| IntroductionHIE is one of the most important causes of newborn death and permanent dysfunction of children's nervous system. At present, the pathogenic mechanisms of HIE is still imperfect. In recent years, some studies suggested that mitochondria played a key role in the pathogenesis of HIE. The main function of mitochondria is energy metabolism, providing energy for all kinds of vital activities. Impairment of mitochondrial energy metabolism after cerebral HI bears a lot in the occurrence of HIE. The reports about time and severity of secondary energy metabolism failure were much different from each other and the studies of HIBD animals were mostly based on newborn pigs. Whereas there are fewer study reports concerned with energy metabolic changes of newborn and (or) newborn animals after cerebral HI in China.Using the HIBD models of newborn rats, the concentration of PGr,ATP,ATP + ADP + AMP in cortical neurons were measured, and the time course of the changes were observed, at the same time, the morphological changes of mitochondria in the corresponding point were observed under transmission electroscope. The purpose of the study is to explore the therapy windows of HIBD and the relationship betweenneuropathological changes and energy metabolism.Methods and materials1. Establishment of HIBD models on newborn rats: Wistar rats, 7 -old -day, llg~18g in weight, were selected without limitation ofsex. Ligated their left carotid to maintain the oxygen concentrations of 8% for 1.5 hours. Randomly divided them into two groups: The sham - operation control group ( n = 6) in which the rats were killed after 4 hours of the sham - operation; and the group with HIBD was divided into 10 subgroups (n =6) , in which the rats were killed respectively after 0,2,4,6,8, 10,12,24,48 and 72 hours of HI.2. Remaining specimen: Left cerebral cortex was separated, section 2 ~ 3mm diameters tissue specimen in the left parieto - occipital cortex, and was fixed in 2.5% glut aldehyde. The remaining of the cortex was placed rapidly into liquid - nitrogen at -196℃ and stored at - 80℃ 24 hours later.3. Transmission electron microscope; After routine process, the mitochondria and cellular ultrastructure were observed under transmission electroscope.4. Measurement of the concentration of PCr,ATP,ADP and AMP in cortical neurons with high performance liquid chromatography.5. Statistics; Variance analysis was used for the examination data with SPSS 10. 0 statistic software. Turkey method was used to test the HI group and the control.Results1. Changes of mitochondral ultrastructureOh after HI; The mitochondria! structure was normal. External membranes were mosdy intact with partly vagueness. There was no obvious enlargement in internal crista cavity. Light electronic high -density areas were found in the matrix.8h after HI: There were some injured mitochondria of different degrees and normal ones in the cytoplasma at the same time. In severely injured mitochondria, external membranes were partly broken, cristas were disordered and matrix distributed unevenly. Meanwhile mitochondria with intact external membranes and small - denatured follicles in internal crista were also found. Mitochondria with intact external membrane and clear crista structure were discovered at the same time.24h after HI; Most of mitochondria resolved with vague structure. Meanwhile nuclear membranes were broken and resolved, nucle-oli could also be found.48h after HI: Mitochondria with denatured follicles were found in cells. Organelles were mostly resolved with vague structure. The borders between cells were not clear, nuclei resolved and nucleoli disappeared.2. Changes of energy metabolismPCr decreased immediately after HI and after 6h of HI reached 50.78% of that in the control ( P < 0. 01). After 10h it resumed to 91. 93% of the normal control ( P > 0. 05) . While 24h later, it decreased again to 58. 06% of the control (P <0. 01).ATP decreased immediately after HI and after 2h of HI it... |
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