目的 制备黄芪甲苷水凝胶，探讨该敷料在大鼠全层皮肤创面修复中的作用。 方法 制备含0、1.5、2.0、2.5 mg/mL黄芪甲苷的甲基丙烯酸酐化明胶水凝胶，分别记为GelMA、GelMA/AS-IV-1.5、GelMA/AS-IV-2.0、GelMA/AS-IV-2.5。在16只大鼠背部各制作6个全层皮肤缺损，分别注射黄芪甲苷溶液（2.5 mg/mL）、GelMA水凝胶、GelMA/AS-IV-1.5水凝胶、GelMA/AS-IV-2.0水凝胶、GelMA/AS-IV-2.5水凝胶，体积均为100 µL，其中1个缺损部位不进行注射治疗（空白对照组）。术后第3、7、14 d观察创面愈合率。苏木精-伊红染色观察创面再上皮化，Masson染色观察创面胶原蛋白沉积，免疫组化染色观察创面血管新生，免疫荧光染色观察创面瘢痕增生、皮肤细胞增殖以及Wnt/β-catenin信号转导通路蛋白表达，RT-qPCR检测增殖细胞核抗原、Wnt/β-catenin信号转导通路相关基因表达，Western blot检测Wnt/β-catenin信号转导通路相关蛋白表达。 结果 术后第14 d，GelMA/AS-IV-1.5水凝胶组、GelMA/AS-IV-2.0水凝胶组创面愈合率高于空白对照组（P<0.05）。GelMA/AS-IV-2.0水凝胶组创面肉芽组织厚度与皮肤再上皮化效果好于其他5组，创面CD31表达量最高，α-平滑肌肌动蛋白表达最低，细胞增殖细胞核抗原的蛋白与mRNA表达量最高，β-catenin、LEF1和c-Myc 的蛋白与mRNA表达最高。 结论 GelMA/AS-IV-2.0水凝胶可通过促进皮肤细胞增殖、胶原蛋白沉积与血管生成以及抑制瘢痕增生等加速创面愈合，并且该过程可能与激活Wnt/β-catenin信号通路有关。
Abstract
Objective To prepare astragaloside IV hydrogel and explore the role of this dressing in the repair of full-thickness skin wounds in rats. Method Methacrylate anhydride modified gelatin hydrogels containing 0, 1.5 mg/mL, 2.0 mg/mL, and 2.5 mg/mL astragaloside IV were prepared, which were respectively denoted AS GelMA, GelMA/AS-IV-1.5, GelMA/AS-IV-2.0, and GelMA/AS-IV-2.5. Six full-thickness skin defects were made on the backs of 16 rats respectively. Astragaloside IV solution (2.5 mg/mL), GelMA hydrogel, GelMA/AS-IV-1.5 hydrogel, GelMA/AS-IV-2.0 hydrogel and GelMA/AS-IV-2.5 hydrogel were injected respectively, with a volume of 100 µL for each. One of the defect sites was not treated with injection (blank control group). The wound healing rate was observed on 3 d, 7 d, and 14 d after the operation. Hematoxylin-eosin staining was used to observe the re-epithelialization of the wound, Masson staining was used to observe the deposition of collagen in the wound, immunohistochemical staining was used to observe the angiogenesis of the wound, and immunofluorescence staining was used to observe scar hyperplasia, skin cell proliferation and the expression of Wnt/β-catenin signal transduction pathway proteins in the wound. The expressions of proliferating cell nuclear antigen and genes related to the Wnt/β-catenin signal transduction pathway were detected by RT-qPCR, and the expressions of proteins related to the Wnt/β-catenin signal transduction pathway were detected by Western blot. Results On 14 d after the operation, the wound healing rates of GelMA/AS-IV-1.5 hydrogel group and GelMA/AS-IV-2.0 hydrogel group were higher than those of blank control group (P<0.05). The thickness of granulation tissue in the wound and the effect of skin re-epithelialization in GelMA/AS-IV-2.0 hydrogel group were better than those in the other five groups. The expression level of CD31 in the wound was the highest, the expression of α -smooth muscle actinin was the lowest, and the protein and mRNA expression levels of cell proliferation nuclear antigen were the highest. The protein and mRNA expressions of β-catenin, LEF1 and c-Myc were the highest. Conclusion GelMA/AS-IV-2.0 hydrogel can accelerate wound healing by promoting skin cell proliferation, collagen deposition and angiogenesis, and inhibiting scar hyperplasia, and this process may be related to the activation of Wnt/β-catenin signaling pathway.
关键词
黄芪甲苷 /
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甲基丙烯酸酐化明胶 /
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水凝胶 /
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创面
Key words
Astragaloside /
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Methacrylate gelatine /
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Hydrogel /
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Wound
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基金
河南省医学科技攻关计划联合共建项目（LHGJ20240853）
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