转基因兔骨髓间充质干细胞对异体跟腱重建前交叉韧带生物力学特性的影响

Biomechanical study of achilles tendon allograft replacement for anterior cruciate ligament with transgenetic mesenchymal stem cells in rabbits

目的分别观察转PDGF-BB、VEGF165和TGFβ1基因的兔骨髓间充质干细胞(mesen chymal stem cells,MSCs)施加于异体跟腱重建兔前交叉韧带(anterior cruciate ligament,ACL)的影响,探讨改善和加强异体移植物重建ACL的途径。方法40只成年新西兰大白兔随机分为5组,每组8只。选用经Gamma射线照射,-80℃储存的同种异体跟腱进行前交叉韧带的重建。1组为对照组,2组为细胞组(MSCs),3组为经逆转录病毒转染稳定表达PDGF-BB的MSCs(PDGF-BB组),4组为经腺病毒转染稳定表达VEGF165的MSCs(VEGF165组),5组为经腺病毒转染稳定表达TGFβ1的MSCs(TGFβ1组)。各组细胞混合l00μL纤维蛋白胶粘附于重建韧带表面,术后12w取材。结果组织学观察发现,VEGF165组新生血管和成纤维细胞浸润高于其他各组。生物力学测试证明,PDGF-BB组和TGFβ1组移植物的最大载荷刚度、吸收能和最大拉张强度明显较对照组增强(P<0.05),细胞组和VEGF165组最大载荷、刚度和吸收能虽高于对照组,但无显著差异(P>0.05),VEGF165组拉断位移大于对照组(P<0.05)。所有实验组移植物、正常ACL和跟腱都在体部断裂。结论转VEGF165基因的MSCs可增加重建韧带的血管,但对生物力学功能无改善;转PDGF-BB基因和TGFβ1基因的MSCs有加速重建韧带成熟的作用。

Objective To observe the effect of rabbit anterior cruciate ligament （ACL） reconstruction with irradiated Achilles tendon allograft, which was added with PDGF-BB,VEGF165 or TGFβ1 genes transduced mesenchymal stem ceiis（MSCs）. Methods Forty male New Zealand white rabbits were divided randomly into five groups with eight animals for each：（1）control group, （2） MSCs group, （3） PDGF-BB transfected MSCs group, （4） VEGF165 transfected MSCs group, （5） TGFβ1 transfected MSCs group. The ACL was replaced with a Achilles tendon allograft. The 2×10^8 cells with 100 μL fibdn sealant was applied on the Achilles allograft. All animals were sacrificed 12 weeks after ACL reconstruction. Results Histologic findings showed that newly formed vessels and infiltrative fibroblasts were more abundant in VEGF165 group than other guoups. Biomechanical properties of PDGF-BB or TGFβ1 groups were significantly better than B control on the max load, the stiffness, the absorbed energy and the maximum tensile strength （P〈0.05）. The max load, the stiffness and the absorbed energy of MSCs or VEGF165 groups were higher than that of control group,while there were no significant differences （P〉0.05）. The snap displacement of VEGF165 group was significantly higher than that of control group （P〈0.05）. The rupture site was all at the body part of the graft, normal ACL and Achilles. Conclusion VEGF165 gene transfected MSCs could significantly promote angiogenesis in the reconstructed ligament, but it did not enhance the mechanical properties of the reconstructed ligament in the rabbit model. Both PDGF-BB gene and TGFβ1 gene transfected MSCs could accelerate the remoedling of the reconstructed ligament.