Nerve Growth Factor Receptor P75 Research Articles

Peripheral glial cell differentiation from neurospheres derived from adipose mesenchymal stem cells

Mesenchymal stem cells derived from bone marrow and adipose tissue are being considered for use in neural repair because they can differentiate after appropriate induction in culture into neurons and glia. The question we asked was if neurospheres could be harvested from adipose-derived stem cells and if they then could differentiate in culture to peripheral glial-like cells. Here, we demonstrate that adipose-derived mesenchymal stem cells can form nestin-positive non-adherent neurosphere cellular aggregates when cultured with basic fibroblast growth factor and epidermal growth factor. Dissociation of these neurospheres and removal of mitogens results in expression of the characteristic Schwann cell markers S100 and p75 nerve growth factor receptor and GFAP. The simultaneous expression of these glia markers are characteristic features of Schwann cells and olfactory ensheathing cells which have unique properties regarding remyelination and enhancement of axonal regeneration. When co-cultured with dorsal root ganglion neurons, the peripheral glial-like cells derived from adipose mesenchymal stem cells aligned with neuritis and stimulated neuritic outgrowth. These results indicate that neurospheres can be generated from adipose-derived mesenchymal stem cells, and upon mitogen withdrawal can differentiate into peripheral glial cells with neurotrophic effects.

Nerve growth factor-mediated paracrine regulation of hepatic stellate cells by multipotent mesenchymal stromal cells

Aims Multipotent mesenchymal stromal cells (MSC) have been reported to prevent the development of liver fibrosis and have emerged as a promising strategy for cell-based therapy. However, the underlying therapeutic mechanism remains unclear. Hepatic stellate cells (SC) activation is a pivotal event in the development of liver fibrosis. Main methods We hypothesized that MSC play an important role in regulating SC proliferation and apoptosis through paracrine mechanisms. To investigate the paracrine interactions between MSC and SC, a co-culture experimental model was developed using human MSC (hMSC) and human SC (hSC). Key findings We demonstrate that hMSC and hSC both express nerve growth factor (NGF) receptor p75. Results acquired from transwell co-culture experiments using hSC and hMSC showed that hMSC secrete NGF, which enhances hSC apoptosis. Transcription factor nuclear factor kappa B (NF-ΚB) and B cell leukemia-xl (Bcl-xl) take part in the process. Significance These findings demonstrated that hMSC indirectly modulate activated hSC in vitro via NGF-mediated signaling cascades and provide a potential mechanism of how transplanted MSC are effective in treating liver fibrosis.

The intracellular domain of the low affinity p75 nerve growth factor receptor is a death effector domain

The death domain superfamily, comprising the death domain, death effector domain, caspase recruitment domain and pyrin domain subfamilies, is one of the largest classes of protein interaction modules, and plays a particularly critical function in the assembly and activation of apoptotic and inflammatory complexes. Members of the death domain superfamily share a common structural feature, the 6-helical bundle fold. However, individual subfamilies exhibit distinct structural and sequence characteristics. The most distinct feature identified in structural studies is that only the death effector domain contains a charge triad, which is formed by the E/D-RxDL motif. However, using sequence alignment and structural comparison, in the present study we found that the p75-NGFR death domain also contains a charge triad. We therefore suggest that the p75-NGFR death domain should be classified as belonging to the death effector domain.

Proteasome inhibition suppresses Schwann cell dedifferentiation in vitro and in vivo

