基于RNA-seq技术探讨硫酸多糖JCS1S2治疗氧诱导视网膜病变相关机制  

Exploring the mechanisms related to the treatment of oxygen-induced retinopathy by sulfated polysaccharide JCS1S2 based on RNA-seq technology

作　　者：白杨 王瑞 王秋元 丁侃 蔡善君 Bai Yang;Wang Rui;Wang Qiuyuan;Ding Kan;Cai Shanjun(Department of Ophthalmology,Affiliated Hospital of Zunyi Medical University,Zunyi Guizhou 563000,China;Key Laboratory of Receptor Research,CAS Shanghai Institute of Materia Medica,Chinese Academy of Sciences,Shanghai 201203,China)

机构地区：[1]遵义医科大学附属医院眼科,贵州遵义563000 [2]中国科学院CAS上海药物研究所受体研究重点实验室,上海201203

出　　处：《遵义医科大学学报》2025年第3期243-251,共9页Journal of Zunyi Medical University

基　　金：国家自然科学基金资助项目(NO:31871261);贵州省科技支撑计划项目(NO:黔科合支撑[2023]265)。

摘　　要：目的 应用高通量转录组测序(RNA-seq)技术检测氧诱导视网膜病变(OIR)大鼠眼球组织基因表达变化,探索硫酸多糖JCS1S2可能的作用机制,并构建相对缺氧细胞模型对通路进行验证。方法 27只SD幼鼠随机分为正常对照组、OIR组和OIR+JCS1S2组,每组9只。对照组与OIR组玻璃体腔内注射4μL PBS,OIR+JCS1S2组玻璃体腔内注射4μL(0.08 mg) JCS1S2,17 d处死幼鼠后行苏木精-伊红(HE)与视网膜铺片进行分析;利用RNA-seq技术筛选出OIR发病基因及相关通路,并体外实验筛选相关蛋白对通路进行验证。结果 形态学观察显示:与正常对照组相比,OIR组幼鼠见大量血管内皮细胞核突破内界膜、无灌注区面积增多,而添加了JCS1S2的OIR组中只有少量的血管内皮细胞核,血管迂曲明显减轻,无灌注区面积减少(P<0.05)。转录组测序结果:时序分析发现通过上调、下调相关基因的表达,通过神经活性配体-受体相互作用,激活血管平滑肌的收缩,药物代谢途径、Toll和IMD信号通路显著表达,提示JCS1S2S药物治疗可能通过激活这些通路来达到治疗疾病的目的。构建细胞模型发现,其机制可能与抑制血清Toll样受体4(TLR4)、p-NF-κB和VEGF蛋白表达有关。结论 JCS1S2可通过降低缺氧对TLR4/p-NF-κB通路的诱导作用,下调VEGF的合成和分泌,从而延缓或阻滞病理性血管重塑。Objective High throughput transcriptome sequencing(RNA-seq)was used to detect the changes of gene expression in eyeball tissue of rats with oxygen-induced retinopathy(OIR),to explore the possible mechanism of sulfated polysaccharide JCS1S2 treatment,and to construct a relative hypoxic cell model to verify the pathway.Methods 27 SD suckling rats were randomly divided into three groups,normal control group,OIR group,and OIR+JCS1S2 group,with 9 rats each,OIR+JCS1S2 group received intravitreal injection 4μl(0.08 mg)JCS1S2.Rats were euthanized on day 17 and eyeball was stained with hematoxylin eosin(HE)and retinal flat-mount preparation analyzed.RNA-seq was used to screen genes and related pathways involved in OIR pathogenesis.Target proteins were screened in vitro to verify the pathways.Results Morphological observation showed that compared with the normal control group,a large number of vascular endothelial nuclei broke through the inner boundary membrane and the area of non-perfusion area increased in OIR group,while only a small number of vascular endothelial nuclei were found in OIR group treated with JCS1S2(P<0.05).Transcriptome sequencing results showed that vascular smooth muscle contraction was activated by up-regulation and down-regulation of related genes,neuroactive ligand-receptor interaction,and significant expression of drug metabolic pathway,Toll and IMD signal pathways,suggesting that JCS1S2S drug therapy may achieve the purpose of treating diseases by activating these pathways.Constructing a cell model revealed that its mechanism may be related to the inhibition of serum Toll like receptor 4(TLR4),p-NF-κB,and VEGF protein expression.Conclusion JCS1S2S can reduce the induction of TLR4/p-NF-κB pathway by hypoxia and down-regulate the synthesis and secretion of VEGF,thus delaying or blocking pathological vascular remodeling.

关 键 词：氧诱导视网膜病变 硫酸多糖JCS1S2 转录组测序 大鼠模型 MÜLLER细胞 

分 类 号：R774.1[医药卫生—眼科]