毛囊干细胞microRNA调控毛乳头细胞诱导毛囊再生能力的实验研究

MicroRNA, a small non-coding non-messenger RNA , is one of the major biological regulators which inhibits gene activity. Previous studies showed that microRNA could regulate bio-functions of stem cells, and played important roles in modulating development of tissues and organs. Our object is to identify the microRNA of hair follicular epithelial stem cells which regulate follicle dermal papilla cells-induced hair follicle regeneration. Using laser capture micro-cut technology, stem cells in different growth cycle of mice will be separated, whose microRNA expression subtypes will then be determined by TaqMan? MicroRNA Assays, in order to establish a microRNA vocabulary of hair follicular epithelial stem cells in different growth cycle of hair follicle, and the target genes are identified and tested. After treatment with miRNA antagomir or miRNA mimics, the morphology and molecular expression of the hair papilla are observed and the growth of the hair follicle is measured in vivo. The hair follicle epithelial stem cells treated with miRNA antagomir or miRNA mimics are co-cultured with hair papilla, and then the hair papilla is transplanted into skin. The ability to dermal papilla cells-inducted follicle regeneration is evaluated. We will identify the microRNA of hair follicular epithelial stem cells which regulate follicle dermal papilla cells-induced hair follicle regeneration by this study. Our study will add to clarify the mechanisms related to functional regulation of hair follicular stem cells, which is of important theoretical significance and clinical therapy prospect. So far, it has not been reported in the literature related to this study about microRNA expression in hair follicular epithelial stem cells in the different hair follicle growth cycle and regulation of function of the hair papilla that induce hair follicle regeneration.

MicroRNA是一种小的非编码非信使RNA，是动物体内主要的调节分子之一。许多研究资料表明干细胞microRNA在调控组织和器官发育方面扮演十分重要的角色。为了筛选调控毛乳头诱导毛囊再生的毛囊上皮干细胞microRNA，我们将采用激光捕捉微切技术分离毛囊上皮干细胞，用TaqMan? MicroRNA Assays建立静止期和生长期毛囊上皮干细胞的microRNA表达谱，并进行靶基因鉴定和验证；体内采用miRNA antagomir及miRNA mimics分别沉默和上调相关的microRNA，观察对毛乳头形态结构、分子表达及毛囊生物学的影响；采用经miRNA antagomir和miRNA mimics处理后毛囊干细胞分别与毛乳头共培养，取共培养的毛乳头体内进行诱导毛囊再生实验，这对阐明毛囊干细胞microRNA在诱导毛囊再生的功能调控机制具有重要的理论意义和临床应用前景。

MicroRNA是一种小的非编码非信使RNA，是动物体内主要的调节分子之一。许多研究资料表明干细胞microRNA在调控组织和器官发育方面扮演十分重要的角色。为了筛选调控毛乳头诱导毛囊再生的毛囊上皮干细胞microRNA，本课题分离培养小鼠触须和背部毛囊不同生长周期的隆突多能干细胞，优化毛囊干细胞的分离培养方法，用于收集microRNA进行检测，采用激光捕获显微切割收集小鼠不同毛囊生长周期隆突干细胞，进行microRNA差异表达分析和靶基因预测。应用microarray及原位杂交等技术发现miR92a在C57/BL小鼠早期胚胎发育中的毛囊上皮选择性高表达，通过脂质体转染法将microRNA92a mimic转染到体外培养的外根鞘细胞，用qRT-PCR可以检测到转染后的外根鞘细胞中microRNA92a的表达量升高。MTS法和划痕实验探索microRNA92a对毛囊外根鞘细胞的增殖和迁移能力实验正在进行中。microRNA92a敲除小鼠和K15转基因工具鼠已经购买并进入SPF环境饲养、育种，后续将用于在个体水平研究microRNA92a对毛囊生长周期的影响。外泌体是细胞分泌的含有microRNA小囊泡，本课题研究了DPC-Exos对毛囊生长周期和毛细胞生长的影响，发现DPC-Exos通过β-catenin和Shh信号通路促进ORSCs的增殖和迁移，诱导生长期，延迟退行期，促进小鼠毛干生长和隆突增大。本课题目前的研究成果已发表SCI论文6篇，对日后进一步探索microRNA在毛囊中的作用奠定了基础。