中华皮肤科杂志 ›› 2025, Vol. 58 ›› Issue (9): 816-824.doi: 10.35541/cjd.20250072

• 论著·色素性皮肤病 • 上一篇    下一篇

[开放获取]   miR-25-5p靶向RAB11B参与调控皮肤色素沉着的机制研究

王雯竹1    杨荷丹1    刘蕴瑶2    孙小洁1    张晓丽1    李秀珍1    谭思齐1    徐浩翔2    杨寅1    林彤2   

  1. 1中国医学科学院  北京协和医学院皮肤病医院激光科，南京  210042；2中国医学科学院  北京协和医学院皮肤病医院  江苏省皮肤病与性病分子生物学重点实验室，南京  210042
  • 收稿日期:2025-02-14 修回日期:2025-07-17 发布日期:2025-09-01
  • 通讯作者: 林彤 E-mail:ddlin@hotmail.com
  • 基金资助:
    国家自然科学基金（82103705）；中国医学科学院医学与健康科技创新工程项目（CIFMS-2021-I2M-1-001）；江苏省科技计划（BE2023675）

Regulation of skin pigmentation by miR-25-5p via targeting RAB11B: a mechanistic study

Wang Wenzhu1, Yang Hedan1, Liu Yunyao2, Sun Xiaojie1, Zhang Xiaoli1, Li Xiuzhen1, Tan Siqi1, Xu Haoxiang2, Yang Yin1, Lin Tong2   

  1. 1Department of Cosmetic Laser Surgery, Hospital for Skin Diseases, Institute of Dermatology, Chinese Academy of Medical Sciences ＆ Peking Union Medical College, Nanjing 210042, China; 2Jiangsu Key Laboratory of Molecular Biology for Skin Diseases and STIs, Hospital for Skin Diseases, Institute of Dermatology, Chinese Academy of Medical Sciences ＆ Peking Union Medical College, Nanjing 210042, China
  • Received:2025-02-14 Revised:2025-07-17 Published:2025-09-01
  • Contact: Lin Tong E-mail:ddlin@hotmail.com
  • Supported by:
    National Natural Science Foundation of China （82103705）; CAMS Innovation Fund for Medical Sciences （CIFMS-2021-I2M-1-001）; Science and Technology Plan Projects in Jiangsu Province （BE2023675）

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摘要： 【摘要】 目的 探讨miRNA（miR）-25-5p对黑素合成的影响及可能机制。方法 通过TargetScan网站预测miR-25-5p的靶基因，采用双荧光素酶报告实验验证miR-25-5p与RAB11B基因的结合位点。选取6 ~ 8周C57BL/6J雌性小鼠，每日采用宽谱中波紫外线（UVB）照射耳背侧皮肤，建立小鼠炎症后色素沉着模型；选取4 ~ 6周雌性棕黄色豚鼠，每日采用UVB照射背部剃毛区皮肤，建立豚鼠炎症后色素沉着模型；其间予动物模型局部皮内注射miR-25-5p模拟物、miR对照模拟物，观察动物模型皮肤表型变化，造模结束后取模型皮肤组织进行Masson-Fontana黑素染色评估黑素含量；免疫组化及qPCR检测模型皮肤组织中RAB11B或酪氨酸酶（TYR）的表达。从健康男性包皮环切术后废弃的正常包皮组织中分离培养人原代黑素细胞，在人原代黑素细胞、人黑色素瘤MNT1细胞中转染miR对照模拟物及miR-25-5p模拟物，采用qPCR检测miR-25-5p及RAB11B mRNA的相对表达量（2-△△Ct）。在人黑色素瘤MNT1细胞中同时过表达miR-25-5p及RAB11B，检测RAB11B及TYR mRNA表达水平变化。组间比较采用t检验或单因素方差分析，组间两两比较采用Tukey检验。结果 通过TargetScan预测及双荧光素酶报告实验证实，RAB11B的3′非翻译区存在miR-25-5p的结合位点。小鼠及豚鼠模型中，外源给予miR-25-5p模拟物后模型局部皮肤色素沉着程度较对照组明显更低，组织病理切片Masson-Fontana染色显示，小鼠、豚鼠miR-25-5p模拟物组真表皮黑素颗粒（x ± s分别为0.050 ± 0.005、0.067 ± 0.015）均低于miR对照模拟物组（0.087 ± 0.008、0.110 ± 0.013，P < 0.05）。免疫组化示，小鼠模型皮肤miR-25-5p模拟物组RAB11B表达水平低于miR对照模拟物组（均P < 0.05）。qPCR显示，豚鼠色素沉着模型miR-25-5p模拟物组皮肤组织RAB11B、TYR mRNA表达水平低于miR对照模拟物组（均P < 0.05）。人原代黑素细胞和MNT1细胞miR-25-5p模拟物组RAB11B mRNA 的表达水平均低于miR对照模拟物组（均P < 0.05）。在人黑色素瘤MNT1细胞中过表达miR-25-5p可抑制TYR mRNA水平，而同时过表达miR-25-5p和RAB11B可回复单独过表达miR-25-5p引起的TYR mRNA表达水平下降。结论 过表达miR-25-5p可改善UVB诱导的炎症后色素沉着，抑制黑素合成，该过程可能与靶向抑制RAB11B的表达有关。

