十四酸甾醇酯调控Wnt/β-catenin信号通路促皮肤创面修复的研究

Cholesterol Myristate Promotes Hair Follicle Stem Cells to Repair Skin Wound by Regulating Wnt/β-catenin Signaling Pathway

目的探讨十四酸甾醇酯(cholesterol myristate)对大鼠皮肤创面修复过程中Wnt/β-catenin通路的调控。方法十四酸甾醇酯作用于大鼠触须部皮肤创面,观察皮肤创面的修复情况,免疫荧光检测皮肤组织中Wnt/β-catenin通路蛋白的表达;体外分离培养SD乳鼠毛囊干细胞(Hair Follicle Stem Cells,HFSCs),免疫荧光检测细胞表面标志物CD34、CK15的表达,CCK-8法检测十四酸甾醇酯对HFSCs增殖的影响,RT-q PCR、Western-blot检测其对HFSCs中Wnt/β-catenin通路相关蛋白的表达。结果术后7 d,十四酸甾醇酯组创面基本愈合,表皮细胞排列紧密,表皮与真皮层连接牢固,可见新生毛细血管;对照组表皮与真皮连接不牢固,表皮排列不规则,仍有炎症浸润。术后4 d,免疫荧光检测十四酸甾醇酯组修复皮肤中β-catenin、Lymphoid enhancing factor 1(LEF1)、cancer-myc(c-myc)、cyclin D1的表达均强于对照组。所培养的细胞表达HFSCs标志物CD34、CK15,十四酸甾醇酯对HFSCs有促增殖作用(P<0.01),且β-catenin、LEF1、c-myc、cyclin D1表达上调。结论十四酸甾醇酯通过激活HFSCs中的Wnt/β-catenin通路促皮肤修复。

Objective To investigate the regulation of Wnt/β-catenin signaling pathway in skin wound repair bycholesterol myristate,one of the active components in plastrum testudinis. Methods SD rat models of whisker skinwere randomly divided into control group and cholesterol myristate group. The effects on the repair of skin woundswere examinated by HE;the expression levels of proteins in Wnt/β-catenin pathway in the tissue were detected byimmunofluorescence detection. The cultivated hair follicle stem cells（HFSCs）from SD neonatal rat were identified byCD34 and CK15, the cell surface markers of HFSCs using immunofluorescence detection. Cell Counting Kit-8（CCK-8）was used to examine the proliferation of HFSCs. RT-q PCR and Western Blot were used to detected m RNAand protein expression levels of factors in Wnt/β-catenin signaling pathway of HFSCs. Results On postoperative 7 d,wounds of cholesterol myristate group were almost healed. The epidermal cells were closely arranged,connectionof epidermis and dermis was firm with visible newborn blood capillary;while in the control group,the epidermallayers arrangement was irregular, connection of epidermal and dermis was still unstable, and there was stillinflammatory infiltration. On postoperative 4 d,β-catenin,LEF1,c-myc,cyclin D1 in the repaired skin detected by immunofluorescence assay in cholesterol myristate group was stronger than those in control group. CD34 and CK15 of both cultured HFSCs were positive. Cholesterol myristate had a proliferation promoting effect on HFSCs（P〈0.01）and upregulation of β-catenin,LEF1,c-myc,cyclin D1. Conclusion Cholesterol myristate promoted the HFSCsproliferation and skin repair by activation of Wnt/β-catenin pathway.