p75NTR Protein Expression Research Articles

Redundancy of p75NTR neurotrophin receptor function in development, growth and fertility in the rat

The p75NTR neurotrophin receptor has positive and negative roles regulating cell survival in the nervous system. Unambiguous interpretation of p75NTR function in vivo has been complicated, however, by residual expression of alternate forms of p75NTR protein in initial p75NTR knock-out mouse models. As rats are the preferred rodent for studying brain and behaviour, and to simplify interpretation of the knock-out phenotype, we report here the generation of a mutant rat devoid of the p75NTR protein. TALEN-mediated recombination in embryonic stem cells (ESCs) was used to flank exon 2 of p75NTR with Lox P sites and produce transgenic rats carrying either un-recombined floxed p75NTREx2-fl, or recombined, exon-2 deleted p75NTREx2-Δ alleles. Crossing p75NTREx2-fl rats with a Cre-deleter strain efficiently removed exon 2 in vivo. Excision of exon 2 causes a frameshift after p75NTR Gly23 and eliminated p75NTR protein expression. Rats lacking p75NTR were healthy, fertile, and histological analysis did not reveal significant changes in cellular density or overall structure in their brains. p75NTR function is therefore largely dispensable for normal development, growth and basal homeostasis in the rat. However, the availability of constitutive and conditional p75NTREx2-Δ rats provides new opportunities to investigate specific roles of p75NTR upon injury and during tissue repair.

Downregulation of OIP5-AS1 affects proNGF-induced pancreatic cancer metastasis by inhibiting p75NTR levels.

We aimed to explore the mechanism by which long non-coding RNA (lncRNA) OIP5-AS1 affects proNGF (precursor nerve growth factor)-induced pancreatic cancer metastasis by targeting the miR-186-5p/NGFR axis. Bioinformatics was used to analyse whether OIP5-AS1 targets miR-186-5p/NGFR and their expression characteristics in pancreatic cancer. OIP5-AS1 and NGFR were overexpressed in pancreatic cancer, and their levels showed a significant positive correlation. Clinical trials also demonstrated that high expression of OIP5-AS1 and NGFR and low expression of miR-186-5p played a pro-cancer role in pancreatic cancer. MiR-186-5p inhibited the migration and invasion of colon cancer cells by targeting NGFR-regulated p75NTR. OIP5-AS1 regulated the action of miR-186-5p on NGFR mRNA and p75NTR by targeting miR-186-5p. Downregulation of NGFR inhibited the expression of p75NTR protein and blocked the role of proNGF in promoting the migration and invasion of pancreatic cancer cells. Animal experiments also showed that the knockdown of miR-186-5p promoted cancer via the expression of NGFR mRNA and p75NTR protein, while the downregulation of proNGF blocked the effects. OIP5-AS1, as a ceRNA, promotes the progression of pancreatic cancer by targeting miR-186-5p/NGFR and affecting the prognosis of patients, which may be related to the action of proNGF.

