Effect of Butein on the Biological Activity of Co-Cultured Human Melanoma Cells A375 and Human Immortalized Keratinocytes HaCaT
Objective To investigate the effect of butein on the biological activity of co-cultured human melanoma cells A375 and human immortalized keratinocytes HaCaT.Methods Co-cultured A375 and HaCaT cells were established and divided into the control group(equal volume of culture medium),the butein low-,medium-and high-dose groups(hereinafter referred to as the low-,medium-and high-dose groups,0.5,1.0,5.0 μg/L).The cell activity was detected by the methyl thiazolyl tetrazolium(MTT)method.The cell cycle and mitochondrial membrane potential were detected by the flow cytometry.The expression levels of stem cell factor(SCF),its receptor c-Kit,microphthalmia-associated transcription factor(MITF)mRNA were detected by the real-time fluorescence quantitative polymerase chain reaction(qPCR).The expression levels of tyrosinase-related protein-1(TRP-1),tyrosinase-related protein-2(TRP-2),SCF,c-Kit and MITF proteins in cells were detected by the Western blot.Results Compared with that in the control group,the proliferation rate in the medium-and high-dose groups significantly increased(P＜0.01).Compared with those in the control group,the proportion of cells at G0/G1 phase in the low-,medium-and high-dose groups significantly decreased,while the proportion of cells at S and G2/M phases significantly increased(P＜0.01);the proportion of cells with high mitochondrial membrane potential significantly increased(P＜0.05 or P＜0.01);the expression levels of SCF,c-Kit,MITF mRNA and the expression levels of SCF,c-Kit,MITF,TRP-1 and TRP-2 proteins significantly increased(P＜0.05 or P＜0.01).Conclusion Butein may affect the proliferation,cell cycle and mitochondrial membrane potential of co-cultured A375 and HaCaT cells by regulating the expression of SCF,c-Kit and MITF.
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