摘要
目的探讨T-2毒素致软骨细胞凋亡与一氧化氮（NO）和Fas凋亡途径的关系。方法采用MTT法检测T-2毒素对软骨细胞存活率的影响；Annexin V／PI法和电镜观察研究T-2毒素致人软骨细胞凋亡的作用；用Griess重氮化法，测定T-2毒素作用于软骨细胞培养上清液中NO含量；Western Blotting检测T-2毒素对人软骨细胞表达一氧化氮合酶（iNOS）和Fas蛋白的影响。统计分析T-2毒素诱导人软骨细胞凋亡与其分泌NO、iNOS和Fas蛋白的相关性。结果T-2毒素在浓度为1～2000ng／mL的范围内，对软骨细胞存活率的作用呈较典型的浓度依赖关系和时间依赖关系，而且随着作用时间的延长，浓度依赖关系更为明显；T-2毒素可引起软骨细胞发生凋亡的典型电镜形态改变，并使早期凋亡率和晚期凋亡率明显增加，在一定范围内呈浓度依赖性；T-2毒素刺激软骨细胞分泌NO和表达iNOS蛋白；T-2毒素使凋亡相关蛋白Fas表达增多；软骨细胞分泌NO和表达iNOS蛋白和Fas蛋白量与T-2毒素诱导人软骨细胞凋亡率均有正相关性。结论T-2毒索引起的软骨细胞凋亡与T-2毒素刺激软骨细胞分泌NO和表达iNOS蛋白增多有关，并与凋亡相关蛋白Fas表达增多有关。
Objective To investigate the relationship of T-2 toxin-induced chondrocytes apoptosis with nitric oxide（NO） and Fas apoptosis pathway. Methods Human chondrocytes cultured in vitro were treated with different concentrations of T-2 toxin at different time （1-5 d）. Cell viability of the treated cells was measured by MTT assay. Apoptotic ultrostructural changes of the treated cells were observed with electron microscopy. Biological changes of apoptosis were detected by annexin V/PI Flow cytometer （FCM）. The levels of NO in culture media were detected by colorimetric method of Griess assay. Nitric oxide synthase （iNOS） and Fas protein were measured by Western blot. Results In this study the results shown the dose-dependent and time-dependent effects of T-2 toxin, within a range of concentration （1-2000 ng/mL） and a period of time （1-5 d）, on the T-2 toxin-treated chondrocytes. Apoptotic body was found in T-2 toxin-treated chondrocytes by electron microscopy. Early-stage apoptosis rate and late-stage apoptosis rate were both increased in T-2 toxln-treated cells when compared with non-treated cells in a dose-dependent manner. The levels of NO in T-2 toxin-treated culture media were higher than that of normal control. Over-expressions of iNOS and Fas protein were detected in T-2 toxin- treated cells. T-2 toxin-induced apoptosis was noted to be significtnly correlated with the level of NO production and the levels of iNOS and Fas protein expression. Conclusion T-2 toxin can enhance NO production and upregulate the expression of iNOS and Fas protein. Both NO and Fas signaling pathway are involved in T-2 toxin- induced apoptosis.
出处
《四川大学学报（医学版）》
CAS
CSCD
北大核心
2006年第4期583-586,共4页
Journal of Sichuan University(Medical Sciences)
基金
国家自然科学基金(批准号30471499)
教育部科技重点项目(重点03152)资助
