健脾解毒方对大肠癌细胞mTOR信号通路相关蛋白表达的影响

摘要

目的：探讨健脾解毒方对大肠癌细胞增殖的影响及可能作用机制。方法采用水提法制作健脾解毒方提取物，利用超高效液相-高分辨飞行时间质谱法分析健脾解毒方的主要成分，MTT法检测健脾解毒方对大肠癌细胞增殖的影响，Graphpad Prism5软件计算IC50值，流式细胞术检测细胞周期，Western Blot技术检测Phospho-mTOR、Phospho-P53和P21的蛋白表达。结果健脾解毒方能够抑制大肠癌细胞增殖，处理24、48、72 h 后四种大肠癌细胞系的 IC50值分别为 HCT116（6.894、5.668、3.648 mg/mL）、LoVo（14.65、8.737、7.849 mg/mL）、SW48（8.029、7.026、5.740 mg/mL）及HT29（13.06、9.646、8.448 mg/mL）；健脾解毒方使大肠癌细胞周期阻滞在G1期；并能够下调Phospho-mTOR蛋白表达（P<0.05），上调Phospho-P53和P21蛋白的表达（P<0.05）,使大肠癌细胞周期阻滞在G1期。结论健脾解毒方可能通过mTOR-P53-P21途径抑制大肠癌细胞的增殖，这可能是健脾解毒方治疗大肠癌的作用机制之一。%Objective To investigate the effects of Jianpi Jiedu Formula (JPJD) on inhibiting colon cancer cell prolif-eration capacity and its possible mechanism. Methods The main components of water extract of JPJD were analyzed by U-PLC-Q-TOF/MS. The effect of JPJD on colon cancer cell proliferation capacity was determined by MTT assays. The IC50 value concentration were calculated by Graphpad Prism5 software. The cell cycle was detected by Flow cytometric method. The protein levels of Phospho-mTOR (P-mTOR), Phospho-P53 (PP53) and P21 were examined by Western Blot. Results MTT assays demonstrated that JPJD can inhibite the colon cancer cell proliferation capacity. The IC50 value concentration of JPJD at 24 h, 48 h and 72 h after treatment were 6.894, 5.668, 3.648 mg/mL in HCT116 cell, 14.650, 8.737, 7.849 mg/mL in LoVo cell, 8.029, 7.029, 5.740 mg/mL in SW48 cell and 13.06, 9.646, 8.448 mg/mL in HT29 cell, re-spectively. JPJD could down-regulate the expression of Phospho-mTOR protein, up-regulate the expression of Phospho-P53 and P21 protein, and keep the cycle of the large intestine cancer cells at G1 phase. Conclusion The JPJD could inhibit HCT116, LoVo, SW48 and HT29 cells proliferation capacity by mTOR-P53-P21 signaling pathways, and it may be the probable mechanism of JPJD on colorectal cancer.