The ubiquitin-proteasome system (UPS), lysosomes, and autophagy are essential protein degradation systems for the regulation of a variety of cellular physiological events including the cellular response to injury. It has recently been reported that the UPS and autophagy mediate the axonal degeneration caused by traumatic insults and the retrieval of nerve growth factors. In the peripheral nerves, axonal degeneration after injury is accompanied by myelin degradation, which is tightly related to the reactive changes of Schwann cells called dedifferentiation. In this study, we examined the role of the UPS, lysosomal proteases, and autophagy in the early phase of Wallerian degeneration of injured peripheral nerves. We found that nerve injury induced an increase in the ubiquitin conjugation and lysosomal-associated membrane protein-1 expression within 1 day without any biochemical evidence for autophagy activation. Using an ex vivo explant culture of the sciatic nerve, we observed that inhibiting proteasomes or lysosomal serine proteases prevented myelin degradation, whereas this was not observed when inhibiting autophagy. Interestingly, proteasome inhibition, but not leupeptin, prevented Schwann cells from inducing dedifferentiation markers such as p75 nerve growth factor receptor and glial fibrillary acidic protein in vitro and in vivo. In addition, proteasome inhibitors induced cell cycle arrest and cellular process formation in cultured Schwann cells. Taken together, these findings indicate that the UPS plays a role in the phenotype changes of Schwann cells in response to nerve injury.

IFRD1 Is a Candidate Gene for SMNA on Chromosome 7q22-q23

We have established strong linkage evidence that supports mapping autosomal-dominant sensory/motor neuropathy with ataxia (SMNA) to chromosome 7q22-q32. SMNA is a rare neurological disorder whose phenotype encompasses both the central and the peripheral nervous system. In order to identify a gene responsible for SMNA, we have undertaken a comprehensive genomic evaluation of the region of linkage, including evaluation for repeat expansion and small deletions or duplications, capillary sequencing of candidate genes, and massively parallel sequencing of all coding exons. We excluded repeat expansion and small deletions or duplications as causative, and through microarray-based hybrid capture and massively parallel short-read sequencing, we identified a nonsynonymous variant in the human interferon-related developmental regulator gene 1 (IFRD1) as a disease-causing candidate. Sequence conservation, animal models, and protein structure evaluation support the involvement of IFRD1 in SMNA. Mutation analysis of IFRD1 in additional patients with similar phenotypes is needed for demonstration of causality and further evaluation of its importance in neurological diseases.

Altered cutaneous nerve regeneration in a simian immunodeficiency virus / macaque intracutaneous axotomy model

To characterize the regenerative pattern of cutaneous nerves in simian immunodeficiency virus (SIV)-infected and uninfected macaques, excisional axotomies were performed in nonglabrous skin at 14-day intervals. Samples were examined after immunostaining for the pan-axonal marker PGP 9.5 and the Schwann cell marker p75 nerve growth factor receptor. Collateral sprouting of axons from adjacent uninjured superficial dermal nerve bundles was the initial response to axotomy. Both horizontal collateral sprouts and dense vertical regeneration of axons from the deeper dermis led to complete, rapid reinnervation of the epidermis at the axotomy site. In contrast to the slower, incomplete reinnervation previously noted in humans after this technique, in both SIV-infected and uninfected macaques epidermal reinnervation was rapid and completed by 56 days postaxotomy. p75 was densely expressed on the Schwann cells of uninjured nerve bundles along the excision line and on epidermal Schwann cell processes. In both SIV-infected and uninfected macaques, Schwann cell process density was highest at the earliest timepoints postaxotomy and then declined at a similar rate. However, SIV-infection delayed epidermal nerve fiber regeneration and remodeling of new sprouts at every timepoint postaxotomy, and SIV-infected animals consistently had lower mean epidermal Schwann cell densities, suggesting that Schwann cell guidance and support of epidermal nerve fiber regeneration may account for altered nerve regeneration. The relatively rapid regeneration time and the completeness of epidermal reinnervation in this macaque model provides a useful platform for assessing the efficacy of neurotrophic or regenerative drugs for sensory neuropathies including those caused by HIV, diabetes mellitus, medications, and toxins.

