低频电磁场调控毛囊微环境中KGF-BMP-Wnt信号网络促进毛囊再生的作用及机制研究

In previous studies, our data had showed that low frequency electromagnetic field promoted hair follicles regeneration and the results was associated with the up-regulation of keratinocyte growth factor (KGF), but the effects of KGF on the microenvironment of hair follicle regeneration and the epithelial–mesenchymal interactions after electromagnetic field exposure is not clear. This present study was designed to determine how low frequency electromagnetic fields exposure induced the activation of KGF-BMP-Wnt signaling mediated by KGF to improve hair follicle regeneration. For this purpose, we chose dermal papilla cells and hair follicle stem cells to make the model of epithelial-mesenchymal interactions. When the model was treat with electromagnetic fields for different times, the proteins were harvested to assess for KGF expression in dermal papilla cells and KGF receptor expression in hair follicle stem cells. Then RNA was isolated from hair follicle and RT-PCR was performed to identify the different of the gene expression of BMP/Wnt signaling compere to the group without electromagnetic fields exposure. Meanwhile, we also confirmed whether electromagnetic fields affected the inter-reaction between KGF and the key factors of the BMP signaling pathways and Wnt signaling pathways such as BMP2/4, Smads, β-catenin, Tcf. Taking together of those data, we demonstrated that the keys nodes of BMP-Wnt signaling mediated by KGF after low frequency electromagnetic fields treatment. Finally, the mice were used to make the model of AGA and CIA, and then those mice divided at random into the control group and low frequency EMF group. After EMF exposure, the skin specimens of the mice were harvested to assess for hairs regrowth, and the relationship between the growth factors including TGF-β,VEGF, EGF and KGF was evaluated. The expression and location of the gene expression of BMP-Wnt signaling network was also tested. Following those results, we inferred that EMF exposure enhance the effect of KGF on hair follicle regeneration though the activation of KGF-BMP-Wnt signaling and its mechanism. The above research project aims to describe the mechanism of low-frequency electromagnetic fields on hair follicle regeneration involving in KGF, provide a theoretical basis for regenerative medicine, organization project and so on.

课题组前期已证明低频电磁场（EMF）促毛囊再生与角质细胞生长因子（KGF）密切相关，但EMF作用下KGF如何影响毛囊间质-上皮微环境未见研究报道。本课题拟选毛乳头细胞-毛囊干细胞为对象，分析EMF干预下KGF及其受体（KGFR）在毛乳头细胞和毛囊干细胞的表达特征，同时检测毛囊干细胞BMP/Wnt信号网络基因表达谱变化，然后探讨KGF与毛囊微环境BMP2/4、Smads、β-catenin、Tcf等关键信号分子的相互作用，确定EMF通过KGF对毛囊微环境BMP/Wnt信号网络调控的关键节点。在雄激素性秃发和化学诱导性脱发模型中，研究EMF作用下KGF与各模型毛囊微环境中TGF-β、VEGF、EGF等相互作用，检测EMF对各模型间质-上皮相互促进效应和调控KGF-BMP-Wnt信号网络对毛囊再生的影响。本研究从调控微环境信号网络角度阐述低频电磁场促毛囊再生的作用机制，为进一步深入研究奠定基础。

在临床应用和基础研究中，低频电磁场促进创面愈合已获得越来越多证据支持；但是，如何重建功能性皮肤仍需深入研究。国内外研究认为，选择合适的种子细胞及其生长微环境的重建是功能性皮肤再生的关键。课题组在前期实验基础上，通过毛发生长模型，观察到50 Hz电磁场促进毛发生长，与电磁场增强毛囊K15+干细胞增殖并增加KGF分泌密切相关；同时，本课题选用毛乳头（DP）细胞-毛囊（表皮）干细胞为研究对象，分析低频电磁场促进间质-上皮相互作用诱导干细胞分化形成毛囊及其对微环境影响机制，结果显示，DP细胞和干细胞的混合物可以作为合适的种子细胞用于功能皮肤重建，而低频电磁场可通过增强DP细胞-表皮（毛囊）干细胞相互作用促进诱导干细胞分化形成毛囊。而在创面愈合模型中，课题组发现Rg1可以改善高糖微环境下人脐静脉内皮细胞增殖，并调控miR-489e3p/Sirt1轴促进创面愈合。以上研究结果进一步明确了重建功能性皮肤的种子细胞及其微环境是构建功能性皮肤的必要条件，并证实低频电磁场增强种子细胞分化形成毛囊的作用，促进低频电磁场在创面治疗和皮肤功能重建领域中的应用；同时，为低频电磁场在病理条件下促功能性皮肤重建的研究奠定了研究基础。