目的 探讨ⅩⅦ型胶原蛋白(COL17)对雄激素性脱发(AGA)模型小鼠毛发生长的作用及机制。 方法 将48只C57BL/6J小鼠建立AGA模型(脱去小鼠背部毛发并外涂二氢睾酮溶液)，采用随机数表法分为6组，每组8只。阴性对照组，脱毛区注射生理盐水(单点注射0.05 ml，共5个点)；阳性对照组，脱毛区外用5%米诺地尔酊，1 ml/d；COL17低、中、高浓度组，在脱毛区分别注射0.5、1.0、2.0 mg/ml的COL17(单点注射0.05 ml，共5个点)；Ⅲ型胶原蛋白(COL3)、COL17(COL3＋COL17)联合高浓度组，在脱毛区注射2.0 mg/ml的COL3和COL17(单点注射0.05 ml，共5个点)。共给药21 d，期间观察记录各组小鼠毛发生长情况；21 d后，取小鼠脱毛区皮肤及皮下组织制作病理切片行HE染色，观察毛囊数量及形态变化；取小鼠脱毛区新鲜皮肤组织行总RNA测序分析，对差异共表达基因进行基因本体论(GO)功能注释、京都基因与基因组百科全书(KEGG)通路分析和基因集富集分析(GSEA)。 结果 给药21 d后，与阴性对照组相比，阳性对照组、COL17高浓度组、COL3＋COL17联合高浓度组小鼠背部脱毛面积明显缩小，HE染色显示其毛囊数量也明显增多。Pearson相关性分析、主成分分析及各组间聚类热图显示，COL17高浓度组与阳性对照组基因相关性较高(R2＝0.95，P＝0.024)，基因表达较为接近，2组有3 882个差异基因(1 705个上调，2 177个下调)，而COL3＋COL17联合高浓度组与阳性对照组基因相关性最高(R2＝0.96，P＝0.001)，基因表达最接近，2组有1 289个差异基因(385个上调，904个下调)；KEGG分析显示，与阴性对照组相比，阳性对照组、COL17高浓度组及COL3＋COL17联合高浓度组小鼠均上调了与毛发生长相关的Wnt信号通路、细胞黏附分子及hedgehog信号通路；GO富集分析提示COL17高浓度组及COL3＋COL17联合高浓度组中皮肤发育、毛发周期循环相关基因上调；GSEA富集分析发现，COL17高浓度组成纤维细胞增殖、白介素1分泌相关基因表达上调，而COL3＋COL17联合高浓度组成纤维细胞迁移、凋亡细胞清除及加速活性氧的代谢相关基因表达上调。 结论 局部注射2.0 mg/ml的COL17对AGA模型小鼠毛发生长具有一定促进作用，且联合注射2.0 mg/ml的COL3后效果更显著，激活Wnt信号通路可能是COL17促毛发生长的主要机制之一。 Objective To investigate the effect and mechanism of type ⅩⅦ collagen (COL17) on hair growth in mice with androgenetic alopecia (AGA). Methods Forty-eight C57BL/6J mice were used to establish AGA model (the back hair of the mice was removed and dihydrotestosterone solution was applied) and divided into 6 groups of 8 mice each by random number table. Negative control group, injection of saline in the depilated area (single point injection of 0.05 ml, 5 points in total) positive control group, topical application of 5% minoxidil tincture in the depilated area, 1 ml/d COL17 low, medium and high concentration groups, injection of 0.5, 1.0 and 2.0 mg/ml COL17 in the depilated area respectively (single point injection of 0.05 ml, 5 points in total) type Ⅲ and ⅩⅦ collagen (COL3+ COL17) combined high concentration group, injection of 2.0 mg/ml COL3 and COL17 in the depilated area (single point injection of 0.05 ml, 5 points in total). The total treatment time was 21 days, during which the hair growth of mice in each group was observed and recorded. After 21 days, the skin and subcutaneous tissue in the depilated area of the mice were taken to make pathological sections for HE staining, and the number and morphological changes of hair follicles were observed fresh skin tissue in the depilated area of the mice was taken for total RNA sequencing analysis, and the differentially co-expressed genes were annotated by gene ontology (GO) functional annotation, Kyoto encyclopedia of genes and genomes (KEGG) pathway analysis and gene set enrichment analysis (GSEA). Results After 21 days of treatment, compared with the negative control group, the depilation area on the back of the mice in the positive control group, COL17 high concentration group, and COL3+ COL17 combined high concentration group was significantly reduced, and HE staining showed that the number of hair follicles was also significantly increased. Pearson correlation analysis, principal component analysis and cluster heat map between groups showed that COL17 high concentration group had high gene correlation with the positive control group (R2=0.95, P=0.024), and the gene expression was relatively close, with 3 882 differentially expressed genes (1 705 up-regulated and 2 177 down-regulated) in the two groups, while COL3+ COL17 combined high concentration group had the highest gene correlation with the positive control group (R2=0.96, P=0.001), and the gene expression was the closest, with 1 289 differentially expressed genes (385 up-regulated and 904 down-regulated). KEGG analysis showed that compared with the negative control group, the positive control group, COL17 high concentration group and COL3+ COL17 combined high concentration group of mice all upregulated Wnt signaling pathway, cell adhesion molecules and hedgehog signaling pathway related to hair growth. GO enrichment analysis suggested that COL17 high concentration group and COL3+ COL17 combined high concentration group had upregulated genes related to skin development and hair cycle. GSEA enrichment analysis found that COL17 high concentration group had upregulated genes related to fibroblast proliferation and interleukin-1 secretion, while COL3+ COL17 combined high concentration group had upregulated genes related to fibroblast migration, clearance of apoptotic cells and accelerated metabolism of reactive oxygen species. Conclusion Local injection of 2.0 mg/ml COL17 has a certain promoting effect on hair growth in AGA model mice, and the effect is more significant after combined injection of 2.0 mg/ml COL3. Activation of Wnt signaling pathway is one of the main mechanisms of COL17 promoting hair growth.
