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目的观察苔黑酚葡萄糖苷对体外培养的正常肝细胞株L02中CYP3A的影响,探讨苔黑酚葡萄糖苷是否通过cAMP-PKA信号通路调控CYP3A的活性。方法分别用cAMP-PKA信号通路中蛋白激酶A(PKA)激动剂环腺苷酸(cAMP)类似物8-溴-环腺苷酸(8-Br-cAMP)及其抑制剂H-89、苔黑酚葡萄糖苷干预L02细胞,MTT法检测细胞活力,完整细胞免疫组化法检测PKA蛋白表达,红霉素-N-脱甲基法检测CYP3A活性,Western blotting检测细胞中孕烷X受体(PXR)蛋白表达。结果与对照组(加入不含药的无血清培养基)相比,苔黑酚葡萄糖苷处理后的细胞CYP3A的活性增强,PKA、PXR蛋白表达增强,与8-Br-cAMP的作用相似。结论苔黑酚葡萄糖苷可通过cAMP-PKA信号通路调控CYP3A的表达,表明其可能是仙茅表现出辛热药性的主要成分之一。
Objective To observe the effect of orino-glucosidase on CYP3A in normal liver cell line L02 cultured in vitro and to explore whether orcin CYG3A activity is regulated by cAMP-PKA signaling pathway. Methods The cAMP-PKA agonist cyclic AMP (8-Br-cAMP) and its inhibitor H-89 Hepatic phenolglucoside intervention L02 cells, MTT assay of cell viability, whole cell immunohistochemistry PKA protein expression, erythromycin-N-demethylation assay CYP3A activity, Western blotting detection of cells in the pregnant X receptor ( PXR) protein expression. Results Compared with the control group (without serum-free medium), the activity of CYP3A was enhanced and the protein expressions of PKA and PXR were enhanced in orcinoligoside-treated cells, which was similar to that of 8-Br-cAMP. CONCLUSION Eustocin glucoside can regulate the expression of CYP3A through cAMP-PKA signaling pathway, indicating that it may be one of the main components of Curculigo pallidum.