尿多酸肽通过调节caspase家族诱导多发性骨髓瘤细胞U266凋亡的实验研究  

Uroacitide-induced multiple myeloma cells of U266 by regulating the caspases

作　　者：刘梅 孙菊 吴迪炯[2] 俞庆宏[2] 刘文宾[2] 胡慧瑾[2] 叶宝东[2] Liu Mei;Sun Ju;Wu Dijiong;Yu Oinghong;Liu Wenbin;Hu Huijin;Ye Baodong(The First School of Clinical Medicine,Zhejiang Chinese Medical University,Hangzhou 310053,China;Department of Hematology,the First Affiliated Hospital of Zhejiang Chinese Medical University,Hangzhou 310006,China)

机构地区：[1]浙江中医药大学第一临床医学院,杭州310053 [2]浙江中医药大学附属第一医院血液科,杭州310006

出　　处：《中华危重症医学杂志（电子版）》2018年第4期256-259,共4页Chinese Journal of Critical Care Medicine:Electronic Edition

基　　金：浙江省卫计委创新人才资助计划项目(1S21702);浙江中医药大学附属第一医院三鹰人才资助计划项目(ZD01601)

摘　　要：目的探讨尿多酸肽抑制多发性骨髓瘤(MM)细胞株U266增值及其机制。方法采用四甲基偶氮唑(MTT)比色法计算不同尿多酸肽浓度(1.0、2.0、3.0、4.0、5.0 mg/g)对U266细胞生长的抑制率,Hoechst33258染色观察尿多酸肽作用后的细胞凋亡情况,Annexin-V/碘化丙啶(PI)检测尿多酸肽作用6、12、24 h后与对照组细胞(加细胞未加药物)的细胞凋亡率。使用Westernblotting法检测caspase 8、caspase 3及其激活物在U266细胞株经尿多酸肽(4 mg/g)作用6、12、24 h后的表达改变。结果随着尿多酸肽浓度提高,U266细胞抑制率显著上升(F=17.276,P <0.001)。经尿多酸肽作用后,Hoechst33258荧光染色提示细胞核浓集及出现凋亡小体。Annexin-V/PI分析显示,尿多酸肽处理6、12、24 h后细胞凋亡率显著高于对照组细胞[(5.53±0.28)%、(9.43±1.78)%、(21.50±2.45)%、(3.03±0.11)%,F=12.242,P <0.001]。同时,随着尿多酸肽作用时间延长,pro-caspase 8、pro-caspase 3表达明显下降(F=18.241,P <0.001;F=4.924,P=0.005),而active-caspase 8、active-caspase 3表达明显上升(F=22.322,P <0.001;F=6.213,P=0.002)。结论尿多酸肽可抑制U266细胞增殖,且可能通过caspase凋亡途径在其中起主要作用。Objective To explore the effect of uroacitide on muhiple myeloma （MM） cells of U266 and its mechanisms. Methods The inhibition ratio of different uroacitides （1.0, 2.0, 3.0, 4.0, 5.0 mg/g） on the growth of U266 cells was calculated by the methyhhiazolyl tetrazolium （MTT） colorimetric assay. The apoptosis of U266 cells was observed by Hoechst33258 staining. Annexin-V / propidium iodide （PI） was used to detect the apoptosis rate of U266 cells with uroacitides at 6, 12, 24 h and cells without uroacitides. The Western-blotting method was used to test expressions of caspase 8, caspase 3 and their activators in U266 cells with uroacitides （4 mg/g） for 6, 12 and 24 h. Results The inhibition rate of U266 cells increased significantly as the concentration of uroacitides increased （F = 17.276, P 〈 0.001）. Condensed nuclei and apoptotic bodies were found in cells with uroacitides through Hoechst33258 fluorescence staining. Annexin-V/PI analysis showed that the apoptosis rate of cells with uroacitides at 6, 12, 24 h was much higher than that of cells without uroacitides [（5.53±0.28）%, （9.43±1.78）%, （21.50±2.45）%, （3.03 ±0.11）%, F= 12.242, P〈 0.001]. Meanwhile, the expressions of pro-caspase 8 and pro-caspase 3 decreased significantly （F= 18.241, P〈 0.001; F= 4.924, P = 0.005）, and expressions

关 键 词：尿多酸肽 U266 半胱氨酸天冬氨酸蛋白酶 多发性骨髓瘤 细胞凋亡 

分 类 号：R733.3[医药卫生—肿瘤]