脂肪干细胞来源的生长因子与口腔黏膜成纤维细胞增殖

Adipose stem cell-derived growth factors and proliferation of oral mucosa fibroblasts

背景:近年来脂肪干细胞被证实对软组织创伤修复具有促进作用,关于其机制的研究中,更多的学者倾向于脂肪干细胞的旁分泌机制,即脂肪干细胞分泌的各种细胞因子对创伤部位细胞的修复功能具有促进作用。但脂肪干细胞分泌的因子种类,每种因子的含量,以及对软组织创伤修复,尤其是口腔黏膜的作用,鲜有报道。目的:观察脂肪干细胞条件培养液中血管内皮生长因子、血小板源性生长因子以及碱性成纤维细胞生长因子对口腔黏膜成纤维细胞增殖的影响。方法:采用蛋白芯片技术,检测脂肪干细胞条件培养液中血管内皮生长因子、血小板源性生长因子以及碱性成纤维细胞生长因子的含量。分别于1,2,3,4,5d时,采用CCK-8法检测0,1,10,20,50,100μg/L血管内皮生长因子、血小板源性生长因子以及碱性成纤维细胞生长因子以及在脂肪干细胞条件培养液中分别加入上述各因子的中和抗体后,对口腔黏膜成纤维细胞增殖的影响。结果与结论:脂肪干细胞条件培养液中血管内皮生长因子、血小板源性生长因子以及碱性成纤维细胞生长因子信号值均大于300。脂肪干细胞条件培养液对成纤维细胞的增殖有明显的促进作用,其中,血小板源性生长因子和碱性成纤维细胞生长因子的促进作用非常明显,且随着因子浓度的变化而呈现峰值性改变,血小板源性生长因子的促进作用在50μg/L时达到峰值,碱性成纤维细胞生长因子的促进作用在1μg/L时达到峰值(P≤0.05),两者的中和抗体均能抑制脂肪干细胞条件培养液对口腔黏膜成纤维细胞的促进作用;而血管内皮细胞生长因子对成纤维细胞增殖无明显的促进作用。实验结果提示脂肪干细胞条件培养液可促进口腔黏膜成纤维细胞的增殖,其中所含有的血小板源性生长因子和碱性成纤维细胞生长因子的促进作用非常明显,且具有浓度依赖性,因此应注意选择适宜的细胞因子浓度,从而有效地促进成纤维细胞增殖。

BACKGROUND： Adipose stem cells have been confirmed to promote the repair of soft tissue after damage, and the action mechanism is possibly related to the paracrine of adipose stem cells, that is adipose stem cells secrete a variety of cytokines, which may promote the restoration of damaged cells. However, little report addressed the types of adipose stem cells secreted factors, contents of each factor, and role in soft tissue repair after damage, especially oral mucosa. OBJECTIVE： To observe the influence of vascular endothelial growth factor, platelet-derived growth factor, and basic fibroblast growth factor in adipose stem cell-conditioned medium on the proliferation of oral mucosa fibroblasts. METHODS： Protein microarray analysis was used to analyze the contents of vascular endothelial growth factor, platelet-derived growth factor, and basic fibroblast growth factor in adipose stem cell-conditioned medium. CCK8 analysis was used to analyze the effects of adipose stem cell-conditioned medium with different concentrations of vascular endothelial growth factor, platelet-derived growth factor, and basic fibroblast growth factor （0, 1, 10, 20,50 and 100 μg/L） or the neutralizing antibody of the three kinds of cytokines on the proliferation of oral mucosa fibroblast on 1, 2, 3, 4 and 5 days. RESULTS AND CONCLUSION： High contents of vascular endothelial growth factor, platelet-derived growth factor, and basic fibroblast growth factor were contained in adipose stem cell-conditioned medium （signal value 300）. Adipose stem cell-conditioned medium could promote the proliferation of oral mucosa fibroblasts, wherein, the promotion effects of platelet-derived growth factor and basic fibroblast growth factor were very significant, and the peak changes could be observed with variation of cytokines concentrations. The optimal concentrations of platelet-derived growth factor and basic fibroblast growth factor were 50 μg/L and 1 μg/L, respectively （P ≤ 0.05）. The neutralizing antibody of platelet-derived growth factor and basic fibroblast growth factor inhibited the promotion effect of adipose stem cell-conditioned medium. On the other hand, vascular endothelial growth factor had no significant effect on the proliferation of oral mucosa fibroblasts. Adipose stem cell-conditioned medium can promote the proliferation of oral mucosa fibroblasts, platelet-derived growth factor and basic fibroblast growth factor in adipose stem cell-conditioned medium have obvious promotion effects, which are dependent on the cytokine concentrations. Therefore, we should pay attention to choose the optimal concentrations of cytokines, thereby effectively promoting the proliferation of fibroblasts.