P0028THE ACTIVATION OF P75NTR RECEPTOR SIGNALING PROMOTES EMT IN ADPKD CELL LINE WT9-12

Abstract Background and Aims ADPKD cystic epithelial cells have been shown to display an undifferentiated phenotype and to undergo epithelial to mesenchymal transition (EMT) changes as part of their phenotype. Thus, in ADPKD kidneys there is upregulation of EMT-related genes and increased fibrosis that support the proliferation of the cyst-lining cells and the progression of ADPKD. In our previous study, we provided that TGFβ1-induced p75NTR receptor activation promotes EMT in renal tubular cells, HK2. p75NTR is the common receptor for all the neurotrophins and their precursors. Since its identification and cloning in the 1980s, p75NTR has been reported to be involved in different and often contradictory functions such as survival, apoptosis, differentiation, proliferation and migration, depending on the cell type and cellular context. Furthermore, p75NTR is known to be overexpressed in some human cancer, where it participates in the stimulation of tumor growth and dissemination. p75NTR, through a neurotrophin-dependent mechanism, dramatically enhances migration and invasion of melanoma and glioma cells. Therefore, starting from these pieces of evidence and since WT9-12 cells exhibit higher constitutive expression of p75NTR and TGFβ1 than that of HK-2, the aim of this study is to carry out in vitro studies to investigate whether p75NTR is involved in EMT of cyst-lining cells and in renal fibrosis in ADPKD. Method Immortalized tubular renal cells homozygous for a truncating PKD1 mutation (WT9-12); Western Blot (WB); Real-Time PCR; Scratch and Trans-well migration assays; Immunofluorescence (IF); RNA interference (RNAi); Transfection assay. Results WB and IF assays showed that in WT9-12 cells TGFβ1 (5ng/ml) increased protein expression of p75NTR and of mesenchymal markers Fibronectin, N-Cadherin, α-SMA, Vimentin while reduced the expression of the epithelial marker E-Cadherin. This biological effect is significantly reversed in cells transfected with RNAi-p75NTR (Figure 1). Functional studies revealed that increased p75NTR protein expression was given to greater p75NTR mRNA stability. Time course assays, performed in cells transfected with RNAi-p75NTR and TGFβ1-treated, demonstrated downregulation of pERK1/2 and pJNK, p75NTR downstream pathways and involved in the non-canonical TGFβ1 pathway promoting pro-fibrotic genes transcription Figure 2A). Results of the analysis of SMAD pathway showed that SMAD3 was not significantly activated in cells untreated with TGFβ1. Moreover, selective ERK1/2 inhibition with PD98059 (10mM) reduced significantly the levels of mesenchymal markers N-Cadherin and α-SMA respect to the controls of untreated cells. Interestingly, in TGFβ1-treated cells in the presence of PD98059 the reduction of mesenchymal markers expression was minor respect to TGFβ1-untreated cells. These findings emphasize the importance of the ERK1/2 pathway in ADPKD. Our results revealed that the ERK1/2 pathway also plays a pivotal role in non-SMAD pathways downstream of TGFβ1 in fibrotic processes in ADPKD. Finally, Scratch and Trans-well migration assays showed a reduced cell motility in p75NTR-knocked-down cells (Figure 2B). Conclusion Although more basic studies are clearly needed to understand the mechanism of action of p75NTR, our results emphasized the crucial role of p75NTR in EMT changes in ADPKD cells which may contribute to the cystic phenotype and to fibrotic process.

The role of p75 neurotrophin receptor in hippocampal neurogenesis impairment after cranial irradiation

Objective To investigate the role of p75 neurotrophin receptor (p75NTR) in the irradiation-induced hippocampal neurogenesis impairment. Methods Thirty Sprague-Dawley rats were subject to whole brain irradiation with a single dose of 10 Gy 4 MeV electron beam. At 1 month after irradiation, the hippocampal tissues of the rats were collected. Western blot was used to detect the changes in the expression level of p75NTR protein. Immunofluorescence confocal laser microscopy was performed to observe the variations in the hippocampal neurogenesis. The stereotatic method was adopted for intra-hippocampal injection of AAV-shp75NTR to specifically knock out p75NTR.The relationship between p75NTR and hippocampal neurogenesis was analyzed. Results Western blot demonstrated that the expression of p75NTR protein was significantly up-regulated by 43.8% after irradiation (P<0.05). Immunofluorescent staining showed that the quantity of BrdU+ NeuN+ cells in rats was significantly decreased by 81.5% at 1 month after irradiation compared with that in the control group (P<0.01). After the specific knockout of p75NTR, hippocampal neurogenesis was obviously protected. Conclusion p75NTR plays a pivotal role in the irradiation-induced hippocampal neurogenesis impairment. Key words: Neurotrophin receptor; Neurogenesis; Hippocampus; Irradiation; Rat

The effects of β-hydroxybutyrate on p75NTR mRNA and protein expression in Aβ-treated SH-SY5Y cells and mechanisms involved

