地塞米松对内毒素诱导的小鼠葡萄膜炎的治疗效果及转录组分析

摘要

Objective To investigate the changes in retinal transcriptome profile of mice with endotoxin-induced uveitis (EIU) following dexamethasone (DEX) treatment and explore the mechanisms underlying the therapeutic effect of DEX.Methods EIU was induced in BALB/c mice by intravitreal injection of 125 ng lipopolysaccharide (LPS),followed by topical application of DEX (0.1％) eye drops every 4 h for 24 h.The anterior chamber inflammation was examined with a slit lamp and the clinical scores were assessed.The morphological changes in the eyes were assessed at 24 h after LPS injection.The retinas were harvested for analysis of transcriptome profile using the next-generation sequencing (NGS)-based RNA sequencing (RNA-seq),and the expressions of the inflammatory cytokines and the differentially expressed genes (DEGs) were verified using real-time PCR.Results DEX alleviated the inflammatory response and reduced the mRNA expressions of IL-6,TNF-α,MCP-1 and ICAM-1 at 24 h after LPS injection.A total of 52 DEGs were identified by RNA-seq.Within these DEGs,37 genes were upregulated and 15 genes were down-regulated in LPS group as compared with DEX+LPS group.No significantly enriched Gene Ontology (GO) terms was noted.Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis showed 6 up-regulated and 2 down-regulated KEGG pathways.RIG-I-like receptor signaling pathway and several immune-and inflammation-related genes including Ifit1,H2-T24,Mx2 and Eif2ak2 were significantly down regulated by DEX.Verification with RT-PCR yielded results consistent with these findings.Conclusion DEX alleviates LPS-induced inflammatory response in the retina of mice,and such protective effect is probably mediated by RIG-I like receptor signal pathway and the immune-and inflammation-related genes.%目的 研究地塞米松(DEX)对内毒素诱导性葡萄膜炎(EIU)的治疗效果及转录组测序探讨其潜在的机制.方法 将125ng脂多糖(LPS)注射入雌性BALB/c小鼠玻璃体内建立EIU模型,每隔4h用0.1％ DEX滴眼,共6次.注射LPS后24h用裂隙灯观察小鼠眼前房炎症情况并进行临床评分.苏木精-伊红染色法观察小鼠眼部形态学变化.RNA-seq对EIU模型组和DEX治疗组小鼠的视网膜进行转录组测序并进行GO和KEGG功能分析.Real-time PCR检测小鼠视网膜炎症细胞因子表达及验证选出的差异基因.结果 连续滴眼6次后,DEX可减轻EIU前房的炎症,并下调炎症因子白介素-6(IL-6)、肿瘤坏死因子-α(TNF-α)、单核细胞趋化蛋白-I(MCP-1)和细胞间黏附分子-1(ICAM-1)mRNA的表达.RNA-seq共检测出52个差异基因,与DEX+LPS组相比,LPS组中上调的基因有37个,下调的基因有15个.差异基因的功能分析未发现显著富集的GO条目.KEGG富集分析显示有6个显著上调的KEGG通路和2个显著下调的KEGG通路.KEGG结果提示DEX治疗后可下调RIG-I类受体信号通路(Ddx58、Ifihl和Isgl5)及一些免疫炎症相关基因(Ifitl、H2-T24、Mx2和Eif2ak2).结论DEX对LPS引起的小鼠内毒素诱导性小鼠葡萄膜炎有保护作用,此保护作用可能与下调RIG-I类受体信号通路及一些免疫炎症相关基因有关.