The p75 neurotrophin receptor influences NT-3 responsiveness of sympathetic neurons in vivo.

To determine the role of the p75 neurotrophin receptor (p75NTR) in sympathetic neuron development, we crossed transgenic mice with mutations in p75NTR, nerve growth factor (NGF) and neurotrophin-3 (NT-3). Neuron number is normal in sympathetic ganglia of adult p75NTR-/- mice. Mice heterozygous for a NGF deletion (NGF+/-) have 50% fewer sympathetic neurons. In the absence of p75NTR (p75NTR-/- NGF+/-), however, neuron number is restored to wild-type levels. When NT-3 levels are reduced (p75NTR-/- NGF+/- NT3 +/-), neuron number decreases compared to p75NTR-/- NGF+/- NT3+/+. Thus, without p75NTR, NT3 substitutes for NGF, suggesting that p75 alters the neurotrophin specificity of TrkA in vivo.     

The p75 neurotrophin receptor and neuronal apoptosis

Although evidence continues to accumulate for the apoptosis-inducing role of the p75 neurotrophin receptor, several outstanding questions remain. One of these concerns the signal transduction pathway of p75, which continues to be elusive. The evidence for the roles of ceramide, c-jun kinase and NF-kappaB is discussed: none of these are able to account satisfactorily for p75 death signalling. Negative modulation of Trk signalling by p75 could account for part of the pro-apoptotic effect, but is unlikely to be a major component. Although recent evidence indicates that the juxtamembrane region is critical for causing cell death, p75 has a well-conserved death domain. This may be important for functions other than killing. In glial cells and some neurons that express p75 but not TrkA, p75 causes cell death in response to nerve growth factor (NGF) binding. In sensory neurons and PC12 cells, p75 appears to signal constitutively. In cholinergic forebrain neurons, p75 expression leads to atrophy and downregulation of cholinergic markers, rather than cell death. The major challenges in p75 research are to define its signalling pathways, and particularly the intracellular proteins with which it interacts. Another major challenge is to develop a model that reconciles the different facets of p75, such as its ability in some situations to assist TrkA to rescue NGF-dependent neurons, but to stimulate apoptosis in others.     

Nerve growth factor survival signaling in cultured hippocampal neurons is mediated through TrkA and requires the common neurotrophin receptor P75

The role of the common neurotrophin receptor p75 (p75NTR) in neuronal survival and cell death remains controversial. On the one hand, p75NTR provides a positive modulatory influence on nerve growth factor (NGF) signaling through the high affinity neurotrophin receptor TrkA, and hence increases NGF survival signaling. However, p75NTR may also signal independently of TrkA, causing cell death or cell survival, depending on the cell type and stage of development. Here we demonstrate that TrkA is expressed in primary cultures of hippocampal neurons and is activated by NGF within 10 min of exposure. In primary hippocampal cultures neuroprotection by NGF against glutamate toxicity was mediated by NF-kappaB and accompanied by an increased expression of neuroprotective NF-kappaB target genes Bcl-2 and Bcl-xl. In mouse hippocampal cells lacking p75NTR (p75NTR-/-) activation of TrkA by NGF was not detectable. Moreover, neuroprotection by NGF against glutamate toxicity was abolished in p75NTR-/- neurons, and the expression of bcl-2 and bcl-xl was markedly reduced as compared to wildtype cells. NGF increased TrkA phosphorylation in hippocampal neurons and provided protection that required phosphoinositol-3-phosphate (PI3)-kinase activity and Akt phosphorylation, whereas the mitogen-activated protein kinases (MAPK), extracellular-regulated kinases (Erk) 1/2, were not involved. P75NTR signaling independent of TrkA, such as increased neutral sphingomyelinase (NSMase) activity causing enhanced levels of ceramide, were not detected after exposure of hippocampal neurons to NGF. Interestingly, inhibition of sphingosine-kinase blocked the neuroprotective effect of NGF, suggesting that sphingosine-1-phosphate was also involved in NGF-mediated survival in our cultured hippocampal neurons. Overall, our results indicate an essential role for p75NTR in supporting NGF-triggered TrkA signaling pathways mediating neuronal survival in hippocampal neurons.     