Objective To investigate the effect and mechanisms of β-hydroxybutyrate (βOHB) regulation on p75NTR expression in Alzheimer's disease (AD) model cells. Methods First, cultured SH-SY5Y cells were exposed to Aβ (final concentrations: 10, 20, 40, 80 μmol/L) with or without 5 mmol/L βOHB pretreatment, and sham-treated cells were used as the control. At 24 h after treatment, the viability of cells was determined by the MTT assay. Secondly, cultured cells were divided into four groups. Cells in the Aβ group were exposed to Aβ (final concentration: 20 μmol/L) with or without 5 mmol/L βOHB pretreatment. Cells in the βOHB group were treated only with 5 mmol/L βOHB, and sham-treated cells were used as the control. At 6 h and 24 h after treatment, the expression of p75NTR, HDAC1/2 mRNA and its protein expression, and p65 protein expression were measured by qRT-PCR or Western blot. Finally, the expression of p75NTR mRNA and protein was analyzed in cultured cells after silencing HDAC1 / 2 with siRNA. Results The viability of cells with 40 μmol/L or 80 μmol/L treatment was lower than that in the control group (P 0.05). Conclusions βOHB up-regulates p75NTR expression by inhibiting HDAC1 of βOHB. It also activates p65 and prevents the decrease of cell viability. Key words: Alzheimer disease; Amyloid beta-protein; Nerve growth factor receptor p75; beta-hydroxybutyrate

The Role of Wnt/β-Catenin Signaling Pathway in the Transdifferentiation from Periodontal Ligament Stem Cells to Schwann Cells.

This study was aimed to investigate the role of Wnt/β-catenin signaling pathway in the differentiation from periodontal ligament stem cells (PDLSCs) to Schwann cells (SCs) and the possible mechanisms. FzB was applied to inhibit the Wnt/β-catenin signaling pathway of differentiated PDLSCs (dPDLSCs), and then immunofluorescence, polymerase chain reaction (PCR), and Western blotting analysis were performed to detect SC marker genes and proteins such as S100, glial fibrillary acidic protein (GFAP), and P75NTR. Results showed that when the Wnt/β-catenin signaling pathway was inhibited, the expression of S100, GFAP, and P75NTR protein significantly decreased in dPDLSCs, p < 0.05, whereas PCR results showed that expression of SC myelinogenesis-related genes krox-20, Oct-6, P0, and PMP-22 was significantly downregulated at the same time, p < 0.05. These results showed that Wnt/β-catenin signaling pathway participated in the differentiation from PDLSCs to SCs, and inhibiting it could inhibit the differentiation process.

Impairment of toll-like receptors 2 and 4 leads to compensatory mechanisms after sciatic nerve axotomy.

BackgroundPeripheral nerve injury results in retrograde cell body-related changes in the spinal motoneurons that will contribute to the regenerative response of their axons. Successful functional recovery also depends on molecular events mediated by innate immune response during Wallerian degeneration in the nerve microenvironment. A previous study in our lab demonstrated that TLR 2 and 4 develop opposite effects on synaptic stability in the spinal cord after peripheral nerve injury. Therefore, we suggested that the better preservation of spinal cord microenvironment would positively influence distal axonal regrowth. In this context, the present work aimed to investigate the influence of TLR2 and TLR4 on regeneration and functional recovery after peripheral nerve injury.MethodsEighty-eight mice were anesthetized and subjected to unilateral sciatic nerve crush (C3H/HeJ, n = 22, C3H/HePas, n = 22; C57Bl6/J, n = 22 and TLR2−/−, n = 22). After the appropriate survival times (3, 7, 14 days, and 5 weeks), all mice were killed and the sciatic nerves and tibialis cranialis muscles were processed for immunohistochemistry and transmission electron microscopy (TEM). Gait analysis, after sciatic nerve crushing, was performed in another set of mice (minimum of n = 8 per group), by using the walking track test (CatWalk system).ResultsTLR4 mutant mice presented greater functional recovery as well as an enhanced p75NTR and neurofilament protein expression as compared to the wild-type strain. Moreover, the better functional recovery in mutant mice was correlated to a greater number of nerve terminal sprouts. Knockout mice for TLR2 exhibited 30 % greater number of degenerated axons in the distal stump of the sciatic nerve and a decreased p75NTR and neurofilament protein expression compared to the wild type. However, the absence of TLR2 receptor did not influence the overall functional recovery. End-point equivalent functional recovery in transgenic mice may be a result of enhanced axonal diameter found at 2 weeks after lesion.ConclusionsAltogether, the present results indicate that the lack of TLR2 or the absence of functional TLR4 does affect the nerve regeneration process; however, such changes are minimized through different compensatory mechanisms, resulting in similar motor function recovery, as compared to wild-type mice. These findings contribute to the concept that innate immune-related molecules influence peripheral nerve regeneration by concurrently participating in processes taking place both at the CNS and PNS.Electronic supplementary materialThe online version of this article (doi:10.1186/s12974-016-0579-6) contains supplementary material, which is available to authorized users.

