Worcester Polytechnic Institute, Department of Biology and Biotechnology, 100 Institute Road, Worcester, MA, 01609, United States.
Worcester Polytechnic Institute, Department of Biology and Biotechnology, 100 Institute Road, Worcester, MA, 01609, United States.
J Dermatol Sci. 2017 Dec;88(3):339-348. doi: 10.1016/j.jdermsci.2017.08.013. Epub 2017 Sep 1.
Previous human and animal studies have demonstrated the ability of exogenously administered basic fibroblast growth factor (FGF2) to act as an antifibrotic agent in the skin. Though the activity of FGF2 as an anti-scarring agent is well-established for fibrotic skin wounds, the mechanisms by which FGF2 exerts these actions are not entirely understood. Canonical FGF2 signaling proceeds in part via FGFR/MAPK pathways in human dermal fibroblasts, and FGF2 has been described to prevent or reverse the fibroblast-to-myofibroblast transition, which is driven by TGFβ signaling and understood to be an important step in the formation of a fibrotic scar in vivo. Thus, we set out to investigate the antagonistic effects of FGF2 on TGFβ signaling as well as the broader effects of MAPK inhibition on the TGFβ-mediated induction of myofibroblast gene expression.
To better understand the effects of FGF2 signaling pathways on myofibroblastic gene expression and cell phenotypes.
Human dermal fibroblasts were cultured in vitro in the presence of FGF2, TGFβ, and/or MAPK inhibitors, and the effects of these agents were investigated by molecular biology techniques including qRT-PCR, immunofluorescence, Western blot, and flow cytometry.
FGF2 inhibited TGFβ-mediated fibroblast activation, resulting in more rapidly proliferating, spindle-shaped cells, compared to the more slowly proliferating, flatter TGFβ-treated cells. Treatment with FGF2 also attenuated TGFβ-mediated increase in expression of myofibroblast markers smooth muscle α-actin, calponin, transgelin, connective tissue growth factor, ED-A fibronectin, and collagen I. FGF2-mediated antagonism of the TGFβ-mediated fibroblast-to-myofibroblast transition was reversed by small molecule inhibition of ERK or JNK, and it was potentiated by inhibition of p38. MAPK inhibition was demonstrated to have qualitatively similar effects even in the absence of exogenous FGF2, and small molecule inhibition of p38 MAPK was sufficient to attenuate TGFβ-mediated fibroblast activation.
Inhibition of select MAPK signaling pathways can reverse or potentiate anti-fibrotic FGF2 effects on human dermal fibroblasts, as well as exert their effects independently of exogenous FGF2 supplementation.
先前的人体和动物研究表明，外源性给予碱性成纤维细胞生长因子（FGF2）可在皮肤中充当抗纤维化剂。尽管 FGF2 作为抗瘢痕形成剂在纤维化皮肤伤口中的活性已得到充分证实，但 FGF2 发挥这些作用的机制尚不完全清楚。在人真皮成纤维细胞中，经典的 FGF2 信号通路部分通过 FGFR/MAPK 通路进行，并且已经描述 FGF2 可预防或逆转由 TGFβ信号驱动的成纤维细胞向肌成纤维细胞的转化，这被认为是体内形成纤维化瘢痕的重要步骤。因此，我们着手研究 FGF2 对 TGFβ信号的拮抗作用以及 MAPK 抑制对 TGFβ介导的肌成纤维细胞基因表达诱导的更广泛影响。
更好地了解 FGF2 信号通路对肌成纤维细胞基因表达和细胞表型的影响。
在体外培养人真皮成纤维细胞，存在 FGF2、TGFβ和/或 MAPK 抑制剂，通过包括 qRT-PCR、免疫荧光、Western blot 和流式细胞术在内的分子生物学技术研究这些试剂的作用。
FGF2 抑制 TGFβ介导的成纤维细胞激活，导致细胞增殖更快，呈纺锤形，与增殖较慢的 TGFβ处理细胞相比更为扁平。用 FGF2 处理还减弱了 TGFβ介导的肌成纤维细胞标志物平滑肌α-肌动蛋白、钙调蛋白、转谷氨酰胺酶、结缔组织生长因子、ED-A 纤连蛋白和 I 型胶原表达的增加。小分子抑制 ERK 或 JNK 逆转了 FGF2 介导的 TGFβ介导的成纤维细胞向肌成纤维细胞的转化，而抑制 p38 则增强了这种转化。即使没有外源性 FGF2 的存在，MAPK 抑制也表现出定性相似的作用，而小分子抑制 p38 MAPK 足以减弱 TGFβ介导的成纤维细胞激活。
抑制特定的 MAPK 信号通路可以逆转或增强 FGF2 对人真皮成纤维细胞的抗纤维化作用，并且可以独立于外源性 FGF2 补充发挥作用。
