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背景:对于神经生长因子的促血管生成作用的研究,目前还处于探索阶段。目的:探讨神经生长因子对再生坐骨神经中血管形成的剂量效应及其机制。设计:随机对照动物实验。单位:解放军第三二四医院神经内科,解放军第三军医大学大坪医院野战外科研究所三室。材料:健康成年Wistar大鼠32只,雌雄不拘,体质量200～250g。硅胶管(上海新亚医用橡胶厂出产);神经生长因子(美国Sigma公司)。方法:实验于2003-08/2005-02在解放军第三军医大学大坪医院野战外科研究所三室完成。实验分组:采用随机数字法将大鼠分成4组:生理盐水组、50,100,500ng神经生长因子组,每组8只。实验干预:切除大鼠后肢坐骨神经,造成10mm缺损,用单通道的硅胶管桥接大鼠坐骨神经缺损,在桥接管内给药。生理盐水组注入5μL生理盐水。50,100,500ng神经生长因子组注入20mg/L的神经生长因子2.5,5,25μL。实验评估:在第30天时,用免疫组织化学的方法检测大鼠再生坐骨神经新生血管内皮细胞中CD34、vWf、血管内皮生长因子、trkA的表达,并作形态计量分析。主要观察指标:各组大鼠神经再生普通病理及组织化学染色观察。结果:32只大鼠全部进入结果分析。①各组大鼠神经再生病理切片结果:术后30d的坐骨神经近远端完全连接,硅胶管内坐骨神经粗壮程度为:500ng神经生长因子组>50ng、100ng组>生理盐水组。各神经生长因子组再生的坐骨神经的纤维数目和排列规则程度要好于生理盐水组,术后30d可见明显的血管生成。②免疫组织化学染色观察:3种不同剂量的神经生长因子组与生理盐水组比较,4种抗原的表达皆有显著性差异(P<0.05);500ng神经生长因子组4种抗原的表达与100ng和50ng神经生长因子组比较增加(CD34:94.2±6.4,74.2±10.9,77.0±11.0;vWf:116.2±20.0,72.0±13.1,68.0±9.7;TrkA:105.4±10.6,57.8±11.5,58.8±6.5;血管内皮生长因子:89.0±3.0,48.6±7.4,35.2±2.9;P<0.05或P<0.01)。结论:神经生长因子能促进再生周围神经的血管生成,且具有剂量效应;其机制可能与神经生长因子促进trkA、血管内皮细胞生长因子的表达密切相关。
Background: The study of the pro-angiogenic effects of nerve growth factor is still in its exploratory stage. Objective: To investigate the dose-effect of nerve growth factor on angiogenesis in regenerative sciatic nerve and its mechanism. Design: Randomized controlled animal experiments. Unit: Department of Neurology, the 324th People's Liberation Army Hospital, Third Military Medical University, Daping Hospital Institute of Field Surgery, Third Hospital. MATERIALS: Thirty-two healthy adult Wistar rats, male and female, weighing 200-250g. Silicone tube (Shanghai New Asia Medical Rubber Factory); nerve growth factor (Sigma, USA). METHODS: The experiment was performed at Room 3, Institute of Field Surgery, Daping Hospital, Third Military Medical University of Chinese PLA from August 2003 to February 2005. Experimental grouping: The rats were divided into 4 groups by random number method: saline group, 50,100,500ng nerve growth factor group, 8 rats in each group. Experimental intervention: Excision of rat hindlimb sciatic nerve, resulting in 10mm defect, with a single channel of silicone tube bridging sciatic nerve defects in the bridge tube administration. The saline group was injected with 5 μL of normal saline. 50, 100, 500 ng nerve growth factor group was infused with 20 mg / L of nerve growth factor 2.5, 5, 25 μL. Experimental evaluation: On the 30th day, the expression of CD34, vWf, VEGF and trkA in rat regenerative sciatic nerve neovascularization endothelial cells were detected by immunohistochemistry and the morphology was measured. MAIN OUTCOME MEASURES: A general pathological and histochemical staining of nerve regeneration in rats in each group. Results: All 32 rats entered the result analysis. ①Nerve regeneration in each group pathological results: 30 days after the sciatic nerve near the distal complete connection, silicone tube sciatic nerve thick: 500ng nerve growth factor group> 50ng, 100ng group> saline group. The number of nerve fibers and the rules of arrangement of sciatic nerve regenerated by each nerve growth factor group were better than that of normal saline group, and obvious angiogenesis was observed 30 days after operation. ② Immunohistochemical staining: Compared with the normal saline group, the expression of the four antigens in the three different doses of nerve growth factor group were significantly different (P <0.05); the expression of four antigens in 500ng nerve growth factor group and 100ng (CD34: 94.2 ± 6.4, 74.2 ± 10.9, 77.0 ± 11.0; vWf: 116.2 ± 20.0, 72.0 ± 13.1, 68.0 ± 9.7; TrkA: 105.4 ± 10.6, 57.8 ± 11.5, 58.8 ± 6.5 ; Vascular endothelial growth factor: 89.0 ± 3.0, 48.6 ± 7.4, 35.2 ± 2.9; P <0.05 or P <0.01). CONCLUSION: NGF can promote the angiogenesis of regenerating peripheral nerve with dose-effect. The mechanism may be closely related with the promotion of trkA and the expression of VEGF by nerve growth factor.