苯海拉明等有机胺类药物干扰MTT还原法的应用  

Diphenhydramine interferes with MTT reduction assay

作　　者：沈耀[1,2] 张晨辉 胡薇薇 陈忠[2] 

机构地区：[1]温州医学院生命科学学院,浙江温州325035 [2]浙江大学药学院,浙江杭州310058

出　　处：《浙江大学学报（医学版）》2011年第2期163-168,共6页Journal of Zhejiang University(Medical Sciences)

基　　金：国家自然科学基金资助项目(30725047);浙江省教育厅资助项目(Y200907400);温州医学院资助项目(QTJ09016)

摘　　要：目的:探讨苯海拉明等有机胺类药物对MTT还原法测定细胞活性准确性的影响。方法:采用原代新生大鼠皮层星形胶质细胞模型,以不同浓度的苯海拉明等有机胺类药物分别作用于星形胶质细胞24 h,以形态学观察、Hoechst 33342和PI核荧光双染法对比MTT染色结果,分析苯海拉明等对MTT还原法应用的影响。结果:苯海拉明(10-4 mol/L)、吡拉明(10-4 mol/L)和卓兰替丁(10-5mol/L)作用于星形胶质细胞能显著增加甲瓒的生成,而核荧光双染法显示,这时星形胶质细胞的增殖和凋亡以及坏死情况并没有发生显著性改变。通过显微镜形态学观察发现,苯海拉明(10-4 mol/L)能显著减少星形胶质细胞膜上的针状晶体———甲瓒。结论:苯海拉明等有机胺类药物通过抑制甲瓒外排而促进胞内MTT被大量还原,从而影响MTT还原法测定细胞活性的准确性。Objective:To determine the effects of organic amine diphenhydramine on the 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide dye(MTT) reduction assay. Methods:The primarily cultured cortical astrocytes were incubated with various concentrations of diphenhydramine for 24 h.To analyze the effects of diphenhydramine and other organic amines on the MTT assay,the data obtained from the MTT assay were compared with the results obtained from morphological observation and hoechst 33342 and propidium iodide(PI) nucleus double staining. Results:The MTT assay showed that diphendramine(10-4mol/L),pyrilamine(10-4mol/L) and zolatidine(10-5mol/L) caused a significant increase in MTT reduction in astrocytes.However there was no proliferation,apoptosis or necrosis detected by hoechst and PI nucleus double staining.Light microscopy revealed that exocytosis of formazan granules was inhibited by diphendramine. Conclusion:Diphenhydramine and other organic amines may enhance MTT reduction by suppression of MTT formazan exocytosis in astrocytes,which may affect the results of cell viability studies.

关 键 词：星形细胞/病理学 细胞活性 苯海拉明/投药和剂量 MTT还原法 

分 类 号：R965[医药卫生—药理学]