Effect of acetyl- l-carnitine on ovarian cancer cells' proliferation, nerve growth factor receptor (Trk-A and p75) expression, and the cytotoxic potential of paclitaxel and carboplatin

Objectives The incidence of chemotherapy induced peripheral neuropathy (CIPN) is 15–25% with platinum and taxanes. CIPN can be permanent and often requires dose reduction or change in chemotherapy. Acetyl- l-carnitine (ALCAR), an ester of l-carnitine, is used to treat CIPN in humans and in animal models. The goals of this study are: 1) examine the effects of ALCAR on ovarian cancer cells, 2) determine if ALCAR affects the cytotoxicity of standard chemotherapy on ovarian cancer cells. Methods OVCAR-3 and SKOV-3 ovarian cancer lines were incubated in ALCAR containing media. Viability, proliferation, and expression of the nerve growth factor receptors (NGFR) Trk-A and p-75 were determined by flow cytometry. Cytotoxicity assays examining ALCAR's effect on paclitaxel and carboplatin were done by flow cytometry and infrared plate-reader. Results Flow cytometry showed no change in percent live ( p = 0.87) or proliferation ( p = 0.95) of OVCAR-3 cells when comparing controls with up to 100 μM ALCAR. However, there was a slight but significant decrease in the proliferation of SKOV-3 cells incubated at higher ALCAR concentrations ( p = < 0.01). Flow cytometry showed no difference in the viability of OVCAR-3 cells when comparing ALCAR: +/− paclitaxel ( p = 1), +/− carboplatin ( p = 0.8), or both ( p = 0.4). Proliferation assays indicated that paclitaxel's cytotoxicity on OVCAR-3 and SKOV-3 cells was unchanged at higher ALCAR concentrations ( p = < 0.01–0.4). ALCAR did not affect the expression of NGFR on OVCAR-3 or SKOV-3 cells. Conclusion ALCAR does not affect the cytotoxicity of paclitaxel or carboplatin. There was no increase in proliferation, or NGFR of OVCAR-3 or SKOV-3 cells exposed to ALCAR.

Fibroblastic Reticular Cell Infection by Hemorrhagic Fever Viruses

Viral hemorrhagic fevers (VHFs) often cause high mortality with high infectivity, multiorgan failure, shock and hemorrhagic diathesis. Fibroblastic reticular cells (FRCs) within secondary lymphoid organs provide a supporting scaffold to T-lymphocyte areas. These cells regulate the movement of various immune cells and soluble molecules that promote T-lymphocyte homeostasis. We previously reported Ebola virus infection of FRCs, but ascribed little significance to this finding. Here, we studied infection of FRCs by Ebola, Marburg and Lassa viruses. We demonstrate that FRCs, or the extracellular 'conduit' of the fibroblastic reticulum of nonhuman primates, are targets of Ebola, Marburg and Lassa viruses. Furthermore, we observed that FRC damage correlates temporally and spatially with lymphocyte damage and that FRCs serve as nidi of fibrin deposition. In addition, we show that nonhuman primate FRCs express p75 NGF receptor and tissue transglutaminase. Our data suggest that viral infection of FRCs may be crucial to the immunological dysfunction and coagulopathy characteristic of VHFs. We further propose that p75 NGF receptor and tissue transglutaminase may be involved in FRC-associated dysfunction during the course of infection.

Dermal sheet preparations in the evaluation of dermal innervation in Parkinson’s disease and multiple system atrophy

Evaluation of dermal nerve fibers in conventional vertical sections is difficult because of the small number of fibers available for examination. In this study, we evaluated dermal sheet mounts for fibers in which the majority of fibers can be visualized. We compared the dermal small fiber density in six Parkinson's disease (PD) and six multiple system atrophy (MSA) patients using dermal sheet preparations (DSP). DSP are based on epidermal-dermal separations and immunostaining of the entire dermis by the nerve growth factor receptor p75 antibody that stains both autonomic and sensory fibers. The small fiber density was reduced in PD compared with MSA (p < 0.0001), suggesting the presence of small fiber neuropathy in PD. DSP offer a unique method of evaluation of dermal nerve fibers. This method can be used to evaluate small nerve fibers in many neurological disorders such as MSA and PD.

