LINC00839调节miR-142-5p/HMGB1轴对LPS诱导的肺泡上皮细胞焦亡的影响

LINC00839 regulates the effect of miR-142-5p/HMGB1 axis on LPS-induced pyroptosis of alveolar epithelial cells

目的探讨LINC00839对miR-142-5p/高迁移率族蛋白1(HMGB1)轴的调节作用,及其对脂多糖(LPS)诱导的肺泡上皮细胞焦亡的影响。方法将人肺泡上皮细胞A549分为Control组、LPS组(10 mg/L的LPS处理)、si-NC组(转染si-NC后用10 mg/L的LPS处理)、si-LINC00839组(转染si-LINC00839后用10 mg/L的LPS处理)、si-LINC00839+inhibitor NC组(转染si-LINC00839和inhibitor NC后用10 mg/L的LPS处理)、si-LINC00839+miR-142-5p inhibitor组(转染si-LINC00839和miR-142-5p inhibitor后用10 mg/L的LPS处理)、si-LINC00839+oe-NC组(转染si-LINC00839和oe-NC后用10 mg/L的LPS处理)、si-LINC00839+oe-HMGB1组(转染si-LINC00839和oe-HMGB1后用10 mg/L的LPS处理)。检测细胞LINC00839、miR-142-5p和HMGB1基因表达、细胞增殖、乳酸脱氢酶(LDH)活性、肿瘤坏死因子-α(TNF-α)、白介素-18(IL-18)和白介素-1β(IL-1β)水平、丙二醛(MDA)含量、过氧化氢酶(CAT)和超氧化物歧化酶(SOD)活性;碘化丙啶(PI)染色法检测细胞膜孔的形成;Western blot检测细胞中HMGB1、核苷酸结合寡聚化结构域样受体蛋白3(NLRP3)、Caspase-1、GSDMD-N、增殖细胞核抗原(PCNA)蛋白表达。结果与Control组相比,LPS组和si-NC组A549细胞LINC00839和HMGB1 mRNA表达、LDH活性、TNF-α、IL-18、IL-1β水平、MDA含量、PI染色阳性率、HMGB1、NLRP3、Caspase-1、GSDMD-N蛋白表达水平升高,miR-142-5p表达、A_(450)(24、48 h)值、SOD和CAT活性、PCNA蛋白表达水平降低,差异均有统计学意义(P均<0.05)。与si-NC组相比,si-LINC00839组A549细胞LINC00839和HMGB1 mRNA表达、LDH活性、TNF-α、IL-18、IL-1β水平、MDA含量、PI染色阳性率、HMGB1、NLRP3、Caspase-1、GSDMD-N蛋白表达水平降低,miR-142-5p表达、A_(450)(24、48 h)值、SOD和CAT活性、PCNA蛋白表达水平升高,差异均有统计学意义(t=18.003、30.209、14.283、9.526、9.242、8.625、8.856、10.386、12.645、10.590、10.943、10.614、11.182、8.492、8.405、7.934、7.436、11.103,P均<0.05)。沉默miR-142-5p表达或上调HMGB1表达均可降低下调LINC00839表达对LPS诱导A549细胞焦亡的改善作用(P<0.05)。结论下调LINC00839可调控miR-142-5p/HMGB1轴减轻LPS诱导的A549细胞焦亡。

Objective To investigate the effect of LINC00839 on the regulation of miR-142-5p/high mobility group protein 1(HMGB1)axis on lipopolysaccharide(LPS)-induced scorch death of alveolar epithelial cells.Methods Human alveolar epithelial cells A549 were divided into control group,LPS group,si-NC group,si-LINC00839 group,si-LINC00839+inhibitor NC group,si-LINC00839+miR-142-5p inhibitor group,si-LINC00839+oe-NC group,si-LINC00839+oe-HMGB1 group.The expression of LINC00839,miR-142-5p and HMGB1 genes,cell proliferation,lactate dehydrogenase(LDH)activity,tumor necrosis factor-α(TNF-α),interleukin-18(IL-18)and interleukin-1β(IL-1β)levels,malondialdehyde(MDA)content,catalase(CAT)and superoxide dismutase(SOD)activity were detected.The formation of cell membrane pores was detected by propyl iodide(PI)staining.Western blot was used to detect HMGB1,NLRP3,Caspase-1,GSDMDN,PCNA protein expression.Results Compared with the control group,the expression levels of LINC00839 and HMGB1 mRNA,LDH activity,TNF-α,IL-18,IL-1β,MDA content,PI staining positive rate,HMGB1,NLRP3,Caspase-1 and GSDMD-N protein in A549 cells of LPS group and si-NC group were increased,the expression of miR-142-5p,A_(450)(24,48 h),SOD and CAT activities,and PCNA protein expression levels were decreased(all P<0.05).Compared with the si-NC group,the expression levels of LINC00839 and HMGB1 mRNA,LDH activity,TNF-α,IL-18,IL-1β,MDA content,PI staining positive rate,HMGB1,NLRP3,Caspase-1 and GSDMD-N protein in A549 cells of si-LINC00839 group were decreased,the expression of miR-142-5p,A_(450)(24,48 h),SOD and CAT activities,and PCNA protein expression levels were increased(t=18.003,30.209,14.283,9.526,9.242,8.625,8.856,10.386,12.645,10.590,10.943,10.614,11.182,8.492,8.405,7.934,7.436,11.103;all P<0.05).Silencing the expression of miR-142-5p or upregulating the expression of HMGB1 could reduce the improvement effect of down-regulating the expression of LINC00839 on LPS-induced pyroptosis in A549 cells(P<0.05).Conclusion Down-regulation of LINC00839 regulated the miR-142-5p/HMGB1 axis to reduce LPS-induced pyroptosis of A549 cells.