Temperature and Inflammation Research Center, Key Laboratory of Colleges and Universities in Sichuan Province, Chengdu Medical College, 610500, China.
Zunyi Medical University, Zunyi, Guizhou 563000, China; Department of Cardiology, Sichuan Academy of Medical Sciences and Sichuan Provincial People's Hospital, Chengdu 610072, China.
Life Sci. 2019 Apr 1;222:195-202. doi: 10.1016/j.lfs.2019.02.048. Epub 2019 Feb 23.
To explore the potential mechanism that the role of the Akt/eNOS/NO pathway in calpain-induced caspase-3 and NF-κB activation during septic apoptosis.
Septic rats were stimulated by LPS (8 mg/kg, i.p.). Myocardial calpain, caspase-3, NO, TNF-α and IL-1β levels were detected by ELISA. The levels of Akt/p-Akt, eNOS/p-eNOS, iNOS proteins and number of apoptotic cells were evaluated by immunohistochemistry, western blot and TUNEL method.
Compared with sham, LPS treatment resulted in 4.1-fold and 1.8-fold increases in myocardial calpain activity and caspase-3 activation, respectively, and a significant increase (6.8-fold) in apoptotic cardiomyocytes was observed. The administration of calpain inhibitors (calpain inhibitor-IV, PD150606 and PD151746) showed that p-Akt and p-eNOS protein levels were correlated with the levels of LPS-induced myocardial calpain and caspase-3 activity. In addition, the quantity of p-Akt protein and NO content were markedly attenuated by wortmannin, a phosphoinositide 3-kinase (PI3K) inhibitor. Pretreatment with L-NAME, an NOS inhibitor, induced a decrease in p-eNOS proteins and apoptosis in myocardial tissues, while iNOS proteins were strongly increased in septic rats.
This study suggests that the Akt/eNOS/NO pathway might lead to a novel pharmacological therapy for cardiomyocytes apoptosis in sepsis.
探讨钙蛋白酶诱导的半胱天冬酶-3 和 NF-κB 激活在脓毒症细胞凋亡中 Akt/eNOS/NO 通路的作用机制。
用 LPS（8mg/kg，腹腔注射）刺激脓毒症大鼠。通过 ELISA 检测心肌钙蛋白酶、半胱天冬酶-3、NO、TNF-α 和 IL-1β水平。通过免疫组织化学、western blot 和 TUNEL 方法评估 Akt/p-Akt、eNOS/p-eNOS、iNOS 蛋白水平和凋亡细胞数量。
与假手术组相比，LPS 处理导致心肌钙蛋白酶活性和半胱天冬酶-3 活性分别增加了 4.1 倍和 1.8 倍，凋亡性心肌细胞数量显著增加（增加了 6.8 倍）。钙蛋白酶抑制剂（钙蛋白酶抑制剂-IV、PD150606 和 PD151746）的给药表明，p-Akt 和 p-eNOS 蛋白水平与 LPS 诱导的心肌钙蛋白酶和半胱天冬酶-3 活性水平相关。此外，PI3K 抑制剂wortmannin 显著降低了 p-Akt 蛋白数量和 NO 含量。NOS 抑制剂 L-NAME 预处理可降低心肌组织中 p-eNOS 蛋白和凋亡，而脓毒症大鼠中 iNOS 蛋白水平显著升高。
本研究表明，Akt/eNOS/NO 通路可能为脓毒症中心肌细胞凋亡提供一种新的药理学治疗方法。
