外源性PTEN基因诱导黏液表皮样癌细胞系凋亡的研究

目的：探讨转染外源性PTEN抑癌基因对黏液表皮样癌细胞系M3SP2的诱导凋亡作用。方法：用含野生型PTEN抑癌基因逆转录病毒载体转染高转移性涎腺黏液表皮样癌细胞系M3SP2,转染空载体亲本细胞为对照细胞。体外细胞凋亡采用苏木精-伊红染色、电镜、TUNEL染色和流式细胞仪检测,裸鼠移植瘤细胞凋亡应用苏木精-伊红染色形态学判定。Bc l-2、P53和H-ras蛋白表达用免疫组化染色检测。结果：试验细胞M3SP2-PTEN与对照细胞M3SP2-pBp比较,出现较多的典型的凋亡细胞特征。试验细胞和对照细胞凋亡率分别为7.5%-13.6%和1.2%-2.3%;裸鼠移植瘤M3SP2-PTEN组和M3SP2-pBp组的凋亡指数分别为17.4%和3.5%。免疫组化染色显示M3SP2-PTEN细胞与对照细胞比较,P53蛋白表达增强,Bc l-2和H-ras蛋白表达减弱。结论：外源性PTEN抑癌基因能显著诱导高转移性涎腺黏液表皮样癌细胞凋亡,并且与P53、Bc l-2和H-ras蛋白表达有一定关系。

Apoptotic Effects of the Exogenous PTEN Tumor Suppressor Gene on Human Mucoepidermoid Carcinoma Cell Line

Objective： To investigate the role of exogenous wild -type PTEN gene inducing apoptosis in mucoepidermoid carcinoma cells and related mechanisms. Methods： Recombinant retrovirus expression vector containing human wild -type PTEN gene packaged with lipofectin were introduced into mucoepidermoid carcinomar cell line M3 SP2 and parental MaSP2 cells transfected with empty vector served as controls. Cellular apoptosis in vitro was determined by hematoxylin and eosin （HE） -staining, TUNEL assay and flow cytometry. Apoptosis index （AI） of xenograft in vivo was evaluated in HE - stained slides. The expression of P53, Bcl - 2 and H - ras protein in cells was analyzed by immunohistochemieal methods. Results： Some typical morphological changes of apoptosis were observed in cells transfected with wild - type PTEN gene （M3SP2 - PTEN） , but not in control cells （M3SP2 -pBp） . The apoptosis rate in M3SP2 - PTEN cells and M3SP2 - pBp cells was 7.5% - 13.6% and 1.2% -2.3% , respectively. AI of xenograft in vivo in PTEN gene transfection group and control group was 17.4% and 3.5% , respectively. The overexpression of PTEN protein in M3SP2 - PTEN ceils was concomitant with an increased level of P533protein and a decreased level of Bcl -2 and H -ras protein, compared with con- trol cells. Condusion： Wild - type PTEN gene transfection can significantly induce apoptosis in mucoepidermoid carcinoma cells in vitro and in vivo. Enhanced expression of PTEN by gene transfer is sufficient to regulate expression of apoptosis - related protein such as P53 , Bcl - 2 and H - ras. These results may provide new clues on the gene therapy in mucoepi- dermoid carcinoma.