Innovative biomarkers detecting spontaneous bacterial peritonitis in ascitic fluid of liver cirrhosis patients

Background:

Spontaneous bacterial peritonitis (SBP), which is defined as bacterial infection of ascitic fluid without any intra-abdominal, surgically treatable focus, is a severe complication in decompensated liver cirrhosis with a one-year mortality of up to 70%. Although SBP pathogenesis is not yet fully understood, bacterial translocation of intestinal bacteria from the gut to mesenteric lymph nodes and into ascitic fluid is regarded as the major underlying process. According to the German S3-Guideline “Ascites, spontaneous bacterial peritonitis, hepatorenal syndrome”, diagnosis of SBP is assured with detection of > 250 polymorphonuclear (PMN) cells per µl. Since antibiotic treatment is indicated at the very onset of SBP, innovative biomarkers for early SBP diagnosis are in great demand.

Methods:

To identify novel potential biomarkers for SBP, ascitic fluid was collected from patients suffering liver cirrhosis at the University Hospital of Regensburg. Protein levels of lactoferrin, C3a, IP-10, IL-6, IL-8 and IL-10 within ascitic fluids were determined by ELISA. Total protein content of ascitic fluid was measured by BCA protein quantification assays.

Results:

In total, 69 ascitic fluid samples from 41 individual patients were analyzed. The range of age in the cohort was between 36 and 77 years at date of paracentesis. 78% (32 patients) were male and 22% (9 patients) were female. Genesis of liver cirrhosis in this study was alcohol-toxic (58.5%), cryptogenic (19.5%), nutritiv-toxic (7.3%), viral (4.9%), autoimmune (2.4%), PBC (2.4%), non-alcoholic steatohepatitis (2,4%) and Budd Chiari Syndrome (2,4%). In 11.6% (8) of the samples diagnosis of SBP was confirmed with the definition of > 250 PMN/µl. The use of a PMN cell count of more than 100 PMN/µl ascitic fluid detected SBP in 13 samples (18,8%). Lactoferrin levels in ascitic fluids of SBP samples (mean 1006 ng/ml) were higher than in ascitic fluids without SBP (mean 75 ng/ml). Follow-up samples indicated that lactoferrin concentrations were elevated in early stages of SBP and coincided with the course of the disease and therapeutic response. C3a levels also correlated with the course of disease, but were reduced in infected ascitic fluids (mean 511 ng/ml) compared to non-SBP samples (mean 975 ng/ml). IP-10 was detected in all samples, but did not display any correlation with disease activity. Also relevant amounts of cytokines IL-6, IL-8 and IL-10 were detected in ascitic fluid of liver cirrhosis patients. However, an association of these parameters with early onset of SBP was not observed.

Conclusion:

Lactoferrin and C3a levels in ascitic fluid of liver cirrhosis patients correlate with the onset and course of SBP. Notably, a lower PMN-cutoff resulted in an enhanced specificity for SBP. High levels of other inflammatory molecules and cytokines detected in ascitic samples highlight the tremendous inflammatory activity in ascitic fluids of liver cirrhosis patients. Thus, these results put SBP in a novel perspective and provide options for earlier diagnosis and treatment of the disease.