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目的:研究8-溴-7-甲氧基白杨素(BrMC)抑制源自人肝细胞癌Huh-7细胞系细胞肿瘤球形成细胞即肝癌干细胞样细胞(LCSLCs)自我更新作用。方法:体外培养人肝癌Huh-7细胞系细胞,干细胞条件培养基悬浮培养法富集和扩增LCSLCs。MTT比色法测定细胞活性;肿瘤球形成法检测自我更新能力;FITC-CD133标记FCM分析CD133表达。结果:干细胞条件培养基悬浮培养6天,Huh-7细胞形成非粘附、三维生长的肝癌球体细胞即LCSLCs。与Huh-7细胞相比,BrMC抑制LCSLCs细胞活性作用更强(P<0.05),并以浓度依赖性方式减少肿瘤球数目(P<0.05),降低CD133蛋白表达(P<0.05)。结论:干细胞条件培养基悬浮培养能分离和富集LC-SLCs;BrMC具有抑制LCSLCs细胞自我更新能力作用,其作用机制与降低干细胞标记物CD133蛋白的表达相关。
OBJECTIVE: To study the effect of 8-bromo-7-methoxychitosan (BrMC) on self-renewal of hepatoma stem cell-like cells (LCSLCs) derived from human hepatocellular carcinoma Huh-7 cell line. Methods: Human hepatocellular carcinoma Huh-7 cell line was cultured in vitro, and stem cell conditioned medium suspension culture was used to enrich and amplify LCSLCs. Cell viability was measured by MTT colorimetric assay; The ability of self-renewal was detected by tumor spheroid formation assay; CD133 expression was analyzed by FITC-CD133 labeled FCM. Results: The stem cell conditioned medium was cultured in suspension for 6 days. Huh-7 cells formed non-adherent and three-dimensional hepatocellular carcinoma cells (LCSLCs). Compared with Huh-7 cells, BrMC inhibited the cytotoxicity of LCSLCs more (P <0.05) and decreased the number of tumor cells (P <0.05) and decreased the expression of CD133 protein in a concentration-dependent manner (P <0.05). CONCLUSION: LC-SLCs can be separated and enriched in suspension culture of stem cell conditioned medium. BrMC can inhibit the self-renewal of LCSLCs cells, and its mechanism may be related to the decrease of CD133 protein expression.