bFGF通过CK2α/α-catenin/β-catenin通路调控下游基因转录促成纤维细胞的迁移增殖和创面修复的机制研究

Skin wound repair is one of the difficulties in clinical therapy. Skin wound repair is a complex process in which fibroblast migration/proliferation is of great significance. Efficacy of bFGF on fibroblast migration/proliferation is well known, however, the underlying mechanism by which bFGF regulates cell migration/proliferation remains unclear. Previous studies revealed that bFGF upregulated the protein level of CK2α and α-catenin and the phosphorylation level of α-catenin, while the phosphorylated α-catenin could inhibit its degradation. Meanwhile, we had already observed that the rate of cell migration was retarded by CK2α inhibitor or α-catenin silence. And interference of α-catenin induced the decrease of nuclear β-catenin. Further through transcriptome analyses, we also had confirmed that nuclear β-catenin in transcriptional level modulate genes which were closely related to cell migration and proliferation. Thus, the objective of this project is to confirm the influence of CK2α, α-catenin and β-catenin regulated by bFGF on fibroblast migration and wound healing, to clarify the relationship between CK2α, α-catenin and β-catenin, and to confirm the connection between downstream genes of β-catenin and cell migration or proliferation. This study will provide new targets for drugs that skin repairs wound.

皮肤创面修复是临床亟待解决的难题之一。创面修复过程复杂，其中成纤维细胞迁移增殖对皮肤创面修复具有重要意义。bFGF促成纤维细胞迁移增殖作用已明确，但其调控机制尚不清楚。前期研究发现，bFGF能上调CK2α和α-catenin蛋白表达及α-catenin的磷酸化，而α-catenin磷酸化能抑制其自身降解。同时，抑制CK2α或干扰α-catenin都明显减缓细胞迁移。干扰α-catenin后细胞核中的β-catenin明显减少，且转录组学技术进一步证实核内β-catenin转录调控的下游基因与细胞迁移增殖息息相关。因此，本项目旨在前期研究基础上，深入探究受bFGF调控的CK2α、α-catenin和β-catenin对细胞迁移增殖和创伤愈合的影响及它们之间的相互作用关系，并确证受β-catenin转录调控的下游基因与细胞迁移增殖的关系。本项目将为开发皮肤创面修复药物提供新理论依据。

创面修复是临床亟待解决的难题之一，创面修复过程复杂，其中成纤维细胞迁移增殖对皮肤创面修复具有重要意义。bFGF促成纤维细胞迁移增殖作用已明确，但其调控机制尚不清楚。本研究的目的是进一步探究bFGF促成纤维细胞迁移增殖的潜在机制。.我们采用LC-MS/MS鉴定成纤维细胞经bFGF处理前后发生磷酸化改变的多肽，发现bFGF能上调9种磷酸化肽段（其中包含α-catenin Ser641），同时下调5种磷酸化多肽的表达。另外还发现LiCl（GSK3β（Ser9）抑制剂）和Bay（p-IκBα抑制剂）均能提高α-catenin Ser641的磷酸化及α-catenin总蛋白的表达水平。此外，我们发现α-catenin Ser641磷酸化水平调控不受ERK1/2的影响。bFGF和蛋白酶体抑制剂MG132能同时促进α-catenin的磷酸化和提高总α-catenin的表达，然而CK2抑制剂TBB则起到相反的抑制效果。上述结果提示α-catenin Ser641发生磷酸化后通过抑制α-catenin泛素化降解使α-catenin积累，免疫沉淀实验进一步验证了这一结论。我们还证实TBB能明显抑制成纤维细胞的迁移。另外在干扰内源性α-catenin表达的情况下，成纤维细胞的增殖、迁移和侵袭水平都受到明显抑制，同时增加了NF-κB p65入核水平，表明激活了NF-κB信号通路。.综上所述，本研究结果显示，bFGF通过磷酸化α-catenin抑制其泛素化降解使α-catenin积累，抑制NF-κB信号通路进而促成纤维细胞迁移增殖，加速皮肤创伤愈合。