circXPO5通过调控GRIN2A的表达在青光眼视觉中枢损伤的保护机制研究

Glaucoma is an ocular disease of the irreversible blindness that is often associated with the elevated intraocular pressure and the loss of retinal ganglion cells (RGCs). However, progressive vision loss is common despite controlling IOP, and the mechanism of vision loss is unclear. Circular RNA (circRNA) is one of non-coding RNA, and it plays an important role in regulation of gene expression. It has been proved that circRNA-microRNA (miRNA) is relevant to protein-coding gene and the biological processes of gene. In the former work of our research, we had discovered the neuronal apoptosis and Alzheimer’s disease-like changes in glaucomatous visual center system in monkey, but the pathogenic mechanism is unknown. Then we detected that the virulence genecore -- GRIN2A and the core circRNA-circXPO5 which has a functional relationship with GRIN2A were both decreased through high throughput sequencing technology on glaucomatous visual center system. Through the biological analysis, we found that circXPO5 as a ceRNA competed with GRIN2A for miR-330-5p binding. And then we have verified the miR-330-5p increased. Our results indicated the decrease of circXPO5 maybe play a protective role neuronal apoptosis of glaucomatous visual center system. We hypothesized that circXPO5 regulate GRIN2A and affect the neuronal apoptosis through sponging miR-330-5p. We will further study on functional verification of the differential gene we detected in molecular and tissue levels to reveal of regulative mechanism of circXPO5, the molecular mechanism of the neuronal apoptosis of glaucomatous visual center system, and the molecular regulative mechanism of circXPO5 on relevant protein of the neuronal apoptosis pathway that relevant with Aβ in glaucomatous visual center system. The results may provide a new idea for developing research therapies that will enhance neuroprotection in glaucoma patients.

circRNA是一种非编码RNA，在基因表达调控中具有重要作用，已证明circRNA-miRNA与蛋白质编码基因有关。我们前期研究发现，高眼压模型的视觉中枢存在神经元凋亡和阿尔茨海默病样改变，其致病机制未明。通过高通量测序筛查致其病基因发现GRIN2A以及与其有功能联系的核心circRNA--circXPO5表达均降低，经生物学分析发现circXPO5和GRIN2A互为ceRNA与miR-330-5p竞争结合。结果提示circXPO5的下降可能在青光眼视觉中枢神经元的凋亡中起着保护作用。据此我们提出假说：circXPO5作为ceRNA通过吸附miR-330-5p而调控GRIN2A的表达，进而影响青光眼视觉中枢神经元的凋亡水平。本课题拟继续深入从分子、细胞、组织层面进行circXPO5的功能验证，明确circXPO5的调控机制，探讨青光眼视觉中枢神经元凋亡的发生机制，为青光眼的治疗提供新思路

circRNA是一种非编码RNA，在基因表达调控中具有重要作用，已证明circRNA-miRNA与蛋白质编码基因有关。我们前期研究发现，恒河猴高眼压模型的视觉中枢存在神经元的凋亡和阿尔茨海默病样改变，其致病机制未明。通过高通量测序筛查其致病基因发现GRIN2A以及与其有功能联系的核心circRNA--circXPO5表达均降低，经生物学分析发现circXPO5和GRIN2A互为ceRNA与miR-330-5p竞争结合。我们的研究结果提示circXPO5的下降可能在青光眼视觉中枢神经元的凋亡中起着保护作用。本课题通过深入从体内和体外实验进行circXPO5的功能和表型验证，结果显示青光眼模型中circXPO5的表达下降，而miR-330-5p和GRIN2A的表达升高，qPCR也进一步验证了circXPO5、miR-330-5p和GRIN2A的表达，结果还发现，主要凋亡关键蛋白cleaved caspase 3和TUNEL染色以及主要自噬关键蛋白Beclin 1、LC3B -II/LC3B-I在青光眼模型的LGN中表达明显升高（P < 0.05），通过电镜技术也发现青光眼模型的上丘（SC）部分神经元出现凋亡小体和自噬溶酶体（神经元的凋亡及自噬），同样，胶质细胞在青光眼模型SC的表达较对照组增强（P < 0.05）。研究提示了circXPO5通过抑制miR-330-5p的表达而调控GRIN2A的水平，进而影响视觉中枢神经元的状态，同时本研究也发现青光眼模型视觉中枢胶质细胞反应增强，神经细胞发生凋亡和自噬活性的升高，该研究结果为青光眼视觉中枢损伤的治疗提供新的研究思路。