Neuronal Ultrastructure And Expression Changes Of Microtubule Associated Protein 2?Tau Protein In Hippocampal CA3 Area Of ApoE Knockout Mice

Objective:(1)To observe the neuron ultrastructural change of hippocampal CA3 Area in apolipoprotein-E knockout(hereinafter referred to as Apo E KO)mice;(2)To observe microtubule-associated protein 2(MAP-2)and Tau protein expression in hippocampal neuron of Apo E KO mice;(3)To discuss the correlation between neuron structure changes of Apo E KO mice and neurodegenerative dysfunction.Methods:10 normally-fed C57BL/6J wild mice were chosen as the control group(C group),and 10 Apo E gene knockout mice were selected as Apo E KO group,fed with a common diet for 3 months.After chloral hydrate anesthesia,inferior vena cava blood sampling was taken and blood lipid changes were detected by spectrophotometer testing.Brain tissues of the two groups of mice were taken out to observe neuronal structural changes in hippocampal CA3 area by HE staining.Neurons austenite content changes in hippocampal CA3 area were detected by means of Nissl's staining.Electron-microscopy technology was applied to observe structural changes of neurons,nerve fibers and synapses.MAP-2 protein and Tau protein expression changes of neurons in hippocampal CA3 area were observed by immunohistochemical technique.Average optical density(OD)values of the above-mentioned two types of proteins were analyzed by computerized image system.Results:(1)Blood lipid change: for serum lipid parameters of C Group mice,plasma cholesterol was(4.08±0.31)mmol/L,plasma triglyceride was(1.86±0.39)mmol/L,low density lipoprotein was(1.27±0.38)mmol/L,whereas for serum lipid parameters of Apo E-KO group mice,plasma cholesterol was(10.56±2.43)mmol/L,plasma triglyceride was(2.64±0.63)mmol/L,low density lipoprotein was(2.51±0.57)mmol/L.Through comparison of serum lipid parameters of the two groups,it could be found that the parameters of Apo E-KO group mice increased dramatically.There were significant differences between the two groups(P <0.05,P<0.01).(2)HE staining results: Neurons of pyramidal layer in hippocampal CA3 area of C group were tightly packed.The cell body was cone-shaped.Neuron cell nucleus was large,round,and lightly stained.Pyramidal cells in hippocampal CA3 area of Apo E-KO group were slightly and disorderly arranged.(3)Nissl's staining results: compared with C group,content of austenite in hippocampal neurons cytoplasm of Apo E-KO group diminished significantly,and the staining was lighter.(4)Electron microscope results: in comparison with C group,hippocampal neurons mitochondria of Apo E-KO group swelled and degenerated and ridge disappeared,myelin sheath of nerve fibers relaxed,the number of synaptic vesicles decreased and synaptic cleft obscured.(5)Immunohistochemical results: MAP-2 immune positive matter were chiefly found in hippocampal neurons cytoplasm and dendrite of the mice,and after immumohistochemical staining,they were yellowish-brown or yellow particles.In C group mice,hippocampal neurons cytoplasm and dendrite were yellow---yellowish particulate matter;and pale-yellow matter was found in Apoe-KO group mice with few stained particles.Phosphorylated Tau protein immune positive matter mainly deposited in hippocampal neurons cytoplasm of the mice,which were stainless or pale-yellow in C group mice,yellow-yellowish-brown in Apo E-KO group mice with increased number of stained particles.(6)Computerized image analysis result: Compared with the average optical density value of austenite content in hippocampal CA3 area neurons of C group mice(0.864±0.027),the average optical density value of austenite content in pyramidal cell of hippocampal CA3 area of Apo E-KO group mice decreased(0.213± 0.076).There was significant difference(P < 0.01);Hippocampal CA3 neurons in C group mice MAP-2 protein content was(0.489±0.065)and phosphorylated Tau protein content was(0.339±0.049).In hippocampal neurons of mice in Apo E-KO group,MAP-2 protein content(0.326±0.081)decreased,and phosphorylated Tau protein content(0.472±0.037)rose,and statistical differences were found in terms of average OD value(P?0.05).Conclusions:(1)Blood lipid level of Apo E gene knockout mice remarkably rose after being normally-fed for 3 months.(2)Hippocampal CA3 pyramidal cells ultrastructure of Apo E gene knockout mice changed,mitochondrion swelled and became deformed,microfilament and microtubule in cytoplasm were disordered.Nerve fiber structure of myelin sheath was incomplete,loosened with decreased density.The number of synaptic vesicles decreased and synaptic cleft obscured.(3)Hippocampal neurons MAP-2 expression of Apo E gene knockout mice reduced in comparison with that of normally-fed mice.And phosphorylated Tau expression was on the rise.Abnormal expression of Microtubule associated protein may be closely correlated with neurodegenerative disease.This study may provide morphological experimental basis for further exploration of the pathogenesis of neurodegenerative diseases.