基于糖酵解的金钗石斛乙醇提取物抗肺癌的药理机制研究

Pharmacological mechanism of anti-lung cancer of Dendrobium nobile ethanol extract through glycolysis

目的 研究金钗石斛乙醇提取物(DNEE)治疗肺癌的药理作用机制。方法 采用人肺癌A549细胞进行体外实验,MTT法检测50~800μg·mLDNEE对A549细胞的抑制率,比色法观察细胞的葡萄糖消耗量和乳酸生成量。以小鼠肺癌LLC细胞皮下移植荷瘤小鼠为疾病模型评估DNEE 200和600 mg·kg^(-1)的体内抗肿瘤作用,测量和绘制荷瘤小鼠体重变化曲线和肿瘤生长曲线。Western blot法检测A549细胞和肿瘤组织中缺氧诱导因子(HIF)-1α、葡萄糖转运蛋白(GLUT)4和己糖激酶(HK)2蛋白表达水平,氯化钴(CoCl2)诱导HIF-1α高表达分析HIF-1α对GLUT4和HK2蛋白表达的影响,分子对接技术探讨DNEE抗肿瘤主要活性成分与HIF-1α、GLUT4和HK2的相互作用。结果 与空白对照组相比,50~800μg·mLDNEE组A549细胞抑制率呈浓度依赖性增加(P <0.05),200、400μg·mLDNEE组A549细胞中葡萄糖消耗量和乳酸生成量显著降低(P <0.05)。与模型组相比,DNEE 200和600 mg·kg^(-1)组LLC细胞皮下移植荷瘤小鼠体重无显著变化(P> 0.05),肿瘤体积显著减小(P <0.05),HIF-1α、GLUT4和HK2表达下调(P <0.05)。moscatin、densiflorol B、gigantol和moscatilin与HIF-1α、GLUT4和HK2形成稳定构象。结论 DNEE具有良好的体内、体外抗肺癌作用,其机制可能与下调HIF-1α、GLUT4和HK2表达从而抑制肿瘤糖酵解有关。

AIM To study the pharmacological mechanism of Dendrobium nobile ethanol extract(DNEE) in the treatment of lung cancer. METHODS Human carcinoma A549 cells were used for in vitro experiments. The inhibition rate of 50 — 800 μg·mLDNEE on A549 cells was detected by MTT assay, and the glucose consumption and lactic acid production were observed by colorimetry. The in vivo anti-tumor effects of DNEE 200 and 600 mg·kg^(-1)were evaluated in mouse carcinoma LLC cells subcutaneously-transplanted tumor mice as a disease model, the body weight change curve and tumor growth curve of tumor-bearing mice were measured and drawn. The expression levels of hypoxia-inducible factor(HIF)-1α,glucose transporter(GLUT) 4 and hexokinase(HK) 2 proteins in A549 cells and tumor tissues were determined by Western blot, the effect of high-expression HIF-1α induced by cobalt chloride on the expression of GLUT4 and HK2 proteins was analyzed, molecular docking technology was utilized to explore the interaction between main anti-tumor compounds in DNEE and HIF-1α, GLUT4, HK2. RESULTS Compared with the control group, the inhibition rate of A549 cells in the 50 — 800 μg·mLDNEE group increased in a concentration-dependent manner(P < 0.05), glucose consumption and lactic acid production in 200 μg·mLDNEE group and 400 μg·mLDNEE group were significantly reduced(P < 0.05).Compared with the model group, the body weight of the tumor-bearing mice in the 200 mg·kg^(-1)DNEE group and 600 mg·kg^(-1)DNEE group had no significantly change(P > 0.05), but the tumor volume was significantly reduced(P < 0.05), and the expression of HIF-1α, GLUT4 and HK2 was down-regulated(P < 0.05). Moscatin, densiflorol B, gigantol and moscatilin formed the stable conformation with HIF-1α, GLUT4 and HK2. CONCLUSION DNEE obtained a good anti-lung cancer effect in vivo and in vitro, and its mechanism may be related to the down-regulation of HIF-1α, GLUT4 and HK2 expression to inhibit tumor glycolysis.