UCP2 基因siRNA 转染对脓毒症血清诱导心肌细胞炎症反应的影响

Effect of UCP2-siRNA on inflammatory response of cardiomyocytes induced by septic serum

摘要:

目的 研究解偶联蛋白2 基因（uncoupling protein 2, UCP2）siRNA 干扰大鼠H9C2 心肌细胞株后对脓毒症血清诱导炎症反应的影响，探讨UCP2 在脓毒症心肌病可能的机制。方法 制备正常大鼠血清和脓毒症大鼠血清。体外培养大鼠心肌细胞株（H9C2），随机分为空白对照组、正常大鼠血清、10%脓毒症大鼠血清刺激12 h 组（脓毒症血清组）、UCP2-siRNA 干扰+10%脓毒症大鼠血清刺激12 h 组（UCP2-siRNA+ 脓毒症血清组）、阴性siRNA 转染+10%脓毒症大鼠血清刺激12 h 组（阴性siRNA+ 脓毒症血清组）。RT-PCR 检测各组TNF-α mRNA，IL-1βmRNA 表达；免疫印迹法检测磷酸化p38MAPK（p-p38MAPK）表达和核转录因子（NF-κB）的表达。结果 UCP2-siRNA+ 脓毒症血清组 p-p38、NF-κB 表达量均较脓毒症血清组明显增高（P<0.05）；UCP2-siRNA+ 脓毒症血清组TNF-α mRNA 表达量与脓毒症血清组相比明显增加（P<0.01），IL-1βmRNA 表达量与脓毒症血清组比较差异无统计学意义。结论 UCP2 参与脓毒症血清诱导的H9C2 心肌细胞p38MAPK 活性和NF-κB 与下游炎症介质的表达调控。

Abstract:

Objective To study the effect of uncoupling protein 2 (UCP2)-siRNA on the inflammatory response of rat cardiomyocytes (H9C2) induced by septic serum and to investigate the possible role of UCP2 in the development of septic cardiomyopathy. Methods Serum samples were separately collected from normal rats and septic rats. Cultured rat cardiac cells (H9C2) were randomly divided into blank control, normal serum, 10% septic serum, UCP2-siRNA+10% septic serum and negative siRNA+10% septic serum groups. Stimulation with 10% septic serum was performed for 12 hours in relevant groups. The mRNA expression of tumor necrosis factor-alpha (TNF-α) and interleukin-1 beta (IL-1β) was measured by RT-PCR. The expression of phosphorylated p38 mitogen-activated protein kinase (p-p38 MAPK) and nuclear factor-kappa B (NF-κB) was measured by Western blot. Results The expression levels of p-p38 and NF-κB in the UCP2-siRNA+10% septic serum group were significantly higher than in the 10% septic serum group (P<0.05). The UCP2-siRNA+10% septic serum group had a significantly higher TNF-α mRNA expression than the 10% septic serum group (P<0.01), but IL-1β mRNA expression showed no significant difference between the two groups. Conclusions UCP2 plays a regulatory role in the activation of p38 MAPK and NF-κB and the expression of downstream inflammatory mediators in H9C2 cells stimulated with septic serum.