient age (P = 0.0132) and pelvic lymph node metastasis (P = 0.0333). The present study suggests that VEGF

Gene Expression Profiling of Transcription Factors of Helicobacter pylori under Different Environmental Conditions.

PubMed

De la Cruz, Miguel A; Ares, Miguel A; von Bargen, Kristine; Panunzi, Leonardo G; Martínez-Cruz, Jessica; Valdez-Salazar, Hilda A; Jiménez-Galicia, César; Torres, Javier

2017-01-01

Helicobacter pylori is a Gram-negative bacterium that colonizes the human gastric mucosa and causes peptic ulcers and gastric carcinoma. H. pylori strain 26695 has a small genome (1.67 Mb), which codes for few known transcriptional regulators that control bacterial metabolism and virulence. We analyzed by qRT-PCR the expression of 16 transcriptional regulators in H. pylori 26695, including the three sigma factors under different environmental conditions. When bacteria were exposed to acidic pH, urea, nickel, or iron, the sigma factors were differentially expressed with a particularly strong induction of fliA . The regulatory genes hrcA, hup , and crdR were highly induced in the presence of urea, nickel, and iron. In terms of biofilm formation fliA, flgR, hp1021, fur, nikR , and crdR were induced in sessile bacteria. Transcriptional expression levels of rpoD, flgR, hspR, hp1043 , and cheY were increased in contact with AGS epithelial cells. Kanamycin, chloramphenicol, and tetracycline increased or decreased expression of regulatory genes, showing that these antibiotics affect the transcription of H. pylori . Our data indicate that environmental cues which may be present in the human stomach modulate H. pylori transcription.

Heterogeneous expression pattern of pro- and anti-apoptotic factors in myeloid progenitor cells of patients with severe congenital neutropenia treated with granulocyte colony-stimulating factor.

PubMed

Cario, Gunnar; Skokowa, Julia; Wang, Zheng; Bucan, Vesna; Zeidler, Cornelia; Stanulla, Martin; Schrappe, Martin; Welte, Karl

2005-04-01

Apoptosis is accelerated in the myeloid progenitor cells of patients with severe congenital neutropenia (CN). Granulocyte colony-stimulating factor (G-CSF) increases neutrophil numbers in most CN patients. The effect of G-CSF on apoptosis in CN was analysed by apoptosis rate and expression of anti- and pro-apoptotic factors. G-CSF-treated patients showed higher apoptosis frequency, lower expression of bcl-2 and bcl-xL, but higher expression of bfl-1/A1 and mcl-1. Caspase 9 was highly expressed in patients and controls after G-CSF administration. Thus, G-CSF acts on apoptosis regulation, but additional mechanisms leading to the increase of neutrophil numbers must be assumed.

NFI Transcription Factors Interact with FOXA1 to Regulate Prostate-Specific Gene Expression

PubMed Central

Elliott, Amicia D.; DeGraff, David J.; Anderson, Philip D.; Anumanthan, Govindaraj; Yamashita, Hironobu; Sun, Qian; Friedman, David B.; Hachey, David L.; Yu, Xiuping; Sheehan, Jonathan H.; Ahn, Jung-Mo; Raj, Ganesh V.; Piston, David W.; Gronostajski, Richard M.; Matusik, Robert J.

2014-01-01

Androgen receptor (AR) action throughout prostate development and in maintenance of the prostatic epithelium is partly controlled by interactions between AR and forkhead box (FOX) transcription factors, particularly FOXA1. We sought to identity additional FOXA1 binding partners that may mediate prostate-specific gene expression. Here we identify the nuclear factor I (NFI) family of transcription factors as novel FOXA1 binding proteins. All four family members (NFIA, NFIB, NFIC, and NFIX) can interact with FOXA1, and knockdown studies in androgen-dependent LNCaP cells determined that modulating expression of NFI family members results in changes in AR target gene expression. This effect is probably mediated by binding of NFI family members to AR target gene promoters, because chromatin immunoprecipitation (ChIP) studies found that NFIB bound to the prostate-specific antigen enhancer. Förster resonance energy transfer studies revealed that FOXA1 is capable of bringing AR and NFIX into proximity, indicating that FOXA1 facilitates the AR and NFI interaction by bridging the complex. To determine the extent to which NFI family members regulate AR/FOXA1 target genes, motif analysis of publicly available data for ChIP followed by sequencing was undertaken. This analysis revealed that 34.4% of peaks bound by AR and FOXA1 contain NFI binding sites. Validation of 8 of these peaks by ChIP revealed that NFI family members can bind 6 of these predicted genomic elements, and 4 of the 8 associated genes undergo gene expression changes as a result of individual NFI knockdown. These observations suggest that NFI regulation of FOXA1/AR action is a frequent event, with individual family members playing distinct roles in AR target gene expression. PMID:24801505

Mangiferin inhibits tumor necrosis factor-α-induced matrix metalloproteinase-9 expression and cellular invasion by suppressing nuclear factor-κB activity.