Localization of the low‐affinity nerve growth factor receptor p75 in human limbal epithelial cells

Biological effects of nerve growth factor (NGF) are mediated through receptors known as nerve growth factor receptors (NGFR), which include p75 and TrkA. This study was initiated after identifying NGFR as an up-regulated gene in the limbus by cDNA microarray analysis and we postulate that its expression may be indicative of a stem/progenitor cell phenotype. Immunohistochemistry was performed on normal human adult (n= 5) and foetal (n= 3) corneal tissue using antibodies directed against p75, TrkA, NGF, p63, ABCG2 and CK3/12. Limbal, conjunctival and pterygium tissue was obtained from patients (n= 10) undergoing pterygium resection and used for immunohistochemical assessment. Paraffin-embedded archival human skin specimens (n= 4) were also evaluated. In vitro expression of NGFR was determined in limbal, conjunctival and pterygium-derived epithelial cells. p75 was selectively expressed by basal epithelial cells in pterygia, conjunctiva and limbus, but was absent in the central cornea. These results were confirmed with two additional p75 specific antibodies. In contrast, TrkA was found in full-thickness pterygium, conjunctival, limbal and corneal epithelium in both adult and foetal eyes. p75 expression was identified in a small percentage, while TrkA was found on the entire population of cultured conjunctival, limbal and pterygium-derived epithelial cells. This receptor was also observed in selective regions of the human epidermis and hair follicle bulge. Our results illustrate the selective expression of p75 in basal pterygium, conjunctival and limbal epithelium, while staining was absent in adult and foetal central cornea. p75 may represent an additional ocular surface epithelial stem/progenitor cell signature gene.

Effect of progestogens and combined oral contraceptives on nerve fibers in peritoneal endometriosis

To investigate how progestogens and combined oral contraceptives change nerve fiber density in peritoneal endometriotic lesions and to identify the types of nerve fibers still present during hormone treatment. Laboratory study using human tissue. University-based laboratory. Hormonally treated and untreated women with endometriosis undergoing laparoscopy, hysteroscopy, and curettage. Biopsy samples from peritoneal endometriotic lesions in hormonally treated and untreated women with endometriosis. Types and density of nerve fibers were immunohistochemically determined in peritoneal endometriotic lesions from hormonally treated and untreated women with endometriosis. The nerve fiber density (mean +/- standard deviation/mm(2)) in peritoneal endometriotic lesions from hormone-treated women with endometriosis (10.6 +/- 2.2/mm(2)) was statistically significantly lower than in peritoneal endometriotic lesions from untreated women with endometriosis (16.3 +/-10.0/mm(2)). Nerve growth factor and nerve growth factor receptor p75 expression in peritoneal endometriotic lesions were slightly reduced in hormone-treated women with endometriosis compared with untreated women with endometriosis. Progestogens and combined oral contraceptives reduced nerve fiber density and nerve growth factor and nerve growth factor receptor p75 expression in peritoneal endometriotic lesions.

Abstract 1838: Neural Crest Stem Cells Supply Intrinsic Cardiac Adrenergic Cells and Contribute to Reinnervation after Myocardial Infarction in Mice

