目的 对鲜地黄、生地黄、熟地黄和地黄叶进行了雌激素样活性筛选。方法 首先通过小鼠子宫增重实验和MCF-7细胞增殖实验对选定的四种药材进行雌激素样活性筛选;接着采用ICI182,780拮抗实验及以pERE-TAL-luc、pβgal-Control、pCXN2-hERα或pCXN2-hERβ共转染的HEK293细胞,以ERE调控的报告基因luc瞬时表达检测进行了雌激素样作用机制初探。结果 鲜地黄水提物和生地黄水提物能够诱导性未成熟雌性小鼠子宫系数的增加,且两者均能够促进MCF-7细胞的增殖,当与雌激素受体阻断剂ICI182,780同时干预时两者对细胞的促增殖作用被抑制;ERE调控的报告基因瞬时表达结果显示,由ERβ介导时,鲜地黄水提物与生地黄水提物的荧光素酶活性均显著高于正常组。结论 鲜地黄水提物和生地黄水提物体内、体外均具有雌激素样活性,且都主要通过ERβ介导发挥。不同炮制品中鲜地黄的雌激素样作用最强,生地黄其次,熟地黄没有雌激素样作用,提示地黄炮制后会使其雌激素样作用发生改变。
Abstract
To screen the estrogenic effects of fresh Radix Rehmanniae, dried Radix Rehmanniae, Radix Rehmanniae Praeparata and the leaves of Rehmannia glutinosa Libosch. METHODS Mouse uterine weight test and MCF-7 cell proliferation assay were used to evaluate the estrogenic effects of the four kinds of Chinese traditional herbs. ICI182,780 antagonnist assay and reporter gene assay were adopted to explore the mechanism of action of fresh and dried Radix Rehmanniae. In reporter gene assay, HEK293 cells were cotranfected with pERE-TAL-luc, pβgal-Control, pCXN2-hERα or pCXN2-hERβ, and the expression of reportr gene luc was controlled by ERE. RESULTS Mouse uterine weight test showed that compared with the control group, fresh and dried Radix Rehmanniae could increase the uterus index of premature female mice, and both of them could promote the proliferation of MCF-7 cells. Co-incubation of MCF-7 cells with estrogen receptor blocker ICI182,780 abolished the inductive effect of the proliferation. The reporter gene controlled by ERE technology showed that when mediated by ERβ, the normalized luciferase activity of the two groups were significantly higher than the activity of the control group. CONCLUSION Fresh and dried Radix Rehmanniae have estrogenic activities which are mainly mediated by ERβ. Fresh Radix Rehmanniae has higher estrogenic activity than dried Radix Rehmanniae.Radix Rehmanniae Praeparata does not have estrogenic activity. The estrogenic activity may change during the processing process of Rehmannia glutinosa Libosch.
关键词
怀地黄 /
雌激素样活性 /
性未成熟小鼠子宫 /
MCF-7细胞 /
报告基因 /
ERβ /
ICI182,780
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Key words
Rehmannia glutinosa Libosch /
estrogenic activity /
uterus /
premature female mice /
MCF-7 cell /
reporter gene /
estrogen receptor beta /
ICI182,780
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参考文献
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脚注
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基金
十二五国家科技支撑计划项目(2011BAI06B02);博士科研基金项目(BSJJ2009-17);河南省2010年科技发展计划项目(102102310343)。
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