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目的:通过加热回流的方法提取蓝萼香茶菜中二萜类物质,测定其含量,研究二萜类物质对乳腺癌癌前病变肝郁肾虚证模型大鼠的干预作用及其作用机制。方法:雌性SD大鼠随机分层分组为正常组、枸橼酸他莫昔芬片组、模型组、蓝萼组。除正常组外其他各组均用二甲基苯蒽(DMBA)结合慢性不可预见应激行为制备乳腺癌癌前病变肝郁肾虚证模型,造模结束后,观察记录各组大鼠的生物表征变化及病理变化验证模型。蓝萼组给予二萜类提取物水溶液5 mg·kg-1灌胃,正常组和模型组给予蒸馏水10 m L·kg-1灌胃,各组均连续灌胃30 d。30 d后,记录各组大鼠肉眼可见肿瘤的发生情况;腹腔取血,使用酶联免疫吸附试验(ELISA)试剂盒检测血清中雌二醇(E2),孕激素(P)水平;采集乳腺织织,用苏木素-伊红(HE)染色观察乳腺组织病理学变化,实时荧光定量PCR(Real-time PCR)及蛋白免疫印迹法(Western blot)检测乳腺组织血管内皮生长因子(VEGF),趋化因子受体4(CXCR4)mRNA及蛋白表达水平。结果:提取得干燥品质量2.162 g,干燥品中二萜类质量分数为26.35%。与模型组比较,蓝萼组血清中E2,P水平明显降低(P<0.05),乳腺组织中VEGF,CXCR4蛋白及mRNA表达水平均明显降低(P<0.05)。结论:蓝萼香茶菜二萜类物质有可能通过抑制VEGF,CXCR4蛋白及mRNA表达,抑制血管生成和趋化因子受体表达,调节大鼠激素水平,从而抑制大鼠乳腺癌癌前病变癌变的发生。
OBJECTIVE: To extract the diterpenoids from Chaxiang Camellia oleifera by heating and refluxing, and to study the effect of diterpene on rats with precancerous lesions of liver-Qi, kidney and kidney of breast cancer. Methods: Female SD rats were randomly divided into normal group, tamoxifen citrate group, model group and blue calyx group. In addition to the normal group, other groups were prepared with DMBA combined with chronic unpredictable stress behavior of breast cancer precancerous lesions of liver stagnation and kidney deficiency model, after modeling, the biological characteristics of the rats were observed and recorded Change and pathological changes validation model. The calyx group was given diterpene extract aqueous solution 5 mg · kg-1 gavage, normal group and model group were given distilled water 10 m L · kg-1 gavage, each group were continuous gavage 30 d. Thirty days later, the occurrence of macroscopic tumor in each group was recorded. Blood was collected from the abdominal cavity and the serum estradiol (E2) and progesterone (P) levels were detected by enzyme-linked immunosorbent assay (ELISA) kit. The tissue histopathology was observed by hematoxylin-eosin (HE) staining. The vascular endothelial growth factor (VEGF) in breast tissue was detected by Real-time PCR and Western blot. Factor 4 (CXCR4) mRNA and protein expression levels. Results: The quality of dried extract was 2.162 g, and the dry content of diterpenoid was 26.35%. Compared with the model group, the levels of E2 and P in the sepal group were significantly decreased (P <0.05), and the levels of VEGF, CXCR4 protein and mRNA in the mammary gland tissue were significantly decreased (P <0.05). CONCLUSION: Diterpenoids from Chaetomium glauca could inhibit the expression of VEGF, CXCR4 protein and mRNA, inhibit angiogenesis and chemokine receptor expression, and regulate hormone levels in rats, thereby inhibiting the carcinogenesis of breast precancerous lesions in rats happened.