Cerebellar Expression of the Neurotrophin Receptor p75 in Naked-Ataxia Mutant Mouse

Spontaneous mutation in the lysosomal acid phosphatase 2 (Acp2) mouse (nax—naked-ataxia mutant mouse) correlates with severe cerebellar defects including ataxia, reduced size and abnormal lobulation as well as Purkinje cell (Pc) degeneration. Loss of Pcs in the nax cerebellum is compartmentalized and harmonized to the classic pattern of gene expression of the cerebellum in the wild type mouse. Usually, degeneration starts in the anterior and posterior zones and continues to the central and nodular zones of cerebellum. Studies have suggested that the p75 neurotrophin receptor (NTR) plays a role in Pc degeneration; thus, in this study, we investigated the p75NTR pattern and protein expression in the cerebellum of the nax mutant mouse. Despite massive Pc degeneration that was observed in the nax mouse cerebellum, p75NTR pattern expression was similar to the HSP25 pattern in nax mice and comparable with wild type sibling cerebellum. In addition, immunoblot analysis of p75NTR protein expression did not show any significant difference between nax and wild type sibling (p > 0.5). In comparison with wild type counterparts, p75NTR pattern expression is aligned with the fundamental cytoarchitecture organization of the cerebellum and is unchanged in the nax mouse cerebellum despite the severe neurodevelopmental disorder accompanied with Pc degeneration.

Molecular changes in the medial prefrontal cortex and nucleus accumbens are associated with blocking the behavioral sensitization to cocaine

Previous studies have demonstrated that cocaine-induced behavioral sensitization is associated with persistent functional and structural alterations in the medial prefrontal cortex (mPFC) and nucleus accumbens (NAc); however, the molecular mechanisms underlying these changes have not been elucidated. In this study, the behavioral sensitization to cocaine was established in Sprague Dawley rats and was measured by locomotion and behavioral rating. The brain tissue homogenization was used for measuring the level of brain-derived neurotrophic factor (BDNF), the expression and activity of integrin-linked kinase (ILK), level of protein kinase B (Akt) phosphorylation at serine 473 and threonine 308, and the expression of p75NTR, TrkA, and TrkB protein. The Results showed that cocaine sensitization was associated with increased BDNF, ILK activity, phospho-Akt Ser473, p75NTR, and TrkB protein levels in the mPFC and NAc core. The combination of pergolide and ondansetron normalized not only behavioral sensitization, but also the increases in these molecular markers. Dual immunofluoresence staining showed that ILK expression is co-distributed with p75NTR and TrkA expression in both the mPFC and NAc core. Results suggested that the BDNF-TrkA/p75NTR-ILK-Akt signaling pathway may be active in cocaine sensitization and associated neural plasticity in the mPFC and NAc core.

Induction of a reactive state in perineuronal satellite glial cells akin to that produced by nerve injury is linked to the level of p75NTR expression in adult sensory neurons

Satellite glial cells (SGCs) surrounding primary sensory neurons are similar to astrocytes of the central nervous system in that they buffer the extracellular environment via potassium and calcium channels and express the intermediate filament glial fibrillary acidic protein (GFAP). Peripheral nerve injury induces a reactive state in SGCs that includes SGC proliferation, increased SGC/SGC coupling via gap junctions, decreased inward rectifying potassium channel 4.1 (Kir 4.1) expression and increased expression of GFAP and the common neurotrophin receptor, p75NTR. In contrast, neuronal p75NTR expression, normally detected in ∼80% of adult rat sensory neurons, decreases in response to peripheral axotomy. Given the differential regulation of p75NTR expression in neurons versus SGCs with injury, we hypothesized that reduced signaling via neuronal p75NTR contributes to the induction of a reactive state in SGCs. We found that reducing neuronal p75NTR protein expression in uninjured sensory neurons by intrathecal subarachnoid infusion of p75NTR-selective anti-sense oligodeoxynucleotides for one week was sufficient to induce a "reactive-like" state in the perineuronal SGCs akin to that normally observed following peripheral nerve injury. This reactive state included significantly increased SGC p75NTR, GFAP and gap junction protein connexin-43 protein expression, increased numbers of SGCs surrounding individual sensory neurons and decreased SGC Kir 4.1 channel expression. Collectively, this supports the tenet that reductions in target-derived trophic support leading to, or as a consequence of, reduced neuronal p75NTR expression plays a critical role in switching the SGC to a reactive state.

