| nicastrin基因沉默的斑马鱼模型构建及其色素异常机制的初步研究 |
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| 引用本文： | 李文锐 贾苇雪 张蕴斌 林麟 李诚让. nicastrin基因沉默的斑马鱼模型构建及其色素异常机制的初步研究[J]. 中华皮肤科杂志, 2021, 54(5): 402-407. DOI: 10.35541/cjd.20200617 |
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| 作者姓名： | 李文锐 贾苇雪 张蕴斌 林麟 李诚让 |
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| 作者单位： | 1中国医学科学院、北京协和医学院皮肤病医院皮肤科江苏省皮肤病与性病分子生物学重点实验室，南京210042；2中国科学院生物化学与细胞生物学研究所，上海200031 |
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| 基金项目： | 国家自然科学基金（81472872）；中国医学科学院医学与健康科技创新工程项目（2016-I2M-1-002） |
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| 摘 要： | 目的:探讨nicastrin（nct）基因对斑马鱼黑素细胞生物学功能的影响。方法:利用吗啉代寡核苷酸（MO）技术针对斑马鱼nct基因mRNA设计nct-MO序列，同时设计对照MO序列（ctrl-MO），并构建5′端为MO靶序列的增强绿色荧光蛋白（EGFP）mRNA，通过显微注射的方式将nct-MO或ctrl-MO与EG...
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| 关 键 词： | 色素沉着异常 化脓性汗腺炎 Nicastrin基因 γ分泌酶 斑马鱼模型 |
| 收稿时间： | 2020-06-18 |
Construction of a nicastrin gene-silenced zebrafish model and a primary study on the mechanism of abnormal pigmentation |
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| Li Wenrui,Jia Weixue,Zhang Yunbin,Lin Lin,Li Chengrang. Construction of a nicastrin gene-silenced zebrafish model and a primary study on the mechanism of abnormal pigmentation[J]. Chinese Journal of Dermatology, 2021, 54(5): 402-407. DOI: 10.35541/cjd.20200617 |
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| Authors: | Li Wenrui Jia Weixue Zhang Yunbin Lin Lin Li Chengrang |
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| Affiliation: | 1Department of Dermatology, Hospital of Dermatology, Chinese Academy of Medical Sciences and Peking Union Medical College, Jiangsu Key Laboratory of Molecular Biology for Skin Diseases and STIs, Nanjing 210042, China; 2Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, Shanghai 200031, China |
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| Abstract: | 【Abstract】 Objective To evaluate the effect of nicastrin （nct） gene on the biological functions of melanocytes in zebrafish. Methods By using a morpholino oligonucleotide （MO） technology, a nct-MO sequence targeting the zebrafish nct mRNA was designed, so was a MO control （ctrl-MO） sequence. Then, the enhanced green fluorescent protein （EGFP） mRNA with MO target sequence at its 5′ end was synthesized, and co-microinjected with the nct-MO or ctrl-MO sequence into the zebrafish embryos to verify the silencing efficiency of nct-MO and observe changes in developmental phenotypes in zebrafish. With wild-type zebrafish as a blank control group, real-time fluorescence-based quantitative PCR （RT-PCR） was conducted to determine the mRNA expression of melanin synthesis- and notch signaling pathway-related genes, including mitfa, tyr, tyrp1a, tyrp1b, dct, pmela, notch1a, notch1b and hey1 genes. One-way analysis of variance was used for the comparison of means among multiple groups, and least significant difference（LSD）-t test for multiple comparisons. Results Eight hours after zebrafish fertilization, green fluorescence was observed in the zebrafish embryos in the ctrl-MO + EGFP mRNA group, but not in the nct-MO + EGFP mRNA group or blank control group. Forty-eight hours after fertilization, the proportion of pigmented area among the whole area of the tail of zebrafish larvae was significantly lower in the nct-MO group （0.169 ± 0.083） than in the ctrl-MO group （0.258 ± 0.042, t = 3.202, P = 0.005）, and disorderly pigment distribution in the tails was observed in the nct-MO group. RT-PCR revealed significant differences in the mRNA expression of pmela, tyrp1a and hey1 genes among the nct-MO group, ctrl-MO group and blank control group （all P < 0.05）, but no significant difference was observed in the mRNA expression of mitfa, tyr, tyrp1b, dct, notch1a or notch1b genes among the 3 groups （all P＞0.05）; the relative expression levels of pmela and tyrp1a mRNAs were significantly lower in the nct-MO group （0.708 ± 0.028, 0.558 ± 0.136, respectively） than in the ctrl-MO group （1.023 ± 0.142, 1.016 ± 0.134, respectively, both P < 0.05）. Conclusion The nct gene may affect biological functions of melanocytes by regulating melanin synthesis in zebrafish. |
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| Keywords: | Pigmentation disorders Hidradenitis suppurativa Nicastrin γ-Secretase Zebrafish models |
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