摘要
目的观察猪脂肪来源干细胞(ADSCs)对成纤维细胞的促增殖、迁移作用。方法取猪颈部皮下脂肪组织,酶消化法获得脂肪干细胞,细胞免疫荧光及流式细胞技术检测细胞表面抗原;原代培养并鉴定猪成纤维细胞;实验分为ADSCs组+成纤维细胞和成纤维细胞+成纤维细胞组,观察ADSCs培养液上清对成纤维细胞的促增殖及迁移作用;测定ADSCs培养液上清中血管内皮生长因子(VEGF)、血小板衍生的生长因子(PDGF-AA)及转化生长因子(TGF)-β1的含量。结果成功培养出猪ADSCs及猪成纤维细胞,ADSCs可促进成纤维细胞的增殖和迁移,ADSCs能分泌VEGF、TGF-β1和PDGF-AA,其浓度分别为(765.63±56.23)pg/ml,(133.42±24.35)pg/ml和(78.43±16.56)pg/ml。结论ADSCs可促进成纤维细胞的增殖和迁移,为ADSCs在创面愈合中的应用提供支持。
Objective This study was designed to evaluate the effect and mechanism of adipose derived stem cells(ADSCs)in the process of promoting fibroblasts proliferation and migration.Methods ADSCs were obtained from back subcutaneous adipose tissue of porcine with collagenase digestion and cultured in vitro.Flow cytometric analysis and cyto immumofluorescence staining technique were carried to detect the immunophenotypes of ADSCs.Porcine skin fibroblasts were cultured by tissue pieces and cyto immumofluorescence staining technique were used to detect the immunophenotypes of cells.The experiment was divided into ADSCs group and fibroblast group,cell to cell co culture experiment and transwell co culture experiment were used to evaluate the proliferation effect of ADSCs supernatant to fibroblasts.MTT assay was used to detect the proliferation effect of conditioned medium of ADSCs to fibroblasts.Also whether the conditioned medium of ADSCs has the stimulatory effect on migration of fibroblasts were determined by in vitro wound healing models.The concentrations of vascular endothelial growth factor(VEGF)、platelet derived growth factor(PDGF-AA)and transforming growth factor(TGF-β1)in conditioned medium of ADSCs were detected by using enzyme linked immunosorbent assay(ELISA).Results The stem cells isolated from subcutaneous adipose tissue were positive for CD44(90.2%)and CD105(99.2%),lacking in CD34(0.3%)and CD45(1.2%).Porcine skin fibroblasts were positive for Hsp47.ADSCs promoted fibroblasts proliferation,not only by cell-to-cell direct contact,which was confirmed by co-culture experiment,but also by paracrine activation through secretory factors,resolved by transwell co-culture and culturing with conditioned medium of ADSCs(P<0.05).Additionally,the fibroblasts showed vigorous proliferate and migratory responses to the conditioned medium of ADSCs.ADSCs produced a variety of growth factors:VEGF(765.63±56.23 pg/ml),TGF-β1(133.42±24.35 pg/ml)and PDGF(78.43±16.56 pg/ml).Conclusion ADSCs can promote the proliferation and migration of fibroblasts.This study can provide some theoretical and experimental basis for the treatment of wound healing.
作者
赵宝成
王振军
韩加刚
马华崇
禹三水
ZHAO Baocheng;WANG Zhenjun;HAN Jiagang(Department of General Surgery,Beijing Chaoyang Hospital,Capital Medical University,Beijing 100020,China)
出处
《临床外科杂志》
2020年第2期155-159,共5页
Journal of Clinical Surgery
基金
首都医科大学附属北京朝阳医院1351人才培养计划项目资助(CYXZ-2017-09).
关键词
脂肪来源干细胞
成纤维细胞
增殖
迁移
adipose derived stem cells
fibroblasts
proliferation
migration
