Research On Lung Injury And Cell Apoptosis In Diet-induced Obesity Mice Under Pneumonia

Obesity has developed into a considerable health problem in the whole world,which identified as a risk factor for adverse outcomes of various diseases.Bacterial pneumonia is an important disease of respiratory system.Apoptosis is a form of programmed cell death,while inflammation and oxidative stress could induce pathological apoptosis.However,the information regarding the difference between the response of obese and normal subjects to pulmonary inflammation is limited and the linking of bacterial pneumonia,obesity and cell apoptosis remains unknown.Here,we investigate the lung injury and mechanism of cell apoptosis between normal and obese mice under bacterial pneumonia through histopathology,microscopic quantitative analysis,immunohistochemistry,flow cytometry,qRT-PCR and Western blot.1.Histopathological Injury Caused by Inflammation and Oxidative Stress in Normal and DIO Mice under acute pneumonia3-week-old male ICR Mice were fed with the control or high-fat diet to establish the Normal and diet-induced obese(DIO)mice,then E.coli or phosphate buffer saline(PBS)was intranasally instilled to reproduce four groups,namely Normal-PBS,DIO-PBS,Normal-E.coli and DIO-E.coli.After infection,serum samples and lung tissues were obtained at 0,12,24,and 72 h.Body weight and lung weight were weighed after euthanasia,and lung index was calculated.Body length was measured to calculate Lee's index.Then,the macrostructure and microstructure of lung were observed.Also,serum triglyceride(TG)and total cholesterol(TC)contents,pulmonary cytokines and adipocytokines contents and oxidative stress level were detected in this study.Results showed that E.coli infection caused an acute suppurative inflammation in the lung with a large number of neutrophils infiltration,increased secretion of mucin and decreased deposition of collegens,as well as,increased lung index and serum TG and TC contents;elevated pulmonary tumor necrosis factor-?,interleukin(IL)-1?,IL-6,IL-8,and leptin levels;and oxidative stress in mice.Interestingly,almost all the above-mentioned parameters peaked at 12 h after infection in the Normal-E.coli group,but after 24 h in the DIO-E.coli group.These results indicated that the DIO mice presented a delayed inflammatory response and oxidative stress in non-fatal acute pneumonia.2.Delayed Apoptosis in lung displayed by Diet-Induced Obesity Mice through Mitochondrial Apoptotic Pathway under acute pneumoniaCell apoptosis rate,mitochondrial transmembrane potential(MMP,??m),TUNEL positive cells,the relative mRNA and protein expression of regulatory factor involved in mitochondrial apoptotic pathway(Bax,Bcl-2,Caspase-9 and Caspase-3)and subcellular localization of these proteins were detected.After infection,E.coli chanllenge caused an elevated cell apoptosis rate,percentage of pulmonary cells depolarized with collapse of the mitochondrial transmembrane potential and numbers of TUNEL-positive cells;and increased mRNA and protein expressions of Bax,Caspase-3 and-9,and decreased mRNA and protein expressions of Bcl-2 in the lung.In the PBS groups,a few Bax,Caspase-9 and Caspase-3 were found occasionally on the alveolar wall,while Bcl-2 positive proteins appeared mainly in the epithelial cells of respiratory bronchioles.After infection,a large numbers of Bax,Caspase-9 and Caspase-3 were visualized in the neutrophils infiltration areas or the alveolar wall in the E.coli groups,but Bcl-2 positive proteins were rarely observed.Interestingly,almost all the above-mentioned parameters peaked before 24 h after infection in the Normal-E.coli group,but after 24 h in the DIO-E.coli group.These results indicated that the DIO mice presented a delayed cell apoptosis in non-fatal acute pneumonia through mitochondrial apoptotic pathway in the lung.3.Diet-induced obesity mice execute pulmonary cell apoptosis via death receptor and endoplasmic reticulum pathways under acute pneumoniaThe relative mRNA and protein expression of regulatory factor involved in endoplasmic reticulum stress-induced apoptotic pathway(Calpain 2,Caspase-12 and JNK)and TNF-? mediated death receptor apoptotic pathway(TNFR1,TNF-? and FADD)and the subcellular localization of these proteins were detected.As shown in this study,after being infected with E.coli,the relative mRNA and protein expressions associated with either endoplasmic reticulum stress-induced apoptotic pathway or TNF-? mediated death receptor apoptotic pathway were all dramatically increased in the DIO mice,while in the Normal mice,only factors involved in TNF-? mediated death receptor apoptotic pathway increased at 72 h after infection.In the PBS groups,all these proteins showed slight positive staining in the cytoplasm of pulmonary epithelia,except for Calpain 2 and Caspase 12 which showed positive staining in the alveolar septum macrophages.However,in the E.coli groups,a large numbers of positive proteins were detected in the cytoplasm of inflammatory cells and pulmonary epithelia in the neutrophil-infiltrated areas.Moreover,the positive expressions of these proteins were higher in the DIO-E.coli group than in the Normal-E.coli group,except TNF-? with a high expression in the two groups.These results indicated that the DIO mice may mainly execute cell apoptosis in the non-fatal acute pneumonia through endoplasmic reticulum stress-induced apoptotic pathway and TNF-? mediated death receptor apoptotic pathway,while Normal mice may chiefly execute cell apoptosis through TNF-? mediated death receptor apoptotic pathway at the late phase afer infection.Conclusion: Bacterial pneumonia could cause inflammatory and oxidative damage to lung in mice,and obese mice had inhibitory or delayed effects to these damages.Moreover,obese mice also had inhibitory or delayed effects on acute nonfatal bacterial pneumonia-induced apoptosis.After infection,Normal mice mainly through the mitochondrial apoptotic pathway evoked apoptosis,and through TNF-? mediated death receptor apoptotic pathway executed apoptosis at the later phase of infection.However,the DIO mice initiated apoptosis through endoplasmic reticulum stress-induced apoptotic pathway or TNF-? mediated death receptor apoptotic pathway.By establishing the dual modes,this study investigated the activation of cell apoptosis and apoptotic pathway in the bacterial pneumonia in the Normal and DIO mice.The results would be meaningful to the future diagnosis and treatment of hospital acquired pneumonia in human medicine involved with obese patient.