Research On The Melanin Synthesis Mechanism Of Three Phytoestrogens Regulation A375 Cells

Objective:Researching the influence and regulatory mechanism of three kinds of phytoestrogens(Wogonin,Bavachin,Bavachalcone)from Eucommia and Psoralea on A375 cells,exploring Chinese medicine phytoestrogens to the regulatory mechanism of melanin synthesis from the level of molecular biology,which provide theoretical basis for the application of Chinese medicine phytoestrogens for treatment of skin pigmentation disorders including chloasma.Methods:The Wogonin,Bavachin,Bavachalcone and estrogen receptor inhibitor ICI182780,ERK pathway inhibitor U0126 in A375 cells,respectively by determined by MTT method,NaOH cracking method and L-dopa oxidation method detect the influence of three kinds of phytoestrogens in A375 cell proliferation,melanin content and tyrosinase activity.Using western blot method detect the influence of three kinds of phytoestrogens to cellular tyrosinase and JNK protein expression.Using RT-PCR method detect the influence of three kinds of phytoestrogens on tyrosinase and its related proteins and key protein kinase mRNA expression in MAPK pathway on A375 cells.And detect estrogen receptor blockers ICI1182780 and ERK pathway blockers U0126 for three kinds plant estrogen effects.Results:Wogonin(10μmol·L-1、1μmol·L-1、10-1μmol·L-1、10-2μmol·L-1、10-3μmol·L-1)、Bavachin(1μmol·L-1、10-1μmol·L-1、10-2μmol·L-1、10-3μmol·L-1)、Bavachalcone(1μmol·L-1、10-1μmol·L-1、10-3μmol·L-1)have no significant effect on A375 cell proliferation,but the amount of melanin synthesis and TYR activity in A375 cells have different degrees of inhibition(P<0.01 or P<0.05).After blocker ICI182780(1 μmol·L-1)and U0126(10μmol·L-1)respectively part of block the effects of Wogonin(10μmol·L-1)、Bavachin(10μmol·L-1)and Bavachalcone(1μmol·L-1),comparing with the drug group,can reverse the downward effect of the drug groups on melanin synthesis and TYR activity(P<0.01 or P<0.05).Wogonin(10μmol·L-1)、Bavachin(10μmol·L-1)and Bavachalcone(1μmol·L-1)have different degree of inhibition of TYR and JNK protein expression,while this effect can be ICI182780 inhibition(P<0.01).Wogonin(10μmol·L-1)、Bavachin(10μmol·L-1)and Bavachalcone(1μmol·L-1)have different degree of inhibition of TYR、TRP-1、TRP-2、ERK 1、ERK2 and JNK2 mRNA expression(P<0.01or P<0.05),while this effect can be ICI182780 and U0126 inhibition(P<0.01or P<0.05).Conclusion:Three kinds of phytoestrogen including Wogonin,Bavachin,Bavachalcone can reduce the amount of TYR activity and protein expression,cut TYR,TRP-1 and TRP-2 mRNA expression,thus inhibiting melanin synthesis of A375 cells.This effect is closely related to ER and its downstream JNK,ERK kinase signaling pathways.