Department of Agricultural Biotechnology, Animal Biotechnology Major, and Research Institute of Agriculture and Life Science, Seoul National University, Seoul, Korea.
Department of Veterinary Physiology, College of Veterinary Medicine, Research Institute for Veterinary Science, and BK21 PLUS Program for Creative Veterinary Science Research, Seoul National University, Seoul, Korea.
Br J Pharmacol. 2018 Sep;175(17):3548-3562. doi: 10.1111/bph.14423. Epub 2018 Jul 26.
The sonic hedgehog pathway (Shh) plays a central role in maintaining stem cell function and behaviour in various processes related to self-renewal and tissue regeneration. However, the therapeutic effect of Shh on mouse embryonic stem cells (mESCs) has not yet been clearly elucidated. Thus, we investigated the effect of Shh on the regulation of mESC behaviour as well as the effect of Shh-pretreated mESCs in skin wound healing.
The underlying mechanisms of Shh signalling pathway in growth and motility of mESCs were investigated using Western blot analysis, a cell proliferation assay and cell migration assay. In addition, the effect of Shh-pretreated mESCs in skin wound healing was determined using a mouse excisional wound splinting model.
Shh disrupted the adherens junction through proteolysis by activating MMPs. In addition, the release of β-catenin from adherens junctions mediated by Shh led to cell cycle-dependent mESC proliferation. Shh-mediated Gli1 expression led to integrin β1 up-regulation, followed by FAK and Src phosphorylation. Furthermore, among the Rho-GTPases, Rac1 and Cdc42 were activated in a Shh-dependent manner while F-actin expression was suppressed by Rac1 and Cdc42 siRNA transfection. Consistent with the in vitro results, the skin wound healing assay revealed that Shh-treated mESCs increased angiogenesis and skin wound repair compared to that in Shh-treated mESCs transfected with integrin β1 siRNA in vivo.
Our results imply that Shh induces adherens junction disruption and integrin β1-dependent F-actin formation by a mechanism involving FAK/Src and Rac1/Cdc42 signalling pathways in mESCs.
sonic hedgehog 通路（Shh）在与自我更新和组织再生相关的各种过程中，对维持干细胞功能和行为起着核心作用。然而，Shh 对小鼠胚胎干细胞（mESCs）的治疗效果尚未得到明确阐明。因此，我们研究了 Shh 对 mESC 行为调控的影响，以及 Shh 预处理的 mESCs 在皮肤创伤愈合中的作用。
通过 Western blot 分析、细胞增殖试验和细胞迁移试验，研究了 Shh 信号通路在 mESCs 生长和迁移中的作用机制。此外，采用小鼠切创夹板模型确定了 Shh 预处理的 mESCs 在皮肤创伤愈合中的作用。
Shh 通过激活 MMPs 破坏粘着连接，从而破坏粘着连接。此外，Shh 介导的 β-catenin 从粘着连接释放导致细胞周期依赖性的 mESC 增殖。Shh 介导的 Gli1 表达导致整合素β1 的上调，随后 FAK 和 Src 磷酸化。此外，在 Rho-GTPases 中，Rac1 和 Cdc42 以 Shh 依赖的方式被激活，而 Rac1 和 Cdc42 siRNA 转染则抑制 F-actin 的表达。与体外结果一致的是，皮肤创伤愈合试验表明，与体内转染整合素β1 siRNA 的 Shh 处理 mESCs 相比，Shh 处理的 mESCs 增加了血管生成和皮肤创伤修复。
我们的结果表明，Shh 通过 FAK/Src 和 Rac1/Cdc42 信号通路诱导 mESCs 粘着连接破坏和整合素β1 依赖性 F-actin 形成。
