Emergency Department, Shenzhen Second People's Hospital, Shenzhen University First Affiliated Hospital, No.3002 Sungang West Road, Shenzhen, 518035, Guangdong, People's Republic of China.
Inflammation. 2021 Oct;44(5):1771-1781. doi: 10.1007/s10753-021-01453-8. Epub 2021 Apr 13.
Cardiac dysfunction in severe sepsis is associated with increased mortality. However, the molecular mechanisms underlying septic heart dysfunction remain unclear. Expression of peroxisome proliferator-activated receptor-γ coactivator 1α (Pgc-1α), concentrations of inflammatory factors, and activation of the nuclear factor kappa-B (NF-κB) signaling pathway were examined in H9c2 cells after a 24-h lipopolysaccharide (LPS) stimulation period using qPCR, enzyme-linked immunosorbent assays (ELISAs), and western blots (WBs), respectively. Pgc-1α was overexpressed and suppressed in cells using a lentivirus vector and siRNA, respectively. The effects of Pgc-1α dysfunction on the release of inflammatory factors and apoptosis were analyzed. Pgc-1α expression was increased after LPS induction for 0.5 h and returned to the pre-induction level at 2 h. Levels of IL-1β, IL-6, and TNF-α increase after LPS induction for 0.5 h and accumulated in the culture supernatants over time. The WBs revealed the highest Pgc-1α and phospho (p)-p65 protein levels after LPS induction for 0.5 h, followed by a decrease; moreover, the cleaved-caspase-3 level increased after LPS induction for 0.5 h and increased gradually thereafter. A functional analysis of Pgc-1α revealed that overexpression of this protein enhanced LPS-induced inflammatory factors and p-p65 levels and inhibited apoptosis during the early stage after LPS induction (0.5 and 4 h). In contrast, the inhibition of Pgc-1α expression inhibited the LPS expression-associated increases in inflammatory factors and p-p65 and promoted apoptosis. Pgc-1α promoted LPS-induced p65 phosphorylation and inflammatory factor release while inhibiting apoptosis.
严重脓毒症患者的心功能障碍与死亡率增加有关。然而,脓毒性心脏功能障碍的分子机制尚不清楚。分别采用 qPCR、酶联免疫吸附测定(ELISA)和 Western blot(WB)检测 H9c2 细胞在脂多糖(LPS)刺激 24 小时后过氧化物酶体增殖物激活受体-γ辅激活因子 1α(Pgc-1α)的表达、炎症因子浓度和核因子 kappa-B(NF-κB)信号通路的激活。分别使用慢病毒载体和 siRNA 过表达和抑制细胞中的 Pgc-1α。分析 Pgc-1α 功能障碍对炎症因子释放和细胞凋亡的影响。LPS 诱导 0.5 小时后 Pgc-1α 表达增加,2 小时后恢复到诱导前水平。LPS 诱导 0.5 小时后,IL-1β、IL-6 和 TNF-α 的水平增加,并随着时间的推移在培养上清液中积累。WB 显示 LPS 诱导 0.5 小时后 Pgc-1α 和磷酸化(p)-p65 蛋白水平最高,随后下降;此外,LPS 诱导 0.5 小时后 cleaved-caspase-3 水平增加,随后逐渐增加。Pgc-1α 的功能分析表明,该蛋白的过表达增强了 LPS 诱导的炎症因子和 p-p65 水平,并在 LPS 诱导后早期(0.5 和 4 小时)抑制细胞凋亡。相反,抑制 Pgc-1α 表达抑制 LPS 表达相关的炎症因子和 p-p65 增加,并促进细胞凋亡。Pgc-1α 促进 LPS 诱导的 p65 磷酸化和炎症因子释放,同时抑制细胞凋亡。