PROTEOMIC STUDY REVEALED DYSREGULATION OF CELL STRUCTURE AND METABOLISM IN SEPTIC PATIENTS SECONDARY TO COMMUNITY ACQUIRED PNEUMONIA

Narendra Kumar Sharma1, Alexandre Kaiji Tashima2, Milena Karina Colo Brunialti1, Éden Ramalho Ferreira3, Ricardo José Soares Torquato2, Renato Arruda Mortara3, Flavia Ribeiro Machado4, Murillo Assunção5, Otelo Rigato6 and Reinaldo Salomao1*

1Division of Infectious Diseases, Escola Paulista de Medicina, Hospital São Paulo, Universidade Federal de Sao Paulo, Sao Paulo, Brazil. 2Departamento de Biochemistry, Escola Paulista de Medicina, Universidade Federal de São Paulo, São Paulo, SP, Brazil.3Department of Microbiology, Immunology and Parasitology, Escola Paulista de Medicina, Universidade Federal de Sao Paulo, Sao Paulo, Brazil. 4Intensive Care Unit, Hospital São Paulo, Escola Paulista de Medicina, Universidade Federal de Sao Paulo, Sao Paulo, Brazil. 5 Intensive Care Unit, Hospital Israelita Albert Einstein, Sao Paulo, Brazil. 6 Intensive Care Unit, Hospital Sirio Libanes, Sao Paulo, Brazil.

Background Sepsis is a life-threatening organ dysfunction and a major cause of mortality worldwide. The major challenge to study sepsis is its diversity such as age, source of infection and etiology. In the present study, we use quantitative proteomics to evaluate host proteome response in septic patients, secondary to community acquired pneumonia (CAP). Methods Briefly, 425 patients were enrolled in our cohort of septic patients. Aiming to avoid heterogeneity of patients, 33 septic patients (20 survivors and 13 non-survivors) with CAP were selected for this study. Healthy volunteers (N=23) matched for age and gender were included as controls. Plasma samples were pooled to representing groups (D0 and D7, survivors and non-survivors, and controls), depleted, trypsin digested, labeled with iTRAQ, fractionated in 40 fractions and analyzed with LC-MS. Raw data was processed with mascot distiller, identified with 1% FDR using swissprot database and further analyzed by bioinformatics tools such as Ingenuity Pathway Analysis. Results We have identified 64 and 79 differentially expressed in survivors, and 68 and 75 proteins in non-survivors, at admission and after 7 days, respectively, compared to controls. Several proteins representing the organization and cellular movement were differentially expressed in all patients, such as KIF27, NF1, MYH9, MYO5B, ALMS1, SYNE1, ASPN (upregulated) and GSN, PLEC, PON1, F2, GCC2 (downregulated). We also identified several other differentially expressed proteins like ACTA1, ACTB, MYH13, MYO5A, MY07A, MYO9A DNAH5, DNAH8, DNAH10, DNAH11, DNAH5, DNAH10 and DNAH11 in the different setting of patients. We validated and changes in mononuclear cells using confocal microscopy and found higher expression of gelsolin and depletion of actin in survivor patients when compare to controls. Interestingly, we observed acute depletion of actin and gelsolin in non-survivors (Figure 1). Functional analysis data shows alteration in several biological functions, like inflammation, acute phase response, blood coagulation, homeostasis and defense response. Focusing in energy metabolism, we identified downregulated apolipoprotein family proteins like APOA1, APOA2, APOA4, APOB, APOC1, APOC2, APOC3, and APOE in septic patients. Changes in apolipoproteins and PON1, GSN, SAA1, LBP, MYO5A, and F2 resulted in alteration in fatty acid metabolism. Conclusion Our results show dysregulation of cytoskeleton and blood cell movement related proteins and alteration in lipid metabolism in septic patients. Further validation of these results is under progress.

Figure 1: Confocal microscopy image of PBMC in healthy volunteers and septic patients.