Cardiac nerve sprouting and heterogeneous reinnervation after myocardial infarction (MI) lead to arrhythmogenesis and sudden cardiac death. Despite its importance, little is known about the contribution of the intrinsic cardiac nervous system on reinnervation. We previously reported that neural crest stem cells (NCSC) reside in the heart, and can differentiate into neuron, glia, smooth muscle cells, and cardiomyocytes. The present study scrutinized the developmental origin of intrinsic cardiac adrenergic (ICA) cells, and investigated whether NCSC supply ICA and contribute to reinnervation after MI. (1) We analyzed two distinct double transgenic mice, encoding protein 0-Cre/Floxed-EGFP (P0-EGFP) and dopamine β -hydroxylase-Cre/Floxed-EGFP (DBH-EGFP), to map neural crest-derived cell fate and catecholaminergic cell fate, respectively. ICA cells, demarcated with tyrosine hydroxylase (TH, catecholaminergic marker)-immunopositive cells within ventricles, co-expressed EGFP in both P0-EGFP and DBH-EGFP mice. Most ICA cells were nucleated and round-shaped, although 5% of the cells extended nerve fibers. (2) NCSC were marked with EGFP in P0-EGFP mice, expressed nestin, musashi-1 and p75 nerve growth factor (NGF) receptor, and showed sphere formation. They frequently resided in the outflow tract, but also existed in all parts of the heart. (3) ICA cell number and TH mRNA expression within ventricles were increased in transgenic mice overexpressing NGF in the heart. (4) P0-EGFP mice were subjected to MI by coronary artery occlusion. EGFP-positive cells were markedly increased after MI, and more so in the border zone area than in the non-infarcted area. (5) Similarly, DBH-EGFP subjected to MI revealed an increase in EGFP-positive cells at the border zone area, and some of them extended neurites among the sprouting nerves. (6) TH and EGFP mRNA were increased in DBH-EGFP ventricles, concomitant with cardiac NGF upregulation. These findings demonstrated that ICA cells were derived from neural crest cells, and dependent on NGF synthesized in the heart. NCSC were mobilized to the infarcted myocardium, differentiated into TH-positive nerves, and contributed to reinnervation after MI.

Differential modulation of nerve growth factor receptor (p75) and cholinergic gene expression in purified p75-expressing and non-expressing basal forebrain neurons by BMP9

The synthesis of acetylcholine and its release from basal forebrain cholinergic neurons (BFCN) that innervate the cerebral cortex and hippocampus are considered essential processes for normal learning, memory and attention. We have developed a purification and cell culture method of BFCN in order to examine the regulation of their cholinergic phenotype. Cells isolated from the septal region of late embryonic mice were purified by fluorescence-activated cell sorting based on their expression of the nerve growth factor receptor (p75), a surface marker for mature BFCN. Consistent with previous reports, p75-postive (p75+) cells were enriched in choline acetyltransferase (ChAT) and the high-affinity choline transporter (ChT), as measured by reverse transcriptase PCR. In culture, these cells maintained their gene expression of p75, ChAT and ChT, while p75-negative (p75−) cells had a low expression of these genes. Incubation of the cells with BMP9 not only increased p75 and ChAT gene expression in p75− cells, but also augmented the expression of these genes in p75+ cells. Conversely, BMP9 decreased ChT gene expression in p75+ cells and had no such effect in p75− cells. Immunostaining confirmed that p75 protein expression was modulated by BMP9 in a similar way as p75 mRNA, and also revealed that only a subset of p75− cells respond to BMP9 in this manner. These data suggest that mature BFCN in culture may express their cholinergic phenotype in the absence of exogenous trophic input, but that BMP9 can further modulate this phenotype. Moreover, BMP9 induces the cholinergic phenotype in a set of basal forebrain non-cholinergic neurons or precursor cells.

Gfi1-mediated Stabilization of GATA3 Protein Is Required for Th2 Cell Differentiation

The differentiation of naive CD4 T cells into Th2 cells requires the T cell receptor-mediated activation of the ERK MAPK cascade. Little is known, however, in regard to how the ERK MAPK cascade regulates Th2 cell differentiation. We herein identified Gfi1 (growth factor independent-1) as a downstream target of the ERK MAPK cascade for Th2 cell differentiation. In the absence of Gfi1, interleukin-5 production and the change of histone modification at the interleukin-5 gene locus were severely impaired. Furthermore, the interferon gamma gene showed a striking activation in the Gfi1(-/-) Th2 cells. An enhanced ubiquitin/proteasome-dependent degradation of GATA3 protein was observed in Gfi1(-/-) Th2 cells, and the overexpression of GATA3 eliminated the defect of Th2 cell function in Gfi1-deficient Th2 cells. These data suggest that the T cell receptor-mediated induction of Gfi1 controls Th2 cell differentiation through the regulation of GATA3 protein stability.

