局部注射转化生长因子β_1质粒与坐骨神经移植后免疫排斥反应的量效关系

Dose-effect correlation of local injection of transforming growth factor beta 1 plasmid to immune rejection following sciatic nerve transplantation

背景：研究表明，转化生长因子β1被应用于周围神经移植来抑制或减弱移植排斥反应，但在某些情况下，转化生长因子又表现出正向免疫调节作用。所以有必要进行量效关系研究获取可靠的数据佐证。目的：从剂量学上观察局部注射转化生长因子β1质粒与免疫排斥反应的量效关系。设计、时间及地点：随机对照动物实验，于2007—06／2008-04在哈尔滨医科大学动物实验中心完成。材料：选用雄性Wistar大鼠20只为供体。清洁级雄性SD大鼠50只为受体，随机分为5组，每组10只：自体神经移植组、空质粒异体神经移植组、低、中、高剂量转化生长因子β1质粒异体神经移植组。pAdTrack—CMV—TGF-β1质粒，pAdEasy—1—Bj51833细胞由华中科技大学同济医学院附属协和医院传染病实验室曾令兰教授惠赠。方法：于手术显微镜下，从犁状肌下孔0．5cm处整齐剪下长约1cm的坐骨神经，将供体神经桥接于神经缺损处，异体神经移植组转化生长因子β1质粒注射剂量为10，20，40μg／只，空质粒异体神经移植组注射空质粒。主要观察指标：术后6周进行运动神经传导速度、病理学、混合淋巴细胞培养和轴突计数检查。结果：高剂量转化生长因子β1质粒异体神经移植组运动神经传导速度、轴突计数与自体神经移植组接近（P〉0．05），并优于低剂量转化生长因子β1质粒异体神经移植组（P〈0．05）。病理学及透射电镜显示，高剂量转化生长因子β1质粒异体神经移植组移植神经段效果接近于新鲜自体神经移植组（P〉0．05），并优于低剂量转化生长因子β1质粒异体神经移植组。高剂量转化生长因子β1质粒异体神经移植组混合淋巴细胞培养、迟发性超敏反应优于低剂量转化生长因子β1质粒异体神经移植组（P〈0．05）。结论：局部注射转化生长因子β1质粒减轻大鼠同种异体坐骨神经移植后免疫排斥反应的作用，在10～40μg范围内，随剂量的增加而增强。

BACKGROUND： Transforming growth factor beta 1 （TGF-β1） has been used for peripheral nerve transplantation to inhibit or reduce rejection, but under some conditions, TGF has a positive regulatory effect. Thus, it is necessary to study dose-effect relationship in order to obtain reliable data evidences. OBJECTIVE： To investigate the dose-effect correlation of immune rejection with local injection of different doses TGF-β1 plasmid. DESIGN, TIME AND SETTING： The randomized control animal experiment was performed at the Animal Experimental Center of Harbin Medical University from June 2007 to April 2008. MATERIALS： Twenty male Wistar rats were used as donors. Fifty clean male Sprague Dawley rats as recipients were randomly assigned into 5 groups, l0 in each group, comprising nerve autograft group, nerve allograft group, low-dose, middle-dose and high-dose TGF- β1 plasmid local injection groups, pAdTrack-CMV-TGF- β1 plasmid, pAdEasy-1-Bj51833 cells were presented by Professor Zeng from Laboratory of Communicable Disease, Union Hospital Affiliated to Tongji Medical College, Huazhong University of Science and Technology. METHODS： A l-cm section of the sciatic nerve was cut off under the infrapiriformis foramen. Donor nerve bridge was connected with nerve defect region. （10, 20, 40 μg） TGF- β1 plasmid was injected into rats in the low-dose, middle-dose and high-dose TGF- β1 plasmid local injection groups. Blank plasmid was injected into rats in the nerve allograft group. MAIN OUTCOME MEASURES： Six weeks after surgery, lab-tests were examined, nerve electrophysiology, pathology, mixed lymphocyte culture and axon counting. RESULTS： Motor nerve conduction velocity and axon number in high-dose TGF- β1 plasmid local injection group was close to nerve autograft group （P 〉 0.05）, and better than low-dose TGF- β1 plasmid local injection group （P 〈 0.05）. Pathology and transmission electron microscope demonstrated that outcome of transplanted nerve segment in the high-dose TGF- β1 plasmid local injection group was close to that in the nerve autograft group （P 〉 0.05）, and better than low-dose TGF- β1plasmid local injection group. The mixed lymphocyte culture and delayed type hypersensitivity in the high-dose TGF- β1 plasmid local injection group were better than in the low-dose TGF- β1 plasmid local injection group （P 〈 0.05）. CONCLUSION： Local injection of TGF- β1 plasmid could reduce immune rejection after sciatic nerve allograft transplantation in rats. The reduction of immune rejection changed synchronously with local injection of different doses TGF- β1 plasmid （10-40 μg）.