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Abstract：
Hypertrophic scar is a fibrotic disease following wound healing and is characterized by excessive extracellular matrix deposition. Autologous microfat grafting proves an effective strategy for the treatment thereof as it could improve the texture of scars and relieve relevant symptoms. This study aims to explore the potential mechanisms underlying the anti-fibrotic effect of microfat on hypertrophic scars.
In this study, we injected microfat into transplanted hypertrophic scars in mouse models and investigated the subsequent histological changes and differential expression of mRNAs therein. As for in vitro studies, we co-cultured microfat and hypertrophic scar fibroblasts (HSFs) and analyzed molecular profile changes in HSFs co-cultured with microfat by RNA sequencing. Moreover, to identify the key transcription factors (TFs) which might be responsible for the anti-fibrotic function of microfat, we screened the differentially expressed TFs and transfected HSFs with lentivirus to overexpress or knockdown certain differentially expressed TFs. Furthermore, comparative secretome analyses were conducted to investigate the proteins secreted by co-cultured microfat; changes in gene expression of HSFs were examined after the administration of the potential anti-fibrotic protein. Finally, the relationship between the key TF in HSFs and the microfat-secreted anti-fibrotic adipokine was analyzed.
The anti-fibrotic effect of microfat was confirmed by in vivo transplanted hypertrophic scar models, as the number of α-SMA-positive myofibroblasts was decreased and the expression of fibrosis-related genes downregulated. Co-cultured microfat suppressed the extracellular matrix production of HSFs in in vitro experiment, and the transcription factor ETV4 was primarily differentially expressed in HSFs when compared with normal skin fibroblasts. Overexpression of ETV4 significantly decreased the expression of fibrosis-related genes in HSFs at both mRNA and protein levels. Fetuin-A secreted by microfat could also downregulate the expression of fibrosis-related genes in HSFs, partially through upregulating ETV4 expression.
Our results demonstrated that transcription factor ETV4 is essential for the anti-fibrotic effect of microfat on hypertrophic scars, and that fetuin-A secreted by microfat could suppress the fibrotic characteristic of HSFs through upregulating ETV4 expression. Microfat wields an alleviative influence over hypertrophic scars via fetuin-A/ETV4 axis.
In this study, we injected microfat into transplanted hypertrophic scars in mouse models and investigated the subsequent histological changes and differential expression of mRNAs therein. As for in vitro studies, we co-cultured microfat and hypertrophic scar fibroblasts (HSFs) and analyzed molecular profile changes in HSFs co-cultured with microfat by RNA sequencing. Moreover, to identify the key transcription factors (TFs) which might be responsible for the anti-fibrotic function of microfat, we screened the differentially expressed TFs and transfected HSFs with lentivirus to overexpress or knockdown certain differentially expressed TFs. Furthermore, comparative secretome analyses were conducted to investigate the proteins secreted by co-cultured microfat; changes in gene expression of HSFs were examined after the administration of the potential anti-fibrotic protein. Finally, the relationship between the key TF in HSFs and the microfat-secreted anti-fibrotic adipokine was analyzed.
The anti-fibrotic effect of microfat was confirmed by in vivo transplanted hypertrophic scar models, as the number of α-SMA-positive myofibroblasts was decreased and the expression of fibrosis-related genes downregulated. Co-cultured microfat suppressed the extracellular matrix production of HSFs in in vitro experiment, and the transcription factor ETV4 was primarily differentially expressed in HSFs when compared with normal skin fibroblasts. Overexpression of ETV4 significantly decreased the expression of fibrosis-related genes in HSFs at both mRNA and protein levels. Fetuin-A secreted by microfat could also downregulate the expression of fibrosis-related genes in HSFs, partially through upregulating ETV4 expression.
Our results demonstrated that transcription factor ETV4 is essential for the anti-fibrotic effect of microfat on hypertrophic scars, and that fetuin-A secreted by microfat could suppress the fibrotic characteristic of HSFs through upregulating ETV4 expression. Microfat wields an alleviative influence over hypertrophic scars via fetuin-A/ETV4 axis.