Evaluation on in vitro biocompatibility of silk fibroin‐based biomaterials with primarily cultured hippocampal neurons

Silk fibroin-based biomaterials have recently found increasing applications in the tissue-engineering field including the generation of artificial nerve guides for peripheral nerve repair. The aim of this study was to investigate the suitability of silk fibroin as a candidate biomaterial for central nervous system (CNS) therapy. We found that substrates made up of silk fibroin fibers supported the survival and growth of the attached hippocampal neurons by using morphological observation. We also cultured the hippocampal neurons in silk fibroin extract for different times, and observed no significant difference occurring in their morphology, cell viability for these cultured hippocampal neurons as compared to those cultured in plain neuronal culture medium. Moreover, immunocytochemistry, RT-PCR, and Western blot analysis revealed that no significant difference was found in mRNA or protein levels of the growth-associated molecules, such as brain-derived neurotrophic factor, growth-associated protein-43, neurofilament, nerve growth factor, and nerve growth factor-receptor P75, between the hippocampal neurons cultured in the silk fibroin extract and in plain neuronal culture medium. Taken together, all the results demonstrate that silk fibroin has good biocompatibility with primarily cultured hippocampal neurons without any significant cytotoxic effects on their cell phenotype and functions, suggesting a potential possible use of silk fibroin for preparing the tissue-engineered nerve guides or drug delivery vehicles to treat CNS injuries or diseases.

Spiral Ganglion Outgrowth and Hearing Development in p75NTR-Deficient Mice

To explore the role of nerve growth factor receptor p75^NTR during the terminal neuronal development of the mammalian cochlea the onset of hearing and the in vitro response of spiral ganglion neurites to neurotrophin 3 (NT-3), which is known to play a critical role during neonatal inner ear development, were investigated in p75^NTR-deficient mice (p75^NTR–/–). Auditory-evoked brain stem response recordings from p75^NTR–/– and wild-type (WT) littermates were measured from postnatal days (PD) 8 to 23. Additionally, spiral ganglion explants from p75^NTR–/– and WT animals were dissected and cultured in an organotypic tissue culture system. In both groups, spiral ganglion neurite outgrowth was analyzed with and without NT-3 supplementation. No significant differences in the onset of hearing of mutant mice compared to the WT mice were detected, and both groups showed a similar development of hearing until PD 23. After stimulation with NT-3, neurite outgrowth was enhanced in both p75^NTR–/– and WT mice. However, neurites from p75^NTR–/– spiral ganglion explants were longer in both culture conditions. Moreover, NT-3 did not significantly enhance neurite number in p75^NTR–/–, as it did in WT mice. P75^NTR has a remarkable influence on spiral ganglion neurite growth behavior. However, p75^NTR does not seem to be essential for the development of basic hearing function in the first 3 postnatal weeks.

C-Jun N-terminal kinases mediate Fas-induced neurite regeneration in PC12 cells

In response to injury, peripheral neuronal cells initiate complex signalling cascades to promote survival and regeneration. In the present study, we used a model of experimental injury in the rat pheochromocytoma cell line PC12 to investigate receptor signals that lead to neurite outgrowth. Nerve growth factor (NGF) dose-dependently induced sprouting and the expression of the NGF receptors Trk tyrosine kinase receptor (TrkA) and p75 neurotrophin receptor (p75NTR) as well as Fas and Fas ligand. Neurite regeneration was decreased by chemical inhibition of TrkA, but not p75NTR, and by the Fas inhibitor protein Fas-Fc. The mitogen-activated protein kinases (MAPKs) extracellular signal-regulated kinase 1/2 (ERK1/2) and c-Jun N-terminal kinases (JNKs) were activated in response to NGF and both significantly contributed to neurite re-growth. Interestingly, otherwise apoptotic Fas ligation supported neuronal recovery exclusively via JNKs and promoted sprouting parallel to NGF. These findings suggest a novel signal integration from the NGF and Fas pathways in the JNK axis of MAPK signalling, where JNKs function as “physiological” mediators of normally apoptotic signals.

