碳化二亚胺交联同种异体肌腱修复兔前交叉韧带损伤的实验研究

Tendon allograft cross-linked and heparinized by EDC/NHS to repair injure to anterior cruciate ligament in rabbits

目的探讨利用碳化二亚胺（EDC）交联的方法对同种异体肌腱进行预处理，改善其修复效果。方法获取兔同种异体跟腱，应用EDC交联并肝素化，应用Ⅱ型胶原酶酶解法检测交联组和未交联组肌腱的降解率并进行比较。应用四氮唑蓝法检测EDC交联（EDC交联组）对细胞增殖率（1、3、7d）的影响，其结果与生理盐水（阴性对照组）及苯酚（阳性对照组）进行比较。取28只雄性SD大鼠，每组14只，随机分为EDC交联组（埋植EDC交联肌腱）和未交联组（埋植未交联肌腱），分别于术后1、4周取材（每一时间点取7只），行HE染色观察免疫反应并进行炎症反应分级比较。用交联绀和术交联组肌腱对兔前交叉韧带损伤进行修复，1、3、6个月取材做光镜和扫描电镜观察，对比不同时间段腱骨愈合情况。结果EDC交联并肝素化组降解率【（6．26±3．16）％】低于未交联组【（30．70±10．24）％】，差异有统计学意义（t=14．200，P=0．025）。培养1、3、7d。阴性对照组和EDC交联组的相对增殖度分别与阳性对照组比较，差异均有统计学意义（P〈0．05，表1），EDC交联组肌腱无明显细胞毒性作用；1、4周未交联组和交联组炎症反应分级，差异有统计学意义（P〈0．05）。移植后组织学和电镜扫描观察可见未交联组愈合比交联组慢。结论EDC交联并肝素化的肌腱，其稳定性增强，抗酶降解能力提高，免疫原性降低，且腱骨愈合加快。

Objective To explore a new way to improve the repairing effect of tendon allograft by pretreatment with ethyldimethylaminopropyl carbodiinfide （EDC） cross-linking. Methods Rabbits＇ tendon allografts were obtained by lyopyilization. The allografts were then cross-linked and heparinized by EDC and N-hydroxy-suecinimide （NHS） respectively. The degradation rate, cytotoxicity and histocompatibility of the cross-linked allografts were detected and compared with those of untreated at different time points. The injured anterior cruciate ligaments （ACL） were repaired by tendon allografts with and without cross-linking pretreatment in rabbits, The tendon-bone healing was observed and compared by light microscopy and scanning electron microscopy （SEM） at 1, 3, 6 months. Results The degradation rate of the pretreated tendon allografts [ （6. 26 ± 3.16）% ] was significantly lower than that of the unpretreated [ （30.70 ± 10.24）% ] （ t = 14. 200, P = 0. 025） . The pretreated tendon allografts were atoxigenic and produced significantly lower inflammatory reaction （ P 〈 0.05）. The pretreated tendon allografts also showed more powerful capability of repairing ACL injury and shorter time of tendon-bone healing. Conclusion Tendon allografts cross-linked and heparinized by EDC/NHS may promote tendon-bone healing with improved stability, degradation and bioeompatibility.