Effect and mechanism of type ⅩⅦ collagen on hair growth in mice with androgenetic alopecia
Objective To investigate the effect and mechanism of type ⅩⅦ collagen (COL17) on hair growth in mice with androgenetic alopecia (AGA). Methods Forty-eight C57BL/6J mice were used to establish AGA model (the back hair of the mice was removed and dihydrotestosterone solution was applied) and divided into 6 groups of 8 mice each by random number table. Negative control group, injection of saline in the depilated area (single point injection of 0.05 ml, 5 points in total) positive control group, topical application of 5% minoxidil tincture in the depilated area, 1 ml/d COL17 low, medium and high concentration groups, injection of 0.5, 1.0 and 2.0 mg/ml COL17 in the depilated area respectively (single point injection of 0.05 ml, 5 points in total) type Ⅲ and ⅩⅦ collagen (COL3+ COL17) combined high concentration group, injection of 2.0 mg/ml COL3 and COL17 in the depilated area (single point injection of 0.05 ml, 5 points in total). The total treatment time was 21 days, during which the hair growth of mice in each group was observed and recorded. After 21 days, the skin and subcutaneous tissue in the depilated area of the mice were taken to make pathological sections for HE staining, and the number and morphological changes of hair follicles were observed fresh skin tissue in the depilated area of the mice was taken for total RNA sequencing analysis, and the differentially co-expressed genes were annotated by gene ontology (GO) functional annotation, Kyoto encyclopedia of genes and genomes (KEGG) pathway analysis and gene set enrichment analysis (GSEA). Results After 21 days of treatment, compared with the negative control group, the depilation area on the back of the mice in the positive control group, COL17 high concentration group, and COL3+ COL17 combined high concentration group was significantly reduced, and HE staining showed that the number of hair follicles was also significantly increased. Pearson correlation analysis, principal component analysis and cluster heat map between groups showed that COL17 high concentration group had high gene correlation with the positive control group (R2=0.95, P=0.024), and the gene expression was relatively close, with 3 882 differentially expressed genes (1 705 up-regulated and 2 177 down-regulated) in the two groups, while COL3+ COL17 combined high concentration group had the highest gene correlation with the positive control group (R2=0.96, P=0.001), and the gene expression was the closest, with 1 289 differentially expressed genes (385 up-regulated and 904 down-regulated). KEGG analysis showed that compared with the negative control group, the positive control group, COL17 high concentration group and COL3+ COL17 combined high concentration group of mice all upregulated Wnt signaling pathway, cell adhesion molecules and hedgehog signaling pathway related to hair growth. GO enrichment analysis suggested that COL17 high concentration group and COL3+ COL17 combined high concentration group had upregulated genes related to skin development and hair cycle. GSEA enrichment analysis found that COL17 high concentration group had upregulated genes related to fibroblast proliferation and interleukin-1 secretion, while COL3+ COL17 combined high concentration group had upregulated genes related to fibroblast migration, clearance of apoptotic cells and accelerated metabolism of reactive oxygen species. Conclusion Local injection of 2.0 mg/ml COL17 has a certain promoting effect on hair growth in AGA model mice, and the effect is more significant after combined injection of 2.0 mg/ml COL3. Activation of Wnt signaling pathway is one of the main mechanisms of COL17 promoting hair growth.
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