Yu Du, Xiao Haijun, Xue Feng, Pan Mingmang, Ju Jinyong, Tang Guo
Third Affiliated Clinical College of Southern Medical University, Guangzhou Guangdong, 510515, P. R. China.
Department of Orthopedics, Fengxian Hospital Affiliated to Southern Medical University.
Zhongguo Xiu Fu Chong Jian Wai Ke Za Zhi. 2016 Sep 8;30(9):1098-1103. doi: 10.7507/1002-1892.20160224.
To explore the role and significance of hypoxia inducible factor lα (HIF-lα) and hypoxia microenvironment in the pathogenesis of post-traumatic heterotopic ossification by detecting the expression of HIF-lα in rat model of heterotopic ossification after Achilles tenotomy.
A total of 140 male Sprague Dawley rats, aged 8-10 weeks, and weighing (210.1±10.6) g, were randomly divided into experimental group (=70) and control group (=70). In experimental group, the Achilles tendon was cut off and clamped to prepare post-traumatic heterotopic ossification model; in control group, only Achilles tendon was exposed. The general condition of rats was observed after operation, and at 2, 3, 4, 5, 6, 7, 8, 10, 12, and 14 days after operation, the Achilles tendon tissue was harvested from 6 rats for gross observation, histological observation, and immunohistochemical staining observation, and real-time fluorescence quantitative PCR and Western blot were used to detect the expressions of HIF-lα gene and protein at different time points in 2 groups. The X-ray films were taken and histological examination was done at 10 weeks after operation to evaluate the formation of heterotopic ossification.
During the experiment, 1 rat died in experimental group at 3 days after operation, and the other rats survived to the end of the experiment. Gross and histological staining showed that the Achilles tendon had no obvious change, with normal tendon structure in control group at each time point. In experimental group, atrophy and necrosis of Achilles tendon stump were observed, with infiltration of inflammatory cells; and the hardness of Achilles tendon tissue gradually increased with the time; there were a large number of irregular connective tissue and cartilage cells. When compared with control group, the HIF-lα mRNA and protein expressions were significantly increased in experimental group at each time point (<0.05). Immunohistochemical staining showed that HIF-lα was positive in experimental group. According to the results of X-ray films and histological examination at 10 weeks after operation, heterotopic ossification was found in experimental group, but no heterotopic ossification in control group.
The expression of HIF-lα significantly increases at early stage of post-traumatic heterotopic ossification after Achilles tenotomy, suggesting that the local hypoxia microenvironment plays an important role in the pathogenesis of heterotopic ossification.
通过检测跟腱切断术后异位骨化大鼠模型中缺氧诱导因子1α（HIF-1α）的表达，探讨HIF-1α及缺氧微环境在创伤后异位骨化发病机制中的作用及意义。
选取140只8-10周龄、体重（210.1±10.6）g的雄性Sprague Dawley大鼠，随机分为实验组（n=70）和对照组（n=70）。实验组切断并钳夹跟腱制备创伤后异位骨化模型；对照组仅暴露跟腱。术后观察大鼠一般情况，于术后2、3、4、5、6、7、8、10、12及14天，每组分别取6只大鼠的跟腱组织进行大体观察、组织学观察及免疫组化染色观察，采用实时荧光定量PCR及Western blot检测两组不同时间点HIF-1α基因及蛋白的表达。术后10周拍摄X线片并进行组织学检查，评估异位骨化形成情况。
实验过程中，实验组1只大鼠术后3天死亡，其余大鼠存活至实验结束。大体及组织学染色显示，对照组各时间点跟腱无明显变化，腱结构正常。实验组可见跟腱残端萎缩、坏死，有炎性细胞浸润；跟腱组织硬度随时间逐渐增加；有大量不规则结缔组织及软骨细胞。与对照组比较，实验组各时间点HIF-1α mRNA及蛋白表达均显著升高（P<0.05）。免疫组化染色显示实验组HIF-1α呈阳性。根据术后10周X线片及组织学检查结果，实验组出现异位骨化，对照组未出现异位骨化。
跟腱切断术后创伤后异位骨化早期HIF-1α表达显著增加，提示局部缺氧微环境在异位骨化发病机制中起重要作用。
