Pan Yuyan, Wang Qiang, Luan Wenjie, Shi Yuedong, Liu Jiaqi, Qi Fazhi
Department of Plastic Surgery, Zhongshan Hospital, Fudan University, Shanghai, China.
Artificial Intelligence Center for Plastic Surgery and Cutaneous Soft Tissue Cancers, Zhongshan Hospital, Fudan University, Shanghai, China.
Ann Transl Med. 2021 Feb;9(4):348. doi: 10.21037/atm-21-176.
Adipose tissue has been proven to play a crucial role in wound healing, while kindlin-2, an integrin-associated protein, has been shown to regulate cell adhesion, migration, and differentiation. This study aimed to explore its involvement in the cell differentiation of 3T3-L1 preadipocytes and its role in wound healing.
Cell adhesion, Cell Counting Kit-8 (CCK-8), Transwell, and wound healing assays, along with adipogenic and osteogenic differentiation induction were performed in 3T3-L1 preadipocytes in which kindlin-2 was knocked down or overexpressed. In vivo, kindlin-2 (+/-) transgenic mice were constructed, and wound healing was analyzed by immunohistochemistry (IHC) in a mouse dorsal wound model. Real-time polymerase chain reaction (RT-PCR) and western blotting were performed to analyze the expression of adipokines and adipogenic markers in mouse wound tissues. Adipogenic differentiation induction of adipose tissue stromal vascular fraction (SVF) were performed, and the expression of adipogenic markers in SVF was detected by western blotting. The target signaling pathway highly related to adipogenic differentiation was explored by computational biology and verified by western blotting.
Knockdown of kindlin-2 was found to inhibit the adhesion, migration, and adipogenic differentiation of 3T3-L1 preadipocytes while promoting their osteogenic differentiation. In contrast, kindlin-2 overexpression resulted in increased adhesion, migration, and adipogenic differentiation of 3T3-L1 preadipocytes while reducing osteogenic differentiation. In vivo, downregulation of kindlin-2 inhibited adipogenesis in kindlin-2 transgenic mice, resulting in delayed wound healing by inhibiting inflammation, angiogenesis, collagen remodeling, and wound contraction. Mechanistically, we found that kindlin-2 could regulate adipogenic differentiation through PI3K/AKT/mTOR signaling pathway.
Our study revealed the essential role that kindlin-2 has in the differentiation and wound healing of 3T3-L1 preadipocytes, which offers a theoretical basis for further research and a novel strategy for wound healing.
脂肪组织已被证明在伤口愈合中起关键作用，而整合素相关蛋白kindlin-2已被证明可调节细胞黏附、迁移和分化。本研究旨在探讨其在3T3-L1前脂肪细胞分化中的作用及其在伤口愈合中的作用。
在敲低或过表达kindlin-2的3T3-L1前脂肪细胞中进行细胞黏附、细胞计数试剂盒-8（CCK-8）、Transwell和伤口愈合试验，以及脂肪生成和成骨分化诱导。在体内，构建kindlin-2（+/-）转基因小鼠，并通过免疫组织化学（IHC）在小鼠背部伤口模型中分析伤口愈合情况。进行实时聚合酶链反应（RT-PCR）和蛋白质印迹法分析小鼠伤口组织中脂肪因子和成脂标志物的表达。对脂肪组织基质血管成分（SVF）进行脂肪生成分化诱导，并通过蛋白质印迹法检测SVF中脂肪生成标志物的表达。通过计算生物学探索与脂肪生成分化高度相关的靶信号通路，并通过蛋白质印迹法进行验证。
发现敲低kindlin-2可抑制3T3-L1前脂肪细胞的黏附、迁移和脂肪生成分化，同时促进其成骨分化。相反，kindlin-2过表达导致3T3-L1前脂肪细胞的黏附、迁移和脂肪生成分化增加，同时减少成骨分化。在体内，kindlin-2的下调抑制了kindlin-2转基因小鼠的脂肪生成，通过抑制炎症、血管生成、胶原重塑和伤口收缩导致伤口愈合延迟。机制上，我们发现kindlin-2可通过PI3K/AKT/mTOR信号通路调节脂肪生成分化。
我们的研究揭示了kindlin-2在3T3-L1前脂肪细胞分化和伤口愈合中的重要作用，为进一步研究提供了理论基础，并为伤口愈合提供了新的策略。
