Background Burn wound healing is a complex process and the role of Wnt ligands varies in this process. Whether and how Wnt4 functions in burn wound healing is not well understood. In this study, we aim to reveal the effects and potential mechanisms of Wnt4 in burn wound healing. Methods First, the expression of Wnt4 during burn wound healing was determined by immunofluorescence, Western blotting and qPCR. Then, Wnt4 was overexpressed in burn wounds. The healing rate and healing quality were analysed by gross photography and haematoxyline and eosin staining. Collagen secretion was observed by Masson staining. Vessel formation and fibroblast distribution were observed by immunostaining. Next, Wnt4 was knocked down in HaCaT cells. The migration of HaCaT cells was analysed by scratch healing and transwell assays. Next, the expression of β-catenin was detected by Western blotting and immunofluorescence. The binding of Frizzled2 and Wnt4 was detected by coimmunoprecipitation and immunofluorescence. Finally, the molecular changes induced by Wnt4 were analysed by RNA sequencing, immunofluorescence, Western blotting and qPCR in HaCaT cells and burn wound healing tissues. Results The expression of Wnt4 was enhanced in burn wound skin. Overexpression of Wnt4 in burn wound skin increased the thickness of epidermis. Collagen secretion, vessel formation and fibroblast distribution were not significantly impacted by Wnt4 overexpression. When Wnt4 was knocked down in HaCaT cells, the ratio of proliferating cells decreased, the ratio of apoptotic cells increased and the ratio of the healing area in the scratch healing assay to the number of migrated cells in the transwell assay decreased. The nuclear translocation of β-catenin decreased in shRNA of Wnt4 mediated by lentivirus-treated HaCaT cells and increased in Wnt4-overexpressing epidermal cells. RNA-sequencing analysis revealed that cell junction-related signalling pathways were significantly impacted by Wnt4 knockdown. The expression of the cell junction proteins was decreased by the overexpression of Wnt4. Conclusions Wnt4 promoted the migration of epidermal cells. Overexpression of Wnt4 increased the thickness of the burn wound. A potential mechanism for this effect is that Wnt4 binds with Frizzled2 and increases the nuclear translocation of β-catenin, thus activating the canonical Wnt signalling pathway and decreasing the cell junction between epidermal cells.

中文翻译：

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Wnt4 通过激活经典 Wnt 信号传导并减少表皮细胞之间的细胞连接来增加烧伤创面的表皮厚度。

背景烧伤创面愈合是一个复杂的过程，Wnt配体在此过程中的作用各不相同。Wnt4 是否以及如何在烧伤伤口愈合中发挥作用尚不清楚。在这项研究中，我们旨在揭示Wnt4在烧伤创面愈合中的作用和潜在机制。方法首先采用免疫荧光、Western blotting和qPCR检测烧伤创面愈合过程中Wnt4的表达。然后，Wnt4 在烧伤创面中过度表达。通过大体摄影和苏木精伊红染色分析愈合率和愈合质量。通过Masson染色观察胶原蛋白分泌。通过免疫染色观察血管形成和成纤维细胞分布。接下来，Wnt4 在 HaCaT 细胞中被敲低。通过划痕愈合和跨孔实验分析 HaCaT 细胞的迁移。下一个，Western blotting和免疫荧光法检测β-catenin的表达。通过免疫共沉淀和免疫荧光检测Frizzled2和Wnt4的结合。最后，通过 RNA 测序、免疫荧光、蛋白质印迹和 qPCR 分析了 HaCaT 细胞和烧伤创面愈合组织中 Wnt4 诱导的分子变化。结果烧伤创面皮肤Wnt4表达增强。Wnt4在烧伤创面皮肤中的过度表达增加了表皮的厚度。Wnt4 过表达不会显着影响胶原蛋白分泌、血管形成和成纤维细胞分布。当 HaCaT 细胞中 Wnt4 被敲低时，增殖细胞的比例下降，凋亡细胞比例增加，划痕愈合实验中愈合面积与Transwell实验中迁移细胞数的比例降低。慢病毒处理的 HaCaT 细胞介导的 Wnt4 shRNA 中 β-catenin 的核转位减少，而在 Wnt4 过表达的表皮细胞中增加。RNA 测序分析表明，Wnt4 敲低对细胞连接相关信号通路有显着影响。Wnt4 的过度表达降低了细胞连接蛋白的表达。结论 Wnt4促进表皮细胞迁移。Wnt4 的过度表达增加了烧伤创面的厚度。这种效应的一个潜在机制是 Wnt4 与 Frizzled2 结合并增加 β-连环蛋白的核转位，