FGF-2基因转染rBMSCs构建组织工程骨移植修复兔股骨头缺血性坏死模型

Construction of tissue engineered bone with FGF-2 gene transfected rBMSCs for repairing femoral head necrosis in rabbit

[目的]评价FGF-2基因转染r BMSCs构建组织工程骨移植修复兔股骨头缺血性坏死的治疗效果。[方法]采用密度梯度离心联合贴壁法体外分离、培养rBMSCs,并检测与XACB的生物相容性。用FGF-2基因过表达慢病毒转染rBMSCs,并检测FGF-2基因在rBMSCs成功过表达后,与XACB复合培养构建组织工程骨。将成功建立的激素型兔股骨头缺血性坏死模型随机分为5组:A组(control)、B组(XACB)、C组(XACB+r BMSCs)、D组(XACB+rBMSCs+Lv-GFP)、E组(XACB+rBMSCs+Lv-FGF2/GFP),进行组织工程骨移植。术后3、6、12周分点取材,进行大体解剖观察、X线片检查、HE染色评估新骨形成面积,评价该组织工程骨修复兔股骨头缺血性坏死模型的治疗效果。[结果]采用密度梯度离心联合贴壁法可获取高度均一的rBMSCs,与XACB的生物相容性良好。FGF-2基因转染rBMSCs(MOI=100),其效率在95%以上,q PCR及Western Blot检测rBMSCs可稳定过表达FGF-2。组织工程骨植入术后6周,大体标本观察及X线片检查显示E组缺损区修复面积在80%以上,术后12周缺损区完全修复,其成骨作用明显强于其他各组,而且X线评分及新骨形成面积也均高于其他各组(P<0.05)。[结论]FGF-2基因过表达慢病毒转染rBMSCs与XACB复合培养构建的组织工程骨可有效促进兔股骨头缺血坏死的修复效果。

[Objective] To evaluate the efficacy of FGF-2 gene over expression lentivirus transfected BMSCs in the construction of tissue-engineered bone graft for avascular necrosis of femoral head in rabbits. [Methods] The tissue-engineered bone was constructed with FGF-2 gene overexpressed lentivirus-transfected BMSCs and XACB. After the model of early avascular necrosis of femoral head was established by LPS combined with hormone in rabbit, the animals were randomly di- vided into 5 groups： including the group A （control）, B （XACB） , C （XACB＋rBMSCs） , D （XACB＋rBMSCs＋Lv-GFP） and E （XACB ＋rBMSCs＋Lv-FGF2/GFP） in term of the materials implanted in the femoral heads. The results were evaluated by gross anatomy and X-ray examination, HE staining to assess the effect of the tissue engineering bone on rabbit avascular necrosis of the femoral head. [Results] FGF-2 gene was transfected into rBMSCs （MOI=100） by lentivirus with efficiency above 95%, because rBMSCs were successfully overexpressing FGF-2 by qPCR and Western Blot. After tissue engineering bone implantation, X-ray examination and gross specimen observation showed that the repair area in the group E was more than 80% at 6 weeks, and the defect was completely repaired at 12 weeks, osteogenesis was stronger than other groups. X-ray score and new bone formation area in the group E were statistically higher than other groups （P〈0.05） . [Conclusion] The tissue engineering bone constructed by FGF-2 gene overexpression lentivirus transfected BMSCs and XACB does effectively promote the repair of avascular necrosis of the femoral head.