Proteomic And Functional Study Of Brain Stem Respiratory Center(PreBotC) In Severe Traumatic Brain Injury

With the development of the modern automobile industry, the incidence of traumatic brain injury in particular severe traumatic brain injury increased year by year. High mortality and disability of severe brain injury has become a whole major issues of common concern in the medical and society as at present, and its one main hub is the early respiratory and cardiac arrest caused by the direct damage to the brain stem. The study confirmed that pre-Botzinger complex (preBotC) is essential for rhythmogenesis as the brain stem respiratory center, which provides a more precise "target" for the study of brain stem inhibition, especially respiratory dysfunction.For complex pathophysiological changes occurred in the respiratory function after brain injury, proteomics with its comprehensive and high-throughput studies the overall protein expression profile as a systems biology approach. By the establishment of severe traumatic brain injury with respiratory insufficiency model in rats, that subject build preBotC quantitative differences in protein through Label-free quantitative technology based on liquid chromatography tandem mass spectrometry(Label-free LC-MS), and made the differences in protein function analysis to further explore the preBotC.â… Establishment of the severe head injury model with respiratory insufficiency in rats and the positioning of the brain stem respiratory center (preBotC).Objective:To research the establishment of the severe head injury model with respiratory insufficiency in rats, and the positioning of the brain stem respiratory center (preBotC). Above researches provided model foundation for the severe traumatic brain injury with respiratory insufficiency and methodological basis for the brainstem respiratory center (preBotC) in further studies.Methods:Using of hydraulic injury equipment in male SD rats(impact force, 2.7-2.8atm), we prepared severe traumatic brain injury model with respiratory dysfunction(experimental group), according to blood gas analysis in 2h after injury. At the same time, we made precise positioning of preBotC through its specific NK1R immunohistochemical stainingin the anatomical plane of the hypoglossal nerve location.Results:After injury, there was the brain stem inhibition of the slow breathing, low blood pressure and heart rate, of which the most significant changes in respiration. Also there showed a brief apnea with the duration of 1-10 minutes immediately after injury. With the animal ventilator-assisted breathing, the 80.0%(36/40) in rats can restore breathing irregular breathing on their owns, and eventually there were 33 animals belongs to severe traumatic brain injury with respiratory insufficiency group by blood gas analysis standards (referred to as'experimental group').Conclusion:Successful establishment of the severe head injury model with respiratory insufficiency in rat,which was with of good stability, good reproducibility and good survive for more than 2 hours through timely and effective aid. Complements of anatomical location and histological preparation about preBotC:By the anatomical location of hypoglossal nerve, NK1R immunohistochemistry could precise position preBotC, so we prepared successfully is the preBotC tissue section included in the hypoglossal nerve.â…¡Comparative proteomic studies in preBotC of severe brain injury rats-Label-free LC-MSObjective:To establish and perfect of the quantitative proteomics processing and detection methods for the deep brain nuclei like preBotC. To establish the comparative proteomics in preBotC of severe brain injury rats. Above researches provided research base to functional analysis of severe head injury preBotC in rats.Methods:After extracting the total protein in samples, we determined its concentration according to Bradford, and detected its distribution by use of polyacrylamide gel electrophoresis (SDS-PAGE). Post the pre-digested of proteins, label-free quantitative techniques based on liquid chromatography-mass spectrometry (Label-free LC-MS) and the database search help us get the comparative proteomics in preBotC of severe brain injury and cluster analysis.Results:The comparative proteomics in preBotC had a total of 74 kinds of proteins (in two-fold difference, p<0.05), which showed 34 species of increased proteins and 40 down of that. Conclusion:Establishment and perfection of the quantitative proteomics processing and detection methods for the deep brain nuclei like preBotC:For high deep nuclei with the less organization and the regional specificity, Label-free LC-MS protein was a good quantitative proteomics detection method, such as covered a wide range of proteins, the molecular weight evenly distributed, and had stability, good reproducibility, quantitative high-precision. Establishment of the comparative proteomics in preBotC of severe brain injury rats:The comparative proteomics in preBotC had a total of 74 kinds of proteins, iwhich showed 34 species of increased proteins and 40 down of that. They participated energy metabolism abnormalities, nerve degeneration, apoptosis, inflammation, oxidative stress and other pathophysiological changes.â…¢Functional analysis of comparative proteomics in preBotC of severe brain injury rats.Objective:The functional analysis of comparative proteomics in preBotC of severe brain injury rats could provide possible new targets for the treatment.Methods:GO analysis (Gene Ontology analysis) made the systematic analysis of significance functions from the comparative proteomics in preBotC. So we got the menu and the functional network (GO Map) corresponding to the comparative proteomics in preBotC.Results:There showed 128 kinds of significance function corresponding to that of 74 proteins by GO analysis in and preBotC, in which 94 increased the functional performance and 34 showed a functional reduction. It also demonstrated that the intracellular protein transport, synaptic vesicle transport, neuron differentiation, adenine metabolism, microtubule depolymerization, regeneration and healing, histamine uptake and transport, melanin metabolism, degenerative protein transport, response to injury stimuli and energy metabolism-related physiological changes.Conclusion:The comparative proteomics in preBotC of severe brain injury rats took part in intracellular protein transport, synaptic vesicle transport, neuron differentiation, adenine metabolism, microtubule depolymerization, regeneration and healing, histamine uptake and transport, melanin metabolism, degenerative protein transport, response to stimulation of the damage, energy metabolism, in which the respiratory function was directly related with of melanin metabolism proteins, microtubule depolymerization proteins such as protein and energy metabolism.