Protective Regulatory Molecular Mechanism Of Lung Proteins In Mice During Torpor

Torpor is a common adaptation strategy for animals to cope with adverse external environments such as cold,food shortage and drought.Animals in torpor will have physiological reactions and behaviors such as hypothermia,decreased metabolic rate,inactivity and inactivity.Although animals in torpor may experience a variety of complex environmental changes such as low temperature,rewarming and hypoxia,they can often recover their physical functions in a short time after waking up,and will not cause permanent damage to organs and tissues.It can be seen that the torpor behavior of animals is of great significance for animals to resist the adverse environment.Due to the weakening of energy supply and the complex changes of environment,various organs of animal torpor will have different degrees of physiological changes over time.It is of great significance to study the physiological and molecular regulation mechanisms of different organs during torpor in order to further explore the self-protection and adaptation strategies of animals under the state of torpor.The lung is an important organ for the body to obtain oxygen.Terrestrial vertebrates mainly exchange oxygen in the external environment with carbon dioxide in the circulatory system through the lung.In addition,the lungs also have the function of preventing the loss of blood and body fluids,as well as resisting infection by pathogens.In torpor state,the oxygen consumption and respiratory rate of mice decreased,and the energy consumed by the expansion and contraction of lungs also decreased.How to make torpor sleeping mice obtain enough oxygen to support the body's energy metabolism,the body has special requirements for the physiological activities and functions of the lungs.In this paper,by establishing the torpor model of C57BL/6 mice at 22?and17?and 12?low temperature,using tissue anatomy method and DIA quantitative proteomics method to study the protection and repair process of mouse lung proteins to explore the molecular regulation mechanism of lung tissue in mouse torpor state.Firstly,this paper established the mouse model in torpor state:the feeding temperature was controlled at 22?,17?and 12?respectively.After about 48 h,40h and 30 h,the mice entered the deep torpor state,extracted the lung tissue,and observed the microstructure and sub microstructure changes of mouse lung cells in torpor state by transmission electron microscope.The results showed that the epidermis of mouse lung tissue in torpor state was thickened,the cells were loosely arranged.At the same time,there were vacuoles in epidermal cells,some cell membranes were damaged,some mitochondria were swollen,and ridges were broken.It shows that the lungs of mice will be damaged during torpor,and the reason is not clear.In order to further explore the molecular mechanism of physiological changes in mouse lung tissue in torpor state,we then used DIA quantitative proteomics technology to comprehensively and systematically study the expression and modification of total protein and phosphorylated protein in mouse lung in torpor state,in order to explain the molecular mechanism of lung injury and repair in torpor state mice.The results showed that the number of lung proteins identified in normal mice(control group,26?)and three groups of torpor mice(22?,17?and 12?)were4405,4318,4358 and 4403 respectively,of which 3471 proteins were identified in each state.The number of differentially expressed proteins was 655(proteins whose expression changed more than twice were regarded as differentially expressed).Through systematic bioinformatics analysis of differentially expressed proteins(GO annotation analysis,KEGG signal pathway analysis,etc.),it was found that some proteins with the effect of causing lung injury had changed,such as the expression of connective tissue growth factor and myeloperoxidase were up-regulated during torpor,neural predictor cell expressed developmentally down-regulated protein 1 was down-regulated in torpor.At this time,when the lung begins to have the tendency of injury and enters the state of injury,some proteins involved in preventing injury and repair,such as avoiding apoptosis,fibrosis and thrombosis,are up-regulated.These proteins include lysocardiolipin acyltransferase 1,flotillin-2 and proteins related to the signal pathway of complex and coagulation cascades(including Alpha-1-antitrypsin 1-2,Alpha-1-antitrypsin 1-4,Complement factor H,etc.).Phosphorylation modification of proteins is a common modification form to regulate a variety of cell physiological functions and molecular regulation.With the innovation of mass spectrometry technology,more and more studies on signal pathways related to protein phosphorylation modification have been reported.In order to explore the molecular regulation mechanism of lung tissue during torpor,the changes of phosphorylation modification level of lung protein during torpor were analyzed by using DIA quantitative proteomics method and Ti O₂enrichment method.The results showed that 5379,5365,5922 and 5187 phosphorylated peptides were identified in normal temperature group and 22?,17?and 12?feeding group,respectively,which belonged to 1590,1636,1707 and 1610 phosphorylated proteins.The number of intersecting phosphorylated peptides found among the four groups was3829(belonging to 1399 phosphorylated proteins).The 3829 intersecting phosphorylated peptides were systematically analyzed by bioinformatics methods.It is found that the phosphorylation modification level of some proteins that may be involved in lung protection function has changed,which has important regulatory significance in promoting cell survival,inhibiting oxidative damage,damage repair and inhibiting apoptosis.For example,the phosphorylation modification level of p21activated kinase 2,heat shock protein beta-1 and catenin alpha-1 was up-regulated,and the phosphorylation modification level of paxillin was down-regulated.Through the in-depth study on the protein regulation strategy of mouse lung during torpor,this study explained the lung injury and possible repair mechanism in torpor state.The results provide a theoretical basis for clarifying the physiological adaptation mechanism of mammalian torpor,and provide a theoretical basis and new ideas for the prevention and treatment of pulmonary thrombosis,pulmonary fibrosis and pulmonary antioxidant stress under torpor.