【摘要】 目的:探讨人cathepsin B(CatB)反义RNA在乳腺癌细胞侵袭与迁移中的作用。方法:构建携CatB反义RNA的重组质粒pBudCE4.1-antiCatB,采用脂质体法将重组质粒瞬时转染人乳腺癌细胞MDA-MB-231。Western blotting法检测MDA-MB-231细胞中CatB蛋白的表达,MTT法检测MDA-MB-231细胞的增殖,细胞-基质黏附实验检测反义CatB对MDA-MB-231细胞黏附能力的影响,体外侵袭、迁移实验分析反义CatB表达对MDA-MB-231细胞体外侵袭和迁移能力的影响。结果:成功构建pBudCE4.1-antiCatB表达载体。转染MDA-MB-231细胞后,与未转染组细胞和转染空载体组细胞相比,转染组MDA-MB-231细胞的CatB蛋白表达水平明显降低[(0.96±0.02)vs(1.98±0.23),(1.84±0.08),P<0.05];转染组细胞的增殖受到明显抑制[(0.255±0.017)vs(0.458±0.033),(0.421±0.022),P<0.01];转染组细胞的黏附(基质胶或纤维黏连蛋白)能力明显下降[(0.054±0.017)vs(0.111±0.018),(0.107±0.017),P<0.01;或(0.052±0.008)vs(0.120±0.014),(0.113±0.009),P<0.01];转染组细胞的侵袭和迁移能力也明显降低[(52.80±7.76)vs(124.00±44.54),(116.80±32.87),P<0.01;(60.25±8.73)vs(132.50±12.15),(119.20±25.13),P<0.01]。结论:CatB反义RNA可抑制乳腺癌细胞体外生长、黏附、迁移和侵袭能力。更多还原

【Abstract】 Objective:To investigate the effect of antisense cathepsin B(CatB) RNA on the invasion and migration of human breast carcinoma cells. Methods: PBudCE4.1-antiCatB recombinant plasmid carrying antisense cathepsin B(CatB) gene was constructed and transfected into breast carcinoma cell line MDA-MB-231 by lipofection system. The expression of CatB protein in MDA-MB-231 cells was detected by Western blotting analysis; the proliferation of MDA-MB-231 cells was determined by MTT assay. Cell-matrix adhesion assay was used to examine the effect of anti-CatB on adhesion ability of MDA-MB-231 cells. The effects of anti-CatB on the invasion and migration abilities of MDA-MB-231 cells were measured by invasion and migration transwell system. Results: The recombinant plasmid PBudCE4.1-antiCatB was successfully constructed. Expression of CatB protein in MDA-MB-231 cells was decreased after PBudCE4.1-antiCatB transfection compared with those in untransfected and mock-vehicle transfected cells([0.96±0.02] vs [1.98±0.23], [1.84±0.08], P<0.05); the proliferation of MDA-MB-231 cells was also inhibited in PBudCE4.1-antiCatB transfected group([0.255±0.017] vs [0.458±0.033], [0.421±0.022], P<0.01); and the adhesion abilities(binding to matrix or fibronectin) were decreased([0.054±0.017] vs [0.111±0.018], [0.107±0.017], P<0.01; [0.052±0.008] vs [0.120±0.014], [0.113±0.009], P<0.01). Transwell asaay showed that the invasion and migration abilities were inhibited in PBudCE4.1-antiCatB transfected group compared with those in the non-transfection and mock-vehicle transfected groups([52.80±7.76] vs [124.00±44.54], [116.80±32.87], P<0.01; [60.25±8.73] vs [132.50±12.15], [119.20±25.13], P<0.01). Conclusion: The expression of antisense CatB RNA can inhibit the growth, adhesion, invasion and migration abilities of MDA-MB-231 cell in vitro.更多还原