摘要
目的:观察左卡尼汀对过氧化氢(H2O2)诱导的大鼠心肌细胞凋亡的保护作用及其机制。方法:利用200μmol/L H2O2刺激12 h,建立体外原代培养新生乳鼠心肌细胞凋亡模型。Ca2+螯合剂1,2-双(2-氨基苯氧基)乙烷-N,N,N',N'-四乙酸(BAPTA)、钙调素依赖蛋白激酶II(CaMKII)特异性抑制剂KN93及左卡尼汀分别于加入H2O2前30 min或1 h加入,以检测这3种药物对H2O2刺激下心肌细胞活力、细胞凋亡、细胞内静息钙浓度([Ca2+]i)及磷酸化CaMKII(p-CaMKII)表达的影响。利用MTT比色法检测心肌细胞活力;流式细胞仪检测细胞凋亡率;利用激光共聚焦扫描检测[Ca2+]i;蛋白质免疫印迹法检测cleaved caspase-3及p-CaMKII的表达。结果:模型组经200μmol/L H2O2作用12 h后,细胞活力显著下降,细胞凋亡率显著增加。BAPTA、KN93及左卡尼汀预处理显著抑制上述细胞损伤。进一步研究发现,H2O2诱导的[Ca2+]i水平升高、cleaved caspase-3及p-CaMKII的表达增加均可被上述3种药物不同程度地抑制。结论:左卡尼汀可抑制H2O2所致的心肌细胞凋亡,该心肌保护作用可能与其抑制Ca2+/CaMKⅡ信号通路有关。
AIM： To examine the inhibitory effect of L-carnitine on hydrogen peroxide（H2O2）-induced apoptosis of rat cardiomyocytes and to further explore the underlying mechanisms.METHODS： Primarily cultured neonatal rat myocardial cells were prepared and challenged by 200 μmol / L H2O2 to induce cell apoptosis.In order to evaluate the effects of Ca2 ＋ chelator 1,2-bis（2-aminophenoxy） ethane-N,N,N＇,N＇-tetraacetic acid（BAPTA）,calmodulin-dependent protein kinase II（CaMKII） inhibitor KN93 and L-carnitine on cell viability,apoptosis,resting intracellular free Ca2 ＋ concentration（[Ca2 ＋]i） and phospho-CaMKII（p-CaMKII） expression,these three agents were added 30 min or 1 h prior to H2O2 stimulation.Cell viability was measured by MTT assay and apoptosis was determined by flow cytomertry.The [Ca2 ＋]i was measured by laser confocal scanning.Cleaved caspase-3 and p-CaMKII expression was detected using Western blotting.RESULTS： Upon 200 μmol / L H2O2 stimulation for 12 h,cell viability decreased and apoptotic rate increased significantly compared with control.Pretreament with L-carnitine,BAPTA and KN93 significantly increased cell viability and decreased apoptosis.Furthermore,intracellular Ca2 ＋ overload triggered by H2O2 could be greatly relieved by L-carnitine and BAPTA pretreatment,but not affected by KN93.H2O2-stimulated cleaved caspase-3 and p-CaMKII expression was also significantly inhibited by all these three agents.CONCLUSION： L-carnitine inhibits H2O2-induced rat cardiomyocyte apoptosis possibly via suppressing Ca2 ＋ / CaMKII signaling pathway.
出处
《中国病理生理杂志》
CAS
CSCD
北大核心
2013年第7期1250-1254,共5页
Chinese Journal of Pathophysiology
基金
辽宁省高等学校优秀人才支持计划(No.2008RC33)
关键词
肉碱
过氧化氢
心肌细胞
细胞凋亡
钙
钙调素依赖蛋白激酶Ⅱ
Carnitine
Hydrogen peroxide
Cardiomyocytes
Apoptosis
Calcium
Calmodulin-dependent protein kinase Ⅱ
