| 胡蔚,查嘉凤,姚贝贝.海艾汤对大鼠真皮成纤维细胞增殖及Wnt/β-catenin信号通路的影响[J].浙江中西医结合杂志,2025,35(10): |
| 海艾汤对大鼠真皮成纤维细胞增殖及Wnt/β-catenin信号通路的影响 |
| Impact of Hai Ai Decoction on Rat Dermal Fibroblast Proliferation and Androgen Receptor and Wnt Expression |
| 投稿时间：2025-03-17 修订日期：2025-08-14 |
| DOI： |
| 中文关键词: 雄性激素源性脱发 海艾汤 5-α还原酶 Wnt/β-catenin信号通路 雄激素受体 |
| 英文关键词:androgenic alopecia Hai Ai Decoction 5-α reductase, Wnt/β-catenin pathway androgen receptor |
| 基金项目:2023年全国中药特色技术传承人才培训项目 |
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| 中文摘要: |
| 目的 探究海艾汤对大鼠真皮成纤维细胞的增殖作用及机制，为其治疗雄性激素源性脱发（androgenic alopecia, AGA）提供理论依据。方法 大鼠灌胃海艾汤制备含药血清，另分离大鼠真皮成纤维细胞进行传代培养，分为空白组、对照组、低、中、高海艾汤血清组、阳性组，用胎牛血清（fetal bovine serum, FBS）、正常血清、2.5%、7.5%、10% 海艾汤含药血清、20 ng/mL碱性成纤维细胞生长因子（basic fibroblast growth factor, bFGF）含药血清干预48 h。CCK-8法测定细胞活性；酶联免疫吸附试验（ELISA）测定细胞上清液血管内皮生长因子（vascular endothelial growth factor, VEGF）含量；荧光定量PCR测定细胞VEGF、成纤维细胞生长因子（fibroblast growth factor, FGF）、5-α还原酶（SRD5A）、雌激素受体（estrogen receptor, ER）mRNA表达；蛋白质印迹（WB）测定肝细胞生长因子（hepatocyte growth factor, HGF）、雄激素受体（androgen receptor, AR）、无翅型MMTV整合位点家族成员3A(Wnt3a)、无翅型MMTV整合位点家族成员5A(Wnt5a)、Axis抑制蛋白2（Axis inhibitor protein2, Axin2）、β-连环蛋白（β-catenin）的蛋白表达。结果 海艾汤血清以浓度依赖性方式显著促进大鼠真皮成纤维细胞生长（P<0.05或P<0.01）。与对照组比较，高、中、低海艾汤血清组和阳性组细胞中VEGF含量显著升高[(138.20±14.68) pg/mL、(151.52±14.98) pg/mL、(176.61±12.51) pg/mL、(172.33±15.70) pg/mL比(89.51±9.43) pg/mL, P<0.01]。qPCR结果显示，与对照组比较，高海艾汤血清组细胞VEGF[(3.62±0.31)比(1.11±0.10), P<0.01]、FGF[(2.48±0.26)比(1.22±0.13), P<0.01]、ER[(2.98±0.32)比(1.21±0.09), P<0.01] mRNA表达显著升高，SRD5A[(0.43±0.08)比(0.96±0.06), P<0.01] mRNA表达显著降低。WB结果显示，与对照组比较，高海艾汤血清组细胞HGF[(2.54±0.27)比(1.11±0.10), P<0.01]、Wnt3a[(0.61±0.07)比(0.99±0.09), P<0.01]、Wnt5a[(1.75±0.17)比(1.11±0.10), P<0.01]、β-catenin[(2.80±0.28)比(1.15±0.11), P<0.01]蛋白表达显著升高，AR[(0.61±0.07)比(0.99±0.09), P<0.01]、Axin2[(0.57±0.06)比(1.08±0.10), P<0.01]蛋白表达显著降低。结论 海艾汤能抑制大鼠真皮成纤维细胞内AR和5-α还原酶表达，促进毛囊生长相关的Wnt/β-catenin信号通路表达，因此可能对AGA具有改善及治疗作用。 |
| 英文摘要: |
| Objective To explore the proliferation effect and mechanism of Hai Ai Decoction on rat dermal fibroblasts, and provide a theoretical basis for the androgenic alopecia (AGA) treatment. Methods Rats were given Hai Ai Decoction to prepare medicated serum, and rat dermal fibroblasts were isolated to subculture. Then divided into the following groups: blank group, control group, low-, medium-, and high-dose Hai Ai Decoction serum groups, and positive control group. The cells were treated with fetal bovine serum (FBS), normal serum, 2.5%, 7.5%, and 10% Hai Ai Decoction medicated serum, or 20 ng/mL basic fibroblast growth factor (bFGF) medicated serum for 48 hours.CCK-8 method was used to detect cell viability, ELISA was used to detect vascular endothelial growth factor(VEGF) content, qPCR was used to detect VEGF, fibroblast growth factor (FGF), 5α-reductase (SRD5A), and estrogen receptoe (ER) mRNA expression, WB was used to detect hepatocyte growth factor (HGF), androgen receptor (AR), Wnt3a, Wnt5a and Axis inhibitor protein2 (Axin2) and β-catenin protein expression. Results Hai Ai Decoction serum significantly promoted the growth of rat dermal fibroblasts in a concentration-dependent manner (P<0.05 or P<0.01). Compared with the control group, the VEGF content in cells was significantly increased in the high-, medium-, and low-dose Hai Ai Decoction serum groups and the positive control group [(138.20±14.68) pg/mL, (151.52±14.98) pg/mL, (176.61±12.51) pg/mL, and (172.33±15.70) pg/mL vs. (89.51±9.43) pg/mL, P<0.01]. The qPCR results showed that, compared with the control group, cells treated with high Hai Ai Decoction serum showed significantly increased mRNA expression of VEGF [(3.62±0.31) vs. (1.11±0.10), P<0.01], FGF [(2.48±0.26) vs. (1.22±0.13), P<0.01], and ER [(2.98±0.32) vs. (1.21±0.09), P<0.01], while SRD5A mRNA expression was significantly decreased [(0.43±0.08) vs. (0.96±0.06), P<0.01]. WB results revealed that the protein expression levels of HGF [(2.54±0.27) vs. (1.11±0.10), P<0.01], Wnt3a [(0.61±0.07) vs. (0.99±0.09), P<0.01], Wnt5a [(1.75±0.17) vs. (1.11±0.10), P<0.01], and β-catenin [(2.80±0.28) vs. (1.15±0.11), P<0.01] were significantly increased, whereas the expression of AR [(0.61±0.07) vs. (0.99±0.09), P<0.01] and Axin2 [(0.57±0.06) vs. (1.08±0.10), P<0.01] was significantly decreased. Conclusion Hai Ai Decoction can inhibit expression of AR and SRD5A, promote expression of Wnt signaling pathway related to hair follicle growth, therefore it may have therapeutic effects on AGA. |
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