人KRT2促进体外培养的羊驼皮肤黑色素细胞的黑色素生成

发布时间：2018-07-04 17:37

本文选题：角蛋白 + 黑色素生成　；参考：《中国生物化学与分子生物学报》2017年03期

【摘要】：基因表达谱显示,绵羊角蛋白2(keratin2,Krt2)的mRNA在不同毛色皮肤的表达不同,暗示Krt2基因可能对皮肤黑色素生成有一定的影响。为探索角蛋白2对体外培养的羊驼皮肤黑色素细胞黑色素生成的影响,首先采用PCR扩增产物测序,结合DNAMAN软件比对分析发现,羊驼Krt2编码序列(c DNA)与NCBI公布的人KRT2高度同源(91%);将人KRT2添加于培养的羊驼皮肤黑色素细胞,观察人KRT2对羊驼皮肤黑色素细胞黑色素的生成作用。免疫组织化学显示,外源性的人KRT2处理羊驼皮肤黑色素细胞72 h后,黑色素细胞的细胞质中Krt2表达增强。实时定量PCR及Western印迹实验揭示,与胎牛血清清蛋白处理的羊驼皮肤黑色素细胞比较,1 ng/m L、10 ng/m L和100 ng/m L人KTR2处理的羊驼皮肤黑色素细胞酪氨酸酶(Tyr)、酪氨酸相关蛋白-1(Tyrp1)、小眼畸形相关转录因子(Mitf)基因表达明显上调(P0.05);尤其在添加10 ng/m L KTR2的细胞中,3个基因的mRNA相对表达水平升高尤其显著,分别是对照细胞的4倍、10倍和12.9倍(P0.01),蛋白质相对表达水平分别是对照细胞的2倍、2.1倍和1.7倍(P0.01)。分光光度法测量A490结果证明,1 ng/m L、10 ng/m L和100 ng/m L人KTR2处理的羊驼皮肤黑色素细胞产生的黑色素含量分别是对照细胞的1.3倍(P0.05)、1.8倍(P0.01)和1.5倍(P0.05)。上述结果说明,人KTR2处理可通过刺激羊驼皮肤黑色素细胞黑色素合成相关信号通路,促进黑色素的合成。
[Abstract]:Gene expression profiles showed that the mRNA expression of keratin2 (keratin2) was different in different skin, suggesting that Krt2 gene might have an effect on melanin production. In order to explore the effect of keratin 2 on melanin production in the skin of alpaca in vitro, the PCR amplification products were sequenced and the results were compared with DNA man software. Human KRT2 coding sequence (c DNA) was highly homologous to human KRT2 published by NCBI (91%), and human KRT2 was added to cultured melanocytes of alpaca skin to observe the effect of human KRT2 on melanin production in alpaca skin. Immunohistochemistry showed that the expression of Krt2 in the cytoplasm of melanocytes increased after 72 h treatment with exogenous human KRT2. Real-time quantitative PCR and Western blot analysis revealed that, Comparison of melanocytes in Alpaca skin treated with Fetal Bovine Serum Albumin 10 ng/m / L and 100 ng/m / L Human KTR2 in Alpaca skin tyrosinase (Tyr), tyrosine Associated protein 1 (Tyrp1) and microocular deformation-Associated transcription Factor (MIF) The relative expression level of three genes was significantly increased in the cells supplemented with 10 ng/m L KTR2 (P0.05), especially in the cells supplemented with 10 ng/m L KTR2. The relative protein expression levels were 2.1 times and 1.7 times of that of the control cells (P0.01), respectively, compared with the control cells (4 times, 10 times and 12.9 times (P0.01), respectively), and the relative protein expression levels of the cells were 2.1 times and 1.7 times of the control cells, respectively. The results of spectrophotometry showed that the melanin content of skin melanocytes treated with 1 ng/m / L 10 ng/m / L and 100 ng/m / L human KTR2 was 1.3-fold (P0.05) and 1.5-fold (P0.05) as compared with the control cells, respectively. These results suggest that human KTR2 treatment can promote melanin synthesis by stimulating melanocyte melanin synthesis signal pathway in alpaca skin.
【作者单位】：山西农业大学动物科技学院;河南科技大学动物科技学院;
【基金】：国家高技术研究发展计划(863计划)(No.2013AA102506)) 公益性行业(农业)科研专项(201303119)资助基金项目山西省自然科学基金(2012011046-6)资助~~
【分类号】：S829.9

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