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目的探讨外源性肝细胞黏附分子(hepatocyte cell adhesion molecule,hepaCAM)转染对人膀胱癌BIU-87细胞株生长及血管内皮生长因子(vascular endothelial growth factors,VEGF)表达的影响。方法使用脂质体2000转染法将携带野生型hepaCAM基因的质粒(pEGFPN2-hepaCAM)及空载质粒(pEGFPN2)转染人膀胱癌BIU-87细胞,建立稳定过表达hepaCAM基因的BIU-87细胞株。以转染空载质粒及未转染的BIU-87细胞株作为对照,采用MTT法检测各组细胞生长能力,逆转录聚合酶链反应(RT-PCR)检测hepaCAM、VEGF mRNA水平的变化,Western blot检测VEGF蛋白水平的变化。结果MTT实验示:hepaCAM基因转染(实验组)BIU-87细胞生长与空载体组和未处理组细胞比较明显受到抑制(P<0.05);hepaCAM基因在膀胱癌BIU-87细胞不表达,转染后hepaCAM mRNA明显增强(P<0.05);实验组VEGF mRNA表达(0.527±0.031)与未处理组(0.803±0.042)、空载组(0.750±0.040)比较明显下降(P<0.05)。Western blot结果示:实验组BIU-87细胞VEGF蛋白表达(0.057±0.015)与未处理组(0.263±0.032)和空载组(0.280±0.036)比较明显下降(P<0.05)。结论转染野生型hepaCAM基因可明显抑制膀胱癌BIU-87细胞的生长能力,并下调VEGF基因及蛋白的表达,hepaCAM基因可能是影响肿瘤的生长及侵袭转移的重要原因之一。
Objective To investigate the effects of hepaCAM transfection on the growth of human bladder cancer cell line BIU-87 and the expression of vascular endothelial growth factor (VEGF). METHODS: Human bladder cancer cell line BIU-87 was transfected with plasmid pEGFPN2-hepaCAM carrying wild-type hepaCAM gene and empty vector pEGFPN2 by lipofectamine 2000, and BIU-87 cells stably overexpressing hepaCAM gene were established Strain. The transfected empty vector and untransfected BIU-87 cell line were used as control. The cell growth ability was detected by MTT assay. The changes of hepaCAM and VEGF mRNA were detected by reverse transcriptase polymerase chain reaction (RT-PCR) blot detection of VEGF protein level changes. Results MTT assay showed that the hepaCAM gene was significantly inhibited in the BIU-87 cells transfected with BIH-87 cells (P <0.05), but not in BIU-87 cells Compared with untreated group (0.803 ± 0.042) and no-load group (0.750 ± 0.040), the mRNA expression of hepaCAM in experimental group was significantly decreased (P <0.05). Western blot results showed that the expression of VEGF protein in BIU-87 cells in experimental group was significantly lower than that in untreated group (0.263 ± 0.032) and no-load group (0.280 ± 0.036) (P <0.05). CONCLUSION: Transfection of wild-type hepaCAM gene can significantly inhibit the growth of bladder cancer cell line BIU-87 and down-regulate the expression of VEGF gene and protein. HepaCAM gene may play an important role in tumor growth and invasion and metastasis.