《中国康复理论与实践》

《中国康复理论与实践》 ›› 2006, Vol. 12 ›› Issue (06): 484-486.

• 基础研究 • 上一篇    下一篇

人退变椎间盘组织细胞凋亡相关基因表达的初步研究

胡明1; 马远征1; 张传森2; 陈道运2; 陈兴1; 才晓军1; 白一冰1   

  1. 1.解放军第三○九医院骨科 北京市 100091;2.第二军医大学人体解剖学教研室 上海市 200433
  • 收稿日期:2005-12-05 出版日期:2006-06-25 发布日期:2006-06-25

Study of Apoptosis and Gene Expression Pattern Related to Human Degenerative Intervertebral Disc Tissue

HU Ming, MA Yuan-zheng, ZHANG Chuan-sen, et al   

  1. The Department of Orthopaedics, PLA309Hospital, Beijing 100091, China
  • Received:2005-12-05 Published:2006-06-25 Online:2006-06-25

PDF (PC)

被引次数

36

摘要： 目的探讨细胞凋亡与腰椎间盘退变的发生机制。方法一步法抽提6例退变及6例正常椎间盘组织的总RNA,分别用cy3,cy5荧光标记,获得两组椎间盘cDNA的探针,与cDNA表达谱芯片杂交,扫描芯片荧光信号图像,对所获得的凋亡相关基因进行生物信息学分析。采用原位末端标记法(TUNEL)和免疫组化方法,检测12例人退变椎间盘及10例正常椎间盘组织的细胞凋亡状态及凋亡相关基因Bcl-2、Bax蛋白的表达。结果在4096条基因中,与细胞凋亡相关类蛋白为10条,其中Bax蛋白相关基因表达增高,Bcl-2蛋白相关基因表达降低。正常组TUNEL检测凋亡指数(AI)为(24.897±3.620),Bcl-2阳性细胞百分率为(31.440±4.150)%,Bax阳性细胞百分率为(29.372±2.588)%,阳性颗粒平均光密度值分别为(0.183±0.010)、(0.203±0.012)和(0.169±0.005);退变组AI为(49.232±3.440),Bcl-2阳性细胞百分率为(18.239±2.470)%,Bax阳性细胞百分率为(52.349±3.764)%,阳性颗粒平均光密度值分别为(0.152±0.003)、(0.310±0.008)和(0.262±0.014),两组间的AI均值、Bcl-2、Bax蛋白表达均有显著性差异(P<0.05)。结论细胞凋亡在椎间盘退变的进程中发挥重要作用,凋亡相关基因Bcl-2和Bax参与了髓核细胞凋亡的发生和发展过程。

关键词: 基因芯片, 基因表达谱, 椎间盘退变, 细胞凋亡, Bcl-2, Bax, 上皮细胞膜蛋白

Abstract: ObjectiveTo investigate the relationship between the degenerative mechanisms of lumbar intervertebral disc (LID) and apoptosis.MethodsThe total RNAs were isolated from human LID tissues. Both the mRNAs from the degeneration and normal LID were reversely transcribed to the cDNAs. The cDNAs were labeled with the incorporations of fluorescent dUTP, for preparing the hybridization probes. The mixed probes were then hybridized to the cDNA microarray. After high-stringent washing, the cDNA microarray was scanned for the fluorescent signals and analyzed by computer image analysis. The apoptotic status and the expression of Bcl-2 and Bax in 12 cases of degenerative LID and 10 cases of normal LID were detected with TdT-mediated dUTP-biotin nick end labeling (TUNEL) and immunohistochemistry methods.ResultsAmong the 4096 targets, there were 10 genes related to apoptosis. The expression related to Bax protein gene was up-regulated and it was down-regulated for Bcl-2 protein. In group of normal LID, the average apoptotic index (AI) was (24.897±3.620); percentage of Bcl-2 positive cells was (31.440±4.150)%; percentage of Bax positive cells was (29.372±2.588)%, average optical density (OD) values of positive particles were (0.183± 0.010 ), ( 0.203 ±0.012) and (0.169±0.005) respectively. In group of degenerative LID, the average AI was (49.232±3.440); percentage of Bcl-2 positive cells was (18.239±2.470)%; percentage of Bax positive cells was (52.349±3.764)%; average OD values of positive particles were (0.152±0.003), (0.310±0.008) and (0.262±0.014) respectively. There were significantly differences in AI and expressions of Bcl-2 and Bax proteins between normal LID and degenerative LID (P<0.05).ConclusionCell apoptosis plays an important role in the process of LID degeneration. Both Bcl-2 and Bax take part in the occurrence and progression of LID.

Key words: DNA microarray, gene expression pattern, intervertebral disc degeneration, apoptosis, Bcl-2, Bax, epithelial membrane protein (EMP)