A proteomic study of sepsis

Introduction

Sepsis, characterized by intense reaction of the organism, due to infection, particularly of the inflammatory and coagulation systems, is a common illness, of high cost treatment and deaths. Thus, new technologies for the detection of sepsis early-stages are urgently needed. Our objective was to identify serum proteomic patterns that would distinguish sepsis patients from healthy controls.

Materials and methods

Six ICU patients that matched the criteria (ACCP/SCCM) for sepsis diagnosis were selected: male/female, 3/3; mean/range age of 81.1/68–89 years; mean LOS_ICU of 15 ± 25 days; 28-day mortality (dead/total/%) = 4/6/66%; mean APACHE II score of 22 (20–26); and mean SOFA = 6 (5–8); VAP (Pseudomonas aeruginosa, three patients) and CRBSI Staphylococcus aureus, Staphylococcus epidermidis, Candida albicans (one each). Serum samples were submitted to isoelectric focusing of proteins in the first dimension with a nonlinear strip of pH 3–10 and in the second to 12.5% 2D-SDS-PAGE. Gels were stained with Coomassie Blue and the final image analyzed using the Image Master Platinum software (Amersham). Selected spots were cut, washed, dried, swollen with trypsin solution and incubated for 16 hours at 37°C. The peptides were extracted, concentrated and analyzed by mass spectrometry in a MALDI-TOF Voyageur DE-PRO instrument (ABI). Peptide maps were processed (MS-Fit, Protein Prospector) for protein identification in the NCBI Data Bank.

Results

Image analyses of the gels showed the presence of spots only in patients with sepsis (Fig. 1, arrows) and the presence of other spots with different intensities in sepsis and controls. Peptide mass fingerprinting of some of these spots identified actin, zinc-finger protein, matrin 3, haptoglobin-2 and apolipoprotein A1.

Figure 1

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Discussion

Actin, a major component of the cytoskeleton, is found in cellular injury as in apoptosis; zinc-finger protein is expressed in cellular injury as a cytoprotective protein with anti-apoptosis activity; matrin 3 functions in transcription and interacts with other proteins of the cellular matrix; apolipoprotein A1 is capable of directly inactivating endotoxin (protein–endotoxin interaction); haptoglobin-2 is present during the acute phase reaction. These findings justify the use of proteomic techniques as a screening tool for the study of all stages of sepsis in high-risk patients.