Bit1介导的星形胶质细胞失巢凋亡在视网膜血管发育重塑中的作用及机制研究

Astrocyte apoptosis are known to be essential for the formation and remodeling of retinal primary vascular plexus. However, the mechanism of astrocyte apoptosis process and is not well known. However, little is known about the apoptosis mechanism and the its effect on retinal vascular remodeling. Bit1(Bcl-2 inhibitor of transcription 1), a mitochondria protein, is mainly associated with anoikis (cell death induced by detachment from extracellular matrix). Our previous work shows specific expression of Bit1 in retinal astrocytes during primary vasculature development. In vitro cell assays reveal that astrocytes undergo apoptosis during anoikis induction. Accordingly, we assume that the Bit1-mediated anoikis in astrocytes trigged by loss of integrin α5β1 ligation from extracellular fibronectin play an important role in retinal vasculature development and remodeling. The anoikis process may be regulated by microenvironmental oxygen tension and may result in decreased VEGF level which effect the survival and proliferation of endothelial cells. This subject aims to study the interaction mechanism of Bit1-mediated anoikis in astrocyte in the retinal developing process through both in vitro and in vivo experiment employing RNAi and overexpress technique. The subject would help explain the potential mechanism of retinal vascular development and provide theoretical evidence for manipulate retinal vascular diseases.

星形胶质细胞的凋亡对于视网膜浅层血管网的发育重塑具有重要作用，但关于其凋亡形式及参与血管网重塑的作用机制尚不十分清楚。我们的前期工作发现在视网膜浅层血管网发育重塑阶段Bit1在星形胶质细胞当中特异性表达。Bit1是失巢凋亡的关键蛋白，体外实验初步揭示星形胶质细胞存在失巢凋亡现象。据此我们提出假设，由整合素α5β1与细胞外基质Fibronectin脱离接触激活了星形胶质细胞中Bit1介导的失巢凋亡，并可能受到微环境氧浓度的调控，通过改变局部VEGF的水平进而影响血管内皮细胞的增殖和凋亡。本项目将从分子的过表达或抑制、细胞实验、整体动物模型等不同层面，通过改变实验氧浓度、采用抑制剂、封闭肽和RNA干扰等多种手段揭示关键蛋白Bit1介导的星形胶质细胞失巢凋亡参与对视网膜血管发育重塑的分子机制，这将有助于更全面地认识视网膜血管发育或新生血管的发生过程，为临床上治疗视网膜血管性疾病提供思路。

星形胶质细胞对视网膜血管网的发育重塑具有重要作用。我们通过研究该阶段星形胶质细胞内特异性表达蛋白Bit1的调控作用，阐述了其对血管网重塑的作用机制。星形胶质细胞网络不仅对血管内皮细胞迁移具有支架引导作用，研究还提示星形胶质细胞释放VEGF促进血管生成并受到Bit1的调控。Bit1在视网膜星形胶质细胞的增殖、迁移及凋亡发挥重要作用，继而通过影响微环境VEGF的水平对视网膜血管内皮细胞的增殖、迁移及小管生成产生作用。常氧条件下，内源性Bit1表达可以维持细胞增殖能力，抑制凋亡发生，同时维持一定水平的VEGF。通过与细胞外基质Fibronectin粘附能上调星形胶质细胞Bit1的表达，进一步促进其增殖，抑制凋亡以及VEGF的表达，从而表现为星形胶质细胞网络的延伸而对血管生成作用受到一定程度抑制。当模拟体内乏氧环境时，星形胶质细胞内Bit1下调，而HIF1α、VEGF显著上升，继而星形胶质细胞来源的VEGF水平上升，促进了血管内皮细胞的增殖、血管生成等，初步揭示出星形胶质细胞Bit1可能通过调控氧浓度诱导的HIF1α-VEGF通路改变微环境VEGF水平，进而影响视网膜血管内皮细胞的增殖、迁移及小管生成等，发挥对视网膜血管网络的重塑作用。通过该研究有助于理解视网膜血管发育的过程，还将为认识病理性微血管异常和新生血管的发生机理提供思路，若能合理地抑制星形胶质细胞来源的VEGF将有可能增加正常血管内皮细胞网络的存活，从而改善视网膜的缺血缺氧并从根本上抑制新生血管的发生，为临床治疗视网膜血管发育性疾病、视网膜新生血管性疾病等提供帮助。