The Role Of Insulin-like Growth Factor Binding Protein-2 In The Progression Of Pneumonia-induced Sepsis

Background:In the guideline of Sepsis 3.0,the latest definition of sepsis is organ dysfunction in crisis life caused by the host’s maladjustment of response to infection^[2].Sepsis is a major cause of human death worldwide^[1].,In Sepsis 3.0 Guide,the latest definition of sepsis is organ dysfunction of life-threatening caused by the host’s maladjustment of response to infection^[2].Studies have shown that one third of intensive care patients are sepsis patients,and the mortality rate is as high as 55.7%^[3].At present,there are no drugs for sepsis in the world.Therefore,sepsis is an important research direction in the field of critical care medicine.Studies have shown that the early diagnosis of sepsis can effectively reduce the mortality of patients with sepsis^[4].According to the new definition,the diagnosis of sepsis depends on a sequential organ failure score（SOFA）,which includes the assessment of the functional status of the respiratory system,coagulation system,liver system,cardiovascular system,central nervous system,and urinary system.This diagnostic criteria means that patients cannot get an accurate diagnosis before organ dysfunction occurs.Research on the pathogenesis of sepsis will help establish an early diagnosis and develop therapeutic drugs.Insulin-like growth factor binding protein-2（IGFBP-2）is one of the members of the insulin-like growth factor binding protein family.Mature IGFBP-2 is formed by the hydrolysis of the signal peptide of the precursor protein,with a molecular weight of about 31.2 k Da.IGFBP-2 acts as a carrier protein by binding to insulin-like growth factors（IGF-I and IGF-II）to regulate cell growth,differentiation,apoptosis,adhesion and movement^[5].At present,IGFBP-2 has been reported to promote the occurrence and metastasis of tumors,the expansion of cancer stem cells,and tumor angiogenesis^[7].However,the role of IGFBP-2 in sepsis is still unclear.In the previous phase,24 plasma samples from 12 patients with sepsis before and after exacerbation were selected from the collected blood sample bank in our laboratory.Protein quantitative mass spectrometry analysis showed that IGFBP-2 was detected in plasma samples from all 24 patients,and the level of IGFBP-2in the moribund was significantly higher than that in the stable,suggesting that IGFBP-2 might be related to the changes of sepsis patients’condition.Relevant studies have shown that the primary infection site of about40-60%of sepsis patients is the lower respiratory tract（i.e.,pneumonia）,followed by abdominal source and urinary tract infection^[14–17].To further explore the role and pathogenesis of IGFBP-2 in sepsis,we collected data from healthy subjects,Community-acquired Pneumonia（CAP）,Pneumonia-induced Sepsis（PIS）and digestive system infection-induced sepsis（DIS）,and to express and apply the recombinant IGFBP-2 protein,in combination with septic animal and cell models,to further explore the role of IGFBP-2 in the disease process of sepsis.Method:I Using sepsis 3.0 as the judgment standard,plasma samples of sepsis patients admitted to the Department of Respiratory and Critical Illness,Huai River Hospital of Henan University and RICU from June 2016 to July 2019 were collected,to screen patients with PIS（SOFA≥2 and primary disease was pneumonia）,DIS（SOFA≥2 and primary disease was digestive system infection）,CAP patients admitted at the same time and healthy subjects.Enzyme-Linked Immunosorbent Assay（ELISA）was used to detect and compare the plasma IGFBP-2 levels in patients of each group.SPSS 26.0 software was used in combination with the clinical data of patients and Spearman correlation analysis was used to assess the correlation between IGFBP-2 levels and each clinical test index.The survival curve was drawn by Kaplan-Meier method and the Cox proportional risk model was used to analyze the related risk factors for death.The ROC curve and AUC area were used to compare the accuracy of IGFBP-2 in judging the prognosis,renal dysfunction,anemia,etc.The analysis was performed using relevant statistical methods such as single-sample K-S test and rank sum test of two independent samples.This study mainly focused on sepsis caused by pulmonary infection.The recombinant mice-derived IGFBP-2（r MSIGFBP-2）protein was expressed and purified,followed by the establishment of a pneumonia-induced mice sepsis（KPS）model,the observation of the effect of the protein on the survival of septic mice,the collection of mouse plasma,bronchoalveolar lavage fluid,fixed lung tissue,and the freezing of various organs（heart,liver,spleen,lung,kidney,and pancreas）in mice.The kit was used to detect injury indicators in plasma,Western Blot was used to detect related injury markers in plasma and lung tissue homogenate,H&E staining of frozen lung sections,and immunohistochemistry of frozen lung sections.In addition,the in vitro cell experiments were conducted to explore the mechanism of IGFBP-2 in the process of pneumonia-induced sepsis disease.Result:1.The IGFBP-2 concentration in Moribund plasma of patients with PIS is significantly higher than that in the Moderate（p=0.001）;2.The level of plasma IGFBP-2 in patients with PIS is significantly higher than that in healthy subjects and patients with CAP(p<10^-4);3.The plasma IGFBP-2 level in patients with PIS is positively correlated with the aggravation of renal dysfunction（p=0.0004）;4.The level of plasma IGFBP-2 has a high AUC value with sepsis,renal dysfunction and prognosis;5.In the pneumonia-induced mice sepsis（KPS）model,increasing the plasma IGFBP-2 level can significantly reduce the mortality of mice;6.IGFBP-2 protein reduces lung injury in KPS mice;7.Injection of recombinant IGFBP-2 can reduce the number and proportion of neutrophils in whole blood of KPS mice;8.In thetaphylococcus aureus pulmonary infection induced mice sepsis（SPS）model,the injection of r Ms IGFBP-2 protein had no significant effect on the survival of the mice;9.IGFBP-2 promotes the proliferation of macrophages;10.IGFBP-2 can bind to Klebsiella and promote macrophages to devour bacteria.Conclusion:Plasma IGFBP-2 level may be a potential marker to predict the occurrence,development and renal dysfunction of sepsis.The increase of plasma IGFBP-2 level may reduce the lung injury caused by Klebsiella pneumoniae and the mortality of sepsis.Stimulating the proliferation and phagocytosis of macrophages may be one of the protective mechanisms of IGFBP-2 in sepsis.The recombinant IGFBP-2 protein has potential value in the treatment of sepsis.