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目的通过观测创伤早期大鼠异位骨化模型中基质金属蛋白酶9(matrix metalloproteinase 9,MMP-9)表达变化,探讨MMP-9对预测异位骨化发生的价值。方法取4~5周龄雄性SD大鼠132只,体重(135.0±6.5)g,随机分为两组(n=66)。实验组通过钳夹并切断跟腱制备异位骨化模型,对照组仅暴露跟腱。术后观察两组大鼠一般情况,2、3、4、5、6、7、8 d两组各取8只大鼠,取跟腱组织行大体、组织学及免疫组织化学染色观察,取血清及跟腱组织分别行ELISA及RT-PCR检测MMP-9蛋白含量及基因表达。通过两组剩余10只大鼠术后5、10周X线片及术后10周组织学观察结果,诊断异位骨化形成情况。结果所有大鼠均存活至实验完成。各时间点对照组大鼠跟腱无明显变化,呈正常跟腱结构;实验组随时间延长,跟腱断端硬度逐渐增加,出现大量不规则结缔组织及软骨细胞,MMP-9免疫组织化学染色呈深染阳性。各时间点实验组MMP-9蛋白含量及基因相对表达量均显著高于对照组(P<0.05);实验组MMP-9蛋白含量及基因相对表达量均总体呈上升趋势(P<0.05)。根据X线片及组织学观察结果,术后10周实验组大鼠跟腱组织均发生异位骨化,对照组均无异位骨化发生。结论在大鼠跟腱异位骨化早期,MMP-9表达明显升高,提示其对预测异位骨化发生有参考意义。
Objective To observe the expression of matrix metalloproteinase 9 (MMP-9) in the heterotopic ossification model of early trauma in rats and to explore the value of MMP-9 in predicting the occurrence of heterotopic ossification. Methods Thirty-two male Sprague-Dawley rats aged 4-5 weeks were randomly divided into two groups (n = 66), weighing 135.0 ± 6.5 g. The experimental group produced heterotopic ossification model by clamping and cutting the Achilles tendon, while the control group only exposed the Achilles tendon. Postoperatively, the general condition of the two groups of rats was observed. At 2, 3, 4, 5, 6, 7 and 8 d, 8 rats in each group were taken, and the Achilles tendon tissues were taken for gross, histological and immunohistochemical staining Serum and Achilles tendon tissue were detected by ELISA and RT-PCR, respectively, and MMP-9 protein content and gene expression. Ectopic ossification was diagnosed by X-ray at 5 and 10 weeks after operation and histology at 10 weeks after surgery. Results All rats survived until the experiment was completed. At each time point, there was no significant change in the Achilles tendon in the control group. The Achilles tendon structure was normal in the experimental group. With the extension of time, the hardness of the Achilles tendon increased gradually and a large number of irregular connective tissue and chondrocytes appeared. The expression of MMP-9 immunohistochemical staining Deep staining positive. The content of MMP-9 and the relative gene expression of the experimental group at each time point were significantly higher than those in the control group (P <0.05). The content of MMP-9 and the relative gene expression of the experimental group showed an overall upward trend (P <0.05). According to X-ray and histological observation, heterotopic ossification of Achilles tendon occurred in the experimental group at 10 weeks after operation, and no heterotopic ossification occurred in the control group. Conclusion In the early stage of heterotopic ossification of Achilles tendon, the expression of MMP-9 is significantly increased in rats, suggesting that it may have reference significance for the prediction of heterotopic ossification.