混编肌腱重建兔膝前交叉韧带关节腔内愈合的组织学研究

Histological studies of intra-articular healing in anterior cruciate ligament reconstruction with auto/allo-graft in rabbit

[目的]比较自体-异体混编肌腱和同种异体肌腱重建兔前交叉韧带在关节腔内的重塑过程。[方法]取新西兰大白兔的双侧后腿趾长伸肌腱作为移植材料装入无菌塑料瓶,深低温-80℃保存14 d,放入-20℃冰箱保存备用。将40只新西兰兔随机平均分成自体异体肌腱混编组(混编组)和同种异体肌腱组(异体组),各20只。取兔右膝关节内侧切口,切除前交叉韧带后,按照正常兔前交叉韧带位置,选取胫骨与股骨骨道进行重建。分别于术后3、8、12、24周对重建肌腱行大体观察后应用HE染色、甲苯胺蓝染色、CD31免疫组织化学染色进行组织学评估比较。[结果]术后3周,两组均有炎性细胞浸润,肌腱内部有坏死现象。术后8周,两组可见大量圆形新生细胞由肌腱表面向中心推进,胶原纤维排列无序,细胞浸润程度以混编组相对较高。12周时,混编组内部大量圆形新生细胞,胶原纤维开始出现有序排列,异体组纤维排列仍紊乱,肌腱中心偶可见无细胞区。24周时,混编组新生细胞呈梭形,胶原纤维沿韧带长轴方向排列有序。异体组细胞仍呈圆形,胶原纤维出现有序排列。甲苯胺蓝染色结果显示:3、8周时,两组均未见异染。12周时混编组甲苯胺蓝染色出现异染,异体组未见异染,术后24周时两组均出现异染。CD31免疫组织化学染色血管计数及HE染色细胞计数在术后3、8、12周均高于异体组(P≤0.05),24周时低于异体组(P≤0.05)。[结论]混编肌腱关节腔内愈合经历缺血坏死期、再血管化及细胞增殖期和韧带重塑期,其塑型改建过程明显快于同种异体移植物。

[ Objective] To compare the intra - articular remodeling processes of anterior cruciate ligament （ACL） recon- struction with auto/allo -graft and allograft in rabbits. [Method] Tendons of the extensor digitorum longus were harvested from both rear legs of New Zealand rabbits as graft materials and were stored in sterilized plastic bottles at -80℃for 14 days, and were then kept in the refrigerator at -20℃. A total of 40 New Zealand rabbits were randomly and equally divided into au- to/allo - graft group and allograft group. The ACL was excised after incision of the right medial knee, and then the ACL was re- constructed through bone tunnels of the tibia and the femur at the normal site of ACL in rabbits. The reconstructed ACL was generally observed before application of hematoxylin and eosin （HE） staining, toluidine blue staining, and CD31 immunohisto- chemical staining to perform histological evaluation and comparison at 3, 8, 12, and 24 weeks after treatment. [ Result] At 3 weeks after treatment, both groups showed inflammatory cell infiltration, as well as necrosis inside the tendon. At 8 weeks after treatment, both groups showed large numbers of round new cells moving from the surface to the center of the tendon; the col- lagenous fibers were arranged in disorder, and the autoJallo - graft group showed a higher degree of cellular infiltration. At 12weeks after treatment, the auto/allo - graft group showed large numbers of round new ceils inside, and the collagenous fibers started to be arranged in order; the allograft group still showed disordered arrangement of collagenous fibers, and cell - free areas could be seen at the center of the tendon. At 24 weeks after treatment, the auto/allo -graft group showed spindle- shaped new cells, and the collagenous fibers werearranged along the long axis of the tendon; the aUograft group still had round cells and orderly arrangement of collagetous fi- bers. Toluidine blue staining showed that at weeks 3 and 8, no metachromasia was seen in the two groups ; at week 12, the au- to/allo -graft group had metachromasia, but no metachromasia was seen in the allograft group; at week 24, both groups showed no metachromasia. CD31 immunohistochemical staining and HE staining showed that the blood vessel count and cell count in the auto/allo-graft group were higher than those in the allograft group at weeks 3, 8, and 12 （P〈0. 05）, but were lower than those in the allograft at week 24 （P〈0. 05 ）. [ Conclusion] Intra- articular healing with auto/allo -graft experiences ischemic necrosis, revascularization, cell proliferation, and ligament reconstruction, a process significantly faster than that with allograft.