Relationship between Cellular Apoptosis and the Expression of p75 Neurotrophin Receptor and Tyrosine Kinase A Receptor in Tissue Surrounding Haematoma in Intracerebral Haemorrhage

Cellular apoptosis and the expression of p75 neurotrophin receptor (p75(NTR)) and tyrosine kinase A receptor (TrkA) were investigated in the tissues surrounding haematoma in patients with intracerebral haemorrhage. Specimens of tissue from near the haematoma (haemorrhagic samples) and tissue from a distant site (control samples) were collected from 14 patients with basal ganglia haemorrhage undergoing surgical intervention. Cellular apoptosis was determined by terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labelling (TUNEL), and p75(NTR) and TrkA location, protein and gene expression were studied using immunohistochemistry, Western blot and real-time polymerase chain reaction, respectively. The percentage of apoptotic cells and expression of p75(NTR), but not of TrkA, were significantly higher in the haemorrhagic samples than in the control samples. There was a positive correlation between the percentage of TUNEL-positive cells and the percentage of p75(NTR)-positive cells. These results suggest that the p75(NTR)-dependent signal transduction pathway plays an important role in apoptosis after intracerebral haemorrhage.

Neurotrophin/receptor expression in urinary bladder of mice with overexpression of NGF in urothelium

Urothelium-specific overexpression of nerve growth factor (NGF) in the urinary bladder of transgenic mice stimulates neuronal sprouting in the urinary bladder, produces increased voiding frequency, and results in increased referred somatic hypersensitivity. Additional NGF-mediated pleiotropic changes might contribute to the increased voiding frequency and pelvic hypersensitivity observed in these transgenic mice, such as modulation of other growth factor/receptor systems. Chronic overexpression of NGF in the urothelium was achieved through the use of a highly urothelium-specific uroplakin II promoter. In the present study, we examined NGF, brain-derived neurotrophic factor (BDNF), and associated receptor [p75(NTR), tyrosine kinase (Trk)A, TrkB] transcript and protein expression in urothelium and detrusor smooth muscle of NGF-overexpressing (OE) and littermate wild-type mice, using real-time quantitative reverse transcription-polymerase chain reaction, ELISAs, and semiquantitation of immunohistochemistry. We focused on these growth factor/receptors given the established roles of NGF/TrkA, NGF/p75(NTR), and BDNF/TrkB systems in bladder function. Increased voiding frequency in NGF-OE mice was confirmed by examining urination patterns. BDNF, TrkA, and TrkB protein expression was significantly (P ≤ 0.01) reduced and p75(NTR) protein expression was significantly (P ≤ 0.01) increased in urinary bladder of NGF-OE mice. The NGF-OE-induced changes in neurotrophic factor/receptor expression in urinary bladder may represent compensatory changes to reduce voiding frequency in the NGF-OE mouse.

PTEN downregulates p75NTR expression by decreasing DNA-binding activity of Sp1

p75NTR is expressed throughout the nervous system and its dysregulation is associated with pathological conditions. We have recently demonstrated a signalling cascade initiated by laminin (LN), which upregulates PTEN and downregulates p75NTR. Here we investigate the mechanism by which PTEN modulates p75NTR. Studies using PTEN mutants show that its protein phosphatase activity directly modulates p75NTR protein expression. Nuclear relocalization of PTEN subsequent to LN stimulation suggests transcriptional control of p75NTR expression, which was confirmed following EMSA and ChIP analysis of Sp1 transcription factor binding activity. LN and PTEN independently decrease the DNA-binding ability of PTEN to the p75NTR promoter. Sp1 regulation of p75NTR occurs via dephosphorylation of Sp1, thus reducing p75NTR transcription and protein expression. This mechanism represents a novel regulatory pathway which controls the expression level of a receptor with broad implications not only for the development of the nervous system but also for progression of pathological conditions.

