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Objective: Neonatal hypoxic ischemic encephalopathy (hypoxic-ischemic encephalopathy, HIE)is a disease caused by asphyxia and hypoxia during the perinatal period ,and is one of the maincauses of neonatal early mortality and chronic neurological damage in the late. Inflammatory caninduce to release a variety of cytokines and inflammatory mediators. In recent years, matrixmetalloproteinases (MMPs), particularly metalloproteinase -2 (MMP-2) and metalloproteinase -9(MMP-9) in hypoxic-ischemic brain injury is proved to be related to HIBD. In this study,hypoxic ischemic brain damage (HIBD) model of neonatal rat is established to observebehavioral changes of rats and pathological changes in brain tissue,and immunohistochemicaldetermination of MMP-2,MMP-9 and TIMP-1 and apoptosis of brain tissue, from the MMP-2,MMP-9 and TIMP-1 on cell apoptosis to explore the pathogenesis of neonatal hypoxic ischemicencephalopathy.Methods: (1)Establishment of neonatal rat HIBD model. (2) Examination of the behavioralCapacity: before , after surgery and during hypoxia , respectively, changes were observed inbehavior of each newborn rat. (3) 72 neonatal Wistar rats of 7 days were randomly divided tosham operation group and HIBD group,according to the time ,the latter were divided intohypoxia-ischemia 6h, 24h, 48h, 5d and 14d of 12.Sham operation group received free thecommon carotid artery in the left neck without hypoxia-ischemia, HIBD group set free and madeleft common carotid artery ligation then back to the maternal side to breastfeed for 2 hours, thenplaced them in hypoxia for 2 hours, neonatal rats in each group were decapitated atcorresponding time points and observation of brain morphological changes in general braintissue in the naked eye, paraffin sections were Stained by HE of the left side of brain tissue tooberve pathological changes ,detected MMP-2, MMP-9 and TIMP-1 expression of the left brainof neonatal rat in Half of each group at different time points by immunohistochemistry, anddetected brain cell apoptosis of the left brain of rats at different time points in the other half ofeach group using flow cytometry.Results: (1)Neonatal rats were of abnormal behavior with varying degrees afterhypoxia-ischemia. (2)There are different degrees of abnormal changes of brain tissue in generalafter hypoxic-ischemic, over time, the extent of brain changes were gradually reduced. (3)HEstaining: sham group brain structure and the cellular level are clear, neurons closely aligned, onlythe individual cell shrinkage, nuclear stained; HIBD 6h group the brain cortex cells is mildswelling, HIBD 24h group cortical cells is swelling obviously, neural disorder, cell gap increases, showing a lot of degeneration of neurons ,a small piece of liquefaction necrosis occurred inHIBD 48h group; the situation of HIBD 5d group is better than HIBD 48h group , the extent ofedema reduced, neuronal survival increased, there are still some degeneration and necrosis ofneuronal ,damage has been restored in HIBD 14d group at a certain level , the survival of nervecells increased significantly.(4) Immunohistochemical staining:Observing by light microscopy,MMP-2,MMP-9 and TIMP-1 positive staining was brown fine particle deposition, which was inthe cerebral cortex and hippocampus, mainly located in the cytoplasm.MMP-2 may be located inthe endothelial cell of ischemia and macrophages,MMP-9 and TIMP-1 may be located in themicroglia. Sham group, the expression of MMP-2 is weak or none.(Figure 7),slightly increasedin HIBD 6h group (Figure 8), in HIBD 5d group reached a peak (Figure 11)and higher thanHIBD 6h group, in addition to HIBD 24h group, compared with the sham group it wasStatistically significant (P <0.05, Table 1). Sham group, there is a certain degree of MMP-9expression (Figure 13), HIBD 6h group slightly increased (Figure 14), peaked at HIBD 24hgroup(Figure 15).In addition to HIBD 14d group, compared with the sham group it wasStatistically significant (P <0.05, Table 1). Sham group, there is a certain degree of TIMP-1expression (Fig. 19), peaked at HIBD 48h group(Figure 22), gradually decreased HIBD 14dgroup to restore the original level, in addition to HIBD 6h group and HIBD 14d group,comparedwith the sham group it was Statistically significant (P <0.05, Table 1). (5) Apoptosis rate ofneurons: brain cells in HIBD 24h group slightly increased ,HIBD 5d group up to peak, HIBD14d group declined a little compared with HIBD 5d group. In addition to HIBD 6h group, eachgroup compared with the sham group was Statistically significant (P <0.05, Table 2).ConclusiConclusion:(1) By observation of the behavioral capacity and the histopathological of neonatalrats brain confirmed that hypoxic-ischemic brain damage model is established successfully, andover time, brain damage reduced. (2) Metalloproteinases expression of HIBD group was changedat a different level,which may attend pathogenesia and repair.
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