蛇毒cystatin基因转染抗人胃腺癌细胞体外侵袭作用的研究

目的:探讨蛇毒半胱氨酸蛋白酶抑制剂(sv-cystatin)对人胃腺癌细胞SGC7901侵袭转移的抑制作用.方法:采用人工拼接方法合成蛇毒cystatin cDNA,构建pcDNA3.1/sv-cystatin真核表达质粒,经脂质体转染将pcDNA3.1/sv-cystatin质粒和pcDNA3.1质粒分别导入胃腺癌细胞系SGC7901:利用RT-PCR和Western blot检测SGC7901细胞中sv-cystatin基因的表达;应用细胞-基质粘附实验、细胞运动实验及重建基底膜侵袭实验分析svcystatin表达对SGC7901细胞粘附、运动和侵袭能力的影响.结果:转染sv-cystatin基因后,SGC/sv-cystatin细胞克隆中可检测到sv-cystatin的明显表达,SGC/sv-cystatin细胞的运动能力和体外穿越重建基底膜的能力明显低于转染空载体细胞和未转染的SGC7901细胞,但其体外粘附能力未见明显变化.结论:sv-cystatin基因的表达可使胃癌SGC7901细胞体外运动能力及侵袭能力明显减弱,提示sv-cystatin具有抑制胃癌细胞侵袭转移的作用.

The Studies on Effect of Snake Venom Cystatin for Invasion and Metastasis of the Human Gastric Carcinoma Cell Line

Objective: To explore the suppressive effects of snake venom cystatin gene(sv-cystatin) on metastatic potential of human gastric carcinoma cell line SGC-7901. Methods: A 297bp DNA fragment ecoding sv-cystatin was designed based on the biased codon usage of yeast. Four partial overlapping cystatin geng fragments were synthesized, and the full length of sv-cystatin cDNA was obtained and linked to pUC18 vector. Expression plasmid pcDNA3.1/sv-cystatin was constructed and transfected into gastric carcinomar cell line SGC-7901 using lipofectamine gene transfer technique. Sv-cystatin gene expression was detected in SGC/sv-cystatin cells by RT-PCR, Western blot assay. Cell-matrix adhesion assay was used to study the adhensive ability of SGC/sv-cystatin cells. The effect of sv-cystatin expression on the invasion and migration of SGC7901 cells was investigated by transwell chamber system. Results: Expression of sv-cystatin was detected after cystatin gene-transfection in SGC/sv-cystatin cells. Matrigel invasion and migration assay showed that the invasion and migration of SGC/sv-cystatin cells were less than those in control groups, sv-cystatin. However, it did not affect the adhesive ability of SGC7901 cells with the matrix. Conclusion: The expression of sv-cystatin could inhibit SGC-7901 cells invision and migration in vitro. It suggests that sv-cystatin may have some role in reduction of gastric carcinoma cell invasion.