普流罗尼克对P-gp底物罗丹明123在Caco-2细胞蓄积的影响

Effect of Pluronic on Cellular Accumulation of Rhodamine 123,a P-gp Substrate in Caco-2 Cells

-目的研究普流罗尼克（PluronicF127，F68，P85，P123）对小肠P-糖蛋白（P-gP）的调控作用。方法采用MTF法检测不同质量浓度的Pluronic对Caco-2细胞生长抑制作用；以P-gP底物罗丹明123（R-123）为荧光探针，评价各种P1uronic辅料和P-gp抑制剂维拉帕米对R-123细胞蓄积的影响，细胞内的R-123浓度采用高效液相-荧光检测。同时考察了Pluronic P123和F127对P-gPATP酶活性的影响。结果除了L61，经Pluronic处理后的细胞生存率都在80％以上，表明在蓄积实验中的细胞活性没有受到PIuronic的影响。在抑制剂维拉帕米和不同浓度的Pluronic（0．001-50mg·mL^-1）作用下，可在不同程度上提高R-123在Caco-2细胞的蓄积作用，同时具有浓度依赖性，在接近或超过临界胶束浓度（CMC）后，细胞蓄积达到最大值，然后随着Pluronic浓度的继续增大，蓄积作用又逐渐减弱。不同浓度的Pluronic P123和F127对P-gPATP酶活性具有抑制作用。结论蓄积实验结果表明，Pluronic F127，F68，1985和P123可以通过抑制P-gP的作用改善药物的吸收，抑制P-即ATP酶活性可能是原因之一。因此，联合使用Pluronic，有望提高P-gP底物药物的口服生物利用度。

OBJECTIVE To determine whether Pluronic, P123, F127, P85, F68 and L61 possess ability to modulate P-glyco- protein （P-gp） mediated efflux of drugs. METHODS Caco-2 cell viability was measured using MTY assay in the presence of Pluronic at different concentrations. To evaluate the effects of Pluronic and verapamil（ inhibitor of P-gp）on cellular drug accumulation in caco-2 cells, rhodamine 123（R-123） , a P-gp substrate, was selected as fluorescent probe. A HPLC-fluorescence method was used to determined the concentration of R-123 in Caco-2 cells. Caco-2 cell membranes were exposed to various Pluronic solutions（ P123 or F127） , and then the ATPase activity of P-gp was assayed by determining liberated inorganic phosphate. RESULTS The viability of Pluronictreated cells was above 80% except 1.61, which suggested that Caeo-2 cells could maintain their viability under the experiment conditions. In the presence of verapamil and Pluronic at concentrations ranging from 0.00 1 -50 mg· mL^-1 , R-123 accumulation were increased significantly to differernt extent. R-123 accumulation reached maximum levels when Pluronic concentrations were close to or above the CMC, Higher Pturonic concentrations resulted in a drop in accumulation compared to that of R-123 control. Furthermore, P123 and F127 treatments decreased P-gp ATPase activity in Caco-2 cell membranes expressing P-gp. CONCLUSION Pluronic （F68, F127, P123 and P85） can modulate drug efflux by inhibiting P-gp activity in Caeo-2 cell membranes. It may be possible to improve the oral bioavailability of P-gp substrates by co-administration of Pluronic.