Effects Of Neutralizing HMGB1 On Influenza Pneumonia In Recipient Mice After Heart Transplantation

Background:Heart transplantation has become the major therapy choice for advanced heart disease.Although the advances of immunosuppressant,such as tacrolimus and cyclosporine,do improved the survival of recipients to a large extent,infections remain a leading cause of post transplantation mortality,especially in the early stage after transplantation.Pneumonia is the major cause of infection in heart transplant patients.Respiratory viruses,such as influenza virus,have worked up as potential cause of pneumonia after transplantation.Recipients of solid-organ transplants with H1N1,the cause of most recent influenza pandemic,are at increased risk of complications compared to the general population,especially in the early post-transplant period.Mortality caused by H1N1 is higher in transplant recipients compared to healthy individuals.Thus,exploring appropriate drugs to reduce morbidity and mortality while prolonging the survival of graft is paramount in recipients.High-mobility group box 1(HMGB1),one of the DAMPs,initially identified as a DNA-binding protein involved in maintaining nucleosome structure and regulating gene transcription.It can be released into the extracellular matrix from immune and non-immune cells in response to varieties of stimuli.Released HMGB1 also contributes to the pathogenesis of numerous diseases,including graft injury and the inflammatory response in heart transplantation.HMGB1 blockaded significantly prolongs the survival of allograft and alleviates the allograft rejection.Preliminary studies in our laboratory showed that neutralizing HMGB1 could maintain recipient mice defenseassociated response against LPS and poly(I:C)challenges.Therefore,we propose the following scientific question: whether neutralizing HMGB1 can prolong the graft survival without affecting the recipient's defense respond to influenza virus.Objective:To investigate the effect of anti-HMGB1 monoclonal antibody on influenza virus induced pneumonia in recipient mice.Methods:In order to simulate the clinical situation,the recipient mice were treated with HMGB1 monoclonal antibody from the day of transplantation,and then infected with H1N1 virus on the 3rd day after heart transplantation.In the experiment,we first observed the whole course of the disease,and then used H&E,Q-PCR,ELISA,Western blot,immunofluorescence,immunohistochemistry,flow cytometry etc technologies to study the immune system and the pathogenesis of HMGB1 during the course of the disease.Results:In the current study,we probed the protective effects of anti-HMGB1 m Ab in the cardiac transplant model challenged with H1N1 virus.1.Blockade of HMGB1 prolonged the survival time of graft and accelerated the recovery of infected recipient mice.2.The administration of HMGB1 m Ab display attenuated severity of inflammatory injury and accumulation of inflammatory cells.The neutralization of HMGB1 mitigated inflammatory injury and reduced lung leucocyte and interstitial macrophages in the lung of H1N1-infected HTx mice.3.Influenza virus infection in recipient mice resulted in the increase of HMGB1.4.H1N1 virus infection induces the translocation and release of HMGB1 in AEC II.5.TLR4 and RAGE,the receptors of HMGB1,were highly expressed in AEC II.6.In AEC II,HMGB1 could activate MAPK pathway through TLR4 signaling,but not RAGE.7.Heart transplantation changed the basic level of HMGB1 in lung of recipients.In vitro,the increase of HMGB1 in microenvironment led to the increase of chemokines released from type ? alveolar epithelial cells after H1N1 infection.This may lead to a higher risk of lung injury in transplant patients in the face of infection.Conclusion:Together,HMGB1 m Ab has potential application value in heart transplant recipients.It not only prolonged the graft survival,but also reduced the pulmonary inflammatory response after infection,protected the resistance of recipients to pathogenic microorganism infection,improved the survival rate and accelerated the recovery after infection.In addition,we found that the weight of the infected recipient mice was different from that of the infected normal mice,suggesting that the infection of the normal mice could not fully reflect the infection of the transplanted mice.We found that the increased level of HMGB1 in the microenvironment could aggravate the level of chemokine KC and CCL2 released by alveolar type II epithelial cells after H1N1 infection,which may lead to a higher risk of lung injury in the challenge of infection.Therefore,blocking HMGB1 provides a new idea for drug use after organ transplantation.Different from the previous immunosuppressants,HMGB1 may be able to inhibit the rejection of transplantation without affecting the recipient's defense against virus infection.