Silk fibroin (SF) is a well-studied biomaterial for tissue engineering applications including wound healing. However, the signaling mechanisms underlying the impact of SF on this phenomenon have not been determined. In this study, through microarray analysis, regulatory genes of NF-ĸB signaling were activated in SF-treated NIH3T3 cells along with other genes. Immunoblot analysis confirmed the activation of the NF-ĸB signaling pathway as SF induced protein expression levels of IKKα, IKKβ, p65, and the degradation of IκBα. The treatment of NIH3T3 cells with SF also increased the expression of cyclin D1, vimentin, fibronectin, and vascular endothelial growth factor (VEGF). The expression of these factors by SF treatment was abrogated when NF-ĸB was inhibited by a pharmacological inhibitor Bay 11-7082. Knockdown of NF-ĸB using siRNA of IKKα and IKKβ also inhibited the SF-induced wound healing response of the NIH3T3 cells in a wound scratch assay. Collectively, these results indicated that SF-induced wound healing through the canonical NF-κB signaling pathway via regulation of the expression of cyclin D1, vimentin, fibronectin, and VEGF by NIH3T3 cells. Using an in vivo study with a partial-thickness excision wound in rats we demonstrated that SF-induced wound healing via NF-κB regulated proteins including cyclin D1, fibronectin, and VEGF. The in vitro and in vivo data suggested that SF induced wound healing via modulation of NF-ĸB signaling regulated proteins.

Statement of Significance

Silk fibroin has been effectively used as a dressing for wound treatment for more than a century. However, mechanistic insight into the basis for wound healing via silk fibroin has not been elucidated. Here we report a key mechanism involved in silk fibroin induced wound healing both in vitro and in vivo. Using genetic- and protein-level analyses, NF-κB signaling was found to regulate silk fibroin-induced wound healing by modulating target proteins. Thus, the NF-κB signaling pathway may be utilized as a therapeutic target during the formulation of silk fibroin-based biomaterials for wound healing and tissue engineering.

中文翻译：

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NF-κB信号是丝素蛋白伤口愈合过程中的关键

丝素蛋白（SF）是一种经过深入研究的生物材料，可用于包括伤口愈合在内的组织工程应用。但是，尚未确定SF影响此现象的信号机制。在这项研究中，通过微阵列分析，SF处理的NIH3T3细胞中的NF-ĸB信号调节基因与其他基因一起被激活。免疫印迹分析证实了SF诱导的IKKα，IKKβ，p65蛋白表达水平和IκBα降解，从而激活了NF-ĸB信号通路。SF处理NIH3T3细胞也可增加细胞周期蛋白D1，波形蛋白，纤连蛋白和血管内皮生长因子（VEGF）的表达。当药理抑制剂Bay 11-7082抑制NF-κB时，通过SF处理消除了这些因子的表达。在伤口刮擦试验中，使用IKKα和IKKβ的siRNA抑制NF-κB也抑制了SF诱导的NIH3T3细胞的伤口愈合反应。总的来说，这些结果表明SF通过NIH3T3细胞调节细胞周期蛋白D1，波形蛋白，纤连蛋白和VEGF的表达，通过典型的NF-κB信号通路诱导伤口愈合。使用在大鼠部分厚度切除伤口的体内研究中，我们证明了SF通过NF-κB调节的蛋白（包括细胞周期蛋白D1，纤连蛋白和VEGF）诱导的伤口愈合。在体外和体内数据表明，SF诱导的伤口经由信令调节蛋白NF-ĸB的调制愈合。

重要声明

丝素蛋白已被有效地用作伤口治疗的敷料已有一个多世纪的历史了。然而，尚未阐明对通过丝素蛋白进行伤口愈合的基础的机理见解。在这里，我们报告了丝素蛋白在体外和体内诱导伤口愈合的关键机制。使用遗传和蛋白质水平的分析，发现NF-κB信号通过调节靶蛋白来调节丝素蛋白诱导的伤口愈合。因此，在基于丝素蛋白的生物材料用于伤口愈合和组织工程的配制期间，NF-κB信号传导途径可用作治疗靶标。