[Construction of injectable tissue engineered adipose tissue with fibrin glue scaffold and human adipose-derived stem cells transfected by lentivirus vector expressing hepatocyte growth factor].

目的: 探讨构建可注射型组织工程脂肪的可行性，为临床软组织缺损修复及美容填充提供简便易行的方法。. Methods: Human adipose-derived stem cells (hADSCs) were extracted from the lipid part of human liposuction aspirate by enzymatic digestion and identified by morphological observation, flow cytometry, and adipogenic induction. The hADSCs underwent transfection by lentivirus vector expressing hepatocyte growth factor and green fluorescent protein (HGF-GFP-LVs) of different multiplicity of infection (MOI, 10, 30, 50, and 100), the transfection efficiency was calculated to determine the optimum MOI. The hADSCs transfected by HGF-GFP-LVs of optimal MOI and being adipogenic inducted were combined with injectable fibrin glue scaffold, and were injected subcutaneously into the right side of the low back of 10 T-cell deficiency BALB/c female nude mice (transfected group); non-HGF-GFP-LVs transfected hADSCs (being adipogenic inducted) combined with injectable fibrin glue scaffold were injected subcutaneously into the left side of the low back (untransfected group); and injectable fibrin glue scaffold were injected subcutaneously into the middle part of the neck (blank control group); 0.4 mL at each point. Twelve weeks later the mice were killed and the implants were taken out. Gross observation, wet weight measurement, HE staining, GFP fluorescence labeling, and immunofluorescence staining were performed to assess the in vivo adipogenic ability of the seed cells and the neovascularization of the grafts. 结果: 经鉴定所培养细胞为 hADSCs。MOI 为10 和 30 时 HGF-GFP-LVs 转染率较低，MOI 为50 和 100 时转染率较高（均>80%），确定最适 MOI 为 50。移植 12 周，转染组和未转染组均获得外观类似脂肪组织的新生物，湿重分别为（32.30±4.06）mg 和（25.27±3.94）mg，差异有统计学意义（ t=3.929， P=0.001）；空白对照组未见新生物。转染组新生脂肪细胞数为（126.93±5.36）个/视野，显著高于未转染组的（71.36±4.52）个/视野（ t=30.700， P=0.000）。荧光显微镜下可见部分单房脂肪细胞发出网状绿色荧光。免疫荧光染色示转染组血管密度为（16.37±2.76）个/视野，显著高于未转染组的（9.13±1.68）个/视野（ t=8.678， P=0.000）。. 结论: 以 hADSCs 为种子细胞，经慢病毒转染 HGF 基因并成脂诱导后与纤维蛋白胶支架复合可在体内成功构建成熟脂肪，能促进移植后的血管新生，从而提高移植物的存活率。.