摘要：
背景：肥厚性瘢痕是伤口愈合后的纤维化疾病，其特征是过度的细胞外基质沉积。自体微脂肪移植证明了其治疗的有效策略，因为它可以改善疤痕的质地并缓解相关症状。本研究旨在探讨微脂肪对增生性瘢痕的抗纤维化作用的潜在机制。
方法：在本研究中，我们在小鼠模型中将微脂肪注射到移植的肥厚性瘢痕中，并研究了随后的组织学变化和其中mRNA的差异表达。至于体外研究，我们共培养了微脂肪和增生性瘢痕成纤维细胞(HSF),并通过RNA测序分析了与微脂肪共培养的HSF的分子谱变化.此外，确定可能负责抗纤维化功能的微脂肪的关键转录因子（TFs），我们筛选了差异表达的TFs并用慢病毒转染HSF以过度表达或敲除某些差异表达的TFs。此外，进行了比较分泌组分析，以研究共培养的微脂肪分泌的蛋白质；在施用潜在的抗纤维化蛋白后，检查了HSF基因表达的变化。最后,分析了HSF中关键TF与微脂分泌的抗纤维化脂肪因子之间的关系。
结果：体内移植的增生性瘢痕模型证实了微脂肪的抗纤维化作用，随着α-SMA阳性肌成纤维细胞数量的减少和纤维化相关基因的表达下调。在体外实验中，共培养的微脂肪抑制了HSF的细胞外基质的产生，与正常皮肤成纤维细胞相比，转录因子ETV4主要在HSF中差异表达。ETV4的过表达在mRNA和蛋白质水平上均显着降低了HSF中纤维化相关基因的表达。微脂肪分泌的胎球蛋白A还可以下调HSF中纤维化相关基因的表达，部分通过上调ETV4表达。
结论：我们的结果表明，转录因子ETV4对于微脂肪对增生性瘢痕的抗纤维化作用至关重要，微脂肪分泌的胎球蛋白A可以通过上调ETV4的表达来抑制HSF的纤维化特性。微脂肪通过胎球蛋白A/ETV4轴对肥厚性瘢痕产生缓解影响。
方法：在本研究中，我们在小鼠模型中将微脂肪注射到移植的肥厚性瘢痕中，并研究了随后的组织学变化和其中mRNA的差异表达。至于体外研究，我们共培养了微脂肪和增生性瘢痕成纤维细胞(HSF),并通过RNA测序分析了与微脂肪共培养的HSF的分子谱变化.此外，确定可能负责抗纤维化功能的微脂肪的关键转录因子（TFs），我们筛选了差异表达的TFs并用慢病毒转染HSF以过度表达或敲除某些差异表达的TFs。此外，进行了比较分泌组分析，以研究共培养的微脂肪分泌的蛋白质；在施用潜在的抗纤维化蛋白后，检查了HSF基因表达的变化。最后,分析了HSF中关键TF与微脂分泌的抗纤维化脂肪因子之间的关系。
结果：体内移植的增生性瘢痕模型证实了微脂肪的抗纤维化作用，随着α-SMA阳性肌成纤维细胞数量的减少和纤维化相关基因的表达下调。在体外实验中，共培养的微脂肪抑制了HSF的细胞外基质的产生，与正常皮肤成纤维细胞相比，转录因子ETV4主要在HSF中差异表达。ETV4的过表达在mRNA和蛋白质水平上均显着降低了HSF中纤维化相关基因的表达。微脂肪分泌的胎球蛋白A还可以下调HSF中纤维化相关基因的表达，部分通过上调ETV4表达。
结论：我们的结果表明，转录因子ETV4对于微脂肪对增生性瘢痕的抗纤维化作用至关重要，微脂肪分泌的胎球蛋白A可以通过上调ETV4的表达来抑制HSF的纤维化特性。微脂肪通过胎球蛋白A/ETV4轴对肥厚性瘢痕产生缓解影响。
