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 study the regulative effects of orcinol glucoside on cytochrome P4503A (CYP3A) of L02 cells and to investigate whether orcinol glucoside regulates the CYP3A activity through cAMP-PKA signal pathway. Methods The L02 cells were treated with 8-Br-cAMP, an analogue of protein kinase A (PKA) agonist (cyclic adenosine monophosphate, cAMP), its inhibitor (H-89), and orcinol glucoside; MTT method was used to detect the cell vitality. The expression of pregnane X receptor (PXR) and PKA protein was detected by Western-blotting and immunohistochemistry, respectively; The activity of CYP3A was detected by erythromycin-N-demethylase. Results Compared with the control group, the activity of CYP3A was increased, the expression of PXR and PKA was increased in L02 cells treated with orcinol glucoside. Conclusion These data support that orcinol glucoside which is a singery effective component in the rhizoma of Curculigo orchioides could regulate CYP3A in L02 cells through cAMP-PKA singal pathway.

国家重点基础研究发展计划(973计划)项目(2007CB512605);陕西省教育厅课题(2010JK493)