miR-548a-3p通过调节NLRP3炎性小体在脓毒症致ARDS中的机制研究

Study of the mechanism of miR-548a-3p in sepsis-induced ARDS by regulating NLRP3 inflammasome

目的 探究微小RNA(miR)-548a-3p通过调节NLRP3炎性小体在脂多糖诱导脓毒症致急性呼吸窘迫综合征(ARDS)中的机制。方法 选取45只小鼠按随机数表法分为对照组、ARDS组和ARDS+miR-548a-3p agomir组,每组各15只,通过鼻腔内滴入6 mg/kg的脂多糖至肺构建ARDS模型。为提高miR-548a-3p的水平,腹腔内注射miR-548a-3p agomir(300μg),检测各组的HE染色情况、肺组织损伤评分、白细胞介素(IL)-1β、IL-18、NLRP3蛋白水平。通过双荧光素酶报告验证miR-548a-3p和NLRP3的靶向关系。通过转染miR-548a-3p mimic过表达miR-548a-3p,通过RT-qPCR和Western blot检测各组NLRP3 mRNA和蛋白水平以及IL-1β和IL-18水平。结果 ARDS组的肺组织损伤评分、IL-1β、IL-18和NLRP3蛋白表达水平均显著高于对照组和ARDS+miR-548a-3p agomir组,差异有统计学意义(P <0.05)。双荧光素酶报告实验证实NLRP3是miR-548a-3p的靶基因,miR-548a-3p mimic组细胞的NLRP3 mRNA和蛋白、IL-1β/pro-IL-1β和IL-18的表达水平低于miR-548a-3p NC组,差异有统计学意义(P <0.05)。结论 miR-548a-3p可通过靶向抑制NLRP3蛋白抑制NLRP3炎性小体的激活,从而缓解ARDS炎性损伤。

Objective In acute respiratory distress syndrome (ARDS), the mechanism of microRNA (miR)-548a-3p is explored caused by lipopolysaccharide-induced sepsis by regulating NLRP3 inflammasome. Methods A total of 45 mice were divided into a control group, an ARDS group and an ARDS+miR-548a-3p agomir group according to the random number table, with 15 mice in each group. The ARDS model was established by intranasal instillation of 6 mg/kg lipopolysaccharide into the lung, and the level of miR-548a-3p was increased by intraperitoneal injection of miR-548a-3p agomir (300 μg). HE staining, lung tissue injury score, interleukin-1β, IL-18 and NLRP3 protein levels were detected in each group. Dual-luciferase reports the targeting relationship between miR-548a- 3p and NLRP3 was confirmed by using them. miR-548a-3p was overexpressed after transfection of miR-548a- 3p mimic. NLRP3 as well as IL-1β and IL-18 levels, were detected by RT-qPCR and Western blot. Results The lung tissue injury score, IL-1β, IL-18 Compared with the control group and the ARDS+miR-548a-3p, the ARDS group had significantly elevated levels of NLRP3 protein expression agomir group, with statistically significant differences (P < 0.05). Dual-luciferase reports indicate that miR-548a-3p can target NLRP3. The expression levels of NLRP3 mRNA and protein, IL-1β/pro-IL-1β and IL-18 in cells of the miR-548a-3p mimic group were significantly lower than those of the miR-548a-3p NC group, with statistically significant differences (P < 0.05). Conclusion miR-548a-3p can inhibit the activation of NLRP3 inflammasome by targeting the inhibition of NLRP3 protein, the inflammatory injury of ARDS was mitigated by doing so.