Expression pattern of p75 neurotrophin receptor protein in human scalp skin and hair follicles: Hair cycle-dependent expression

The p75 neurotrophin receptor (p75NTR) is a death factor (apoptosis-promoting protein) that belongs to the tumor necrosis factor receptor superfamily of membrane proteins. In the murine hair follicle (HF) model, p75NTR plays a critical role during HF morphogenesis, functioning as a receptor that negatively controls HF development. p75NTR signaling is involved in the control of keratinocyte apoptosis during catagen. To date, knowledge about the expression pattern of p75NTR protein in human scalp skin and HFs is limited. In this investigation we hypothesized that p75NTR protein is expressed in human scalp skin and its expression in HFs fluctuates with the transitions from anagen --> catagen --> telogen stages. To test this hypothesis, the immunoreactivity of p75NTR protein was examined in human scalp skin by immunofluorescent and immunoalkaline phosphatase methods. A total of 50 normal-appearing human scalp skin biopsy specimens were examined (healthy women age 53-57 years). In each case, 50 HFs were analyzed (35, 10, and 5 follicles in anagen, catagen, and telogen, respectively). We found variations in p75NTR protein expression with HF cycling. p75NTR expression was negligible in early, mid, and mature anagen and weak during late anagen. p75NTR expression was moderate during anagen-catagen transition. It was strong in both catagen and telogen HF. Also, p75NTR protein expression was strong in the stratum corneum (epidermis), dermal fibroblasts, blood vessels, nerve endings, adipocytes, and both sebaceous and sweat glands. Our knowledge about other proteins (prosurvival and pro-apoptotic molecules) interacting with p75 is incomplete. Our investigation reports, for the first time, the expression patterns of p75NTR in human scalp skin and HFs. p75NTR protein expression exhibited significant hair cycle-dependent fluctuation, suggesting a possible role in human HF biology.

Pan‐neurotrophin receptor p75NTR expression is strongly induced in lesional atopic mast cells

Neurotrophins such as nerve growth factor or brain-derived neurotrophic factor influence neuronal proliferation and differentiation via the low-affinity pan-neurotrophin receptor p75NTR that may play a pivotal role in linking the immune with the nervous system. Because the precise regulation of p75NTR gene transcription in mast cells under states of allergic inflammation has not been investigated in detail so far, the present studies assessed the gene regulation and expression of this receptor. Transcriptional expression of p75NTR in human skin was studied in isolated cutaneous cells by means of RT-PCR. In situ lesional mast cell p75NTR expression was analysed by immunohistochemistry. The p75NTR mRNA expression was found in isolated human skin mast cells and keratinocytes. Lower mRNA levels were present in fibroblasts and melanocytes but no transcripts were found in endothelial cells. The p75NTR protein expression was found in situ in lesional and non-lesional mast cells. A significantly increased expression of p75NTR protein was found in atopic dermatitis lesional mast cells when compared with control mast cell expression (P<0.05). The demonstration of an increased level of p75NTR gene transcription in lesional mast cells points to an induction of low-affinity neurotrophin receptor sensitivity of mast cells under states of allergic inflammation. Topically administered neurotrophin receptor-modulating compounds may act as anti-inflammatory mediators in cutaneous allergic inflammation.

Neurotrophin-induced upregulation of p75NTR via a protein kinase C-delta-dependent mechanism

