口腔黏膜创伤修复中生长因子对牙龈成纤维细胞的影响及调节机制

Effects and mechanisms of growth factors on gingival fibroblasts during oral mucosal wound healing

目的:观察口腔黏膜创伤修复中不同生长因子对牙龈成纤维细胞迁移和增殖的影响,并探讨其可能的机制。方法:牙龈标本采集于复旦大学附属眼耳鼻喉科医院口腔科牙拔除术或助萌手术的患者。无菌条件下,用酶消化法培养人牙龈成纤维细胞,通过划痕实验、Transwell实验、CCK-8法和实时定量PCR等检测方法,比较表皮生长因子(EGF)、碱性成纤维细胞生长因子(bFGF)、转化生长因子(TGF)-β1、TGF-β3等不同生长因子对牙龈成纤维细胞在迁移、增殖及细胞外基质分子Ⅰ型及Ⅲ型胶胶原蛋白(CollagenⅠ及CollagenⅢ)、1型及13型基质金属蛋白酶(MMP-1及MMP-13)基因表达的差异。结果:划痕实验及Transwell实验的结果显示,EGF、bFGF、TGF-β1、TGF-β3均能够显著增加牙龈成纤维细胞的迁移(均P<0.05)。EGF在10～100ng/mL、TGF-β1仅在1ng/mL及TGF-β3在10～50ng/mL对牙龈成纤维细胞的增殖有明显的促进作用(P<0.05)。与对照组相比,EGF和bFGF显著上调了牙龈成纤维细胞CollagenⅢ、MMP-1及MMP-13基因的表达水平(P<0.05);TGF-β1,TGF-β3显著上调了牙龈成纤维细胞MMP-1及MMP-13基因的表达水平(P<0.05)。结论:EGF、bFGF、TGF-β1、TGF-β3对牙龈成纤维细胞迁移和增殖的促进作用可能是通过调控细胞外基质分子基因的表达来实现的。

Objective：To observe the effects of different growth factors on the migration and proliferation behaviors of the gingival fibroblasts during oral mucosal wound healing and to explore the possible mechanisms involved.Methods：Healthy gingival samples were obtained from patients with extraction or assisted surgery in Department of Stomatology of Eye Ear Nose and Throat Hospital of Fudan University.Human primary gingival cells were cultured by enzyme digestion method.Wound scratch assay,Transwell,CCK-8 and real time RT-PCR were used to analyze the effects of epidermal growth factor（EGF）,basic fibroblast growth factor（bFGF）,transforming growth factor（TGF）-β1 and TGF-β3 on the migration,proliferation,and the expressions of Collagen Ⅰ,Collagen Ⅲ,matrix metalloproteinase（MMP）-1 and MMP-13 in the gingival fibroblasts.Results：The EGF,bFGF,TGF-β1 and TGF-β3 could significantly increased the migration of gingival fibroblasts.EGF at 10-100 ng/mL,TGF-β1 at 1 ng/mL and TGF-β3 at 10-50 ng/mL obviously promoted the proliferation of gingival fibroblasts（P〈0.05）.Compared with the control group,EGF or bFGF treatment remarkably increased the expression levels of Collagen Ⅲ,MMP-1 and MMP-13（P〈0.05）,while TGF-β1 or TGF-β3 treatment significantly increased the expressions of MMP-1 and MMP-13（P〈0.05）.Conclusion：EGF,bFGF,TGF-β1 and TGF-β3 promoted the migration and proliferation of gingival fibroblasts possibly via regulating the expressions of extracellular matrix genes.