| Objectives (1)To observe the microstructural and ultrastructural changes of the neurons of hippocampus of diabetic rats. (2)To observe the expressive changes of iNOS, NT-3, Bcl-2 and Bax of the neurons of hippocampus of diabetic rats. (3)To observe the effect of insulin therapy on the expressive changes of iNOS, NT-3, Bcl-2 and Bax of the neurons of hippocampus of diabetic rats. (4)To study the effect of hyperglycaemia on structural and functional changes of the neurons of hippocampus and its mechanism; To investigate the potential effect of iNOS, NT-3, Bcl-2 and Bax on the formation of diabetes and diabetic encephalopathy; To study the effect of insulin therapy on structural and functional changes of neurons of hippocampus of diabetic rats and its mechanism. Methods 30 Sprague-Dawley rats were randomly divided into one-month and three-month control groups, one-month and three-month diabetes mellitus groups and one-month and three-month insulin therapy groups. There are 5 rats in each group. Rats of diabetes mellitus and insulin therapy groups were injected intraperitonealy with streptozocin(STZ) to make experimental diabetes mellitus models. Rats were sacrificed for removing brains after transcardiac perfusion fixation after weighing and assaying blood glucose by the end of one and three months respectively, ultrastructural changes of the neurons of hippocampus were observed by transmission electron microscrope (TEM); The histological changes were observed by hematoxylin eosin (HE) and nissal staining; The expressive change of iNOS, NT-3 ,Bcl-2 and Bax of neurons of hippocampus were studied by immunohistochemical SABC. The optic density of iNOS,NT-3, Bcl-2 and Bax immunoreactive neurons of hippocampus was analyzed by computerized image pattern analysis. All experimental results were statisticallyanalyzed with SPSS 10.0.Results ? The weight and blood glucose of diabetic were (229.83±55.44) g, (23.31±6.81) mmol/L; and those of control group were (305.50±34.67 ) g, (7.23±0.84 ) mmol/L. The difference between two groups was significant (PO.01). Compared to that of control group, weight (354.67±31.94) g and blood glucose (6.44±1.21) mmol/L of insulin therapy group have no significant difference (P>0.05) , which indicate we made the experimental model successfully. (D Under light microscope, hippocampal cortex of rats in control groups consisted of molecular layer, pyramidal layer and polymorphic layer. Most cells in pyramidal layer were regular pyramidal cells. Dentate gyrus also consisted of three layers, i.e. molecular layer, granular cell layer and polymorphic layer, Within which granular cell layer consisted of several strata of compact granular cells with small and round nuclei. Compared to control group, the microstructures of hippocampus of diabetic and insulin therapy groups changed little. (3) Compared to that of control group, quantity and chromoscopy of Nissle body of the neurons of hippocampus of diabetic and insulin groups have no significant difference. ? Under TEM ,the ultrastructure of neurons of rats' hippocampus in diabetic group presented a series of pathologic changes in contrast to control group, with rough endoplasmic reticula expanded and ruptured, ribosomal granules broke away, mitochondria swelled and its crista broke, Golgi complexes swelled and its polarity disappeared, the synapses degenerated, which contained small clear and large dense cored synaptic vesicles; presynaptic membrane and postsynaptic membrane were unclear, and the numbers of small clear synaptic vesicles slightly decreased, myelin sheaths swelled, broke up and its interspace enlarged, and so did the capillary endothelia. The ultrastructure of neurons of hippocampus of insulin therapy group have no significant change. ?There is no iNOS immunoreactive neuron of the hippocampus of the control group. But in one-month diabetic group, iNOS immunoreactive neurons were located in pyramidal layer of hippocampus and in granular cell layer of dentategyrus by immunohistochemical SABC staining. But the number and chromoscopy of iNOS immunoreactive neurons was less than that of three-month diabetic group. There were iNOS immunoreactive neurons in every section of hippocampus of three-month diabetic group. iNOS neurons were more in quantity and much denser in distribution than that of one-month diabetic group, especially in CA3 section. There are few iNOS immunoreactive neurons of the hippocampus of insulin therapy group. NT-3 immunoreactive neurons in control group were distributed densely in the pyramidal layer of hippocampus and in the granular cell layer of dentate gyrus. NT-3 neurons were highly positive in cytoplasm and nuclei negative. In one-month diabetic group, NT-3 neurons are weakly positive and less in quantity. In three-month diabetic group, NT-3 neurons are even weakly positive and less in quantity, especially in CA1 area. Compared to that of control group, the quantity and positive of NT-3 neurons in the insulin therapy groups have no significant difference. Neither Bax nor Bcl-2 expressed in the neurons of hippocampus of control groups, only Bcl-2 expressed in the neurons of hippocampus of one-month diabetic group. Bcl-2 neurons were middle positive in cytoplasm and nuclei negative. There are both Bax and Bcl-2 middle positive neurons of the hippocampus of three-month diabetic group. No Bax/Bcl-2 immunoreactive neurons can be seen in the hippocampus of insulin therapy group. (6) Compared to control group, mean optical density level of iNOS, Bcl-2 and NT-3 immunoreactive neurons of rats hippocampus increased and decreased respectively and this variety related to the course of diabetes. All the differences were significant, especially in CA3 or CA1 regions.Conclusions (1)A series of pathologic ultrastructural changes of neurons of hippocampus of rats could be induced by diabetes. (2)The iNOS, NT-3, Bcl-2 and Bax immunoreactive neurons of hippocampus of diabetic groups had significant change. (3) With the development of the disease, the expression of iNOS increased but that of NT-3 of neurons of hippocampus of diabetic rats decreased. the change of Bcl-2/Bax... |
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