摘要
目的探讨硫化氢(H2S)是否通过抑制p38MAPK保护PC12细胞对抗化学性缺氧诱导的损伤。方法应用化学性低氧模拟剂氯化钴(CoCl2)处理PC12细胞建立化学性缺氧损伤模型。应用CCK-8比色法检测细胞存活率;Hochest33258核染色法观察细胞凋亡的形态学改变;双氯荧光素(DCFH-DA)染色荧光显微镜照像检测细胞内的活性氧(ROS);罗丹明123(RH123)染色荧光显微镜照像检测线粒体膜电位(MMP);Western blot法检测p38MAPK蛋白的表达水平。结果应用600μmol·L-1CoCl2处理PC12细胞2h可使磷酸化(p)p38明显增多;在应用600μmol·L-1CoCl2处理PC12细胞前30min,应用400μmol·L-1硫氢化钠(NaHS,H2S的供体)预处理细胞不仅可明显的抑制CoCl2诱导的p-p38MAPK表达的增多,还能保护PC12细胞对抗600μmol·L-1CoCl2引起的损伤,使细胞存活率升高,凋亡细胞和胞内ROS水平明显降低,MMP丢失减小;在CoCl2损伤PC12细胞前60min应用p38抑制剂SB302580(20μmol·L-1)预处理也能产生类似NaHS预处理的细胞保护作用。结论 p38MAPK介导CoCl2引起PC12细胞的损伤作用;H2S通过抑制p38MAPK的表达及氧化应激反应保护PC12细胞对抗化学性缺氧引起的损伤作用。
Aim To investigate whether hydrogen sulfide（H 2 S） protected PC12 cells against chemical hypoxia-induced injury by inhibiting p38 MAPK.Methods PC12 cells were treated with cobalt chloride（CoCl 2 ） to set up a chemical hypoxia-induced cellular injury model.Cell viability was tested by cell counter kit（CCK-8） ;morphological changes of apoptotic cells were detected by Hochest 332580 staining;intracellular level of reactive oxygen species（ROS） was measured by DCFH-DA staining and photofluorograph;mitochondrial membrane potential（MMP） was observed by rhodamine 123（RH123） staining and photofluorography;the expression of p38MAPK was determined by Western blot assay.Results Exposure of PC12 cells to 600μmol· L -1 CoCl2 for 2 h significantly enhanced phosphorylated（p） p38MAPK expression.Pretreatment with 400 μmol·L -1 NaHS（a donor of H 2 S） for 30 min prior to exposure of PC12 cells to 600μmol·L -1 CoCl2 not only inhibited CoCl2 induced increase in expression of p38MAPK,but also protected PC12 cells against injuries induced by 600μmol·L -1 CoCl2,enhancing cell viability and decreasing amount of apoptotic cells and intracellular ROS level,as well as loss of MMP.Similarly,pretreatment with SB302580（20μmol·L-1） ,an inhibitor of p38MAPK,for 60 min prior to exposure of PC12 cells to CoCl2 also conferred the same cytoprotective effect of H2 S.Conclusions p38MAPK mediates CoCl2-induced injuries in PC12 cells.H2 S protects PC12 cells against chemical hypoxia-induced injury by inhibiting p38MAPK expression and oxidative stress.
出处
《中国药理学通报》
CAS
CSCD
北大核心
2010年第10期1339-1344,共6页
Chinese Pharmacological Bulletin
基金
广东省科技计划资助项目(No2006B60501024
2007B080701030)
关键词
硫化氢
预处理
氯化钴
P38MAPK
活性氧
线粒体膜电位
凋亡
hydrogen sulfide
pretreatment
cobalt chloride
p38MAPK
reactive oxygen species
mitochondrial membrane potential
apoptosis
