| Project Background Wars made casualties,traumatic brain injury(TBI)is one of the main damage form types in wounded soldiers, moreover its an important cause to death.its expression forms, illness development is rapid, high mortality, prognostic difference big, and many other features.Its expression in a variety of forms,develop rapidly,high mortality and prognostic extremely differences,et al.TBI Has been gradually became a major field that Studied by military and local medical school, or institute of institutions.With the progress of science and technology,traffic developed, leading to its incidence rate rising gradually. Although we cannot avoid the direct action that related to TBI, its a siginificant mean looking for one method to reduce extremely secondary damage by TBI.Such as,inflammatory stress, oxidative stress, and endoplasmic reticulum stress and so on.The method is important in improving the treatment of TBI and in significantly reducing the mortality and morbidity. Hypoxia preconditioning(Hypoxic preconditioning, HPC) is a method that through a appropriate hypoxia training ahead of time, which can significantly enhance the body’s, especially the brain’s various tolerance for ischemia, hypoxia and other blow then happened suddenly.In the end protecting the brain tissue.Phosphorylation of eukaryotic translation initiation factor alpha 2 is phosphorylated by e IF2 alpha during the Endoplasmic Reticulum Stress(ERS),which is an important molecular and played an significant role in the beginning of protein synthesis in endoplasmic reticulum.When e IF2 alpha phosphorylated lead to protein synthesis blocked.With the endoplasmic reticulum stress continues,(P)e IF2 alpha turned to mediate the transcription of apoptosis gene, caused neurons apoptosis or necrosis.growth arrest and DNA damageinducible protein 34(GADD34),is an important molecule during the process of ndoplasmic reticulum stress(ERS),participated the restart of protein synthesis during the process of ERS.Studying the influence of HPC to the change of ERS and its apoptosis after TBI,maybe reducing the neuron injury, apoptosis,and improving behavioral cognitive function.In order to reduce the mortality and morbidity providin theory support for the clinical treatment of TBI.Part one: The experimental research of hypoxia preconditioning affect the expression of endoplasmic reticulum stress protein P-e IF2 alpha in rats with traumatic brain injuryObjective(1) To observe the pathological change in rats after traumatic brain injury,and the influence to that change by hypoxia preconditioning;(2)To explore the influence of hypoxia preconditioning on the expression of endoplasmic reticulum stress protein P-e IF2 alpha and the change of neuron apoptosis.Method 208 Sprague-Dawley adult male rats were divided,using the method of random Numbers table,into four groups,that is control(Con) group, ischemia preconditioning(HPC) group, the traumatic brain injury(TBI) group, and hypoxia preconditioning traumatic brain injury(HPCT) group. The group of TBI and HPCT were further divided into eight subgroups:1 h, 3 h, 6 h, 12 h and 24 h, 3 d, 7 d, 14 d. The rats in the group of Con just Scalp incision and suture then put to death.while,the animal model of TBI making by the improved Feeney ’s free traumatic brian injury fall.Impactive location:in front 3.0 millimeter of anterior fontanelle,and in the left side of the midline 3.0 millimeter,at the same time closed to the coronal suture.Drilled a bone window about 5.0 millimeter in diam. Impactive weight and height:Using a weight of 50 gram in high of 25 centimeter free fall to impact the bone window.Hitting the depth of about 3-4 millimeter.Giving a appropriate training in advance(- 50 KPa, 3 h/d, continuous 3d) by a low pressure oxygen chamber to make a hypoxia preconditioning rat model.By using HE staining and making fresh brain tissue ultra-thin sliced, then to observe,under light microscope and electron microscope,the pathological changes of rat brian tissue in each group; Immunohistochemical staining method and Western blottin were used to determined the expression of protein P-e IF2 alpha for each group. TUNEL staining to detect the change of the neuron apoptosis.Result( 1) There was no difference between Con group and HPC group in pathological changes which observed by light microscope and electron microscope.While,HPCT group has a lighter pathological changes than that in TBI,which had significantly reduce neurons damage and significantly reduce the ultrastructure.(2) Immunohistochemical staining and Western blotting method testing found that P-e IF2 alpha protein expressed in 3 h after injury, then rised gradually to expresse in 6h-12 h, peak at 24 h, after decreased in 3d-14d; Among them,the expression of P-e IF2 alpha protein in HPCT group at 6h-7d after injury lower significantly reduced than that of TBI group.The difference between the two groups have statistical significance(P<0.05). Neuron apoptosis were detect by TUNEL staining found that TBI group and HPCT groupappeared at 6h after injury,then trend to increase in 12h-1d, achieve its peak at 3d, then to decrease at 7d-14 d. The apoptosis cells declined obviously in 12h-7d after injury in HPCT than that in TBI group,the apoptosis rate between the two groups was statistically significant(P < 0.05).Conclusion(1) Hypoxia preconditioning can significantly reduce the brain tissue of TBI pathological change,and better to maintain the integrity of the ultrastructure of neurons, reduced endoplasmic reticulum injury.(2) Hypoxia preconditioning may through selective inhibition of the expression of endoplasmic reticulum stress protein P-e IF2 alpha in rat cortex after TBI,reducing the damage caused by ERS and the apoptosis of nerve cells.In order to promote cell survival and maintain the stability of the neural function.Part two: The experimental research of hypoxia preconditioning influence the expression of GADD34 and behavioristics in hippocampus of rats after traumatic brain injuryObjective Determineed the influence of hypoxia preconditioning on the expression of GADD34 and behavior change in hippocampus of rat.To reseach the mechanism of hypoxia preconditioning reducing endoplasmic reticulum stress injury in TBI.Method 48 clean New Zealand SD male rats,were randomly divided into contr ol(Con) group, hypoxic preconditioning(HPC) group, and traumatic brain injur y(TBI) group and hypoxia preconditioning craniocerebral trauma(HPCT) group. traumatic brain injury rat medol mede by the improved traumatic brain injury free fall device.preconditioning the rats in a low pressure oxygen chamber to m ake hypoxia preconditioning rat medol.Put the four groups of rats within 24 h af ter injury to death and taken specimens. Using the improved Sugrwara method t o evaluate the behavior of rats in 24 h after injury. Using the method of RE-PC R、Western blot and immunohistochemical staining to determine the changes abo ut the expression of GADD34 m RNA and protein.and TUNEL staining method was used to detect the to detect the changes of apoptosis neurons expression.Result The GADD34 m RNA and protein detected respectively by the method of RT-PCR, Western blotting and immunohistochemical staining,which is higher expresion in HPCT group than that in TBI group in the rat hippocampus.The difference between the two groups have statistical significance(P<0.05).Moreover,rats at 24 h after injury in HPCT group had significantly higher behavior score than that in TBI group(P<0.05). TUNEL staining found that apoptotic cells of HPCT group decreased significantly compeared with TBI group in rat hippocampus, and apoptosis rate dramaticly declined, the defference between the two groups was statistically significant(P<0.05).Conclusion Hypoxia preconditioning can reduce rats hippocampal behavior damage after TBI,and its mechanism more likely through to increase the expression of GADD34 in hippocampal, reduced hippocampal neuron apoptosis, relieved ERS injury. plaing a significant role in brain protection. |
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