目的 研究小槐花Desmodium caudatum抑制肿瘤细胞增殖和转移的作用，并探讨其对Wnt/β-连环蛋白（β-catenin）和腺苷酸活化蛋白激酶（AMPK）信号通路的调控作用。方法 采用不同条件制备8种小槐花提取物，MTT法检测小槐花提取物1～8对HCT-116、SW480细胞增殖的影响；结晶紫染色检测小槐花提取物2、3对结肠癌HCT-116、SW480细胞集落形成的影响；划痕实验检测小槐花提取物2、3对HCT-116、SW480细胞迁移的影响；从活性较高的提取物2中提取分离出5个黄酮类化合物（1～5），MTT法检测化合物1～5对HCT-116、SW480、HepG2、RAW264.7、LX-2细胞增殖的影响；结晶紫染色检测化合物1 （8-异戊烯基槲皮素，0.5、1.0、2.0、5.0、10.0 μmol· L^-1）对HCT-116和SW480细胞集落形成的影响，划痕实验检测8-异戊烯基槲皮素对HCT-116和SW480细胞转移的影响。Western blotting实验检测8-异戊烯基槲皮素对HCT-116和SW480细胞中AMPK和Wnt/β-catenin信号通路相关蛋白表达的影响；荧光素酶报告基因检测实验观察8-异戊烯基槲皮素对β-catenin与T细胞因子（TCF）结合能力的影响。结果 与对照组比较，小槐花根茎提取物2和3以及8-异戊烯基槲皮素能显著抑制结肠癌细胞的增殖和迁移（P<0.05、0.01、0.001）。与对照组比较，8-异戊烯基槲皮素能够显著上调p-AMPK蛋白的表达（P<0.05、0.001），显著下调AMPK、乙酰辅酶A羧化酶（ACC）、肉毒碱棕榈酰基转移酶1A （CPT-1A）和脂肪酸合成酶（FAS）蛋白的表达（P<0.05、0.01、0.001），表明其能够促进癌细胞AMPK信号通路的激活。与对照组比较，8-异戊烯基槲皮素显著下调β-catenin以及其下游糖原合酶激酶3β（GSK-3β）、核内原癌基因（c-Myc）和G₁/S-特异性周期蛋白-D1 （Cyclin D1）的蛋白表达（P<0.05、0.01、0.001），同时显著降低TCF报告基因结合位点（野生型）和荧光素酶开放阅读框TOPflash（pGL3-OT）活性（P<0.05、0.01），但不影响FOPflash（含突变位点）活性，表明其抑制β-catenin的表达和β-catenin/TCF复合物的结合。结论 小槐花黄酮8-异戊烯基槲皮素通过调控Wnt/β-catenin和AMPK信号通路抑制结肠癌细胞增殖和转移。

Objectives To investigate the mechanism of Desmodium caudatum in inhibiting the proliferation andmetastasis of tumor cells and its regulation on Wnt/β-catenin and AMPK signaling pathways. Methods Eight extracts of D. caudatum were prepared under different conditions. MTT assay was used to detect the effect of extract 1-8 on the proliferation of HCT-116 and SW480 cells. Crystal violet staining was used to detect the effect of extract 2 and 3 on colony formation of colon cancer HCT-116 and SW480 cells. Effects of extract 2 and 3 on HCT-116 and SW480 cells migration detected by scratch test. Western blotting assay was used to detect the effect of 8-prenylquercetin on AMPK and Wnt/β-catenin signaling pathway related protein expression in HCT-116 and SW480 cells. The effect of 8-prenylquercetin on the binding ability of β-catenin and T cell factor (TCF) by luciferase reporter gene test. Results Compared with control group, extracts 2 and 3 and 8-prenylquercetin significantly inhibited the proliferation and migration of colon cancer cells. Compared with control group, 8-prenylquercetin could significantly up-regulate the expression of p-AMPK protein (P < 0.05 and 0.001) and down-regulate the expression of AMPK, ACC, p-ACC, CPT-1A and FAS proteins (P < 0.05, 0.01 and 0.001), indicating that it could promote the activation of AMPK signaling pathway in cancer cells. In addition, compared with control group, 8-prenylquercetin downregulated the expression of β-catenin and its downstream proteins pGSK 3β, c-Myc and Cyclin D1 (P < 0.05, 0.01 and 0.001). Compared with control group, 8-prenylquercetin significantly reduced TOPflash activity in a concentration dependent manner, but not FOPflash activity, indicating that it inhibited β-catenin expression and β-catenin/TCF complex binding. Conclusion 8-prenylquercetin a flavonoid from D. caudatum, inhibits the proliferation and metastasis of tumor cells by regulating Wnt/β-catenin and AMPK signaling pathways.

R285.5

江苏省双创团队项目（JSSCTD202133）；天津市自然科学基金项目（18JCYBJC94800）