PubMed

Dilshara, Matharage Gayani; Kang, Chang-Hee; Choi, Yung Hyun; Kim, Gi-Young

2015-10-01

We investigated the effects of mangiferin on the expression and activity of metalloproteinase (MMP)-9 and the invasion of tumor necrosis factor (TNF)-α-stimulated human LNCaP prostate carcinoma cells. Reverse-transcription polymerase chain reaction (RT-PCR) and western blot analysis showed that mangiferin significantly reversed TNF-α-induced mRNA and protein expression of MMP-9 expression. Zymography data confirmed that stimulation of cells with TNF-α significantly increased MMP-9 activity. However, mangiferin substantially reduced the TNF-α-induced activity of MMP-9. Additionally, a matrigel invasion assay showed that mangiferin significantly reduced TNF-α-induced invasion of LNCaP cells. Compared to untreated controls, TNF-α-stimulated LNCaP cells showed a significant increase in nuclear factor-κB (NF-κB) luciferase activity. However, mangiferin treatment markedly decreased TNF-α-induced NF-κB luciferase activity. Furthermore, mangiferin suppressed nuclear translocation of the NF-κB subunits p65 and p50. Collectively, our results indicate that mangiferin is a potential anti-invasive agent that acts by suppressing NF-κB-mediated MMP-9 expression.

Building gene expression signatures indicative of transcription factor activation to predict AOP modulation

EPA Science Inventory

Building gene expression signatures indicative of transcription factor activation to predict AOP modulation Adverse outcome pathways (AOPs) are a framework for predicting quantitative relationships between molecular initiatin...

Analysis of Obesity-Related Factors and their Association with Aromatase Expression in Canine Malignant Mammary Tumours.

PubMed

Shin, J-I; Lim, H-Y; Kim, H-W; Seung, B-J; Ju, J-H; Sur, J-H

2016-07-01

This study was designed to investigate the role of obesity in canine malignant mammary tumours (CMMTs), by assessing aromatase expression and the regulatory roles of immune mediators such as cyclo-oxygenase-2 (COX2), prostaglandin E2 (PGE2), nuclear factor kappa beta (NF-κB), hypoxia inducible factor-1α (HIF-1α) and adipokines (i.e. leptin) in lean, optimal body weight, overweight and obese animals. Clinicopathological data, including the breed, body weight, body condition score and age and neutering status, were collected, together with histopathological characteristics (i.e. histological types, grading and lymphatic invasion). To determine the expression of each factor, immunohistochemistry was conducted with 60 samples of malignant CMMTs. CMMTs from overweight and obese animals had significantly elevated levels of PGE2, and aromatase expression correlated significantly with PGE2, NF-κB and leptin expression. However, no significant difference was observed in terms of histopathological characteristics. The results suggest that PGE2, a known obesity-related immune mediator, could be upregulated in CMMTs from overweight and obese animals. In addition, PGE2, NF-κB and leptin influenced the expression of aromatase, as observed in women. Copyright © 2016 Elsevier Ltd. All rights reserved.

Expression of Fushi tarazu factor 1 homolog and Pit-1 genes in the pituitaries of pre-spawning chum and sockeye salmon.

PubMed

Higa, M; Ando, H; Urano, A

2001-06-01

Fushi tarazu factor-1 (FTZ-F1) and Pit-1 are major pituitary transcription factors, controlling expression of genes coding for gonadotropin (GTH) subunits and growth hormone/prolactin/somatolactin family hormone, respectively. As a first step to investigate physiological factors regulating gene expression of these transcription factors, we determined their mRNA levels in the pituitaries of chum salmon (Oncorhynchus keta) at different stages of sexual maturation. FTZ-F1 gene expression was increased in males at the stage before spermiation, where the levels of GTH alpha and IIbeta subunit mRNAs were elevated. Pit-1 mRNA showed maximum levels at the final stage of sexual maturation in both sexes, when expression of somatolactin gene peaked. To clarify whether gonadotropin-releasing hormone (GnRH) is involved in these increases in FTZ-F1 and Pit-1 gene expression, we examined effects of GnRH analog (GnRHa) administration on their gene expression in maturing sockeye salmon (Oncorhynchus nerka). GnRHa stimulated Pit-1 gene expression in females only, but failed to stimulate FTZ-F1 gene expression in both sexes. The up-regulated expression of FTZ-F1 and Pit-1 genes at the pre-spawning stages suggest that the two transcription factors have roles in sexual maturation of salmonids. Physiological factors regulating gene expression of FTZ-F1 and Pit-1 are discussed in this review.