NGF-mediated sensitization of the excitability of rat sensory neurons is prevented by a blocking antibody to the p75 neurotrophin receptor

Nerve growth factor (NGF) can play a causal role in the initiation of hyperalgesia. Recent work demonstrates that NGF can act directly on nociceptive sensory neurons to augment their sensitivity to a variety of stimuli. Based on the existing literature, it is not clear whether this sensitization is mediated by the high-affinity TrkA receptor or the low-affinity p75 neurotrophin receptor. We examined whether a blocking antibody to the p75 neurotrophin receptor can prevent the NGF-induced enhancement of excitability in capsaicin-sensitive small-diameter sensory neurons that have been isolated from the adult rat. In this report, pretreatment with the p75 blocking antibody completely prevents the NGF-induced increase in the number of action potentials evoked by a ramp of depolarizing current as well as the suppression of a delayed rectifier-type of potassium current(s) in these neurons. Although the sensitization by NGF was blocked, the antibody had no effect on the capacity of ceramide, a putative downstream signaling molecule, to either enhance the excitability or inhibit the potassium current. These results indicate that NGF can increase the excitability of nociceptive sensory neurons through activation of the p75 neurotrophin receptor and its consequent liberation of ceramide from neuronal sphingomyelins.     

Identification of the neurotrophin receptors p75 and trk in a series of Wilms' tumors.

The molecular mechanisms underlying the pathogenesis of Wilms' tumor (WT) are poorly understood, although a variety of growth factors including platelet-derived growth factor and insulin-like growth factor are expressed and are thought to contribute to tumor development. In earlier studies, WT cells in culture were found to express the low affinity nerve growth factor receptor, p75. These WT cells were capable of responding to the neurotrophin (NT) NGF, suggesting that NT may be involved in WT pathogenesis. We have examined a group of WT immunohistochemically with antibodies recognizing known trk receptor proteins, the p75 receptor, and the NTs, NGF and NT-3. Confirmatory immunoprecipitation and Western blots were then performed on representative WT samples from the study group. The p75 receptor was found predominantly in the epithelial and blastemal components where high levels of NT were also identified. The trk A and B receptors were primarily within stromal components, whereas the trk C and C' receptors were present within epithelial structures. Western blot analyses confirmed the presence of the respective receptor proteins with variations correlating in some cases with histological type. The selective presence of NT receptors and growth factors in this series of WT implies autocrine/paracrine mechanisms for tumor development.     

慢病毒介导沉默P75神经营养因子受体联合神经生长因子过表达促进大鼠骨髓间充质干细胞的增殖

文题释义：P75神经营养因子受体(P75 Neurotrophin receptor,P75^NTR)：属于一种Ⅰ型糖蛋白,在非神经系统组织及多种肿瘤细胞中表达。p75^NTR与Trk受体通过不同的信号传递对细胞增殖与凋亡发挥着双重的生物效应。神经生长因子：在神经生长因子信号通路中,p75^NTR可以与神经生长因子相结合,引起神经酰胺的释放而激活JNK通路来调节细胞的增殖和凋亡。  摘要背景：P75神经营养因子受体(P75 Neurotrophin receptor,P75^NTR)是神经生长因子(nerve growth factor,NGF)的受体之一。在各种细胞组织中发挥着促进增殖或凋亡的双重作用,且P75^NTR在骨折不愈合处存在高表达,而过量的NGF可关闭P75^NTR受体,从而挽救受损的细胞。因此研究沉默P75^NTR联合NGF过表达共转染对骨髓间充质干细胞增殖的影响,可为临床治疗骨折不愈合提供新思路。目的：观察慢病毒介导沉默P75^NTR联合NGF过表达共转染对SD大鼠骨髓间充质干细胞增殖的影响。方法：体外培养至第3代SD大鼠骨髓间充质干细胞分为空白对照组、阴性对照组、沉默p75^NTR组、NGF过表达组、沉默p75^NTR联合NGF过表达组。将慢病毒介导沉默P75^NTR、过表达NGF转染至大鼠骨髓间充质干细胞,倒置荧光学显微镜观察慢病毒转染第3天细胞形态变化,流式细胞仪检测转染效率及Western blot方法检测P75^NTR和NGF的表达,最后用MTT法和CCK-8法检测各组细胞增殖活性。结果与结论：①共转染慢病毒后的细胞生长、分布良好;双基因慢病毒载体的转染效率已超过70%;②与空白对照组和阴性对照组比较,沉默P75^NTR联合NGF过表达组P75^NTR蛋白表达明显下调,NGF蛋白表达明显增加;③与空白对照组与阴性对照组相比,沉默P75^NTR联合NGF过表达组、沉默P75^NTR组、NGF过表达组细胞增殖明显加快,差异有显著性意义(P < 0.05),其中沉默P75^NTR联合NGF过表达组增殖最快;④结果显示：沉默P75^NTR联合NGF过表达共转染可促进SD大鼠骨髓间充质干细胞的增殖。ORCID: 0000-0003-3826-7213(王宁) 中国组织工程研究杂志出版内容重点：干细胞;骨髓干细胞;造血干细胞;脂肪干细胞;肿瘤干细胞;胚胎干细胞;脐带脐血干细胞;干细胞诱导;干细胞分化;组织工程     