Quantitative Assessment of Neuronal Differentiation in Three-dimensional Collagen Gels Using Enhanced Green Fluorescence Protein Expressing PC12 Pheochromocytoma Cells

There is a paucity of quantitative methods for evaluating the morphological differentiation of neuronal cells in a three-dimensional (3-D) system to assist in quality control of neural tissue engineering constructs for use in reparative medicine. Neuronal cells tend to aggregate in the 3-D scaffolds, hindering the application of two-dimensional (2-D) morphological methods to quantitate neuronal differentiation. To address this problem, we developed a stable transfectant green fluorescence protein (GFP)-PC12 neuronal cell model, in which the differentiation process in 3-D can be monitored with high sensitivity by fluorescence microscopy. Under 2-D conditions, the green cells showed collagen adherence, round morphology, proliferation properties, expression of the nerve growth factor (NGF) receptors TrkA and p75(NTR), stimulation of extracellular signal-regulated kinase phosphorylation by NGF and were able to differentiate in a dose-dependent manner upon NGF treatment, like wild-type (wt)-PC12 cells. When grown within 3-D collagen gels, upon NGF treatment, the GFP-PC12 cells differentiated, expressing long neurite outgrowths. We describe here a new validated method to measure NGF-induced differentiation in 3-D. Having properties similar to those of wt-PC12 and an ability to grow and differentiate in 3-D structures, these highly visualized GFP-expressing PC12 cells may serve as an ideal model for investigating various aspects of differentiation to serve in neural engineering.

Alterations of nerve-growth factor and p75NTR expressions in urinary bladder of fructose-fed obese rats

The study investigated the effects of metabolic syndrome on urinary bladder nerve-growth factor (NGF) and p75(NTR) expression in fructose-fed obese rats. Age-matched male Wistar rats were divided into four groups; group I: normal control rats; group II: 6-week fructose-fed rats (FR); group III: 9-week FR; and group IV: 12-week FR. In vivo cystometry under anesthesia was performed. In vitro bladder NGF levels were measured by enzyme-linked immunosorbent assay (ELISA). Expressions of the mRNAs that encoded NGF and NGF receptor p75(NTR) in control and 9-week FR bladder were studied using the method of reverse transcription combined with polymerase chain reaction (RT-PCR). The three groups of FR showed significant increases in body weight, blood pressure, plasma glucose, insulin and triglyceride levels when compared to control rats. The bladder NGF levels in the 9-week and 12-week FR were significantly lower than the control (66.8+/-5.4, 49.4+/-7.1 and 95.2+/-6.5 pg/microg protein; p<0.05 and p<0.01 respectively, for n=8 in each group). The bladder emptying function was deteriorated in these two groups of FR as shown by the significantly increased residual volume and decreased emptying fraction. The mRNA expressions of bladder NGF and p75(NTR) in the 9-week FR were significantly decreased when compared to the control group (p<0.05 and p<0.001 respectively, n=8 in each group). In conclusion, long-term metabolic syndrome in FR significantly decreases bladder NGF and p75(NTR) expression. These alterations are associated with deterioration in bladder emptying function.

Troy/Taj and its role in CNS axon regeneration

Binding of myelin inhibitors to the NgR1/p75/LINGO-1 signaling complex activates RhoA to mediate the inhibition of axonal outgrowth. The nerve growth factor receptor p75, a TNF family receptor, is absent or poorly expressed in certain types of neurons that respond to myelin inhibitors, thereby prompting speculation that other TNF family receptors are involved in the NgR1 complex. Troy/Taj is an orphan TNF family receptor that is broadly expressed in postnatal and adult neurons. Troy binds to NgR1 and can functionally replace p75 in the p75/NgR1/LINGO-1 complex to activate RhoA and block neurite outgrowth in the presence of myelin inhibitors. Neurons from Troy-deficient mice are more resistant to the suppressive action of the myelin inhibitors. The discovery of TROY function in axon growth is an important step for understanding the complex regulation of axonal regeneration by diverse members of the TNF receptor family.