经腹腔暴露2,5-己二酮致大鼠脊髓组织细胞凋亡的研究

Effect of exposure to 2,5-hexanedione on apoptosis of spinal cord cells in rats

目的观察经腹腔暴露2,5-己二酮(2,5-HD)大鼠脊髓组织细胞凋亡及相关调控蛋白的表达,探讨其致毒机制。方法将40只SD大鼠按体重随机分为4组,每组10只。将2,5-HD溶于0.9%生理盐水,经腹腔注射染毒。低剂量组,中剂量组和高剂量组染毒剂量分别为100 mg/kg,200 mg/kg和400 mg/kg,注射量为0.3 mL/100 g,对照组以相同剂量的生理盐水腹腔注射,每天1次,每周5 d,共染毒5周。染毒过程中,每周对大鼠进行神经行为学评价。通过TUNEL-Hoechst双染技术观察脊髓组织细胞凋亡;Western Blot检测脊髓组织凋亡相关蛋白Bcl-2和Bax表达。结果神经行为学观察显示,在染毒第5周,100 mg/kg,200 mg/kg和400 mg/kg 2,5-HD染毒组大鼠的步态评分值分别为1.80,2.80和3.94,明显高于对照组(P<0.01)。TUNEL-Hoechst双染结果显示,在染毒组大鼠脊髓切片可见绿色阳性细胞,其凋亡指数(AI)高于对照组(P<0.01),且随着染毒剂量增高而增加。Western Blot检测结果显示,2,5-HD染毒大鼠脊髓组织Bax蛋白表达明显高于对照组(P<0.01),而Bcl-2蛋白表达明显低于对照组(P<0.01)。结论经腹腔暴露2,5-HD可诱导大鼠脊髓组织细胞凋亡,其机制与上调Bax蛋白表达和下调Bcl-2蛋白表达有关。

Objective To observe the effect of exposure to 2,5-hexanedione （2,5-HD） by intraperitoneal on apoptosis and on the relative protein expression in rat spinal cord to explore the mechanism of 2,5 -HD-induced neurotoxicity . Methods Forty adult male SD rats were divided randomly into one control and three experimental groups ,each 10 animals. 2,5-HD was administered by intraperitoneal injection to experimental groups with a daily dose of 100, 200, 400 mg/kg for continuous 5 weeks （5 times per week）.The control group was treated by saline.Neurobehavioral index were deter-mined per week.On the fifth week after 2,5-HD exposure, the rats were sacrificed to extract spinal cord tissue .Apoptosis of spinal cord cells was examined by TUNEL -Hoechst .The expressions of Bax and Bcl -2 protein were examined by west-ern blotting.Results Neurobehavioral observation showed that at the fifth week , the gait score of three experimental groups were 1.80, 2.80 and 3.94 respectively, which were higher than that of the control group significantly （P〈0.01）.The apoptosis indices of the spinal cord cells in the rats exposed to 2,5-HD were significantly higher than that in control group （P〈0.01）.Western blotting results showed that expression of Bax protein increased significantly in the spinal cord of rats in 2,5-HD groups （P〈0.01）, and expression of Bcl -2 protein decreased significantly in the spinal cord of rats in 2,5-HD groups （P〈0.01）.Conclusion Exposure to 2,5-HD by intraperitoneal could induce cells to apoptosis in rat spinal cord by up-regulating expression of Bax and down-regulating expression of Bcl -2 protein .