关键词: 皮肤色素沉着, 模型, 动物, 黑素合成, miR-25-5p, RAB11B, 酪氨酸酶

Abstract: 【Abstract】 Objective To investigate the role of microRNA-25-5p （miR-25-5p） in melanogenesis, and to explore its underlying mechanisms. Methods Target genes of miR-25-5p were predicted using the TargetScan database. The interaction between miR-25-5p and the 3' untranslated region （3' UTR） of the RAB11B gene （a member of RAS oncogene family） was validated through a dual-luciferase reporter assay. Post-inflammatory hyperpigmentation （PIH） models were established in female C57BL/6J mice （6 - 8 weeks old） and female brown guinea pigs （4 - 6 weeks old） through daily broadband ultraviolet B （UVB） irradiation on the dorsal skin of the mouse ear or shaved dorsal skin of guinea pigs, while untreated mice and untreated dorsal skin areas of guinea pigs served as control groups. During modeling, these experimental animals received intradermal injections of a miR-25-5p agomir or a miR control agomir. Changes in skin pigmentation were observed, and skin tissue samples were harvested for further analysis after modeling. Melanin content in skin tissues was evaluated using Masson-Fontana staining. Expression of RAB11B and tyrosinase （TYR） in skin tissues was determined using immunohistochemical staining and quantitative real-time PCR （qPCR）. Primary human melanocytes were isolated from discarded normal foreskin tissues of healthy males after circumcision. Both primary human melanocytes and human MNT1 melanoma cells were transfected with miR-25-5p mimics or miR control mimics. Relative expression levels of miR-25-5p and RAB11B mRNA were quantified by qPCR using the 2-ΔΔCt calculation method. In MNT1 cells, miR-25-5p and RAB11B were co-overexpressed to assess their effect on the mRNA expression of RAB11B and TYR. Statistical analysis was conducted using t test or one-way analysis of variance followed by Tukey's post hoc test for multiple comparisons. Results The bioinformatic prediction and dual-luciferase reporter assay confirmed a binding site for miR-25-5p in the 3′ UTR of the RAB11B gene. In both animal models, the treatment with the miR-25-5p agomir significantly reduced local skin pigmentation compared to the control groups; Masson-Fontana staining showed a marked decrease in the density of melanin granules in the epidermis and dermis in the miR-25-5p agomir groups compared with the miR control agomir groups （mice: 0.050 ± 0.005 vs. 0.087 ± 0.008; guinea pigs: 0.067 ± 0.015 vs. 0.110 ± 0.013; both P < 0.05）. Immunohistochemical staining revealed significantly lower expression of RAB11B in mouse skin tissues in the miR-25-5p agomir group than in those in the miR control agomir group （both P < 0.05）. qPCR revealed significantly lower mRNA expression of RAB11B and TYR in skin tissues of guinea pigs in the miR-25-5p agomir group than in those in the miR control agomir group （both P < 0.05）. Similarly, RAB11B mRNA expression significantly decreased in the miR-25-5p mimics group compared with the miR control mimics group in primary human melanocytes and MNT1 cells （both P < 0.05）. In human MNT1 melanoma cells, miR-25-5p overexpression could suppress TYR mRNA expression, whereas co-overexpression of miR-25-5p and RAB11B could reverse this suppression. Conclusion Overexpression of miR-25-5p could alleviate UVB-induced post-inflammatory hyperpigmentation and inhibit melanogenesis, likely by targeted suppression of RAB11B expression.

Key words: Skin pigmentation, Models, animal, Melanogenesis, miR-25-5p, RAB11B, Tyrosinase

引用本文

王雯竹 杨荷丹 刘蕴瑶 孙小洁 张晓丽 李秀珍 谭思齐 徐浩翔 杨寅 林彤. [开放获取]   miR-25-5p靶向RAB11B参与调控皮肤色素沉着的机制研究[J]. 中华皮肤科杂志, 2025,58(9):816-824. doi:10.35541/cjd.20250072

Wang Wenzhu, Yang Hedan, Liu Yunyao, Sun Xiaojie, Zhang Xiaoli, Li Xiuzhen, Tan Siqi, Xu Haoxiang, Yang Yin, Lin Tong. Regulation of skin pigmentation by miR-25-5p via targeting RAB11B: a mechanistic study [J]. Chinese Journal of Dermatology, 2025, 58(9): 816-824.doi:10.35541/cjd.20250072