Expression Of Autophagy-Related Protein,AKT,mTOR And LC3A In The Development Of Mice Hippocampus

ObjectiveTo observe the expression of autophagy-related proteins,serine/threonine kinase AKT,mammalian target of rapamycin(mTOR)and microtubuleassociated protein 1 light chain 3A(LC3A),in the hippocampus of mice on embryonic(E)18 days(day,d),postnatal(P)1 d,7 d,14 d,21 d and 28 d.This study can provide basic experimental evidence for further study on the role of autophagy in the hippocampus development.MethodsKunming mice were fed with a female-male ratio of 1:2.The day when the vaginal suppository of female mice fell off was the E0 d,and the day when the offsprings were born was P 0 d.Hippocampal tissue were taken on E18 d,P1 d,P7 d,P14 d,P21 d and P28 d.Hematoxylin-eosin(HE)and 4',6-diamidino-2-phenylindole(DAPI)staining were used to detect the morphological changes of hippocampus.Immunohistochemistry,western blotting and image analysis were used to check the expression of AKT,mTOR and LC3 A protein in the hippocampus of mice.ResultsHE and DAPI staining showed that both molecular layer and polymorphous layer of dentate gyrus(DG)and Ammon' corn(CA)were gradually thickened from E18 d to P21 d.Size of cells in stratum granulosum and pyramidal layers increased bigger and cells became darker gradually in this period.The number of cells increased gradually at first,peaked on P7 d,then decreased gradually(P<0.01).On P28 d,the number of cells was unchanged compared with P21 d(P>0.05).Immunohistochemistry revealed that AKT,mTOR and LC3 A protein mainly expressed in the cytoplasm of cells in the pyramidal cell layer of the hippocampus CA area and the granular cell layer of the DG area.On E18 d,the AKT,mTOR and LC3 A positive cells in the CA and DG areas of the hippocampus were sparsely arranged,with light staining and low optical density value.From E18 d to P7 d,the AKT,mTOR and LC3 A positive cells were dense gradually.The staining deepened,the optical density increased gradually.The peak appeared on P7d(P<0.05).From P14 d to P21 d,the density of AKT,mTOR and LC3 A positive cells decreased,the staining lightened and the optical density decreased gradually(P<0.05).On P28 d,the density of positive cells and the staining were similar to those on P21 d and there was no significant difference optical in density(P>0.05)except LC3 A expression(P<0.05).Western blot detection revealed that AKT,mTOR and LC3 A protein expressed at all stages.The expression of three proteins gradually increased from E18 d to P7 d and reached the peak on P7 d(P<0.01).From P14 d to P21 d,the expression gradually decreased(P<0.01).On P28 d,both mTOR and LC3 A expression kept cutting down(P<0.01)while the AKT expression was similar to that on P21d(P>0.05).ConclusionsAutophagy-related proteins AKT,mTOR and LC3 A are highly expressed during the development of mouse hippocampus.Autophagy are closely related to mouse hippocampus development.