染料木黄酮对光老化成纤维细胞增殖作用的影响  

Effect of genistein on proliferation of photo-aged fibroblasts

作　　者：谭春花[1] 罗晓燕[1] 王华[1] 

机构地区：[1]重庆医科大学附属儿童医院皮肤科,重庆400014

出　　处：《中国生物制品学杂志》2014年第1期84-88,共5页Chinese Journal of Biologicals

基　　金：2011年中华医学会-欧莱雅中国人健康皮肤/毛发研究项目(S2011-012)

摘　　要：目的探讨染料木黄酮对光老化成纤维细胞增殖作用的影响。方法经酶消化法分离培养真皮成纤维细胞,并进行细胞传代,免疫细胞化学鉴定细胞。实验分为3组:正常细胞对照组:未经处理的真皮成纤维细胞;8-甲氧补骨脂素(8-methoxypsoralan,8-MOP)/长波紫外线(ultraviolet A,UVA)对照组:用含50 ng/ml 8-MOP的培养基避光孵育细胞24 h,再以9 J/cm2UVA照射;8-MOP/UVA+染料木黄酮(0、1.25、2.5、5、10、20μg/ml)组:用含50 ng/ml8-MOP及各浓度染料木黄酮的培养基共同孵育细胞24 h,再以9 J/cm2UVA照射24 h。MTT法检测各组细胞增殖活力,流式细胞术检测各组细胞细胞周期,光镜下观察各组细胞分裂和分裂后表型,实时荧光定量PCR法检测各组细胞中基质金属蛋白酶-1、3水平。结果皮肤成纤维细胞阳性率达99%以上;8-MOP/UVA+染料木黄酮1.25、2.5μg/ml组细胞增殖率分别为8-MOP/UVA对照组的112.6%(P>0.05)和146.6%(P<0.05),染料木黄酮的光保护作用随浓度的升高而增强,然而随着浓度的继续升高,细胞增值率逐渐降低,当浓度达20μg/ml时,细胞增殖率仅为8-MOP/UVA对照组的19.4%(P<0.01),选择染料木黄酮的最佳保护浓度为2.5μg/ml;8-MOP/UVA对照组细胞细胞周期阻滞于S期,而8-MOP/UVA+染料木黄酮2.5μg/ml组细胞细胞周期则由S期进入G2/M期;正常细胞对照组、8-MOP/UVA组及8-MOP/UVA+染料木黄酮2.5μg/ml组分裂细胞表型占总细胞数目的比率分别为(99.7±0.58)%、(7.7±1.15)%、(57.7±3.51)%,各组间差异有统计学意义(P<0.01);8-MOP/UVA+染料木黄酮2.5μg/ml组细胞基质金属蛋白酶-1、3水平均明显低于8-MOP/UVA对照组,而高于正常细胞对照组,各组间差异均有统计学意义(P<0.05)。结论染料木黄酮可在一定程度上解除8-MOP/UVA所致光老化成纤维细胞的生长抑制作用。Objective To investigate the effect of genistein on proliferation of photo-aged fibroblasts. Methods Dermal fibroblasts were isolated and cultured by enzyme digestion, then subeultured and subjected to immunocytochemieal assay. The fibroblasts were divided into normal control, 8-methoxypsoralan （8-MOP）/ultraviolet A （UVA） control and 8-MOP/ UVA ＋ genistein groups. The fibroblasts in normal control group were untreated, while those in 8-MOP/UVA group were incubated in medium containing 50 ng/ml containing 8-MOP for 24 h, protected from light, and subjected to radiation with UVA at a dosage of 9 J /em2. However, the fibroblasts in 8-MOP / UVA ＋ genistein group were incubated in media containing 50 ng/ml 8-MOP and various concentrations （0, 1.25, 2. 5, 5, 10 and 20 μg/ml） of genistein for 24 h re- spectively, then subjected to a radiation with UVA at a dosage of 9 J / cm2 for 24 h. The proliferative activities of fibrob- lasts in various groups were determined by MTT assay, while cell cycle by flow cytometry, the mitotic and postmitotic phenotypes by optical microscopy, and the expression levels of matrix metal proteases（MMP）-I and -3 by real-time fluores- cent quantitative PCR. Results The positive rate of dermal fibroblasts was more than 99%. The proliferation rates of fi- broblasts treated with 8-MOP/UVA ＋ genistein at concentrations of 1. 25 and 2. 5μg/ ml group were 112. 6% （P 〉 0. 05） and 146. 6 （P 〈 0.05） of those in 8-MOP/UVA control group respectively. The photoprotection of genistein was enhanced with the increasing concentration, while the proliferation rate of fibroblasts decreased gradually and was only 19.4% of that in 8-MOP/UVA control group （P 〈 0. 01） when the concentration was 20 μg/ml. The optimal genistein concentration for photoprotection was 2. 5μg / ml. The cell cycle of fibroblasts in 8-MOP / UVA control group was arrest-ed at S phase, while that in 8-MOP/UVA ＋ 2. 5 μg/ml genistein group entered from S phase to G2/M phase. The mitot- ic fibroblas

关 键 词：染料木黄酮 8-甲氧补骨脂素 紫外线 光老化 成纤维细胞 

分 类 号：Q253[生物学—细胞生物学]