摘要
目的探讨紫铆花素对人黑色素瘤细胞A375与人永生化角质形成细胞HaCaT共培养细胞生物学活性的影响。方法建立A375细胞与HaCaT细胞的共培养细胞体系,实验分为对照组(等体积培养液),紫铆花素低、中、高剂量组(以下简称低、中、高剂量组,0.5,1.0,5.0μg/L)。采用四甲基偶氮噻唑蓝(MTT)法检测细胞活性;采用流式细胞仪检测细胞的周期分布和线粒体膜电位;采用实时荧光定量聚合酶链反应(qPCR)法检测干细胞因子(SCF)和其受体c-Kit,以及小眼畸形相关转录因子(MITF)的mRNA表达水平;采用Western blot法检测细胞酪氨酸酶相关蛋白1,2(TRP-1,TRP-2)及SCF,c-Kit,MITF的蛋白表达水平。结果与对照组比较,中、高剂量组细胞的增殖率均显著升高(P<0.01);与对照组比较,各剂量组G0/G1期细胞比例均显著降低,S期和G_(2)/M期细胞比例均显著升高(P<0.01),高线粒体膜电位细胞比例均显著升高(P<0.05或P<0.01),SCF,c-Kit,MITF的mRNA表达水平及SCF,c-Kit,MITF,TRP-1,TRP-2的蛋白表达水平均显著升高(P<0.05或P<0.01)。结论紫铆花素可能通过调节SCF,cKit,MITF的表达而影响A375细胞与HaCaT细胞共培养细胞的增殖、细胞周期和线粒体膜电位等细胞生物学活性。
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,0.5,1.0,5.0μg/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 G_(2)/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.
作者
高莉
谭雪
罗静莺
闫明
GAO Li;TAN Xue;LUO Jingying;YAN Ming(Key Laboratory of Traditional Uyghur Medicine Prescription,Institute of Xinjiang Traditional Uyghur Medicine,Urumqi,Xinjiang,China 830011)
出处
《中国药业》
CAS
2024年第6期36-39,共4页
China Pharmaceuticals
基金
国家自然科学基金[81260689]
新疆维吾尔自治区自然科学基金重点项目[2022D01D25]。