siRNA沉默p75神经营养因子受体逆转胶质瘤恶性表型

目的 利用小干扰RNA(siRNA)技术沉默与胶质瘤细胞凋亡和侵袭密切相关的p75神经营养因子受体(p75NTR)基因,观察其逆转胶质瘤恶性表型的治疗效果.方法 设计靶向p75NTR基因的siRNA片段,脂质体转染人U251胶质瘤细胞系,逆转录聚合酶链反应(RT-PCR)和免疫细胞化学方法 检测p75NTR的mRNA和蛋白表达;Transwell细胞侵袭实验检测U251细胞的侵袭力;软琼脂集落形成实验检测细胞集落形成能力;建立裸鼠颅内U251胶质瘤荷瘤模型,原位重复注射siRNA-p75NTR/脂质体复合物3次,MRI检测颅内瘤体积,免疫细胞化学SP法检测p75NTR、神经生长因子(NGF)和细胞周期蛋白(cyclin)D2表达,用TUNEL法做移植瘤细胞原位细胞凋亡检测.结果 转染siRNA基因片段的U251细胞p75NTR mRNA和蛋白质表达水平显著下降(P<0.05),细胞侵袭能力及软琼脂集落形成能力均显著降低(P<0.05);体内实验显示p75NTR的表达程度与NGF的表达呈正相关,与cyclin D2表达及原位细胞凋亡数量呈负相关,MRI示瘤体积增长缓慢,边界轮廓清晰,动物生存期明显延长(P<0.05).结论 靶向p75NTR的siRNA技术可有效降低胶质瘤细胞的侵袭及增殖能力,诱导肿瘤细胞凋亡.     

siRNA沉默p75神经营养因子受体逆转胶质瘤恶性表型

目的 利用小干扰RNA(siRNA)技术沉默与胶质瘤细胞凋亡和侵袭密切相关的p75神经营养因子受体(p75NTR)基因,观察其逆转胶质瘤恶性表型的治疗效果.方法 设计靶向p75NTR基因的siRNA片段,脂质体转染人U251胶质瘤细胞系,逆转录聚合酶链反应(RT-PCR)和免疫细胞化学方法 检测p75NTR的mRNA和蛋白表达;Transwell细胞侵袭实验检测U251细胞的侵袭力;软琼脂集落形成实验检测细胞集落形成能力;建立裸鼠颅内U251胶质瘤荷瘤模型,原位重复注射siRNA-p75NTR/脂质体复合物3次,MRI检测颅内瘤体积,免疫细胞化学SP法检测p75NTR、神经生长因子(NGF)和细胞周期蛋白(cyclin)D2表达,用TUNEL法做移植瘤细胞原位细胞凋亡检测.结果 转染siRNA基因片段的U251细胞p75NTR mRNA和蛋白质表达水平显著下降(P<0.05),细胞侵袭能力及软琼脂集落形成能力均显著降低(P<0.05);体内实验显示p75NTR的表达程度与NGF的表达呈正相关,与cyclin D2表达及原位细胞凋亡数量呈负相关,MRI示瘤体积增长缓慢,边界轮廓清晰,动物生存期明显延长(P<0.05).结论 靶向p75NTR的siRNA技术可有效降低胶质瘤细胞的侵袭及增殖能力,诱导肿瘤细胞凋亡.     