Histone Deacetylase Inhibitors Repress Tumoral Expression of the Proinvasive Factor RUNX2.

PubMed

Sancisi, Valentina; Gandolfi, Greta; Ambrosetti, Davide Carlo; Ciarrocchi, Alessia

2015-05-01

Aberrant reactivation of embryonic pathways occurs commonly in cancer. The transcription factor RUNX2 plays a fundamental role during embryogenesis and is aberrantly reactivated during progression and metastasization of different types of human tumors. In this study, we attempted to dissect the molecular mechanisms governing RUNX2 expression and its aberrant reactivation. We identified a new regulatory enhancer element, located within the RUNX2 gene, which is responsible for the activation of the RUNX2 promoter and for the regulation of its expression in cancer cells. Furthermore, we have shown that treatment with the anticancer compounds histone deacetylase inhibitor (HDACi) results in a profound inhibition of RUNX2 expression, which is determined by the disruption of the transcription-activating complex on the identified enhancer. These data envisage a possible targeting strategy to counteract the oncongenic function of RUNX2 in cancer cells and provide evidence that the cytotoxic activity of HDACi in cancer is not only dependent on the reactivation of silenced oncosuppressors but also on the repression of oncogenic factors that are necessary for survival and progression. ©2015 American Association for Cancer Research.

Involvement of human decidual cell-expressed tissue factor in uterine hemostasis and abruption.

PubMed

Lockwood, C J; Paidas, M; Murk, W K; Kayisli, U A; Gopinath, A; Huang, S J; Krikun, G; Schatz, F

2009-11-01

Vascular injury increases access and binding of plasma-derived factor VII to perivascular cell membrane-bound tissue factor (TF). The resulting TF/VIIa complex promotes hemostasis by cleaving pro-thrombin to thrombin leading to the fibrin clot. In human pregnancy, decidual cell-expressed TF prevents decidual hemorrhage (abruption). During placentation, trophoblasts remodel decidual spiral arteries into high conductance vessels. Shallow trophoblast invasion impedes decidual vascular conversion, producing an inadequate uteroplacental blood flow that elicits abruption-related placental ischemia. Thrombin induces several biological effects via cell surface protease activated receptors. In first trimester human DCs thrombin increases synthesis of sFlt-1, which elicits placental ischemia by impeding angiogenesis-related decidual vascular remodeling. During pregnacy, the fibrillar collagen-rich amnion and choriodecidua extracellular matrix (ECM) provides greater than additive tensile strength and structural integrity. Thrombin acts as an autocrine/paracrine mediator that degrades these ECMs by augmenting decidual cell expression of: 1) matrix metalloproteinases and 2) interleukin-8, a key mediator of abruption-associated decidual infiltration of neutrophils, which express several ECM degrading proteases. Among the cell types at the maternal fetal interface at term, TF expression is highest in decidual cells indicating that this TF meets the hemostatic demands of labor and delivery. TF expression in cultured term decidual cells is enhanced by progestin and thrombin suggesting that the maintenance of elevated circulating progesterone provides hemostatic protection and that abruption-generated thrombin acts in an autocrine/paracrine fashion on decidual cells to promote hemostasis via enhanced TF expression.

Involvement of human decidual cell-expressed tissue factor in uterine hemostasis and abruption

PubMed Central

Lockwood, C.J.; Paidas, M.; Murk, W.K.; Kayisli, U.A.; Gopinath, A.; Krikun, G.; Huang, S.J.; Schatz, F.

