Brn3a和Brn3b对小鼠视网膜神经节细胞凋亡及保护的作用机制研究

Glaucoma is an irreversible blind-causing disease characterized by the programmed death of retinal ganglion cells (RGC). Its pathogenesis is currently unknown. Previous studies using glaucoma models revealed a significant down-regulation of transcription factors (TFs) Brn3a and Brn3b before the programmed apoptosis of RGC. Over-expression of Brn3b plays a neuro-protective role in ocular hypertension rat model in vitro. Suggesting an intimate correlation of these TFs with the pathogenesis and treatment of glaucoma. Brn3b is the key factor during RGC development, and Brn3a functioned equivalently with Brn3b. However, the mechanism how Brn3a and Brn3b regulate the RGC development and survival is unknown. In our project we will employ conditional knockout models that we have generated and over-express Brn3a and Brn3b using AAV vectors. RNA-seq will be performed to search the gene expression level change and RT-PCR and Western blot will be performed to confirm the gene expression level and protein alteration. Chip-seq will be used to seek the regulation pathway. This project will expand regulation pathways that controlling the development and survival of RGC, and further explore the mechanism of programmed RGC apoptosis.

青光眼是以视网膜神经节细胞（RGC）程序性死亡为特点的不可逆性致盲眼病，发病机制不明确。青光眼模型研究显示RGC死亡前，Brn3a和Brn3b表达显著降低；同时体外研究发现 Brn3b的过表达对青光眼条件下的RGC具有保护作用，提示两者对青光眼发病和治疗具有重要意义。目前已知Brn3b为RGC正常发育存活所必需，且Brn3a与Brn3b在功能上相互平等，但两者对RGC凋亡和保护的作用机制尚不清楚。为了明确该问题，本项目将在前期建立小鼠Brn3a和Brn3b条件性基因敲除模型的基础上，利用腺相关病毒载体建立Brn3a和Brn3b过表达模型。通过高通量RNA-seq观察Brn3a、Brn3b表达缺失和过表达情况下的基因表达差异，明确其下游基因；同时使用ChIP-seq确定Brn3a和Brn3b与下游基因的作用关系，探索RGC存活与凋亡调控通路及程序性死亡机制，对青光眼的诊断和治疗具有重要意义。