Lgr5和CD44标记的肿瘤干细胞通过高表达骨桥蛋白调控胃癌中血管的形成

Cancer stem cells labeled by Lgr5 and CD44 regulate angiogenesis through overexpression of osteopontin

目的 检测骨桥蛋白在胃癌组织、胃癌细胞株及胃癌干细胞中的表达水平,并初步探讨其与血管形成之间的关系.方法 运用无血清悬浮培养法分别从胃癌细胞株SGC-7901和BGC-823中筛选出悬浮性成球干细胞（SC）,并通过检测胃癌干细胞表面标志物Lgr5和CD44来判断是否筛选成功.采用RT-PCR和Western blot法分别检测骨桥蛋白（OPN）在胃癌组织、原代贴壁细胞株（AC）以及SC中mRNA和蛋白水平的表达,最后通过血管形成实验来检测OPN的血管形成能力.结果 与癌旁组织相比,5例胃癌组织中有4例高表达OPN.SC高表达Lgr5和CD4,表明无血清悬浮培养法成功地筛选出了肿瘤干细胞.另外,不管在mRNA和蛋白质水平,OPN在SC中的表达水平都要远高于AC（P〈0.05）.血管形成实验表明OPN具有很强的血管生成能力（P〈0.05）.结论 胃癌干细胞通过高表达OPN来调控新生血管的形成,进而影响胃癌细胞的增殖和侵袭等.

Objective To explore the levels of osteopontin （OPN） in gastric cancer tissues,gastric cancer cell lines and stem cells （SCs）,and investigate the relationship between OPN and angiogenesis.Methods Gastric cancer cell lines SGC-7901 and BGC-823CSCs were used to screen out SCs in suspension by serum free medium followed by detection of the surface markers of gastric cancer SCs Lgr5 and CD44.The levels of OPN protein and mRNA in gastric cancer tissues,adherence cells （ACs） and SCs were examined by Western blotting and RT-PCR.The angiogenic activity of OPN was evaluated by the tube formation assay.Results Compared with adjacent non-cancerous tissues,OPN was highly expressed in four out of five gastric cancer tissues in the current study.High expression of Lgr5 and CD44 in gastric cancer SCs indicated the successful screening of the targeted cells.Moreover,the levels of OPN protein and mRNA were higher in SCs than those of ACs （P〈0.05）.The tube formation assay demonstrated that OPN had strong angiogenic activity （P〈0.05）.Conclusions Gastric cancer SCs can affect the proliferation and invasion of gastric cancer cells by regulating angiogenesis through overexpression of OPN.