2009-01-01

Vascular injury increases access and binding of plasma-derived factor VII to perivascular cell membrane-bound tissue factor (TF). The resulting TF/VIIa complex promotes hemostasis by cleaving pro-thrombin to thrombin leading to the fibrin clot. In human pregnancy, decidual cell-expressed TF prevents decidual hemorrhage (abruption). During placentation, trophoblasts remodel decidual spiral arteries into high conductance vessels. Shallow trophoblast invasion impedes decidual vascular conversion, producing an inadequate uteroplacental blood flow that elicits abruption-related placental ischemia. Thrombin induces several biological effects via cell surface protease activated receptors. In first trimester human DCs thrombin increases synthesis of sFlt-1, which elicits placental ischemia by impeding angiogenesis-related decidual vascular remodeling. During pregnacy, the fibrillar collagen-rich amnion and choriodecidua extracellular matrix (ECM) provides greater than additive tensile strength and structural integrity. Thrombin acts as an autocrine/paracrine mediator that degrades these ECMs by augmenting decidual cell expression of: 1) matrix metalloproteinases and 2) interleukin-8, a key mediator of abruption-associated decidual infiltration of neutrophils, which express several ECM degrading proteases. Our recent observations that: 1) among the cell types at the maternal fetal interface at term TF expression is highest in decidual cells indicates that this TF meets the hemostatic demands of labor and delivery; 2) TF expression in cultured term decidual cells is enhanced by progestin and thrombin suggest that maintenance of elevated circulating progesterone at term provides hemostatic protection, whereas abruption-generated thrombin can act in autocrine/paracrine fashion on DCs to promote hemostasis via enhanced TF expression. PMID:19720393

Combined caveolin-1 and epidermal growth factor receptor expression as a prognostic marker for breast cancer.

PubMed

Liang, Ya-Nan; Liu, Yu; Wang, Letian; Yao, Guodong; Li, Xiaobo; Meng, Xiangning; Wang, Fan; Li, Ming; Tong, Dandan; Geng, Jingshu

2018-06-01

Previous studies have indicated that caveolin-1 (Cav-1) is able to bind the signal transduction factor epidermal growth factor receptor (EGFR) to regulate its tyrosine kinase activity. The aim of the present study was to evaluate the clinical significance of Cav-1 gene expression in association with the expression of EGFR in patients with breast cancer. Primary breast cancer samples from 306 patients were analyzed for Cav-1 and EGFR expression using immunohistochemistry, and clinical significance was assessed using multivariate Cox regression analysis, Kaplan-Meier estimator curves and the log-rank test. Stromal Cav-1 was downregulated in 38.56% (118/306) of tumor tissues, whereas cytoplasmic EGFR and Cav-1 were overexpressed in 53.92% (165/306) and 44.12% (135/306) of breast cancer tissues, respectively. EGFR expression was positively associated with cytoplasmic Cav-1 and not associated with stromal Cav-1 expression in breast cancer samples; however, low expression of stromal Cav-1 was negatively associated with cytoplasmic Cav-1 expression in total tumor tissues, and analogous results were identified in the chemotherapy group. Multivariate Cox's proportional hazards model analysis revealed that, for patients in the estrogen receptor (ER)(+) group, the expression of stromal Cav-1 alone was a significant prognostic marker of breast cancer. However, in the chemotherapy, human epidermal growth factor receptor 2 (HER-2)(-), HER-2(+) and ER(-) groups, the use of combined markers was more effective prognostic marker. Stromal Cav-1 has a tumor suppressor function, and the combined marker stromal Cav-1/EGFR expression was identified as an improved prognostic marker in the diagnosis of breast cancer. Parenchymal expression of Cav-1 is able to promote EGFR signaling in breast cancer, potentially being required for EGFR-mediated initiation of mitosis.

The Fur-Iron Complex Modulates Expression of the Quorum-Sensing Master Regulator, SmcR, To Control Expression of Virulence Factors in Vibrio vulnificus

PubMed Central

Kim, In Hwang; Wen, Yancheng; Son, Jee-Soo; Lee, Kyu-Ho

2013-01-01

The gene vvpE, encoding the virulence factor elastase, is a member of the quorum-sensing regulon in Vibrio vulnificus and displays enhanced expression at high cell density. We observed that this gene was repressed under iron-rich conditions and that the repression was due to a Fur (ferric uptake regulator)-dependent repression of smcR, a gene encoding a quorum-sensing master regulator with similarity to luxR in Vibrio harveyi. A gel mobility shift assay and a footprinting experiment demonstrated that the Fur-iron complex binds directly to two regions upstream of smcR (−82 to −36 and −2 to +27, with respect to the transcription start site) with differing affinities. However, binding of the Fur-iron complex is reversible enough to allow expression of smcR to be induced by quorum sensing at high cell density under iron-rich conditions. Under iron-limiting conditions, Fur fails to bind either region and the expression of smcR is regulated solely by quorum sensing. These results suggest that two biologically important environmental signals, iron and quorum sensing, converge to direct the expression of smcR, which then coordinates the expression of virulence factors. PMID:23716618

Characterization of the stability and bio-functionality of tethered proteins on bioengineered scaffolds: implications for stem cell biology and tissue repair.

