| Background: Epilepsy is a chronic brain disease in which the neurons in the brain are highly synchronized and discharge abnormally for a variety of reasons.It is characterized by high morbidity,disability rate and mortality.Repeated seizures can aggravate the damage of brain neurons,and autophagy has been reported to be involved in the pathophysiological mechanism of brain injury after epilepsy.Moderate autophagy plays an indispensable role in maintaining homeostasis of the body,but over-activated autophagy will lead to the death of cells.Studies have shown that autophagy may be the cause of death of hippocampal neurons after epilepsy and is closely related to brain injury after epilepsy.Antiepileptic drugs are mainly used in clinical treatment,but there is a lack of effective brain protectant at present.Therefore,it is a new direction that treat autophagy as a target to find effective therapeutic drugs which cannot be ignored in future research.Gastrodin,the most important bioactive ingredient in gastrodia elata,is used in the treatment of headache,dizziness and other diseases.Studies have shown that gastrodin alleviates hippocampal neuronal injury by inhibiting autophagy in bilateral common carotid artery ligation induced models.This suggests that gastrodin may reduce brain injury after epilepsy by inhibiting autophagy,but the specific role and mechanism still need to be further studied.The purpose of this study was to observe the expression characteristics of autophagy in status epilepticus and the effect of gastrodin on brain injury after status epilepticus,so as to provide new ideas and experimental basis for exploring the pathogenesis of epileptic brain injury and clinical treatment of epilepsy.Methods:(1)In this experiment,fifty-six male C57 BL / 6 mice were randomly divided intocontrol group and SE group with 4 mice in each group respectively by establishing the model of pilocarpine induced SE model.Hippocamps were collected at 6h,12 h,24h,48 h,72h and 96 h after SE.The mRNA and protein levels of Beclin-1 and LC3 in hippocampus were detected by qRT-PCR and Western blot.(2)Forty male C57 BL / 6 mice were selected and the models were made according to the above methods.The mice were randomly divided into the control group,SE group,GAS50 mg/kg group,GAS100 mg/k group,GAS150 mg/kg group with 4 mice in each group.0.2g gastrodin injection was dissolved in 83 ml normal saline and the mice were given intraperitoneal injection according to the dose of 50mg/kg,100mg/kg,150mg/kg respectively with 2 times/day after SE.Hippocampus tissues were taken at the time when autophagy activity was highest after SE.The mRNA and protein levels of Beclin-1 and LC3 in the hippocampus were detected by qRT-PCR and Western blot.(3)Thirty-two C57 BL / 6 mice were selected and the models were made according to the above methods.The mice were randomly divided into control group,SE group,gastrodin group and WM group with 4 mice in each group.WM was dissolved in dimethylsulfoxide(DMSO)and intraperitoneal injection was performed at a dose of0.5mg/kg 30 min before the administration of atropine sulphate.Hippocampus tissues were taken at the time when autophagy activity was highest after SE.The mRNA and protein levels of Beclin-1 and LC3 in the hippocampus were detected by qRT-PCR and Western blot.Results:(1)Compared with the control group,the mRNA expression of beclin-1 reached the highest level at 72 h after SE,and began to decline at 96 h after SE(P < 0.05);Compared with the control group,the protein expression of beclin-1 reached the highest at 72 h after SE(P < 0.05).(2)Compared with the control group,the mRNA expression of LC3 reached the highest level at 72 h after SE,and began to decrease at 96 h after SE(P < 0.05);Compared with the control group,the ratio of LC3II/I reached the highest at 72 h after SE(P < 0.05).(3)Compared with the control group,the mRNA expression of beclin-1 was significantly increased in the SE group;Compared with the SE group,The mRNA expression of beclin-1 began to decrease in GAS100mg/kg group and decreased to the lowest in GAS150mg/kg group(P<0.05);Compared with the control group,the protein expression of beclin-1 was significantly increased in the SE group;Compared with the SE group,the protein expression of beclin-1 decreased to the lowest in GAS150mg/kg group(P<0.05).(4)Compared with the control group,the mRNA expression of LC3 in the SE group was significantly increased;Compared with the SE group,the mRNA expression of LC3 began to decrease in GAS100mg/kg group and decreased to the lowest in GAS150mg/kg group(P<0.05);Compared with the control group,the ratio of LC3II/I increased significantly in the SE group;Compared with the SE group,the ratio of LC3II/I began to decrease in the GAS100mg/kg group and decreased to the lowest in the GAS150mg/kg group(P<0.05).(5)Compared with the control group,the mRNA expression of beclin-1 was increased significantly in the SE group;Compared with the SE group,the mRNA expression of beclin-1 was decreased significantly in GAS150mg/kg group;Compared with the GAS150mg/kg group,the mRNA expression of beclin-1 was increased in the WM group and was lower than that in the SE group(P<0.05);Compared with the control group,the protein expression of beclin-1 was increased significantly in the SE group;Compared with the SE group,the protein expression of beclin-1 was decreased significantly in GAS150mg/kg group;Compared with GAS150mg/kg group,the protein expression of beclin-1 was increased significantly in the WM group,but was still lower than that in SE group(P<0.05).(6)Compared with the control group,the mRNA expression of LC3 was increased significantly in the SE group;Compared with the SE group,the mRNA expression of LC3 was reduced significantly in the GAS150mg/kg group;Compared with the GAS150mg/kg group,the mRNA expression of LC3 increased in the WM group and was lower than that in the SE group(P<0.05);Compared with the control group,the ratio of LC3II/I wasincreased significantly in the SE group;Compared with the SE group,the ratio of LC3II/I was reduced significantly in GAS150mg/kg group;Compared with the GAS150mg/kg group,the ratio of LC3II/I increased significantly in the WM group,but was still lower than that in SE group(P<0.05).Conclusion:(1)The express of Beclin-1 and LC3 increased in SE mice,and the degree of increase was time-dependent,suggesting that autophagy may be involved in post-SE brain injury.(2)Gastrodin can reduce the expression of beclin-1 and LC3,and the degree of reduction is related to the dose of gastrodin,suggesting that gastrodin may have a protective effect on brain injury after SE.(3)WM can reduce the effect of gastrodin on autophagy active factors beclin-1 and LC3,suggesting that gastrodin may play a role in brain injury after SE by regulating autophagy. |
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