为了研究内皮素-3(Endothelin-3,EDN3)对羊驼皮肤黑色素细胞内毛色形成相关基因的影响。本研究在体外培养的羊驼皮肤黑色素细胞中添加不同浓度(0、10-9、10-8、10-7 mol·L-1)的EDN3,通过MTT法、紫外分光光度法、qRT-PCR和Western blotting技术分别检测黑色素细胞活力、黑色素产量、相关基因和蛋白(包括内皮素受体B(Endothelin receptor B,EDNRB)、KIT、小眼畸形相关转录因子(Microphthalmia-associtated transcriptionfactor,MITF)和酪氨酸酶(Tyrosinase,TYR))的表达情况。结果表明,在羊驼皮肤黑色素细胞添加EDN3 72h后,黑色素细胞呈双树突或三树突状,且细胞数量明显增加;在添加适当浓度10-8 mol·L-1时,细胞具有明显的增殖,细胞内EDNRB、KIT、MITF和TRY在转录水平和蛋白水平的表达量被上调,黑色素细胞产生黑色素的量也被上调,且与对照组相比,差异显著(P〈0.05)。EDN3对羊驼黑色素细胞内黑色素的产生具有重要的调节作用。
The study aimed to investigate the effects of endothelin-3 (EDN3) on the expression of hair color genes of alpaca. The activity of melanocyte, the production of melanin in melanocytes and the expressions of related genes and proteins including Endothelin receptor B (EDNRB), KIT, Microphthalmia-associtated transcription factor (MITF) and Tyrosinase (TYR) in alpaca melanoeytes in cultures with addition of different dose of EDN3(0, 10-9 , 10-8 , 10-7mol · L-1) , were examined by MTT, ultraviolet spectrophotometry, qRT-PCR and Western blotting, respec- tively. The results showed that the amount of melanocytes was increased with the increasing double- or tridendrites At the addition dose of 10-8 tool· L-1 for 72 h after adding EDN3, compared to control group, the proliferation of melanocytes was obvious and the expressions of EDNRB, KIT, MITF and TYR significantly up-regulated at the level of both transcription and translation in melanocytes(P〈0.05). Moreover, the production of melanin in melanocytes sig- nificantly increased(P〈0.05). These findings indicate that EDN3 play an important role in regu- lating melanins production in melanocytes in alpaca.