脊髓损伤大鼠关节软骨转化生长因子β和肿瘤坏死因子α表达的变化

Changes of the expressions of transforming growth factor beta and tumor necrosis factor alpha in articular cartilage of rats with spinal cord injury

目的:观察脊髓损伤模型大鼠膝关节软骨组织中转化生长因子β和肿瘤坏死因子α的表达变化,分析其在软骨代谢变化中的意义,为关节软骨的修复重建方面提供理论基础。方法:实验于2004-05/12在西安交通大学医学院骨病教研室完成。选择清洁级健康成年SD雌性大鼠36只,按随机数字表法分为脊髓损伤组和对照组,各18只。按改良Allen打击法建立大鼠脊髓中度损伤模型,对照组仅行T10椎板切除术。伤后1,3,6周麻醉下处死,两组每时间点均为6只。用ABC法进行转化生长因子β、肿瘤坏死因子α免疫组化染色及通过倒置光学显微镜及成像系统行灰度值扫描,对大鼠膝关节软骨组织中转化生长因子β、肿瘤坏死因子α的表达变化进行观察并采用SPSS12.0软件对结果进行统计分析。结果:脊髓损伤组在术后6d因泌尿系感染死亡1只,对照组组术后3d因硬膜外血肿死亡1只,后补齐进入结果分析大鼠36只。①免疫组化观察阳性表达信号为红色颗粒状,主要定位于胞浆和胞膜部位。肿瘤坏死因子α、转化生长因子β主要分布关节软骨表层中,阴性对照均呈现弱表达状态。②脊髓损伤后第1,3周时肿瘤坏死因子α阳性表达强度高于对照组穴181.13±7.34,193.19±8.64;178.02±5.05,190.78±8.59;P<0.05雪,第6周时与对照组比较,差异有非常显著性意义(175.56±6.05,192.98±8.26,P<0.01)。③在脊髓损伤1周时转化生长因子β表达与对照组相比差异无显著性意义(P>0.05),第3,6周时表达差异有显著性意义穴182.25±4.73,191.40±6.87;176.85±7.68,191.41±8.53;P<0.05雪。结论:脊髓损伤后肿瘤坏死因子α的表达具有时间上的早期性,转化生长因子β的表达则随着脊髓神经功能的恢复而增强,两者在第3周时关节软骨中表达均增强。

AIM： To observe the change of expressions of transforming growth factor （TGF） β and tumor necrosis factor （TNF） α in rat articular cartilage in the spinal cord injury （SCI） models and analyze its meaning in the variation of cartilage metabolism so as to provide rationale in the clinical articular cartilage re-establishment. METHODS： The experiment was conducted in Department of Orthopaedics, Medical School of Xi＇an Jiaotong University from May to December 2004. Totally 36 healthy mature female SD rats of clean degree were randomly divided into 2 groups： SCI group and control group with 18 rats in each group. Moderate injury models were established with modified Allen method. T10 laminectomy was performed in rats of the control group. The rats were killed at weeks 1, 3 and 6 after injury, 6 rats in each time point. TGF-β and TNF-α were stained with immunohistochemical staining （ABC method） and gray scanning was conducted with inverse photon microscope and imaging system. TGF-β and TNF-α in the knee joint cartilage of rats were observed, and the result and statistics processing were analyzed with the SPSS 12.0 software. RESULTS： One rat died in the SCI group at day 6 after operation, because of urinary system infection and one rat died in the control group at day 3 after operation; because of epidural hematoma, and then another 2 rats were included. A total of 36 rats were involved in the result analysis. ① Positive signals were expressed as red granules in the immunohistochemical staining, mainly pitched at the cellular plasm and the cellular membrane. TGF-β and TNF-α were mainly distributed in the surface layer of the articular cartilage, with a weak expression of the negative control. ② In SCI group the positive expression intensity of TNF-α was evidently higher than in the control group at weeks 1 and 3 （181.13±7.34, 193.19±8.64; 178.02±5.05, 190.78±8.59; P 〈 0.05）. At week 6 there were significant differences as compared with the control group （175.56±6.05, 192.98±8.26, P 〈 0.01）. ③ TGF-β expression in the SCI group had no remarkable difference as compared with the control group at week 1 （P 〉 0,05）. There were remarkably significant difference at weeks 3 and 6 （182.25±4.73, 191.40±6.87; 176.85±7.68, 191.41±8.53; P 〈 0.05）. CONCLUSION： The expression of TNF-α possesses of earlier period in time after SCI and the expression of TGF-β strengthens with revive of spinal neural function. Both expressions are strengthening in the articular cartilage at the third week.