少突胶质谱系细胞钾离子通道Kv1.3在多发性硬化中对髓鞘再生的作用及机制研究

In central nervous system, neuronal axons are myelinated with oligodendrocytes (Ol) and damage of such a myelin sheath is a striking pathological feature of many demyelinating diseases, including multiple sclerosis. There is no effective way to stimulate and promote axonal remyelination because the oligodendrocyte progenitor cells (OPC) cannot differentiate to mature myelin-producing Ol. It has been demonstrated that activation of voltage-dependent K+ channels (Kv) involves in the regulation of OPC/Ol differentiation and apoptosis. Our preliminary results showed that Kv1.3 channels and currents were highly expressed in early stage of oligodendrocyte progenitor cells and decreased in mature Ols; Kv1.3 activity is closely related to p38 MAPK and AKT signals, which are the regulators of Ol differentiation. It is our hypothesis that activation of Kv1.3 channels negatively regulates Ol differentiation and, impairs their physiological functions in remyelination. The proposed study will utilize patch-clamp, siRNA knockdown, overexpression by plasmid transfection, and electron microscopy methodologies to investigate the underlying mechanisms of the role Kv1.3 played in remyelination in primary cultured Ol, cultured brain slices and demyelination animal models. In addition, we want to explore whether the Kv1.3 could be a potential target for the development of therapeutic strategies to combat against demyelinating diseases, in particular, the multiple sclerosis.

中枢神经系统的髓鞘由少突胶质细胞（Ol）包绕轴突形成。髓鞘损伤是以多发性硬化为代表的脱髓鞘疾病的标志性病理改变，而目前尚缺乏促进髓鞘再生的有效方法。病理条件下幼稚的少突胶质前体细胞（OPC）无法分化成熟是受损髓鞘不能修复的关键问题所在。电压门控性钾离子通道（Kv）参与调节OPC/Ol的分化及凋亡。我们的预实验结果显示Kv1.3在OPC中高表达，而在成熟Ol中表达降低；且Kv1.3活性与参与Ol分化调控的p38 MAPK及AKT信号密切相关。因此我们设想Kv1.3负性调节Ol的分化成熟，并损伤髓鞘再生。此课题将通过原代培养Ol、脑组织切片培养及脱髓鞘动物模型，采用膜片钳、siRNA沉默、转染过表达、电子显微镜等手段，从分子细胞、组织及动物水平探讨Kv1.3在髓鞘再生中的作用，明确Kv1.3调节Ol分化成熟的机制，为促髓鞘再生的治疗提供新思路。

中枢神经系统的髓鞘由少突胶质细胞（OL）包裹轴突形成，由此保证动作电位的快速传导。髓鞘损伤是以多发性硬化为代表的众多脱髓鞘疾病的显著病理特征，虽然普遍的观点认为自身免疫反应参与了疾病的脱髓鞘过程，导致髓鞘损伤继而引发脱髓鞘，但其发病机制并不完全清楚。电压门控钾离子通道Kv1.3是OL中主要的钾离子通道，参与细胞周期的调控，影响OL细胞功能。我们以Kv1.3为切入点，构建了细胞水平白介素17（IL-17）诱导的OL损伤和动物在体水平溶血卵磷脂（LPC）诱导的髓鞘损伤模型，试图探究Kv1.3抑制剂5-(4-phenoxybutoxy) psoralen (PAP)对OL/髓鞘是否具有保护作用。结果显示Kv1.3电流随着细胞发育成熟逐渐降低；IL-17的应用增加了Kv1.3的表达，导致AKT活化减少，增殖受抑，这些反应可以被Kv1.3的抑制剂PAP拮抗; AKT通路的激活能有效对抗IL-17引起的细胞损伤，而p38通路的激活不具有此种效应。因此, Kv1.3作为多发性硬化的潜在治疗靶点，可能归因于对OL的保护而非仅作用于免疫抑制。