摘要
目的制备大鼠骨髓来源内皮祖细胞(BM-EPCs)组织工程学细胞膜片并行生物学功能研究。方法建立大鼠BM-EPCs培养体系,获取培养至14 d的EPCs。应用流式细胞术行细胞表面标志物检测并Dil标记的乙酰化低密度脂蛋白(Dil-acLDL)和异硫氰酸荧光素(FITC)标记的荆豆凝集素-1(FITC-UEA-1)共同染色方法鉴定BM-EPCs。应用Nunc UpCell Surface培养皿制备BM-EPCs组织工程学细胞膜片,测量膜片厚度并应用CCK-8法检测膜片中细胞代谢功能,体外血管形成实验检测内皮分化功能,间接酶联免疫吸附试验(ELISA)法检测细胞膜片旁分泌基质细胞衍生因子1α(SDF-1α)、血管内皮生长因子(VEGF)、表皮细胞生长因子(EGF)水平。结果通过建立大鼠BM-EPCs培养体系并成功获得EPCs膜片。体外实验结果显示,与EPC单细胞组比较,EPC膜片组具有更强的增殖能力及血管形成能力,且具有更强的旁分泌SDF-1α、VEGF作用,差异均有统计学意义[(4.86±0.09)pg/mL vs.(5.32±0.12)pg/mL、(162.88±0.96)pg/mL vs.(194.72±6.53)pg/mL,P<0.01]。两组细胞EGF水平比较,差异无统计学意义[(281.50±9.96)pg/mL vs.(294.16±8.98)pg/mL,P>0.05]。结论EPCs膜片具有更强的细胞增殖能力、血管形成能力及旁分泌作用,可为干细胞移植方式提供参考。
Objective To prepare the endothelial progenitor cell(EPC)sheet of rat bone marrow-derived endothelial progenitor cells(BM-EPCs),and to study its biological function.Methods The rat BM-EPCs culture system was established.EPCs cutured for 14 d were obtained.The flow cytometry was used to conduct the cell surface marker detection.`BM-EPCs was identified by the co-staining method with Dil-acLDL and FITC-UEA.The Nunc UpCell Surface culture dish was used to prepare the BM-EPCs tissue engineering cell sheet,the thickness of the sheet was measured and the CCK-8 method was used to detect the cell metabolism function in the cell sheet,the in vitro angiogenesis test was used to detect endothelial differentiation function and the indirect enzyme-linked immunosorbent assay(ELISA)was used to detect the levels of paracrine factors(SDF-1α,VEGF and EGF)in the culture supernatant.Results The culture system of BM-EPCs was established and the EPC sheet was successfully obtained.The in vitro experiments results showed that compared with the EPCs single-cell group,the EPCs sheet group had stronger proliferative capacity and angiogenicability(P<0.05),moreover had stronger effects of paracrine SDF-1αand VEGF factors,and the differences were statistically significant[(4.86±0.09)pg/mL vs.(5.32±0.12)pg/mL,(162.88±0.96)pg/mL vs.(194.72±6.53)pg/mL,P<0.01].There was no statistically significant difference in the EGF level between the two groups[(281.50±9.96)pg/mL vs.(294.16±8.98)pg/mL,P>0.05)].Conclusion The EPCs sheet has stronger cell proliferation ability,angiogenic ability and paracrine effect,which provides a reference for stem cell transplantation.
作者
薛奋龙
简锴陶
XUE Fenlong;JIAN Kaitao(Department of Cardiovascular Surgery,Tianjin Municipal First Central Hospital,Tianjin 300192,China;Department of Cardiovascular Surgery,Tianjin Municipal Chest Hospital,Tianjin 300051,China;Department of Cardiovascular Surgery,Shanghai DeltaHealth Hospital,Shanghai 200336,China)
出处
《重庆医学》
CAS
2021年第10期1647-1652,共6页
Chongqing medicine
基金
天津市自然科学基金项目(16JCYBJC23300)。
关键词
细胞膜片
内皮细胞
干细胞
血管生成
组织工程学
cell sheet
endothelial cells
stem cells
angiogenesis
tissue engineering
