周围神经放射线照射后原位再植再生功能与组织学的恢复

Regenerative function and histological restoration of peripheral nerve segments replanted in situ after radiation

目的:观察大鼠体外坐骨神经节段经放射线照射后原位再植再生功能与组织学影响。方法:实验于2004-05/09在哈尔滨医科大学附属肿瘤医院动物实验中心完成,选择清洁级5月龄雌性大白鼠26只,其中2只编号为1、2号。余24只随机分为3组,对照组,单纯手术组和4500cGy照射组,每组8只。制备模型,单纯手术组切取右腿坐骨神经中段10mm后原位再植;4500cGy照射组坐骨神经节段离体以4500cGy高能射线一次照射后原位再植。1号鼠神经节段同单纯手术组处理,2号鼠同自体神经移植4500cGy照射组处理,完毕后即行标本固定,光镜、电镜观察。对照组鼠不给以干预处理,各组大鼠术后饲养12周进行一般情况观察及大鼠坐骨神经功能测定、胫前肌定量分析与组织学评价。结果:纳入实验大鼠26只,无动物死亡,均进入结果观测与分析。①术后12周单纯手术组和4500cGy照射组的坐骨神经功能指数显著低于对照组正常对照值[(-33.41±2.99),(-11.23±2.38),q=22.62,P<0.05];[(-34.63±2.91),(-11.23±2.38),q=23.87,P<0.05]。神经纤维数量多于对照组正常值[(192.50±21.00),(203.00±16.94),(174.13±10.18)×102根/mm2]。②组织学变化的光镜观察结果:1号鼠神经结构完整,神经纤维排列规则,髓鞘无肿胀;2号鼠4500cGy高能X射线照射后,神经纤维轻度空泡样变,轴索见境界尚清,髓鞘轻度肿胀,无髓鞘脱失。术后12周末对照组为正常神经组织;单纯手术组神经束毛细血管增生不明显,髓鞘无明显肿胀。4500cGy照射组神经纤维粗细基本一致,排列尚规则,少量炎性细胞浸润和纤维组织增生。③坐骨神经的组织学变化的电镜观察结果:1号鼠髓鞘多呈椭圆形,明暗相间板层结构清晰可见,轴索结构完整,神经膜细胞内见丰富的线粒体、粗面内质网等结构。2号鼠可见髓鞘板层结构轻度空泡样变,细胞核基本完好。术后12周末对照组神经呈椭圆形,髓鞘壁薄厚一致,板层结构致密,轴浆内富含神经微丝和微管,许旺细胞核少见;单纯手术组和4500cGy照射组均可见神经轴突发育良好,呈近似椭圆形,髓鞘壁薄,许旺细胞核多见,轴索内见线粒体。结论:经4500cGy高能射线一次照射大白鼠坐骨神经体外节段原位再植,神经干再生结构与功能恢复完全,可最大限度保存神经干再生能力。实验条件下,体外放射灭活处理法作为神经保存的方法是可行的。

AIM： To observe the function and histology changes of rats after the in vitro sciatic nerve segment treated with radiation. METHODS＂ The experiment was done in the Animal Central Laboratory, the Tumor Hospital affiliated to Harbin Medical University from May to September 2004. Twenty-six female rats of clean grade aged 5 months were selected, two of the rats were marked with No. 1 and 2, and others were divided randomly into control group （n=8）, simple operation group （n=8） and 4 500 cGy radiation group （n=8）. Animal models were made. The sciatic nerves in middle segment on the right side in the simple operation group were cut for 10 mm and replanted in situ directly; The segments were replanted in situ after radiated in dose of 4 500 cGy in the 4 500 cGy radiation group. The No.1 rat was treated as those in the simple operation group and the No.2 rat was treated as those in the 4 500 cGy radiation group, and then the specimen fixation, light microscope and electron microscope were conducted immediately. Rats in the control group were not treated. All the rats in the three groups raised for 12 weeks were observed for sciatic nerve function, quantitative analysis of tibial muscle and evaluation histologically. RESULTS： All the 26 rats were involved in the analysis of result. ① The sciatic function index at 12 weeks after operation was siguificanfly lower in the simple operation group and 4500cGy radiation group than in the control group [（-33.41±2.99）, （-11.23±2.38）, q=22.62, P〈0.05; （-34.63±2.91）, （-11,23±2.38）, q=23.87, P〈0.05]. The number of myelinated fibers was in the simple operation group and 4500cGy radiation group than the normal value in the control group [（192.50±21.00）, （203.00±16.94）, （174.13±10.18）×10^2/mm^2].② Histological observation under light microscope： The nerve structure of No.1 rat was complete, neurofibra arranged regularly, and there was no swelling in myelin sheath; Neurofibra in No.2 rat radiation exposure in dose of 4500 cGy was little vacuole degeneration and axis cylinder milieu was clear. Myelin sheath was little swell and not amyelination. Sciatic nerve in the control group was normal nerve tissue at 12 weeks; Blood capillary proliferation in nerve tract and myelin sheath swell were all not obvious. Neurofibra caliber was uniformit and arranged regularly with small inflammatory cell infiltration and fibroplasias in the 4500 cGy radiation group. ③ Histological changes of sciatic nerve under electron microscope： For the No.1 rat, myelin sheath was oval, layers could be observed clearly, and uxihary fibers were integrated. Many chondriosomes and rough endcplasmic reticulum were observed in the nerve corpuscles; For the No.2 rat, the layers of myelin sheath were vacuole degeneration and cell nucleus was generally good. At the end of the 12^th week, neurofibra in the control group was oval and myelin sheath thickness was uniformity. Pyknolayers, much neurofilament and neurotubule in axoplasm were observed, but Schwann cell nucleus was less. In the simple operation group and 4 500 cGy radiation group, cylindraxile developed well, and was similar to oval. Myelin sheath was thin, more Sehwann cell nuclei were observed, and chondriosome was seen in cylindraxile. CONCLUSION： 4500 cGy has been proved to be the ideal dose to preserve the ability of peripheral nerve regeneration while killing the tumor cells. Nerve inactivation treatment by radiation can be used under the experimental condition.