17β雌二醇对大鼠坐骨神经损伤的保护

Protective effect of 17 beta-estradiol against sciatic nerve injury in rats

目的:探讨预先应用17β雌二醇对大鼠坐骨神经损伤的治疗作用,为周围神经损伤的临床治疗提供实验依据。方法:实验于2004-10/2005-08在锦州医学院实验室完成。选择成年雄性SD大鼠54只,随机数字表法分为3组,假手术组18只,仅暴露不损伤神经;手术对照组18只,制作坐骨神经钳夹伤模型;17β雌二醇处理组18只,损伤前1周给予17β雌二醇(1mg/kg)腹腔注射,1次/d。余二组在手术前1周仅腹腔注射同体积的蓖麻油,1次/d。①于术后2,4,7,14,21,28d进行坐骨神经功能指数评定,坐骨神经功能指数=-38.3(实验侧足印长度-正常侧足印长度)/正常侧足印长度+109.5(实验侧足印宽度-正常侧足印宽度)/正常侧足印宽度+13.3(实验侧足趾宽度-正常侧足趾宽度)/正常侧足趾宽度-8.8。坐骨神经功能指数以0为正常值,-100为神经完全离断指标。②术后7,14,28d各组取损伤近段坐骨神经甲苯胺蓝染色光镜下观察,醋酸铀和枸橼酸铅染色电镜下观察神经纤维超微结构。计算横断面单位面积的平均神经纤维个数和直径、轴突直径及髓鞘厚度。③术后7,14,21,28d各组大鼠取损伤近段坐骨神经进行S-100蛋白检测。结果:纳入动物54只,均进入结果分析。①假手术组术后2,28d坐骨神经功能指数值分别为-6.42和-5.71,差异无显著性意义。手术对照组和17β雌二醇处理组术后4,7,14,21d坐骨神经功能开始恢复,17β雌二醇处理组在术后21,28d坐骨神经功能指数值分别为-16.73和-16.14,而手术对照组在术后28d坐骨神经功能指数值为-16.32,可见17β雌二醇处理组大鼠功能恢复比手术对照组早1周左右。②光镜下观察术后14d17β雌二醇处理组损伤近段坐骨神经新生纤维较多,轴突直径较大且形状规则,纤维密度大、髓鞘较厚,明显高于手术对照组。术后28d手术对照组与17β雌二醇处理组变化基本一致,接近假手术组。③术后14d17β雌二醇处理组损伤近段坐骨神经电镜观察见大量新生神经纤维密集,许旺细胞包绕,轴突大小接近、形态均一,鞘内细胞器较多且清晰,明显优于手术对照组。术后28d手术对照组、17β雌二醇处理组新生神经纤维均基本成熟,差异不明显,已接近假手术组。④在周围神经损伤中S-100仅表达于许旺细胞中。术后14d17β雌二醇处理组阳性显色深且范围宽,与手术对照组比较差异有显著性意义。术后21,28d手术对照组与17β雌二醇处理组又无明显差异。结论:17β雌二醇能加速大鼠坐骨神经损伤后功能的恢复以及神经纤维的再生修复。

AIM： To explore therapeutical effect of 17β estradiol on sciatic nerve injury of rats and provide experimental basis for clinical treatment of peripheral nerve injury. METHODS： The experiment was conducted at Laboratory of Jinzhou Medical University from October. 2004 to August 2005. Totally 54 adult male SD rats were divided randomly into 3 groups： sham operation group with 18 rats （only exposed but no injured nerve）, operation control group with 18 rats （establishing sciatic nerve ring clamp models）, 17β estradiol treatment group with 18 rats （one week before injury 1 mg/kg 17β estradiol was given by intraperitoneal injection, once a day）. The rats in the other two groups were treated with ficin oil of the same volume one week before operation by intrapefitoneal injection, once a day. ①Functional index of sciatic nerve was assessed at days 2, 4, 7, 14, 21 and 28 after operation. Functional index of sciatic nerve was equal to -38.3 （length of pedogram of experimental side-length of pedogram of normal side）/length of pedogram of normal side＋109.5 （width of pedogram of experimental side-width of pedogram of normal side）/width of pedogram of normal side＋13.3 （width of toes of experimental side-width of toes of normal side）/width of toes of normal side-8.8. In functional index of sciatic nerve, 0 indicated normal value, and -100 indicated complete nerve mutilation index. ②Proximal injured sciatic nerve was determined with toluidine blue staining and observed under light microscope at days 7, 14 and 28 after operation. Ultrastructure of nerve fiber was determined with uranyl acetate arid lead citrate staining and observed under electron microscope. Number and diameter of mean nerve fibre in unit area of transverse plane, diameter of axon and depth of myelin sheath were calculated. ③Proximal injured sciatic nerve of rats were determined with S-100 protein examination at days 7, 14, 21 and 28 after operation. RESULTS： A total of included 54 animals were involved in the result analysis. ①Functional indexes of sciatic nerve in the sham operation group were -6.42 and -5.71 at days 2 and 28 after operation, respectively, and the difference was insignificant. The function of sciatic nerve began to recover in the operation control group and 17β estradiol treatment group at days 4, 7, 14 and 21 after operation. Functional indexes of sciatic nerve in the 17β estradiol treatment group were -16.73 and -16.14 at days 21 and 28 after operation, while it was -16.32 at day 28 after operation in the operation control group. It was showed that functional recovery of rats in the 17β estradiol treatment group was earlier about 1 week as compared with the operation control group. ② Under light microscope there were many new fibers at proximal injured sciatic nerve in the 17β estradiol treatment group at day 14 after operation; diameter of axon was long with regular shape, and the density of fiber was thick with thick myelin sheath, which were siguificantly higher than those in the operation control group. At day 28 after operation the change was mostly coincident between the operation control group and 17βestradiol treatment group, which was close to .that of the sham operation group. ③Under electron microscope at proximal injured sciatic nerve in the 17β estradiol treatment group at day 14 after operation we found that there were a mass of new nerve fiber, Schwann cell encasement, similar size axon, similar shape, and the cellular organ in sheath was more and clear, which were markedly better than those in the operation control group. At day 28 after operation the new nerve fiber was mostly mature in the operation control group and 17β estradiol treatment group, and the difference was insignificant, which was near to that in the sham operation group. ④S-100 only appeared in Schwann cell in the peripheral nerve injury. At day 14 after operation the positive colouration was dark and wide in the 17β estradiol treatment group, which had significant difference with the operation control group. There was no significant difference between the operation control group and the 17β estradiol treatment group at days 21 and 28 after operation. CONCLUSION： 17β estradiol can accelerate the functional recovery and regenerative reparation of nerve fiber after sciatic nerve injury in rats.