Neurotrophins exert their biological effects via p75NTR and Trk receptors. Functional interplay between these two receptors has been widely explored with respect to p75NTR enhancing the activation and signalling of Trk, but few studies address the bidirectional aspects. We have previously demonstrated that the expression of p75NTR can be differentially modulated by different Trk receptor mutations. Here we investigate the mechanism of Nerve Growth Factor (NGF)-induced upregulation of p75NTR expression. We utilize pharmacological inhibition to investigate the role of various TrkA-associated signalling intermediates in this regulatory cascade. Notably, the inhibition of phospholipase C-γ (PLC-γ) using U73122, prevented the NGF-induced upregulation of p75NTR protein and mRNA. The inhibition of protein kinase C-δ (PKC-δ) activation by rottlerin, a selective PKC-δ inhibitor, and by small interfering RNA (siRNA) directed against PKC-δ also inhibited this NGF-induced upregulation. Finally, we also show that in cerebellar granule neurons, BDNF acting via TrkB increases p75NTR expression in a PKC-δ dependent manner. These results indicate the importance of Trk-dependent PLC-γ and PKC-δ activation for downstream regulation of p75NTR protein expression in response to neurotrophin stimulation, a process that has implications to the survival and growth of the developing nervous system.

Neurotrophin-3 administration alters neurotrophin, neurotrophin receptor and nestin mRNA expression in rat dorsal root ganglia following axotomy

In the months following transection of adult rat peripheral nerve some sensory neurons undergo apoptosis. Two weeks after sciatic nerve transection some neurons in the L4 and L5 dorsal root ganglia begin to show immunoreactivity for nestin, a filament protein expressed by neuronal precursors and immature neurons, which is stimulated by neurotrophin-3 (NT-3) administration. The aim of this study was to examine whether NT-3 administration could be compensating for decreased production of neurotrophins or their receptors after axotomy, and to determine the effect on nestin synthesis. The levels of mRNA in the ipsilateral and contralateral L4 and L5 dorsal root ganglia were analyzed using real-time polymerase chain reaction, 1 day, 1, 2 and 4 weeks after unilateral sciatic nerve transection and NT-3 or vehicle administration via s.c. micro-osmotic pumps. In situ hybridization was used to identify which cells and neurons expressed mRNAs of interest, and the expression of full-length trkC and p75 NTR protein was investigated using immunohistochemistry. Systemic NT-3 treatment increased the expression of brain-derived neurotrophic factor, nestin, trkA, trkB and trkC mRNA in ipsilateral ganglia compared with vehicle-treated animals. Some satellite cells surrounding neurons expressed trkA and trkC mRNA and trkC immunoreactivity. NT-3 administration did not affect neurotrophin mRNA levels in the contralateral ganglia, but decreased the expression of trkA mRNA and increased the expression of trkB mRNA and p75 NTR mRNA and protein. These data suggest that systemically administered NT-3 may counteract the decrease, or even increase, neurotrophin responsiveness in both ipsi- and contralateral ganglia after nerve injury.

The Aryl Propionic AcidR-Flurbiprofen Selectively Induces p75NTR-Dependent Decreased Survival of Prostate Tumor Cells

Epidemiologic studies show that patients chronically consuming nonsteroidal anti-inflammatory drugs (NSAID) for arthritis exhibit a reduced incidence of prostate cancer. In addition, some NSAIDs show anticancer activity in vitro. NSAIDs exert their anti-inflammatory effects by inhibiting cyclooxygenase (COX) activity; however, evidence suggests that COX-independent mechanisms mediate decreased prostate cancer cell survival. Hence, we examined the effect of selected aryl propionic acid NSAIDs and structurally related compounds on the decreased survival of prostate cancer cell lines PC-3, DU-145, and LNCaP by induction of the p75(NTR) protein. p75(NTR) has been shown to function as a tumor suppressor in the prostate by virtue of its intracellular death domain that can initiate apoptosis and inhibit growth. The most efficacious compounds for induction of p75(NTR) and decreased survival, in rank-order, were R-flurbiprofen, ibuprofen, oxaprozin, fenoprofen, naproxen, and ketoprofen. Because R-flurbiprofen and ibuprofen exhibited the greatest efficacy, we examined their dose-dependent specificity of induction for p75(NTR) relative to other members of the death receptor family. Whereas treatment with R-flurbiprofen or ibuprofen resulted in a massive induction of p75(NTR) protein levels, the expression of Fas, p55(TNFR), DR3, DR4, DR5, and DR6 remained largely unchanged. Moreover, transfection of either cell line before R-flurbiprofen or ibuprofen treatment with a dominant negative form of p75(NTR) to antagonize p75(NTR) activity or p75(NTR) small interfering RNA to prevent p75(NTR) protein expression rescued both cell lines from decreased survival. Hence, R-flurbiprofen and ibuprofen selectively induce p75(NTR)-dependent decreased survival of prostate cancer cells independently of COX inhibition.

