NO介导TNF-α诱导的培养大鼠缺氧/复氧心肌细胞的保护作用

Nitric oxide-mediated the cardioprotection of tumor necrosis factor-alpha on cultured neonatal rat cardiomyocytes during hypoxia/reoxygenation

目的 :研究一氧化氮 (NO)在肿瘤坏死因子α(TNF -α)诱导的缺氧 /复氧心肌细胞的保护作用。方法 :在培养的大鼠乳鼠心肌细胞上 ,建立缺氧 /复氧模型 ,测定复氧后培养液中乳酸脱氢酶 (LDH)的释放量和细胞内锰超氧化物歧化酶 (Mn -SOD)活性。结果 :(1)用TNF -α(10 5U/L)预处理 ,缺氧 /复氧后心肌细胞内Mn -SOD活性增高、LDH释放量减少 ;(2 )NO供体硝普钠 (SNP) (5 μmol/L)和前体L -精氨酸 (L -Arg) (5mmol/L)预处理心肌细胞3h ,缺氧 /复氧后细胞内Mn -SOD活性增高、LDH释放量减少 ,用Nω -硝基 -L -精氨酸甲酯 (L -NAME) (10 0 μmol/L)预处理减弱了TNF -α的抑制缺氧 /复氧心肌细胞LDH释放和诱导Mn -SOD活性增高的作用 ;(3)可溶性鸟苷酸环化酶 (sGC)抑制剂ODQ(10 μmol/L)和CPK抑制剂chelerythrine (5 μmol/L)可分别减弱SNP、L -Arg和TNF -α对缺氧/复氧心肌细胞的保护作用 ;(4 )TNF -α对缺氧 /复氧心肌细胞的保护作用可被抗氧化剂 2 -巯基丙酰氨基乙酸 (2 -MPG ,4 0 0 μmol/L)削弱 ,但是酪氨酸蛋白激酶 (TK)抑制剂 4 ,5 ,7-三羟基异黄酮 (genistein ,5 0 μmol/L)预处理对TNF-α诱导的心肌缺氧 /复氧保护无影响。结论 :NO参与TNF -α对缺氧 /复氧损伤心肌细胞的保护作用 。

AIM: To investigate the role of nitric oxide synthase (NOS), soluble guanylyl cyclase (sGC) and protein kinase C (PKC) signaling in tumor necrosis factor-α (TNF-α)-induced cardioprotection against hypoxia/reoxygenation (H/R) injury. METHODS: Neonatal rat ventricular myocytes were pretreated with TNF-α or sodium nitroprusside (SNP) or L-arginine (L-Arg), respectively, for 12 h and then subjected to continuous hypoxia for 12 h, followed by reoxygenation for 6 h. The manganese superoxide dismutase (Mn-SOD) activity of the cells was measured after H/R. Myocyte injury was determined by the release of lactic dehydrogenase (LDH). RESULTS: TNF-α (10~5 (U/L)) significantly increased the Mn-SOD activity and decreased release of LDH from ventricular myocytes. The cardioprotection against H/R injury was induced by the pretreatment with SNP (5 μmol/L) or L-Arg (5 mmol/L), which was blocked by ODQ (10 μmol/L), the specific sGC inhibitor, and Chel (5 μmol/L), the specific PKC inhibitor. Pretreatment with L-NAME (100 μmol/L), ODQ, Chel, antoxidant 2-MPG (400 μmol/L) or tyrosine kinase inhibitor genistein (50 μmol/L) attenuated the increased Mn-SOD activity and reduced LDH level induced by TNF-α. CONCLUSION: The results suggest that NO may play a role in TNF-α-induced cardioprotection, which is mediated by sGC and PKC. [