Wang Zhijie, Huang Yinhua, Chu Feixue, Ji Shangli, Liao Kai, Cui Zekai, Chen Jiansu, Tang Shibo
Aier School of Ophthalmology, Central South University, Changsha, People's Republic of China.
Aier Eye Institute, Aier Eye Hospital Group, Changsha, People's Republic of China.
J Inflamm Res. 2021 Nov 11;14:5901-5918. doi: 10.2147/JIR.S326091. eCollection 2021.
Retinal inflammation is involved in the pathogenesis of several retinal diseases. As one of the core clock genes, has been reported to suppress inflammation in many diseases. We investigated whether pharmacological activation of can inhibit retinal inflammation and delineated the mechanisms of in alleviating microglia activation.
Lipopolysaccharide (LPS) induced mice models were used to examine the effects of SR9009 (agonist of NR1D1) treatment on inflammatory phenotypes in vivo. Anti-inflammatory effects of and associated mechanisms were investigated in the BV2 microglia cell line, and in primary retinal microglia in vitro.
SR9009 treatment alleviated LPS-induced inflammatory cell infiltration, elevated cytokine levels and morphological changes of the microglia in mice models. In LPS-stimulated BV2 cells and primary retinal microglia, SR9009 suppressed cytokine expressions by inhibiting the NF-κB signaling pathway. Moreover, SR9009 treatment increased the levels of the M2 phenotype marker (CD206) and the proportions of ramified microglia. Suppression of with siRNA reversed the inhibitory effects of SR9009 on cytokine production in BV2 cells. RNA-seq analysis showed that genes that were upregulated following knockdown were enriched in inflammatory-associated biological processes. Subsequently, ChIP-seq of NR1D1 in BV2 was performed, and the results were integrated with RNA-seq results using the Binding and Expression Target Analysis (BETA) tool. Luciferase assays, electrophoretic mobility shift assay (EMSA), qPCR and Western blotting assays revealed that NR1D1 binds the promoter of to suppress its transcription. Notably, overexpressed in activated microglia could partly abolish the anti-inflammatory effects of .
The clock gene protects against retinal inflammation and microglia activation in part by suppressing transcription.
视网膜炎症参与多种视网膜疾病的发病机制。作为核心生物钟基因之一，据报道在许多疾病中具有抑制炎症的作用。我们研究了NR1D1的药理学激活是否能抑制视网膜炎症，并阐明了NR1D1减轻小胶质细胞激活的机制。
使用脂多糖（LPS）诱导的小鼠模型来检测SR9009（NR1D1激动剂）治疗对体内炎症表型的影响。在BV2小胶质细胞系和原代视网膜小胶质细胞中体外研究NR1D1的抗炎作用及相关机制。
SR9009治疗减轻了LPS诱导的小鼠模型中的炎症细胞浸润、细胞因子水平升高和小胶质细胞的形态变化。在LPS刺激的BV2细胞和原代视网膜小胶质细胞中，SR9009通过抑制NF-κB信号通路抑制细胞因子表达。此外，SR9009治疗增加了M2表型标志物（CD206）的水平和分支状小胶质细胞的比例。用siRNA抑制NR1D1可逆转SR9009对BV2细胞中细胞因子产生的抑制作用。RNA测序分析表明，NR1D1敲低后上调的基因富集于炎症相关的生物学过程。随后，对BV2中的NR1D1进行染色质免疫沉淀测序（ChIP-seq），并使用结合和表达靶标分析（BETA）工具将结果与RNA测序结果整合。荧光素酶测定、电泳迁移率变动分析（EMSA）、qPCR和蛋白质免疫印迹分析表明，NR1D1与[基因名称未提及]的启动子结合以抑制其转录。值得注意的是，在活化的小胶质细胞中过表达[基因名称未提及]可部分消除NR1D1的抗炎作用。
生物钟基因NR1D1部分通过抑制[基因名称未提及]转录来预防视网膜炎症和小胶质细胞激活。
