Role of NGF signaling in the bladder after spinal cord injury

 描述（由申请人提供）：我们最近发表了NGF受体p75是脊髓损伤的潜在治疗靶点：脊髓损伤（SCI）后的运动功能在口服递送LM 11 A-31（一种设计为靶向p75的小分子）后得到改善。通过观察到病理性proNGF与p75结合的阻断，证实了LM 11 A-31对p75的有效体内靶向作用，这表明该药物可用于p75在与proNGF或其他原神经营养因子相互作用后发挥类似病理作用的其他系统中。我们已经发表了ProNGF水平在CNS损伤后迅速增加，被释放到脑脊液中。与结合TrkA和p75的成熟NGF不同，proNGF选择性地结合p75，从而激活p75的下游凋亡级联。 神经生长因子与脊髓损伤后的膀胱功能障碍以及膀胱过度活动症和间质性膀胱炎/膀胱疼痛综合征有关。在这些条件下和SCI后，在尿液中观察到NGF水平的显著增加，表明NGF可以是一般膀胱功能障碍的生物标志物，并且可以靶向改善膀胱功能。尽管长期的兴趣，然而，研究，旨在中和神经生长因子的行动导致了混合的结果。我们相信，在我们专注于神经生长因子和相关的神经营养因子作为治疗目标，以减轻膀胱功能障碍之前，更详细地了解神经生长因子及其受体在膀胱生物学中发挥的作用将是必要的。 我们已经发现，proNGF和proBDNF在SCI后不久释放到尿液中，而成熟的NGF在几天后释放。这些原神经营养因子似乎诱导负责建立渗透性屏障的伞细胞的凋亡，因为LM 11 A-31施用导致SCI后伞细胞的保留。因此，我们推测，在伞细胞中表达的p75在SCI后伞细胞的凋亡中发挥作用。除了伞细胞，我们发现p75也在脊髓损伤后尿路上皮增生期间尿路上皮的增殖祖细胞中表达。LM 11 A-31给药减弱了这种增生反应，表明前神经营养素和p75在调节SCI诱导的增生中发挥了额外的作用。因此，我们假设p75在脊髓损伤后尿道鞘的再生中起双重作用，（1）在脊髓损伤后不久诱导伞细胞凋亡，（2）通过调节细胞凋亡，（3）通过调节细胞凋亡。 祖细胞的增殖，从而补充丢失的伞细胞。在此假设下，我们建议确定p75是否诱导SCI后尿道鞘中伞细胞的死亡（目标1），这将通过条件性p75小鼠来解决，其中p75在伞细胞中被选择性地删除;为了确定p75在SCI后尿道增殖中的作用（目的2），这将通过分析SCI后的小鼠来解决，其中p75在尿路上皮祖细胞中缺失;并确定LM 11 A-31是否改善SCI后的膀胱功能（目标3），这将使用新的光学映射方法来解决，单单位传入神经记录、渗透性测量和膀胱测压图（CMG），同时进行尿道括约肌肌电图（EMG）。我们相信这项研究的结果将改变我们目前对神经营养因子在膀胱中作用的理解，并为现有的努力提供动力，以神经营养因子为目标，改善各种泌尿系统疾病的膀胱功能。

 DESCRIPTION (provided by applicant): We have recently published that a NGF receptor, p75, is a potential therapeutic target for the spinal cord injured: motor function after spinal cor injury (SCI) improved after oral delivery of LM11A-31, a small molecule that was designed to target p75. Effective in vivo targeting of p75 by LM11A-31 was confirmed by observation of a block in binding of a pathological proNGF to p75, suggesting that the drug can be used in other systems where p75 exerts similar pathological effects upon interaction with proNGF or other proneurotrophins. We have published that ProNGF levels increase rapidly after injury to the CNS, being released into the cerebrospinal fluid. Unlike mature NGF that binds both TrkA and p75, proNGF binds p75 selectively, thereby activating p75's downstream apoptotic cascade. NGF has been implicated in bladder dysfunction after SCI as well as in overactive bladder and interstitial cystitis/painful bladder syndromes. Under these conditions and after SCI, a significan increase in NGF levels is observed in the urine, suggesting that NGF can be a biomarker for general bladder dysfunction and may be targeted to improve bladder function. Despite the long-standing interest, however, studies that aimed at neutralizing NGF action has led to mixed results. We believe a more detailed understanding of the roles that NGF and its receptors play in bladder biology will be necessary before we focus on NGF and related neurotrophins as therapeutic target for alleviating bladder dysfunction. We have found that proNGF and proBDNF are released into the urine shortly after SCI, while mature NGF is released days later. These proneurotrophins appear to induce apoptosis of umbrella cells that are responsible for establishing the permeability barrier, as LM11A-31 administration led to retention of umbrella cells after SCI. We thus hypothesize that p75 that is expressed in umbrella cells play a role in apoptosis of umbrella cells after SCI. In addition to umbrella cells, we found that p75 is also expressed in proliferating progenitors of the urothelium during the period of urothelial hyperplasia after SCI. LM11A-31 administration attenuated this hyperplasic response, suggesting that proneurotrophins and p75 play an additional role in regulating SCI-induced hyperplasia. We thus hypothesize that p75 plays dual roles in the regeneration of the urothelium after SCI, (1) by inducing apoptosis of the umbrella cells shortly after SCI, and (2) by regulating proliferation of the progenitor cells, thereby replenishing the lost umbrella cells. Under this hypothesis, we propose to determine whether p75 induces death of umbrella cells in the urothelium after SCI (Aim 1), which will be addressed by conditional p75 mice, in which p75 is selectively deleted in umbrella cells; to determine the role of p75 in proliferation of the urothelium after SCI (Aim 2), which will be addressed by analyzing mice after SCI in which p75 is deleted in urothelial progenitors; and to determine whether LM11A-31 improves bladder function after SCI (Aim 3), which will be addressed using novel optical mapping approaches, single-unit afferent nerve recording, permeability measurements and cystometrograms (CMG) with simultaneous urethral sphincter electromyograms (EMG). We believe the outcome of this study will change our current understanding on neurotrophin action in the bladder and energize the existing effort to target neurotrophin in improving bladder function in various urological conditions.