内源性二氧化硫通过次磺化修饰核转录因子κB(NF-κB)p65抑制小鼠视网膜光感受器细胞凋亡的研究

Photoreceptor apoptosis is an important pathological basis for irreversible vision loss caused by age-related macular degeneration. The study of its unclear regulatory mechanism is of great significance for the prevention and treatment of age-related macular degeneration. In our previous study, we found the existence of endogenous sulfur dioxide, known as a famous gasotransmitter, and it’s synthetize pathway in mouse photoreceptor cells. On this basis, this project intends to use light-induced and oxidative stress-induced retinal photoreceptor damage as a model to observe the changes in endogenous sulfur dioxide pathway during photoreceptor apoptosis; use overexpression or knockout of sulfur dioxide synthetize enzymme gene to intervene sulfur dioxide production at the animal and cellular levels to explore the regulation effect of endogenous sulfur dioxide on photoreceptor apoptosis. Furthermore, taking NF-κB p65 as a target, we intend to use chemically modified probes, gene-directed mutagenesis and mutational gene knock-in to explore whether endogenous sulfur dioxide may down-regulates NF-κB gene binding activity and nuclear transcription level and therefore inhibits photoreceptor apoptosis via sulfenylating NF-κB p65 at cystein 38 both in vivo and in vitro. This study will reveal the regulation and molecular mechanism of endogenous sulfur dioxide pathway on photoreceptor apoptosis, and provide a new target for photoreceptor cell protection of age-related macular degeneration.

光感受器细胞凋亡是年龄相关性黄斑变性导致不可逆视功能损伤的重要病理基础，深入阐明其尚不明确的调控机制，对年龄相关性黄斑变性的防治具有重要意义。申请者前期研究发现小鼠光感受器细胞内存在气体信号分子二氧化硫调控体系。在此基础上，本项目拟以光损伤及氧化应激诱导视网膜光感受器细胞损伤为模型，观察光感受器细胞凋亡过程中内源性二氧化硫体系变化；通过二氧化硫合成酶AAT基因过表达或敲除，干预二氧化硫生成，探究内源性二氧化硫对光感受器细胞凋亡的调控作用。进而以NF-κB p65为靶点，通过化学修饰探针、基因定点突变和突变基因敲入，从动物水平及细胞水平揭示内源性二氧化硫通过次磺化修饰NF-κB p65第38位半胱氨酸，下调其基因结合活性及核转位水平，进而抑制光感受器细胞凋亡。本研究将深入阐明内源性二氧化硫体系对光感受器细胞凋亡的调控作用及分子机制，为年龄相关性黄斑变性中的光感受器细胞损伤保护提供新靶点。

年龄相关性黄斑变性(AMD)是我国主要的致盲性眼病之一。视网膜光感受器细胞凋亡是AMD 患者最终发生视力损害的主要病理改变，阐明其发生机制已经成为AMD等退行性视网膜疾病研究领域的重要科学问题。本项目从气体信号分子二氧化硫（SO2）切入，研究发现小鼠视网膜组织及视网膜光感受器细胞中可检测到内源性SO2及其生成关键酶AAT的表达；H2O2诱导视网膜光感受器细胞凋亡模型中内源性SO2/AAT体系下调，补充SO2供体可抑制视网膜光感受器细胞凋亡；TNF-α诱导视网膜光感受器细胞凋亡模型中内源性SO2/AAT体系上调，AAT抑制剂HDX可加重视网膜光感受器细胞凋亡；内源性SO2通过对caspase-3的Cys163位点次磺化修饰，抑制caspase-3活性，进而抑制视网膜光感受器细胞凋亡。综上，本项目研究成果揭示了内源性SO2是视网膜光感受器细胞的重要保护因子，可拮抗H2O2及TNF-α损伤因素诱导的细胞凋亡，并阐明了内源性SO2的保护机制。研究成果有望为改善AMD病变、拮抗视网膜光感受器细胞损伤提供新思路和新策略。围绕上述研究成果，发表SCI论文1篇，项目负责人晋升中级职称。