Inhibition Effect Of The Natural Active Component Ginkgolide B On P-glycoprotein

Ginkgolide B (GB) is one of an important active ingredient in ginkgo biloba leaf extract, and it has obvious protective effects on cardiovascular and cerebrovascular diseases. GB prodrug (PGB) synthesized by our engineering center, is characterized by its brain targeting in our previous study. Weakened P-glycoprotein (P-gp) efflux capacity is beneficial to improve the brain targeting of PGB, whether P-gp could regulate the brain targeting of PGB remains unknown. In this thesis, we will study the P-gp affinity in vitro, the expression level of P-gp, the transport function of P-gp in the presence of PGB and GB, which will help to explore the regulation mechanism of P-gp in the brain targeting of PGB. The results are as follows:(1) PGB has a slightly higher affinity with P-gp in vitro than that of GB.P-gp ATPase analysis showed that PGB and GB can stimulate P-gp ATPase activity to some degree in a time-dependent fashion and in a concentration-dependent manner. The data of ATPase activity is processed by two methods which are double reciprocal and nonlinear fitting methods, the affinity is represented by Vmax/Km.PGB has slightly higher affinity with P-gp than that of GB, which implies that the inhibition of PGB on P-gp efflux is stronger than that of GB. It is to say that PGB is easily to be transported out of the brain when PGB and decomposed GB fromPGB existed in brain simultaneously, which weakened or blocked the efflux of GB. As a result, decomposed GB has a more accumulation in the brain, thus brain-targeting of PGB is enhanced in some extent.(2) PGB has a stronger inhibition on the expression of P-gp than that of GB in rat brain microvascular endothelial cells (rBMECs).In the concentration range of 5-100 ?mol-L-1, an obvious inhibition on the expression of P-gp was found in PGB-treated or GB-treated group compared with control group in rBMECs (P<0.01). Also the inhibition showed a concentration-dependent manner. Under the same treatment concentration, the inhibition effect of PGB on the expression of P-gp was stronger than that of GB (P< 0.01).(3) PGB has significant inhibitory effect on the expression of P-gp in the hippocampus of rats.P-gp has a higher expression in hippocampus of normal rats. Compared with the control group, tail vein injection of GB (10 mg/kg) has almost no effect on the expression level of P-gp in the hippocampus of rats after 0.1-8 h of administration. However, the injection of PGB (12.83 mg/kg) could significantly suppress the expression of P-gp in hippocampus of rats (P< 0.01). On the other hand, PGB has stronger inhibition on the expression of P-gp than that of GB (P<0.01).(4) PGB significantly inhibited P-gp efflux function of rat brain.Rhodamine 123 (Rho123), the reprensentative P-gp substrate, is used as a fluorescence probe in this study. The effect of GB, PGB on P-gp efflux function of rat brain was assessed. Compared with the control group, pretreatment with PGB can effectively improve the concentration of Rho123 in the brain and the Kp value. The Kp value was increased from 0.097 to 0.224 when the administration continued for 0.5 h or 1.5 h. Also, PGB can inhibit the P-gp efflux function in rat brain, and the inhibition was stronger than that of GB (P< 0.01).