大鼠脊髓缺血再灌注损伤后转化生长因子β1的表达

Expression of transforming growth factor beta 1 in rats with spinal ischemia reperfusion injury

目的:观察具有多种生物学功能的多肽生长因子转化生长因子β1及其mRNA在脊髓缺血再灌注损伤后的表达变化。方法:实验于2003-11/2004-04在兰州大学第二医院骨科研究所进行。健康Wistar大鼠42只,随机分成6组,正常组,伤后3,6,12,24和48h组,每组7只。正常组不做模型。其他5组大鼠建立脊髓缺血再灌注损伤模型,夹闭腹主动脉40min后放开。应用改良Tarlov评分对模型进行评估(共5分,0分为完全瘫痪;5分为正常)。观察正常大鼠脊髓及缺血再灌注损伤后各时间点大鼠脊髓转化生长因子β1及其mRNA的分布和含量变化(灰度值变化与正常组比较,其灰度值越低表示阳性细胞越多),检测方法采用免疫组织化学技术和原位杂交技术。结果:42只大鼠均进入结果分析。①正常大鼠脊髓不表达转化生长因子β1。②大鼠脊髓神经功能评分:于伤后3h显著下降,以后逐渐恢复。③大鼠脊髓转化生长因子β1蛋白表达结果:免疫组织化学染色显示,损伤后3h蛋白的表达开始增加(149.26±12.97),损伤后24h表达强度达高峰(104.95±9.40)。④大鼠脊髓转化生长因子β1mRNA表达结果:原位杂交图像分析显示,损伤24h表达阳性细胞达高峰,其灰度值明显低于正常组(79.08±10.42,140.40±10.85,P<0.01)。⑤大鼠脊髓转化生长因子β1结果:光镜观察脊髓缺血再灌注损伤后出现大量的转化生长因子β1阳性细胞;主要是小胶质细胞、星形胶质细胞、巨噬细胞和神经元。结论:大鼠脊髓缺血再灌注损伤后转化生长因子β1表达显著增加,脊髓的神经功能随时间延长逐渐恢复,说明大鼠脊髓损伤的同时,也启动了其内源性保护机制,转化生长因子β1的高表达可能与其保护作用有关。

AIM： To study the expression changes of transforming growth factor betal （TGF-β1） and its mRNA from polypeptide growth factors （PGFs） after spinal ischemia reperfusion injury. METHODS： The experiment was conducted in Institute of Orthopaedics of Second Hospital of Lanzhou University from November 2003 to April 2004. Healthy Wistar rats were divided randomly into normal group （n=6） with 7 in each group, and experimental group （n=35） including 3, 6, 12, 24 and 48 hours group. The rats were given no operation in normal group. Spinal ischemia reperfusion injury models were founded in other five groups, open abdominal and clamping rat abdominal aorta for 40 minutes. Tarlov standard was used to evaluate the models. Totally 5 points, 0 means pamplegia, and 5 was normal.The distribution and content changes of TGF-β1 and its mRNA in spinal cord segments were observed by means of immunohistochemistry and in situ hybridization. RESULTS： All 42 rats were involved in the results analysis. ① Expression of TGF-β1 was negative in the normal spinal cord. ② The evaluation of spinal nerve function： It was significantly decreased 3 hours after injury, and recovered gradually. ③ Protein expression of myeltransforming growth factor： Staining display of immunohistochemistry, protein began to increase at 3 hours after injury （149.26±12.97）. After 24-hour injury protein expressed to peak （104.95± 9.40）. ④ The protein expression results of TGF β1： The expression intensity of positive cells was at 24 hours after injury in situ hybridization image analysis. There was significant difference between experimental and control groups （79.08±10.42, 140.40±10.85, P 〈 0.01）. ⑤ The results of transforming growth factor β1： TGF β1 positive cells after spinal cord ischemia reperfusion injury were observed by light microscope, including microglia, astrocyte, histocyte and CONCLUSION： TGF β1 expression after spinal cord ischemia reperfusion injury significantly increases. Recovery of spinal nerve function with elongation of time clarifies that with the spinal injury, endogenous protective mechanism is started. Significant increase in the expression of TGF-β1 may have a certain relation with the protective effect.