siRNA沉默p75神经营养因子受体逆转胶质瘤恶性表型

目的 利用小干扰RNA(siRNA)技术沉默与胶质瘤细胞凋亡和侵袭密切相关的p75神经营养因子受体(p75NTR)基因,观察其逆转胶质瘤恶性表型的治疗效果.方法 设计靶向p75NTR基因的siRNA片段,脂质体转染人U251胶质瘤细胞系,逆转录聚合酶链反应(RT-PCR)和免疫细胞化学方法 检测p75NTR的mRNA和蛋白表达;Transwell细胞侵袭实验检测U251细胞的侵袭力;软琼脂集落形成实验检测细胞集落形成能力;建立裸鼠颅内U251胶质瘤荷瘤模型,原位重复注射siRNA-p75NTR/脂质体复合物3次,MRI检测颅内瘤体积,免疫细胞化学SP法检测p75NTR、神经生长因子(NGF)和细胞周期蛋白(cyclin)D2表达,用TUNEL法做移植瘤细胞原位细胞凋亡检测.结果 转染siRNA基因片段的U251细胞p75NTR mRNA和蛋白质表达水平显著下降(P<0.05),细胞侵袭能力及软琼脂集落形成能力均显著降低(P<0.05);体内实验显示p75NTR的表达程度与NGF的表达呈正相关,与cyclin D2表达及原位细胞凋亡数量呈负相关,MRI示瘤体积增长缓慢,边界轮廓清晰,动物生存期明显延长(P<0.05).结论 靶向p75NTR的siRNA技术可有效降低胶质瘤细胞的侵袭及增殖能力,诱导肿瘤细胞凋亡.     

p75 neurotrophin receptor regulates basal and fluoxetine-stimulated hippocampal neurogenesis

It is widely acknowledged that neurogenesis occurs in the adult hippocampus under normal conditions and that the rate can be regulated by environmental factors, including antidepressant drugs, with concomitant effects on behaviour. Using a quick and sensitive flow cytometry method that can assess changes in the number of bromodeoxyuridine (BrdU)-positive cells in hippocampus, in combination with traditional histological cell counts in the dentate gyrus, we report that mice lacking the p75 neurotrophin receptor gene (p75^NTR−/−) have significantly reduced hippocampal neurogenesis. Chronic treatment with the antidepressant fluoxetine stimulated hippocampal cell proliferation in p75^NTR−/− animals, but it did not result in an increase above basal levels of the number of newly born neurons in the dentate gyrus. These results indicate that p75^NTR acts as a regulator of fluoxetine-stimulated as well as basal adult hippocampal neurogenesis.     

Distribution of the neurotrophin receptors p75 and trkB in peripheral mechanoreceptors; observations on changes after injury

The neurotrophin family mediates effects of growth, cell differentiation and cell death through low- and high-affinity transmembrane receptors. The Pacinian corpuscle (PC) is the largest peripheral mechanoreceptor in mammals and was studied by immuno-histochemistry and immuno-electron microscopy with regard to the distribution of neurotrophin receptors, p75; p140 trkA, p145 trkB and 145 trkC. TrkA- and trkC-like immunoreactivity (IR) was not expressed in rat and cat PCs. Developing and adult animals expressed p75 and trkB in lamellar cells of the PC. The inner core cells, thought to be specialised Schwann cells, demonstrated an injury-induced increased immuno-labelling for trk B. Perineurial-derived outer core cells were reactive to p75 after injury similar to the perineurium of distal nerve stumps. Inner core cells of PCs behaved as leptomeningeal cells with regard to trkB. Outer core lamellar cells of PCs behaved as perineurial cells with regard to p75. A role for brain-derived neurotrophic factor is proposed in the development and nerve regeneration of PCs via an anterograde messenger transfer through p75 and trkB.     

