Objective: To investigate the effect of microRNA (miR-148b) targeting decoy receptor 3 (DcR3) on macrophage polarization in sepsis. Methods: Experimental study. From December 2019 to December 2022, serum microRNA expression was detected in 3 patients with sepsis and 3 healthy controls in the clinical laboratory of Songjiang Hospital Affiliated to Shanghai Jiao Tong University School of Medicine. Phorbol 12-myristate 13-acetate (PMA) was used to induce the differentiation of human acute monocytic leukemia cells THP-1 into macrophages, and then lipopolysaccharide (LPS) was added to stimulate the establishment of a sepsis cell model, and the expression changes of miR-148b and DcR3 were detected by RT-PCR and Western blot. Overexpression of DcR3 was used to detect the expression levels of TNF-α, CD163 and IL-10 in macrophages stimulated by LPS (100 ng/ml). Overexpression of miR-148b was used to observe the changes of molecular markers of macrophage polarization. The targeting regulation effect of miR-148b on DcR3 was determined by dual-luciferase reporter assay. t test was used to analyze whether there were statistical differences among the groups. Results: The expression of miR-148b was down-regulated (P<0.05) and the expression of DcR3 was up-regulated (P<0.01) in THP-1 macrophages stimulated by LPS. Overexpression of DcR3 inhibited the expression of TNF-α (P<0.05) and promoted the expression of CD163 (P<0.01) and IL-10 (P<0.01). When miR-148b mimics was added, the opposite effect was observed. The dual-luciferase reporter assay confirmed that miR-148b targets and binds to DcR3, inhibiting its transcription and expression. The results of flow cytometry showed that DcR3 could reverse the promoting effect of miR-148b on the CD86/CD163 ratio of macrophages (P<0.05). Conclusion: miR-148b inhibits the expression of DcR3, thereby inhibiting M2 polarization in LPS-stimulated macrophage cells.
目的： 探讨脓毒症中微小RNA-148b（miR-148b）靶向抑制诱骗受体3（DcR3）对巨噬细胞极化的影响。 方法： 2019年12月至2022年12月期间，在上海交通大学医学院附属松江医院（筹）检验科，检测3例脓毒症患者及3名健康对照血清microRNA表达。采用佛波酯（PMA）诱导人急性单核细胞白血病细胞THP-1分化为巨噬细胞，继而加入脂多糖（LPS）刺激建立脓毒症细胞模型，检测miR-148b和DcR3表达变化。过表达DcR3检测LPS（100 ng/ml）刺激巨噬细胞TNF-α、CD163及IL-10的表达水平。过表达miR-148b观察巨噬细胞极化分子标志物的变化。采用双荧光素酶报告基因实验测定miR-148b对DcR3的靶向调控作用。通过t检验分析各组间是否具有统计学差异。 结果： LPS刺激THP-1巨噬细胞miR-148b表达下调（P<0.05），DcR3表达升高（P<0.01）；过表达DcR3抑制TNF-α的表达（P<0.05），促进CD163（P<0.01）和IL-10（P<0.01）的表达，而加入miR-148b模拟物则相反；双荧光素酶报告基因实验证实miR-148b靶向结合DcR3，抑制其转录和表达，流式细胞检测结果显示DcR3可逆转miR-148b对巨噬细胞CD86/CD163比值的促进作用（P<0.05）。 结论： miR-148b靶向抑制DcR3的表达，从而抑制LPS诱导的巨噬细胞模型M2极化。.