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目的:探讨人血管能抑素(canstatin)对小鼠Lewis肺癌移植瘤生长、转移和血管新生的影响。方法:将pCMV-Script/canstatin及空载体pCMV-Script通过电穿孔的方法转染A549细胞,G418筛选获得阳性克隆。RT-PCR检测转染后细胞中canstatin mRNA的表达,Western blotting检测转染后细胞中canstatin蛋白的表达。建立Lewis肺癌小鼠移植瘤模型,观察pCMV-Script/canstatin组A549细胞培养上清对小鼠Lewis肺癌移植瘤的治疗作用,免疫组化检测各治疗组荷瘤小鼠移植瘤的微血管密度。结果:pCMV-Script/canstatin转染A549细胞在G418筛选后成功形成克隆,转染的A549细胞能有效表达can-statin mRNA和蛋白。pCMV-Script/canstatin治疗组小鼠肿瘤体积明显小于pCMV-Script组和NS组[(1.47±0.21)cm~3vs(2.43±0.15)cm~3、(2.53±0.18)cm~3,P<0.01);pCMV-Script/canstatin组、pCMV-Script组和NS组的肺转移结节数分别为(3.00±1.00)、(7.80±1.48)、(7.60±2.41)个,pCMV-Script/canstatin组肿瘤转移受到显著的抑制(P<0.01);pCMV-Script/canstatin组小鼠的肿瘤组织微血管数明显少于pCMV-Script组和NS组[(84.40±8.83)vs(188.68±11.15)、(190.24±12.91)个,P<0.01]。结论:pCMV-Script/canstatin能在A549细胞中表达并分泌至细胞外,canstatin可明显抑制Lewis肺癌移植瘤的生长、转移和血管新生。
Objective: To investigate the effect of canstatin on the growth, metastasis and angiogenesis of Lewis lung carcinoma in mice. METHODS: A549 cells were transfected with pCMV-Script / canstatin and empty vector pCMV-Script by electroporation. Positive clones were screened by G418. The expression of canstatin mRNA was detected by RT-PCR and the expression of canstatin protein was detected by Western blotting. Lewis lung cancer mouse xenograft model was established. The effect of culture supernatant of A549 cells in pCMV-Script / canstatin group on Lewis lung cancer xenografts in mice was observed. The microvessel density of tumor-bearing mice in each treatment group was detected by immunohistochemistry. RESULTS: A549 cells transfected with pCMV-Script / canstatin successfully cloned after G418 selection. The transfected A549 cells could express can-statin mRNA and protein efficiently. The tumor volume in pCMV-Script / canstatin-treated mice was significantly lower than that in pCMV-Script group and NS group [(1.47 ± 0.21) cm ~ 3 vs (2.43 ± 0.15) cm ~ (3.00 ± 1.00), (7.80 ± 1.48) and (7.60 ± 2.41) pCMV-Script / canstatin, pCMV-Script and NS groups, respectively (P <0.01). The number of microvessel in tumor tissue in pCMV-Script / canstatin group was significantly lower than that in pCMV-Script group and NS group [(84.40 ± 8.83) vs (188.68 ± 11.15), (190.24 ± 12.91) months, P <0.01]. Conclusion: pCMV-Script / canstatin can be expressed in vitro and secreted to the outside of cells. Canstatin can significantly inhibit the growth, metastasis and angiogenesis of Lewis lung carcinoma.