The Role of Sodium Channel Nav1.7 in Osteoarthritis - Resubmission - 1

骨关节炎（OA）是最常见的关节疾病，目前还没有有效的预防或治疗方法。 减缓关节退化因此，鉴定新的OA相关分子可能提供宝贵的信息， 形成对寻找新的治疗目标的OA。我们的RNAseq筛选新的，差异性的 在OA中表达的基因导致Nav1.7（由SCN 9A编码）作为新的OA相关分子的分离。 Nav1.7由于其与一系列遗传性人类疼痛障碍的相关性而具有特别重要的意义。一 据报道，SCN 9A的核苷酸多态性与OA患者疼痛敏感性增加相关。 我们的初步数据表明，Nav1.7在两种人的浅层软骨细胞中表达增加， OA软骨和来自小鼠OA模型的软骨。我们很高兴地发现，阻止Nav1.7 降低人软骨细胞的膜电位。据我们所知，这是第一个证据证明- 说明钠通道在非兴奋性软骨细胞中具有电生理功能。超极化 在软骨细胞中由Nav1.7阻断导致的分泌/跨膜转运的改变 蛋白质。事实上，直接阻断Nav1.7和与条件培养基一起孵育收集的细胞都可以被激活。 Nav 1.7处理的软骨细胞增强了软骨细胞的抗凋亡作用，并抑制了IL-1β诱导的软骨细胞凋亡。 禁忌主义。此外，一系列的蛋白质组学筛选从Nav1.7阻断剂处理的细胞中分离出HSP 70和中期因子。 条件培养基作为Nav1.7在软骨细胞中的两种潜在关键介质。更重要的是，当地和 口服Nav1.7特异性抑制剂可预防OA并减少OA相关疼痛， ly和化学诱导的OA模型。该申请的假设是Nav1.7在以下方面起着关键作用： 软骨细胞代谢和OA通过调节膜电位和软骨分泌蛋白质组谱， 细胞。具体目的是：（1）阐明Nav1.7调节细胞凋亡的分子和细胞机制; 延缓软骨细胞代谢。我们将确定SA#1A）Nav1.7阻断、过表达和 缺失软骨细胞代谢; SA#1B）介导Nav1.7功能的Nav1.7靶基因， 软骨细胞; SA#1C）Nav1.7阻断剂处理的条件培养基中的分子决定簇， SA#1D）参与Nav1.7调节的Nav1.7辅因子， 软骨细胞。(2)明确Nav1.7在OA发生和进展中的重要性，以及 涉及的潜在机制。我们将确定SA#2A）Nav1.7整体消融对OA的影响， 发生和进展; SA#2B）软骨细胞表达的Nav1.7对OA的重要性; SA#2C）依赖性 Nav1.7阻断对HSP 70和中期因子介导的对OA的保护作用的影响;以及SA#2D）治疗性 药理学Nav1.7阻断对OA和OA相关疼痛的影响。研究报告不仅将- 万斯通过阐明Nav1.7对软骨细胞和OA的调节而进入该领域，但也可能导致软骨细胞的发展。 Nav1.7抑制剂作为治疗OA的新型疾病修饰药物，而不仅仅是止痛药。

Osteoarthritis (OA) is the most common joint disease and currently there is no effective means of preventing or slowing joint degeneration. Thus, identification of new OA-associated molecule(s) may provide invaluable in- formation toward the search for novel therapeutic targets for OA. Our RNAseq screen for novel, differentially expressed genes in OA led to the isolation of Nav1.7 (encoded by SCN9A) as a novel OA-associated molecule. Nav1.7 is of particular significance due to its correlation with a spectrum of hereditary human pain disorders. A nucleotide polymorphism of SCN9A was reported to be correlated with increased pain sensitivity in OA patients. Our preliminary data demonstrated that Nav1.7 was increased in superficial zone chondrocytes in both human OA cartilage and cartilage from a mouse OA model. We were excited to find that blocking Nav1.7 dramatically reduced membrane potential in human chondrocytes. To our knowledge, this is the first evidence demonstrat- ing that sodium channel has electrophysiological function in non-excitable chondrocytes. Hyperpolarization caused by Nav1.7 blockade in chondrocytes results in the alternations of secretome/cross-membrane transport of the proteins. Indeed, both direct blockade of Nav1.7 and incubation with the conditioned medium collected from Nav1.7 inhibitor-treated chondrocytes enhanced chondrocyte anabolism and inhibited IL-1β induced ca- tabolism. Further, a series of proteomics screens isolated HSP70 and midkine from the Nav1.7 blocker-treated conditioned medium as two potential key mediators of Nav1.7 in chondrocytes. More significantly, local and oral delivery of Nav1.7-specific inhibitor protected against OA and reduced OA-associated pain in both surgical- ly- and chemically-induced OA models. The hypothesis of the application is that Nav1.7 plays a pivotal role in chondrocyte metabolism and OA through regulating membrane potential and secretome profiling of chondro- cytes. The Specific Aims are: (1) To elucidate the molecular and cellular mechanisms by which Nav1.7 regu- lates chondrocyte metabolism. We will determine SA#1A) the effects of Nav1.7 blockade, overexpression and deletion on chondrocyte metabolism; SA#1B) the target genes of Nav1.7 that mediate the functions of Nav1.7 in chondrocytes; SA#1C) the molecular determinants in Nav1.7 blocker-treated conditioned medium which medi- ate Nav1.7 regulation of chondrocytes; and SA#1D) the co-factor(s) of Nav1.7 that are involved in Nav1.7 regu- lations of chondrocytes. (2) To define the importance of Nav1.7 in the initiation and progression of OA, and the underlying mechanisms involved. We will determine SA#2A) the effects of global ablation of Nav1.7 on OA ini- tiation and progression; SA#2B) the importance of chondrocyte-expressed Nav1.7 to OA; SA#2C) the depend- ence on HSP70 and midkine of Nav1.7 blockade mediated protection against OA; and SA#2D) the therapeutic effects of pharmacological Nav1.7 blockade on OA and OA-associated pain. Proposed studies will not only ad- vance the field by elucidating Nav1.7 regulation of chondrocytes and OA, but may also lead to the development of Nav1.7 inhibitors as novel diseases-modifying drugs for treating OA rather than just pain relievers.