Effects of lead acetate on expression of brain-derived neurotropic factor and P75NTR in rat brain

To study the effects of lead acetate on the expression of brain-derived neurotropic factor (BDNF) and its receptor P75NTR in rat brain. Lead acetate was given to SD rats by intraperitoneal injection (ip) for 5 days at the dosage of 25, 50 and 100mg/kg body weight respectively. The contents of lead in serum, cerebral cortex and hippocampus were measured by atomic absorption spectrophotochemistry. The levels of BDNF mRNA and protein expression in cerebral cortex and hippocampus were observed by RT-PCR and immunohistochemistry, respectively. The levels of P75NTR protein expression in rat brain were measured by immunohistochemistry. Compared with the control, the contents of lead were significantly increased in serum, cerebral cortex and hippocampus in the treatment groups respectively (P < 0.01, P < 0.05). The BDNF mRNA expression in the cerebral cortex (0.52 +/- 0.05, 0.33 +/- 0.03) and hippocampus (0.77 +/- 0.10, 0.92 +/- 0.08) of 50, 100 mg/kg treated groups was significantly higher than that of the control group (0.52 +/- 0.05, 0.33 +/- 0.03), respectively (P < 0.05). The results of immunohistochemistry showed that the area density of BDNF protein in cerebral cortex of every treatment group (0.040 +/- 0.027, 0.048 +/- 0.027, 0.086 +/- 0.040) was significantly increased whereas the average gray value (187.11 +/- 11.15, 180.53 +/- 5.82, 180.15 +/- 8.01) was significantly lower than that of the control (0.026 +/- 0.005, 204.98 +/- 3.45) (P < 0.05, P < 0.01). The area density of BDNF protein in hippocampus of every treatment group was 0.040 +/- 0.027, 0.048 +/- 0.027, 0.086 +/- 0.040, respectively, which was significantly increased compared with the control (0.045 +/- 0.019, P < 0.05). The average gray value of BDNF protein in hippocampus (181.03 +/- 5.16, 171.25 +/- 12.65) of 50, 100 mg/kg were significantly lower than that of the control (198.98 +/- 6.40, P < 0.01). There was no positive expression of P75NTR protein in the control and 25 mg/kg body weight groups. The positive expression of P75NTR protein was detected in 50 and 100 mg/kg body weight groups. Lead can increase the BDNF and P75NTR expression in rat brain which might play an important role in the neural damage and repair.

Expression of glial progenitor markers p75NTR and S100 protein in the developing mouse parathyroid gland

Drosophila glial cells missing (Drosophila Gcm) is a transcription factor that is required for the differentiation of glial cells. Gcm2, a mouse homologue of Drosophila Gcm, is a master regulatory gene of parathyroid development and is expressed in the parathyroid rudiment. We have found that the mouse parathyroid exhibits the glial progenitor markers, p75(NTR) and S100 protein, during fetal development. At embryonic day 11.5 (E11.5), a bulge of the parathyroid rudiment is formed in the cranial part of the third pharyngeal pouch. The rudiment exhibits immunoreactivity for p75(NTR) and S100 protein, in addition to secretory protein 1/chromogranin A. While the thymus rudiment, which arises from the caudal part of the third pharyngeal pouch, is moving downwards, the parathyroid is attached to the top of thymus. The parathyroid comes into contact with the thyroid lobe at E13.5 and then separates from the thymus. The parathyroid maintains intense immunoreactivity for p75(NTR) and S100 protein during the migration and development in the thyroid lobe. The co-localization of p75(NTR) and S100 in the developing parathyroid cells has been confirmed by confocal microscopy. Other glial markers, viz. glial fibrillary acidic protein, Sox10, vimentin and nestin, are not expressed in the parathyroid at any stage of development. The neural progenitor markers, neurofilament 160 and TuJ1, are also absent from the parathyroid. Taken together, we suggest that Gcm2 supplies only some glial progenitor characteristics to the parathyroid rudiment.