周细胞在视网膜损伤内源性修复中的作用

Pericytes share similarity with mesenchymal stem cells (MSC). Our previous report reveals MSC promote injury repair in retina via glial modulation. A similar role may exist in pericytes accordingly. In this proposal, we are interested in how the interaction between perivascular cells and Müller glia may impact on endogenerous repair in retinal injury. Pericytes will be cultured and purified to develope a coculture system, in which the interaction between pericytes and Müller glia will be investigated by analysing changes of proliferation, transdifferentiation and STAT3 signal pathway in both cells. Recombinant lentivirous targeting pericytes will be designed to inhibit pericyte function and intravitreously injected. The injected animals will subsequently undergo light-induced retinal injury..Morphological changes and apoptosis in retina will be identified transversally and longitudinally under confocal laser scaning microscope to evaluate the role that pericytes play in light-induced retinal injury. Müller cell count and protein expression regarding PCNA、Nestin、Rhodopsin and p-STAT3 will be detected for an insight into how pericytes affect Müller glia from the aspects of cell proliferation, transdifferentiation and STAT3 signal pathway. Taken together, we will outline possible mechanism by which pericytes involve in retinal repair and interact with Müller glia. This research proposal highlighted potential application of pericytes for endogenous repair in retinal injury.

周细胞与间充质干细胞（MSC）相似，我们前期研究提示MSC通过胶质细胞调节视网膜损伤修复，周细胞可能有类似作用。故本项目拟研究周细胞与Müller细胞相互作用对视网膜自身修复的影响。体外培养、扩增并纯化视网膜周细胞，采用共培养模式探讨损伤后视网膜周细胞与Müller细胞的增殖、转分化及STAT3信号通路变化。利用重组慢病毒转染视网膜以抑制周细胞功能，建立视网膜光损伤模型，共聚焦显微镜下纵向及横向扫描观察视网膜组织结构、血管形态及细胞凋亡情况，评估周细胞在参与视网膜光损伤修复的作用，并进行Müller细胞计数、PCNA、Nestin、Rhodopsin及p-STAT3表达检测，探讨视网膜周细胞对Müller细胞增殖、转分化及STAT3信号通路的影响。探讨视网膜周细参与视网膜修复的可能机制及其与Müller细胞间的作用方式，为进一步利用周细胞促进视网膜损伤内源性修复提供基础。

周细胞是视网膜微血管的重要组成细胞，参与血-视网膜屏障构建，在视网膜网膜的损伤修复中也发挥重要作用。鉴于周细胞与间充质干细胞在细胞表型、生物学特性的相似性及我们前期发现的间充质干细胞参与视网膜组织损伤修复过程，本项目研究了视网膜周细胞在光损伤视网膜内源性修复中的作用，研究提取了视网膜周细胞，建立了视网膜光损伤动物模型，通过眼内注射转染视网膜周细胞，下调PDGFR-β表达以构建周细胞功能障碍模型，研究发现视网膜光损伤诱导了组织内PCNA高表达，视网膜细胞增殖活动加强可能参与了组织内源性修复，在PDGFR-β表达下调介导的周细胞功能障碍中视网膜应对光损伤内源性修复的能力下降，细胞增殖活动减弱，PCNA表达下调，而Nestin表达无明显变化，同时视网膜光损伤诱导的感光细胞大量损害，Rhodopsin表达下调，在PDGFR-β表达下调介导的周细胞功能障碍中，Rhodopsin表达进一步下调，表明周细胞功能障碍加重视网膜损伤的进程，为探讨相关机制，研究着手于视网膜STAT3信号通路，发现在光损伤模型中视网膜STAT3通路蛋白磷酸化水平升高，启动了内源性修复，而PDGFR-β下调介导的周细胞功能障碍抑制了视网膜STAT3蛋白磷酸化，组织病理学观察表明视网膜结构损害加重，提示周细胞功能下调抑制了其参与内源性修复的过程，研究为进一步利用周细胞促进视网膜损伤内源性修复提供了基础数据。