基于mTOR自噬信号通路探讨竹节参对脊髓损伤的保护作用

Study on the protective effect of Panax japonicus on spinal cord injury based on mTOR autophagy signal pathway

目的探讨竹节参对脊髓损伤(SCI)大鼠神经元的保护作用是否通过抑制mTOR信号通路激活细胞自噬而实现的。方法 64只SPF级SD雄性大鼠,随机分为假手术组、SCI组、雷帕霉素组、竹节参组。除假手术组外,其余3组均采用Allen’s打击方法制备SCI模型。假手术组及SCI组给予生理盐水灌胃,竹节参组给予竹节参水煎液灌胃,在造模前各组分别给予相应处理持续3天,雷帕霉素组在造模后0.5h内给予腹腔注射给药;造模3h、6h后采用改良Tarlov评分、HE染色、免疫组化、Western blot方法,观察竹节参对运动功能、脊髓前角区神经元的形态结构及mTOR自噬信号通路的影响。结果与模型组比,同一时间段内竹节参组运动功能得到改善(P<0.05),神经元细胞形态破坏减轻,脊髓神经元Beclin1、LC3表达均升高(P<0.05),mTOR表达均降低(P<0.05),且竹节参组与雷帕霉素组Beclin1、LC3的差异均存在统计学意义(P<0.05)。结论竹节参能够改善SCI模型大鼠下肢运动功能,其保护作用可能是通过抑制mTOR信号通路激活细胞自噬而实现的。

Objective In order to investigate investigate the protective effect of Panax japonicus on neurons in SCI rats, whether its protective effect was achieved by inhibiting mTOR signaling pathway and activating autophagy.Methods 64 male Sprague-Dawley rats were randomly divided into sham operation group, model group, rapamycin and Panax japonicus treatment group.Except for the sham operation group, the other three groups were treated with Allen’s method to copy the SCI model.Then, the model group and the sham operation group were given saline daily, Panax japonicus treatment group were given Panax japonicus daily.Before operation, each group was given corresponding treatment for 3 days.Rapamycin group was given intraperitoneal injection within 0.5 h after operation.The treatment effect was evaluated by modified Tarlov scores 3 and 6 hours after operation.The rats were killed and the spinal cord was taken, which the morphological changes of spinal cord neurons were observed by HE staining.The expression of Beclin1, LC3 and mTOR positive cells in spinal cord was detected by immunohistochemistry.The expression of Beclin and mTOR in spinal cord was detected western blot.Results In 3 and 6 hours periods, compared with the model group, the improved Tarlov of Panax japonicus group and rapamycin group increased in the same period(P<0.05);The morphology of neurons was complete, nucleolus and nuclear membrane were fuzzy, and the number of neurons was increased;The expression of Beclin1, LC3 positive cells in spinal cord decreased, while the expression of mTOR positive cells increased(P<0.05);The expression of Beclin1 protein was up-regulated in spinal cord tissues, while the expression of mTOR protein was down-regulated(P<0.05).Conclusion Panax japonicus can improve the motor function of lower limbs in SCI model rats, and its protective effect may be achieved by inhibiting mTOR signaling pathway and activating autophagy.