| Neonatal hypoxic-ischemic encephalopathy is one of the common cerebral damage,which can cause a series of neurological diseases,such as CP?MR?epilepsy,mental retardation,etc.,even leading to neonatal death.During perinatal,it is caused by various factors,which include hypoxia or decreasing cerebral blood flow.At present,it is not clear about HIBD's pathogenesis,no complete and unified clinical treatment.Therefore,further study of damage mechanism and reliable and effective prevention and control measures have also been more and more people's attention.It has become one of the hot topics of encephalopathy in children medical research.Traditional Chinese medicine has been a breakthrough in the treatment of children with encephalopathy,especially acupuncture had achieved more significant clinical effect.In recent years,autophagy is the hot direction of the study of neurological diseases,which has a close relationship with cerebral ischemia and brain injury.In the continuous research of autophagy mechanism,we found that autophagy and neuronal damage' s repairment have a certain correlation.Does acupuncture have improved the behavior of the HIBD model rats?Whether can through the mitochondrial autophagy mechanism to protect the damaged neurons?Its research is still progressing.ObjectiveThis paper is divided into two purposes.One is to study the effect of acupuncture,which is the expression of LC3 and p62 in HIBD model rats,and to explore the mechanism of acupuncture on HIBD,and whether it can affect the damaged neurons through the mitochondrial autophagic pathway.The expression of LC3 was observed at different time points.The autophagy level was confirmed by ultrastructure,pathological examination and molecular level.The effects of acupuncture intervention on mitochondrial autophagy can provide a more scientific basis theory.MethodsThe experiment included two parts.Experiment 1:HIBD model was established by delaying cesarean section.There are in the normal group,the normal and acupuncture group,the model group,the model and acupuncture group.On the 14th day after the birth of the newborn rats,rats were treated with acupuncture intervention.The normal group and the model group were not acupuncture,at the same time with acupuncture group.After acupuncture intervention,the open field test and Morris water maze test observe the behavioral ability of newborn rats.Beheaded in the brain,western blot and immunohistochemistry technique detect LC3 and p62.Experiment 2:The same with experiment 1 in modeling methods and intervention methods.It included normal group,model group,acupuncture group,and setting up four time points,namely D1,D14,D21,D28.Transmission electron microscopy observed autophagy body on the corresponding time.Western blotting detect protein LC3,and TTC stain detect necrosis and brain damage.ResultsExperiment 11.The behavior test results:Compared with the normal group,it is significant increased in model group from the central region,and the number of crossing the platform was significantly decreased.Between the normal group and the normal and acupuncture group had no significant difference(P>0.05).Compared with the model group,the model and acupuncture group shortened the central region,but there was no difference(P>0.05).The number of crossing the platform was significantly increased(P<0.05).2.Western Blot test results:The LC3II in hippocampus of normal and normal and acupunvture group are lower.The LC3 of normal group and normal and acupuncture group are higher,and p62 decreased.Compared with the model group,the model and acupuncture group was no significant difference(P>0.05),the model one had significant difference(P<0.05).Compared with model group,LC3 II is significantly increased,and p62 is decreased obviously(P<0.05).3.Immunohistochemical test results:The positive expression of LC3 ? is mostly in the cytoplasm,brown or tan.LC3 ? in the normalgroup and and normal and acupuncture group was lower,and a litter positive cells was observed.Compared with the model group,there was no significant difference in normal and acupuncture group(P>0.05),model group was significantly higher(P<0.05).Compared with model group,the model and acupunture group was significantly increased(P<0.05).Experiment 21.Weight results:Groups had no difference in D1(P>0.05).The body weight has increased in D14 and D28,and the normal group was the fastest,the model group was slower.But acupuncture group is higher than the model one.After comparison,which had significant differences(P<0.05).2.Electron microscopy results:Normal group saw complete neuronal cells in D1,the model group can be seen scattered cells,irregular shape,cell interstitial voids,some nuclear membrane damage.Normal group can be seen a single multilayer structure sample of autophagy body in D14,the model group can see more markedly swollen mitochondria and scattered distribution of autophagy body.There were no autophagosomes in normal group in D21,and a small number of autophagosomes in the model group.The mitochondria and lysosomes were encapsulated.The acupuncture group was significantly higher than other group.The neuron cells were seen in the normal group in D28.The model group did not possess the characteristics of the neuron cells.The acupuncture group showed obvious swelling of the mitochondria but no autophagy.3.TTC test results:Normal brain tissue showed normal red in D1,the model group showed a large area of pale-white infarct area.There was significantly different in the percentage of infarct area(P<0.05).The area of brain tissue of normal group showed uniform red in D28,and the area of white tissue was seen in the model group.In the infarct size,both are significant difference(P<0.05).Compared with the model one,acupuncture group was significantly decreased(P<0.05).4.Western Blot test results:Compared with normal group,model group was higher in D1 and D14(P<0.05).LC3 ? in model group and acupuncture group were higher than normal group in D21,D28(P<0.05),acupuncture group is higher than the model one(P<0.05).At all time points,normal group had no significant difference.Model group was higher in D1,which was decreased significantly in other days.But no different(P>0.05).Acupuncture group expressed lower in D14 and increased significantly in D21 and D28.But no difference(P>0.05).Conclusion1.HIBD model was successfully prepared by delayed cesarean section.2.Acupuncture can improve the behavior of HIBD rats.3.Acupuncture can promote the weight of HIBD.4.Intrauterine distress caused by HIBD can induce autophagy related protein expression in hippocampus.After acupuncture treatment,LC3 ? was significantly increased,p62 decreased,which suggest acupuncture could activate autophagy,and improve autophagy level.5.At the same time,autophagy has different levels in different groups,acupuncture intervention can promote HIBD improvement,which caused by neuronal damage.At different points,the autophagy levels in the same group are different,and length of acupuncture intervention had no significant effect on the expression of autophagy.But needs futher study. |
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