PubMed

Wang, Ting-Yi; Bruggeman, Kiara A F; Sheean, Rebecca K; Turner, Bradley J; Nisbet, David R; Parish, Clare L

2014-05-23

Various engineering applications have been utilized to deliver molecules and compounds in both innate and biological settings. In the context of biological applications, the timely delivery of molecules can be critical for cellular and organ function. As such, previous studies have demonstrated the superiority of long-term protein delivery, by way of protein tethering onto bioengineered scaffolds, compared with conventional delivery of soluble protein in vitro and in vivo. Despite such benefits little knowledge exists regarding the stability, release kinetics, longevity, activation of intracellular pathway, and functionality of these proteins over time. By way of example, here we examined the stability, degradation and functionality of a protein, glial-derived neurotrophic factor (GDNF), which is known to influence neuronal survival, differentiation, and neurite morphogenesis. Enzyme-linked immunosorbent assays (ELISA) revealed that GDNF, covalently tethered onto polycaprolactone (PCL) electrospun nanofibrous scaffolds, remained present on the scaffold surface for 120 days, with no evidence of protein leaching or degradation. The tethered GDNF protein remained functional and capable of activating downstream signaling cascades, as revealed by its capacity to phosphorylate intracellular Erk in a neural cell line. Furthermore, immobilization of GDNF protein promoted cell survival and differentiation in culture at both 3 and 7 days, further validating prolonged functionality of the protein, well beyond the minutes to hours timeframe observed for soluble proteins under the same culture conditions. This study provides important evidence of the stability and functionality kinetics of tethered molecules. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

Parental Expression of Disappointment: Should It Be a Factor in Hoffman's Model of Parental Discipline?

ERIC Educational Resources Information Center

Patrick, Renee B.; Gibbs, John C.

2007-01-01

The authors addressed whether parental expression of disappointment should be included as a distinct factor in M. L. Hoffman's (2000) well-established typology of parenting styles (induction, love withdrawal, power assertion). Hoffman's 3-factor model, along with a more inclusive 4-factor model (induction, love withdrawal, power assertion, and…

Genome-wide identification and expression analysis of TCP transcription factors in Gossypium raimondii.

PubMed

Ma, Jun; Wang, Qinglian; Sun, Runrun; Xie, Fuliang; Jones, Don C; Zhang, Baohong

2014-10-16

Plant-specific TEOSINTE-BRANCHED1/CYCLOIDEA/PCF (TCP) transcription factors play versatile functions in multiple aspects of plant growth and development. However, no systematical study has been performed in cotton. In this study, we performed for the first time the genome-wide identification and expression analysis of the TCP transcription factor family in Gossypium raimondii. A total of 38 non-redundant cotton TCP encoding genes were identified. The TCP transcription factors were divided into eleven subgroups based on phylogenetic analysis. Most TCP genes within the same subfamily demonstrated similar exon and intron organization and the motif structures were highly conserved among the subfamilies. Additionally, the chromosomal distribution pattern revealed that TCP genes were unevenly distributed across 11 out of the 13 chromosomes; segmental duplication is a predominant duplication event for TCP genes and the major contributor to the expansion of TCP gene family in G. raimondii. Moreover, the expression profiles of TCP genes shed light on their functional divergence.

Genome-wide identification and expression analysis of TCP transcription factors in Gossypium raimondii

PubMed Central

Ma, Jun; Wang, Qinglian; Sun, Runrun; Xie, Fuliang; Jones, Don C.; Zhang, Baohong

2014-01-01

Plant-specific TEOSINTE-BRANCHED1/CYCLOIDEA/PCF (TCP) transcription factors play versatile functions in multiple aspects of plant growth and development. However, no systematical study has been performed in cotton. In this study, we performed for the first time the genome-wide identification and expression analysis of the TCP transcription factor family in Gossypium raimondii. A total of 38 non-redundant cotton TCP encoding genes were identified. The TCP transcription factors were divided into eleven subgroups based on phylogenetic analysis. Most TCP genes within the same subfamily demonstrated similar exon and intron organization and the motif structures were highly conserved among the subfamilies. Additionally, the chromosomal distribution pattern revealed that TCP genes were unevenly distributed across 11 out of the 13 chromosomes; segmental duplication is a predominant duplication event for TCP genes and the major contributor to the expansion of TCP gene family in G. raimondii. Moreover, the expression profiles of TCP genes shed light on their functional divergence. PMID:25322260

The Role of Multiple Transcription Factors In Archaeal Gene Expression

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

Charles J. Daniels

2008-09-23