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Staurosporine诱导小鼠心肌细胞凋亡模型的构建 (PDF)

《心脏杂志》[ISSN:1009-7236/CN:61-1268/R]

期数:
2004年第1期
页码:
4-10
栏目:
实验研究
出版日期:
2004-01-01

文章信息/Info

Title:
Molecular identification of mouse cardiomyocytes apoptosis induced by staurosporine
作者:
王晓明1; 2;  臧益民2;  高 峰2;  龚卫琴1; 2;  李源1
第四军医大学: 1.西京医院老年病1科, 2.生理学教研室, 陕西 西安 710032
Author(s):
WANG Xiao-ming1; 2;  ZANG Yi-min2;  GAO Feng2;  GONG Wei-qin1;  LI Yuan2
1.Department of Geriatrics Xijing Hospital, 2.Department of Physiology, Fourth Military Medical University, Xi'an Shaanxi 710032, China
关键词:
心肌细胞;  小鼠;  细胞凋亡;  staurosporine;  caspase-3;  乳酸脱氢酶
Keywords:
cardiomyocytes;  mouse;  apoptosis;  staurosporine;  caspase-3; LDH
分类号:
Q255; R33.13
DOI:
-
文献标识码:
A
摘要:
目的: Staurosporine可诱导不同细胞类型的细胞凋亡,但尚缺乏其对小鼠心肌细胞作用影响的报道。本研究旨在观察Staurosporine是否也可引起小鼠心肌细胞的凋亡,并对其模型进行分子水平的鉴定。方法: 用Staurosporine处理原代培养的小鼠心肌细胞的直接刺激后,观察心肌细胞的形态学变化、DNA片段、半胱天冬蛋白酶(caspase)-3活性、细胞存活率以及细胞膜完整性。结果: ① Staurosporine处理过的心肌细胞具有明显的凋亡形态学特征:细胞的皱缩、核的浓缩及DNA梯改变。② Staurosporine 4 mol/L对心肌细胞作用8 h后,与正常对照组比较细胞存活率降至31.2 % (与对照组比,P<0.01),且细胞存活率与Staurosporine的作用浓度和作用时间呈依赖关 系;同时测定细胞培养液中能反映细胞膜完整性的胞浆内容物乳酸脱氢酶(LDH)水平,发现在Staurosporine作用1～8 h的各个时间点,LDH的漏出水平最高未超过10 %(与对照组相比,P>0.05)这种高的细胞死亡率与低的细胞膜的损伤率,恰恰反映了细胞死亡的性质是凋亡性死亡。③ Staurosporine能激活心肌细胞内的Caspese-3的活性,当使用广谱的 Caspese抑制剂ZVAD-fmk预处理培养的小鼠心肌细胞后,能够有效的阻止上述细胞凋亡的发生,从而避免了Staurosporine诱导的细胞死亡。结论:staurosporine能够快速、有效的诱导小鼠心肌细胞凋亡,caspase-3在诱导的细胞凋亡过程中起着至关重要的作用。
Abstract:
AIM: Staurosporine has been shown to induce apoptosis in some types of cell. The present study was designed to establish and identify a model of mouse cardiomyocyte apoptosis nduced by staurosporine. METHODS: Neonatal mouse cardiomyocytes were cultured and treated with or withouts taurosporine. Cellularmor phological characteristics, DNA fragmentations, caspase-3 activity, cellviability and cell membran eintegrity were observed. RESULTS: Remarable morphological characteristics of cardiomyocytes apoptosis, including cell shrinage, condensed chromatin and DNA fragmentations (DNA laddering) were observed. After 8 h exposure to 4 ?mol/L staurosporine, the cell viability of cardiomyocytes was 31.2 % (P<0.01, vs control group) and which was time and concentration (1～16 mol/L) dependent. Myocyte lactate dehydrogenase (LDH) leaage were changed insignificantly in staurosporine reated cells (1 to 8 h) as compared with the control. In addition, staurosporine induced myocyte apoptosis as inhibited by irreversible tripeptide inhibitor of caspases ZVAD-fmk. CONCLUSION: These data suggest that staurosporine can rapidly and efficiently induce apoptotic death of mouse cardiomyocytes Caspase-3 may play a critical role in the process of staurosporine induced cardiomyocyte apoptosis.

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备注/Memo

备注/Memo:
收稿日期：2003-1-10.
更新日期/Last Update: 2004-01-01