To investigate the mechanism of action of Srg3 in acute lung injury caused by sepsis. First, a sepsis-induced acute lung injury rat model was established using cecal ligation and puncture (CLP). RNA sequencing (RNA-seq) was used to screen for highly expressed genes in sepsis-induced acute lung injury (ALI), and the results showed that Srg3 was significantly upregulated. Then, SWI3-related gene 3 (Srg3) was knocked down using AAV9 vector in vivo, and changes in ALI symptoms in rats were analyzed. In vitro experiments were conducted by establishing a cell model using lipopolysaccharide (LPS)-induced BEAS-2B cells and coculturing them with phorbol 12-myristate 13-acetate (PMA)-treated THP-1 cells to analyze macrophage polarization. Next, downstream signaling pathways regulated by Srg3 and transcription factors involved in regulating Srg3 expression were analyzed using the KEGG database. Finally, gain-of-loss functional validation experiments were performed to analyze the role of downstream signaling pathways regulated by Srg3 and transcription factors involved in regulating Srg3 expression in sepsis-induced acute lung injury. Srg3 was significantly upregulated in sepsis-induced acute lung injury, and knocking down Srg3 significantly improved the symptoms of ALI in rats. Furthermore, in vitro experiments showed that knocking down Srg3 significantly weakened the inhibitory effect of LPS on the viability of BEAS-2B cells and promoted alternative activation phenotype (M2) macrophage polarization. Subsequent experiments showed that Srg3 can regulate the activation of the NF-κB signaling pathway and promote ferroptosis. Specific activation of the NF-κB signaling pathway or ferroptosis significantly weakened the effect of Srg3 knockdown. It was then found that Srg3 can be transcriptionally activated by interferon regulatory factor 7 (Irf7), and specific inhibition of Irf7 significantly improved the symptoms of ALI. Irf7 transcriptionally activates the expression of Srg3, which can promote ferroptosis and activate classical activation phenotype (M1) macrophage polarization by regulating the NF-κB signaling pathway, thereby exacerbating the symptoms of septic lung injury.

中文翻译：

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Irf7调节Srg3表达及铁死亡轴加重脓毒症急性肺损伤

探讨Srg3在脓毒症所致急性肺损伤中的作用机制。首先，采用盲肠结扎穿刺（CLP）建立脓毒症诱导的急性肺损伤大鼠模型。利用RNA测序（RNA-seq）筛选脓毒症引起的急性肺损伤（ALI）中高表达的基因，结果显示Srg3显着上调。然后，使用AAV9载体在体内敲低SWI3相关基因3（Srg3），并分析大鼠ALI症状的变化。通过使用脂多糖（LPS）诱导的BEAS-2B细胞建立细胞模型并将其与佛波醇12-肉豆蔻酸酯13-乙酸酯（PMA）处理的THP-1细胞共培养来进行体外实验以分析巨噬细胞极化。接下来，利用KEGG数据库分析Srg3调控的下游信号通路以及参与调控Srg3表达的转录因子。最后，进行增益-损失功能验证实验，分析Srg3调节的下游信号通路和参与调节Srg3表达的转录因子在脓毒症诱导的急性肺损伤中的作用。Srg3 在脓毒症引起的急性肺损伤中显着上调，敲除 Srg3 可以显着改善大鼠的 ALI 症状。此外，体外实验表明，敲低Srg3显着削弱了LPS对BEAS-2B细胞活力的抑制作用，并促进替代激活表型（M2）巨噬细胞极化。后续实验表明，Srg3可以调节NF-κB信号通路的激活，促进铁死亡。NF-κB信号通路的特异性激活或铁死亡显着削弱了Srg3敲低的效果。随后发现Srg3可以被干扰素调节因子7（Irf7）转录激活，特异性抑制Irf7可以显着改善ALI的症状。Irf7转录激活Srg3的表达，Srg3可通过调节NF-κB信号通路促进铁死亡并激活经典激活表型（M1）巨噬细胞极化，从而加剧脓毒性肺损伤的症状。