α-MSH和lpa-miR-nov-66在羊驼黑色素细胞内的作用研究
发布时间：2019-06-14 20:53
【摘要】：lpa-miR-nov-66是羊驼皮肤中发现的新miRNA,在黑色素生成过程中起重要的调控作用。a-MSH是一种重要的神经内分泌激素,通过调控MC1R影响皮肤黑色素的生成和分布。本实验通过在黑色素细胞内转染lpa-miR-nov-66和外源性添加一定浓度的a-MSH,分析二者在羊驼黑色素细胞内对黑色素生成的影响,以探讨二者的相互作用。主要实验结果如下：1用实时细胞检测仪监测细胞增殖,实验35小时后,与对照组相比,各实验组表现出不同的增殖情况,但没有显著差异。2用qRT-PCR技术和Western blotting检测基因情况和和蛋白表达量,结果如下：添加a-MSH后.lpa-miR-nov-66表达量是正常羊驼黑色素细胞的0.24±0.012倍,并呈差异极显著(P0.01)；与对照组相比,转染组、添加组、转染添加组中MITF的基因和蛋白表达量分别下降0.16±0.014倍/0.90±0.07倍、2.03±0.051倍/1.24±0.09倍、0.30±0.013倍/1.12±0.06倍,并呈差异极显著(P0.01); TYR的表达量分别下降了0.73±0.033倍/0.85±0.05倍、上调了4.67±0.14倍/1.69±0.14倍、2.54±0.21倍/1.27±0.11倍,并呈差异极显著(P0.01)。3用ELISA方法检测细胞内cGMP和cAMP含量的变化情况,结果发现：与对照组相比,转染组、添加组、转染添加组中cGMP和cAMP的含量分别是对照组的0.149±0.012倍、2.68±0.17倍和2.267±0.12倍：0.57±0.042倍、2.07±0.19倍和1.36±0.124倍,且均呈差异极显著(P0.01)。4用酶标法检测各实验组中黑色素产量变化,结果发现：与对照组相比,黑色素的产量分别是对照组的0.58±0.043倍、3.27±0.27倍和1.81±0.16倍,且呈差异极显著(P0.01)。由此得出结论：lpa-miR-nov-66和a-MSH协同作用,通过调控羊驼黑色素细胞中毛色形成的cAMP路径,从而影响黑色素的产生。
[Abstract]:Lpa-miR-nov-66 is a new miRNA, found in alpaca skin, which plays an important role in melanogenesis. A-MSH is an important neuroendocrine hormone, which affects the production and distribution of melanin in alpaca skin by regulating MC1R. In this study, the effects of lpa-miR-nov-66 and exogenous a / MSH on melanin production in alpaca melanocytes were analyzed in order to explore the interaction between them. The main results were as follows: 1 the proliferation of cells was monitored by real-time cell detector. 35 hours later, compared with the control group, each experimental group showed different proliferation, but there was no significant difference. 2 the gene and protein expression were detected by qRT-PCR and Western blotting, the results were as follows: after adding a-MSH, the expression of LPA-miR-nov-66 was 0.24 卤0.012 times higher than that of normal alpaca melanocytes. The difference was very significant (P 0.01). Compared with the control group, the expression of MITF gene and protein decreased by 0.16 卤0.014 times / 0.90 卤0.07 times, 2.03 卤0.051 times / 1.24 卤0.09 times, 0.30 卤0.013 times / 1.12 卤0.06 times, respectively, and the difference was very significant (P 0.01). The expression of TYR decreased by 0.73 卤0.033 times / 0. 85 times, increased by 4. 67 卤0. 14 times / 1. 69 卤0. 14 times, 2. 54 卤0. 21 times / 1. 27 卤0. 11 times, and the difference was very significant (P01). 3 the changes of cGMP and cAMP contents in the cells were detected by ELISA method, and the results showed that compared with the control group, the transfection group, the addition group, The contents of cGMP and cAMP in the control group were 0.149 卤0.012 times, 2.68 卤0.17 times and 2.267 卤0.12 times as much as those in the control group: 0.57 卤0.042 times, 2.07 卤0.19 times and 1.36 卤0.124 times, respectively, and there were significant differences (P 0.01). 4 the melanin production in each experimental group was detected by enzyme labeling method, and the results showed that the melanin yield was 0.58 卤0.043 times higher than that in the control group, respectively. 3.27 卤0.27 times and 1.81 卤0.16 times, and the difference was very significant (P 0.01). It is concluded that the synergistic effect of lpa-miR-nov-66 and a-MSH affects the production of melanin by regulating the cAMP pathway of hair color formation in alpaca melanocytes.
【学位授予单位】：山西农业大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：S829.9
本文编号：2499687
[Abstract]:Lpa-miR-nov-66 is a new miRNA, found in alpaca skin, which plays an important role in melanogenesis. A-MSH is an important neuroendocrine hormone, which affects the production and distribution of melanin in alpaca skin by regulating MC1R. In this study, the effects of lpa-miR-nov-66 and exogenous a / MSH on melanin production in alpaca melanocytes were analyzed in order to explore the interaction between them. The main results were as follows: 1 the proliferation of cells was monitored by real-time cell detector. 35 hours later, compared with the control group, each experimental group showed different proliferation, but there was no significant difference. 2 the gene and protein expression were detected by qRT-PCR and Western blotting, the results were as follows: after adding a-MSH, the expression of LPA-miR-nov-66 was 0.24 卤0.012 times higher than that of normal alpaca melanocytes. The difference was very significant (P 0.01). Compared with the control group, the expression of MITF gene and protein decreased by 0.16 卤0.014 times / 0.90 卤0.07 times, 2.03 卤0.051 times / 1.24 卤0.09 times, 0.30 卤0.013 times / 1.12 卤0.06 times, respectively, and the difference was very significant (P 0.01). The expression of TYR decreased by 0.73 卤0.033 times / 0. 85 times, increased by 4. 67 卤0. 14 times / 1. 69 卤0. 14 times, 2. 54 卤0. 21 times / 1. 27 卤0. 11 times, and the difference was very significant (P01). 3 the changes of cGMP and cAMP contents in the cells were detected by ELISA method, and the results showed that compared with the control group, the transfection group, the addition group, The contents of cGMP and cAMP in the control group were 0.149 卤0.012 times, 2.68 卤0.17 times and 2.267 卤0.12 times as much as those in the control group: 0.57 卤0.042 times, 2.07 卤0.19 times and 1.36 卤0.124 times, respectively, and there were significant differences (P 0.01). 4 the melanin production in each experimental group was detected by enzyme labeling method, and the results showed that the melanin yield was 0.58 卤0.043 times higher than that in the control group, respectively. 3.27 卤0.27 times and 1.81 卤0.16 times, and the difference was very significant (P 0.01). It is concluded that the synergistic effect of lpa-miR-nov-66 and a-MSH affects the production of melanin by regulating the cAMP pathway of hair color formation in alpaca melanocytes.
【学位授予单位】：山西农业大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：S829.9
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1 潘国焰;林荣;吴兵;洪美满;陈乘波;黄雪娥;;环磷酸鸟苷/蛋白激酶G信号通路的激活与再灌注损伤挽救激酶信号通路的关系[J];重庆医科大学学报;2013年12期
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