Nav1.9突变导致人类痛觉异常的分子机制

Pain was a medical puzzle. The pathologic mechanisms have remained elusive. Many ion channel genes have been associated with human genetic pain disorders. The SCN11A gene, encoding the ninth member of ɑ subunits of sodium channels (Nav1.9), is preferentially expressed in nociceptive neurons and plays important role in pain by verification of in vitro and vivo experiments. However, it was unclear whether mutations in SCN11A are associated with human genetic pain disorders. Untill 2013, Nav1.9 had been recognized as an important pain channel in humans reported by two independent research groups. One report by Leipold et al. showed that two individuals felt no pain carrying with the heterozygous mutation (L811P) in SCN11A; and another work by our research group studied two large Chinese families with many members affected by similar episodic pain symptoms, inherited in an autosomal-dominant pattern. Furthermore, we found a different mutation in each family in SCN11A/Nav1.9, R225C, and A808G, and Nav1.9 channels carrying either mutation passed more excitatory current than wild-type channels which was similar to the L811P mutation. However, new problems why similar gain-of-function mutations in the same SCN11A cause different pain syndromes are thrown up. In this study, we are aiming to further identify the Nav1.9 mutational category through mutation analysis in SCN11A by collecting more patients with pain disorders. We will then employ multidisciplinary approaches, including patch-clamp, molecular biology, protein biochemistry, immunocytochemical staining, fluorescence imaging, and knockout/knockin mouse strategy, to elucidate the molecular mechanisms underlying different pain disorders caused by SCN11A mutations. Therefore the proposed research is critically important for the further understanding of Nav1.9 functions in pain sensation, and also provides insights into analgesic drug developments based on Nav1.9 target.

疼痛一直是困扰医学界难题，发病机理不是十分清楚。许多离子通道基因突变与人类遗传性疼痛关联。Nav1.9是由SCN11A基因编码的电压门控型钠离子通道ɑ亚基第9个成员，它在伤害性感受器中呈特异分布，一些体内、外实验证明Nav1.9在疼痛中具有重要作用，但一直没有在人类疼痛异常患者中发现Nav1.9突变。2013年本课题组和Leipold等几乎同时报道Nav1.9突变与人类疼痛异常相关。但Nav1.9类似获得性功能突变所引起临床结果相反，Leipold等报道无痛，而我们发现的是发作性疼痛。为什么？本研究拟收集更多的疼痛异常患者对SCN11A进行突变检测，进一步确定突变类型；从体外（研究这些突变的电生理特性）和体内（KO和KI小鼠研究相似的功能突变导致的不同临床表现机制）水平研究Nav1.9突变导致疼痛异常发生的分子机制，为开发以Nav1.9为药靶的止痛药物奠定基础。

疼痛是一种机体警戒信号，能有效防止机体进一步的伤害，并有助于损伤组织的愈合。但机体损伤和疾病引起适应不良导致的慢性疼痛影响世界近五分之一的人口，严重影响人们的生活质量。2013年，本课题组首次鉴定出Nav1.9变异与人类发作性疼痛相关。后来陆续发现Nav1.9变异可导致发作性疼痛，先天性无痛症和痛性周围神经病。但没有报道以Nav1.9为靶点的治疗药物。本研究将通过Nav1.9-Knock in小鼠探索Nav1.9变异导致发作性疼痛的机制，以及以Nav1.9为靶点筛选多肽类药物以及互作蛋白，为后续为开发以Nav1.9为靶点止痛药物奠定基础。.在该项目资助下，发现Nav1.9突变患者饮酒触发疼痛致病原因是患者同时携带Nav1.9突变和ALDH2的SNP位点p.Glu504Lys, 进一步成功构建了Nav1.9-KI小鼠模型，以及证明COX2抑制剂可以部分缓解Nav1.9-KI(Scn11aA796G/A796G )小鼠疼痛超敏；同时通过Nav1.9-KI小鼠模型发现PKG-Crip2通路参与调控Nav1.9突变导致疼痛信号传导；另一方面，发现蜘蛛毒素HpTx1通过增大Nav1.9电流可以使Nav1.7功能缺失小鼠DRG神经元兴奋性增强，进而恢复Nav1.7功能缺失小鼠痛觉，以及揭示了Nav1.7、Nav1.8和 Nav1.9三个通道之间共同精细调节神经元兴奋性的机制；最后，从小鼠DRG cDNA文库中筛选得到与Nav1.9互作的蛋白——精氨酸甲基转移酶Prmt7，证明Prmt7使Nav1.9电流增加，而抑制Prmt7活性可以使Nav1.9电流减小和降低DRG神经元兴奋性，因此对治疗Nav1.9突变相关疼痛提供了新思路。.该项目实施过程中参加10人次国内学术交流会议和获得1项授权国家发明专利，在该项目支持下，发表论文11篇，标注该项目影响因子>10的SCI论文1篇（nature communications），其他SCI论文10篇。培养博士生5名、硕士生3名（均已毕业）和1位博士后。