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目的:探讨不同浓度消癌解毒方含药血清对人结肠癌细胞增殖及糖酵解过程的影响及其作用机制。方法:8只新西兰兔随机分为含药血清组(消癌解毒方40 g·kg·~(-1)d~(-1))和空白血清组(等体积生理盐水),每组兔子连续灌胃3 d,制备消癌解毒方含药血清和空白血清。5%,10%,15%消癌解毒方含药血清处理人结肠癌HT-29与HCT-116细胞后,采用噻唑蓝比色法测定其对HT-29与HCT-116细胞生长的抑制作用,乳酸试剂盒检测HT-29与HCT-116细胞的乳酸生成量,葡萄糖试剂盒检测HT-29与HCT-116细胞的葡萄糖生成量,蛋白免疫印迹法(Western blot)检测己糖激酶(HK2),丙酮酸脱氢酶激酶(PDK1),乳酸脱氢酶(LDHA)和低氧诱导因子-1α(HIF-1α)蛋白表达。结果:消癌解毒方含药血清能够抑制HT-29与HCT-116细胞增殖能力,15%消癌解毒方含药血清对人结肠癌HT-29与HCT-116细胞的抑制率分别为42.7%,50.2%。Western blot结果显示,与空白组比较,消癌解毒方含药血清可降低HK2,PDK1,LDHA,HIF-1α蛋白相对表达量(P<0.05),且随浓度的增加表达降低。乳酸及葡萄糖试剂盒检测结果显示,与空白组比较,消癌解毒方含药血清可降低HT-29与HCT-116细胞的乳酸及葡萄糖水平(P<0.05),且随浓度的增加乳酸及葡萄糖水平降低。结论:消癌解毒方含药血清可以通过抑制人结肠癌HT-29与HCT-116细胞糖酵解的过程从而抑制肿瘤细胞的增殖,其抑制糖酵解过程的机制可能与降低HIF-1α表达,进而减少糖酵解相关酶HK2,PDK1,LDHA表达有关。
OBJECTIVE: To investigate the effects of different concentrations of Xiaojian Jiedu Fang containing serum on proliferation and glycolysis of human colon cancer cells and its mechanism. Methods: Eight New Zealand rabbits were randomly divided into drug-containing serum group (40g · kg · d -1) and blank serum group (equal volume of normal saline) Gavage 3 d, preparation of Xiaojianjiedu Fang containing serum and blank serum. The inhibitory effects of 5%, 10%, 15% Xiaojian Jiedu Fang containing serum on human colon cancer HT-29 and HCT-116 cells were determined by thiazolyl blue colorimetric assay. Lactic acid kit was used to detect the production of lactate in HT-29 and HCT-116 cells. Glucose kit was used to detect glucose production in HT-29 and HCT-116 cells. Western blot was used to detect hexokinase (HK2) , Pyruvate dehydrogenase kinase (PDK1), lactate dehydrogenase (LDHA) and hypoxia inducible factor-1α (HIF-1α) protein expression. Results: The serum containing Xiaojian Jiedu Decoction could inhibit the proliferation of HT-29 and HCT-116 cells, and the inhibitory rates of 15% Xiaojian Jiedu decoction-containing serum on human colon cancer HT-29 and HCT-116 cells were 42.7% , 50.2%. Western blot results showed that compared with the blank group, the drug-containing serum of JJJD reduced the relative expression of HK2, PDK1, LDHA and HIF-1α protein (P <0.05), and decreased with increasing concentration. Lactic acid and glucose kit test results showed that, compared with the blank group, Xiaoji Jiedu Fang containing serum can reduce the lactate and glucose levels of HT-29 and HCT-116 cells (P <0.05), and with the increase of concentration of lactic acid and glucose Lower level. Conclusion: The drug-containing serum of JJJT can inhibit the proliferation of tumor cells by inhibiting the process of glycolysis of human colon cancer HT-29 and HCT-116 cells. The mechanism of inhibiting the glycolysis may be related to the decrease of HIF-1α expression , Thereby reducing the glycolytic enzymes HK2, PDK1, LDHA expression.