T2毒素对软骨细胞合成一氧化氮(NO)影响.docVIP

T2毒素对软骨细胞合成一氧化氮(NO)影响 　 作者：杨占田，陈静宏，楚雍烈，曹峻岭，王治伦，师钟丽，郭雄 毕业论文 【关键词】; 一氧化氮合酶 ; 毕业论文 　　摘要：目的; 探讨T2毒素对软骨细胞合成一氧化氮(NO)的影响。方法; 胎儿软骨细胞体外培养，用MTT法检测软骨细胞存活率；用Griess重氮化法测定软骨细胞培养上清液中NO含量；用Western blot检测软骨细胞一氧化氮合酶(iNOS)的表达。结果; T2毒素在质量浓度在1-2000mu;g/L的范围内，软骨细胞存活率对其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； T2毒素刺激软骨细胞分泌NO和表达iNOS蛋白。结论; T2毒素引起的软骨细胞损伤和生长抑制与T2毒素刺激软骨细胞表达iNOS蛋白和分泌NO增多有关。 毕业论文 　　关键词：T2毒素；软骨细胞；一氧化氮；一氧化氮合酶 　 　ABSTRACT: Objective; To investigate the effects of T2 toxin on nitric oxide (NO) production and nitric oxide synthase (iNOS) expression in chondrocytes. Methods; Human chondrocytes cultured in vitro were treated with different concentrations of T2 toxin in different time (1-5 day). Cell viability of the treated cells was measured by MTT assay; The levels of NO in culture media were detected by colorimetric method of Griess assay; iNOS protein was measured by Western blot. Results; Dosedependent and time dependent effects of T2 toxin were found on T2 toxin treated chondrocytes within a certain range of concentration (1-2000mu;g/L) and a periods of time (1-5 day); The levels of NO in T2 toxin treated culture media were higher than that of normal control group; Overexpression of iNOS was detected in T2 toxin treated cells. Conclusion; T2 toxin can enhance NO production and upregulate expression of iNOS. NO plays an important role in T2 toxin induced growth inhibitory and toxic effects. 毕业论文 　　KEY WORDS: T2 toxin; chondrocytes; nitric oxide; nitric oxide synthase 毕业论文 　　T2毒素的病因学意义是目前大骨节病研究领域的热点。大量研究发现T2毒素可致软骨细胞损伤[1]。本研究采用胎儿软骨细胞体外培养，观察T2毒素对软骨细胞分泌一氧化氮(nitric oxide, NO)及其诱导型一氧化氮合酶(nitric oxide synthase, iNOS)的影响，旨在探讨T2毒素致软骨细胞损伤的分子机理。 毕业论文 　　1; 材料与方法 毕业论文 　　1.1; 试剂与仪器; DMEM培养基(GIBCO)、胰蛋白酶、透明质酸酶和II型胶原酶(Sigma)、 iNOS单克隆抗体(博士德生物工程公司)、四甲基偶氮唑(MTT)、二甲基亚砜(DMSO)、N1萘乙二胺盐酸盐和磺胺(华美生物工程公司)， T2毒素由中国军事医学科学院毒物药物研究所杨进生教授赠送，752型紫外分光光度计(上海精密科学仪器有限公司)，EI x800型通用酶标仪(基因公司)。 毕业论文 　　1.2; 软骨细胞培养及分组; 中孕引产胎儿的软骨细胞原代培养方法同文献[2]。第一代软骨细胞培养48h后随机分为对照组、T2毒素处理组，将不同质量浓度T2毒素(1、5、10、20、50、100、10