Autophagy And Its Underlying Mechanism In Mouse Thymus Induced By Influenza Virus Infection

Influenza A virus is a single-stranded negative-sense RNA virus which can infect human,chickens,pigs and other animals,and cause the respiratory infection.Influenza pandemic had broken out several times from "Spanish Flu Pandemic"occurred in 1918 to the influenza pandemic in 2009 and caused high morbidity and mortality.In addition,there’re many poultry died of avian influenza infection every year.Therefore,influenza virus poses a great threat to human and animal health.Influenza virus not only cause huge loss in breeding industry throughout the world,but also forms a great challenge to public health.Autophagy is a fundamental cellular homeostatic mechanism,while it is also known as a defensive system in eukaryotic cells.In the studying of the relationship between autophagy and virus replication,people had found that autophagy is involved in the infection of many virus in which virus sometimes can take advantage of autophagy to promote virus replication.Some studies had shown that influenza virus can induce autophagy in both infected cells and infected animal lungs.But until now there is no concrete research reports about whether influenza A virus can induce autophagy in thymus of infected animals and whether autophagy is involed in the thymic atrophy or not.In this study,we first made ultrathin slices of thymus of mice infected with high titers of influenza virus.Using electron microscope analysis,we found that there were accumulated autophagosomes in the thymocytes of mice infected with influenza virus for 24 hours.Furthermore,we detected the expression of LC3 protein in the thymus of infected mice,and found that the expression of LC3-II in the thymus was significantly increased after infection with influenza virus for 24 hours,which was consistent with the previous observation in lungs.However,the expression of LC3-II was not seen in spleen of infected mice.These results indicate that influenza A virus could induce autophagy in thymus tissue of infected mice at early infection stage.After treated with autophagy inhibitor,the mice were sacrificed and observed that the acute thymic atrophy caused by influenza virus infection had been ameliorated.To further investigate the pathological features of the thymus,hematoxylin and eosin(H&E)staining was performed.There was less severe pathological damage in the infected mice treated with autophagy inhibitor as compared with that in control mice.These results suggest that the autophagy induced by influenza virus infection in the thymus may be one of important causes of the thymic atrophy.On the other hand,the expression of anti-apoptosis proteins was examined in the thymus of influenza virus infected mice.We found that expression of Bcl2 and Bcl-xl was markedly decreased at late infection stage,indicating that influenza A virus could induce apoptosis in mouse thymus.However,there was no significant difference in the expression of anti-apoptosis proteins in the spleen of infected and uninfected mice.Given the expression change of LC3-Ⅱ in the thymus and spleen of infected mice,we speculated that the apoptosis induced by influenza virus infection in mouse thymus is likely related to autophagy.However,this remains an ongoing task.Finally,to understand the mechanism by which influenza virus induced autophagy,we analyzed the relevant signaling pathway.We found that the extent of Akt and mTOR phosphorylation was decreased in the thymus of virus infected mice,suggesting that the activation of Akt and mTOR in thymus is suppressed by the infection.These data indicated that influenza virus may regulate autophagy in the thymus of infected mice by targeting Akt-mTOR signaling pathway.Because influenza virus infection can trigger secretion of glucocorticoid in host,we injected mice with dexamethasone and found that dexamethasone could also induce autophagy and apoptosis in mice thymus.Interestingly,the phosphorylation of Akt and mTOR was inhibited by dexamethasone treatment as well.Together,these results reveal that influenza virus can induce autophagy in the thymus of infected mice,which is one of the possible reasons causing thymic atrophy.Importantly,glucocorticoid triggered by influenza virus infection likely plays a key role in the induction of autophagy.