Neurotrophin signaling through the p75 neurotrophin receptor

The neurotrophins are growth factors that play critical roles in the development, maintenance, survival, and death of the nervous system. The signal transducing systems that mediate the diverse biological functions of the neurotrophins are initiated by their interactions with two categories of cell surface receptors, the Trk family of tyrosine kinases and the p75 neurotrophin receptor (p75NTR). While the Trk receptors are responsible for most of the survival and growth properties of the neurotrophins, the actions of p75NTR fall into two categories. First, p75NTR is a Trk co-receptor that can enhance or suppress neurotrophin-mediated Trk receptor activity. Second, p75NTR autonomously activates signaling cascades that result in the induction of apoptosis or in the promotion of survival. The signaling cascades activated by p75NTR remain elusive, but structural and functional differences between p75NTR and other tumor necrosis factor receptor (TNFR) superfamily members suggest that p75NTR employs distinct signaling pathways. p75NTR has been shown to activate the NF-kappaB, Akt, and JNK pathways and interacts with several adaptor proteins. Of these, NRAGE, NADE, and NRIF have been associated with the induction of apoptosis, and FAP-1, RIP2, and TRAF6 appear to promote cellular survival. It remains a major challenge to link the various p75NTR binding proteins to specific p75NTR-dependent functions, but the identification of p75NTR interactors and signaling pathways has sparked new directions in p75NTR research, and will provide a better understanding of this enigmatic receptor.     

Nerve growth factor activates aorta endothelial cells causing PI3K/Akt- and ERK-dependent migration

Nerve growth factor (NGF) is a well-known neurotrophin. We determined whether NGF can activate endothelial cell migration and signalling that underlie angiogenic processes. We showed that aorta endothelial cells express mRNA for both the receptor tyrosine kinase TrkA and the p75 neurotrophin receptor (p75NTR) that associates with TrkA when signalling occurs. Pig aortic endothelial cells migrated when exposed to an NGF gradient, due to the simultaneous activation of the phosphatidylinositol 3-kinase and extracellular signal-regulated kinase signalling pathways. Furthermore, morphological changes were found in migrating cells: they appear with elongated structures with a smaller cell volume than control cells. Our data show that NGF is an activator of endothelial cells and suggest that NGF plays a role in mediating angiogenesis.     

Pro-NGF isolated from the human brain affected by Alzheimer's disease induces neuronal apoptosis mediated by p75NTR

The pro-form of nerve growth factor (pro-NGF) has been shown to be a high affinity ligand for p75NTR and to induce apoptosis through this receptor. It has been reported that pro-NGF, rather than mature NGF, is the predominant form of this neurotrophin in human brain. In the present work we studied the potential involvement of pro-NGF purified from human brains affected by Alzheimer's disease (AD), where it is especially abundant, in the neuronal apoptosis observed in this disease. Western blot analysis of human brain tissue showed the existence of several pro-NGF forms. Some of these pro-NGF forms were significantly increased in AD brain cortex in a disease stage-dependent manner. Pro-NGF, purified by chromatography from human AD brains, induced apoptotic cell death in sympathetic neurons and in a p75NTR stably transfected cell line. Blocking p75NTR in cell culture abolished neuronal apoptosis caused by pro-NGF. p75NTR-transfected cells underwent apoptosis in the presence of pro-NGF while control wild-type cells did not. Taken together, these results indicate that pro-NGF purified from AD human brains can induce apoptosis in neuronal cell cultures through its interaction with the p75NTR receptor.     

Brain-derived neurotrophic factor enhances the excitability of rat sensory neurons through activation of the p75 neurotrophin receptor and the sphingomyelin pathway

Neurotrophin-mediated signalling cascades can be initiated by activation of either the p75 neurotrophin receptor (p75(NTR)) or the more selective tyrosine kinase receptors. Previously, we demonstrated that nerve growth factor (NGF) increased the excitability of sensory neurons through activation of p75(NTR) to liberate sphingosine 1-phosphate. If neurotrophins can modulate the excitability of small diameter sensory neurons through activation of p75(NTR), then brain-derived neurotrophic factor (BDNF) should produce the same sensitizing action as did NGF. In this report, we show that focally applied BDNF increases the number of action potentials (APs) evoked by a ramp of depolarizing current by reducing the rheobase without altering the firing threshold. This increased excitability results, in part, from the capacity of BDNF to enhance a tetrodotoxin-resistant sodium current (TTX-R I(Na)) and to suppress a delayed rectifier-like potassium current (I(K)). The idea that BDNF acts via p75(NTR) is supported by the following observations. The sensitizing action of BDNF is prevented by pretreatment with a blocking antibody to p75(NTR) or an inhibitor of sphingosine kinase (dimethylsphingosine), but not by inhibitors of tyrosine kinase receptors (K252a or AG879). Furthermore, using single-cell RT-PCR, neurons that were sensitized by BDNF expressed the mRNA for p75(NTR) but not TrkB. These results demonstrate that neurotrophins can modulate the excitability of small diameter capsaicin-sensitive sensory neurons through the activation of p75(NTR) and its downstream sphingomyelin signalling cascade. Neurotrophins released upon activation of a variety of immuno-competent cells may be important mediators that give rise to the enhanced neuronal sensitivity associated with the inflammatory response.     

p75 Neurotrophin Receptor-Mediated Signaling Promotes Human Hair Follicle Regression (Catagen)

Nerve growth factor (NGF) and its apoptosis-promoting low-affinity receptor (p75NTR) regulate murine hair cycling. However, it is unknown whether human hair growth is also controlled through p75NTR, its high-affinity ligand pro-NGF, and/or the growth-promoting high-affinity NGF receptor tyrosine kinase A (TrkA). In microdissected human scalp anagen hair bulbs, mRNA for NGF, pro-NGF, p75NTR, and TrkA was transcribed. Immunohistomorphometry and in situ hybridization detected strong NGF and pro-NGF expression in terminally differentiating inner root sheath keratinocytes, whereas TrkA was co-expressed with p75NTR in basal and suprabasal outer root sheath keratinocytes. During spontaneous catagen development of organ-cultured human anagen hair follicles, p75NTR mRNA levels rose, and p75NTR and pro-NGF immunoreactivity increased dramatically in involuting compartments primarily devoid of TrkA expression. Here, TUNEL(+) apoptotic cells showed prominent p75NTR expression. Joint pro-NGF/NGF administration inhibited hair shaft elongation and accelerated catagen development in culture, which was antagonized by co-administration of p75NTR-blocking antibodies. In addition, mRNA and protein expression of transforming growth factor-beta2 increased early during spontaneous catagen development, and its neutralization blocked pro-NGF/NGF-dependent hair growth inhibition. Our findings suggest that pro-NGF/NGF interacts with transforming growth factor-beta2 and p75NTR to terminate anagen in human hair follicles, implying that p75NTR blockade may alleviate hair growth disorders characterized by excessive catagen development.     

The p75 neurotrophin receptor in human development and disease

The functional effects of nerve growth factor (NGF) and its precursor, pro-NGF, are thought to be mediated through binding of these ligands to one or both of their receptors, TrkA and p75NTR. While the signaling pathways and downstream effects of NGF binding to TrkA are reasonably well known, those related to the binding of NGF and pro-NGF to p75NTR are less well understood. Furthermore, p75NTR appears to play functional roles that are unrelated to its ability to bind NGF and pro-NGF, some of which are ligand-independent and others of which are dependent upon binding to other neurotrophins. As these functional roles and their biochemical mechanisms become better known, the importance of p75NTR, related receptors, and both extracellular ligands and intracellular interactors and effectors for human development and health has become increasingly apparent. A complete understanding of p75NTR and its cellular partners is best served by approaching the remaining questions from both sides, with studies of function in normal states and studies of dysfunction in aberrant states mutually informing one another.