Tamas Tornai

High prevalence of IgA class anti-neutrophil cytoplasmic antibodies (ANCA) is associated with increased risk of bacterial infection in patients with cirrhosis

BACKGROUND & AIMS Anti-neutrophil cytoplasmic antibodies (ANCA) are a non-uniform family of antibodies recognizing diverse components of neutrophil granulocytes. ANCA formation might be induced by protracted bacterial infections or probably reflect an abnormal immune response to commensal microorganisms. Bacterial infections are common complications in cirrhosis with high incidence of episodes caused by enteric organisms, therefore, we sought to study the presence and clinical importance of ANCA in cirrhosis. METHODS Sera of 385 patients with cirrhosis of different etiologies were assayed for ANCA of IgG, IgA, IgA1, IgA2, and secretory IgA subtypes by indirect immunofluorescence and ELISAs. The control group comprised 202 patients with chronic liver diseases without cirrhosis and 100 healthy subjects. In cirrhosis, a 2-year follow-up, observational study was conducted to assess a possible association between the presence of ANCA and clinically significant bacterial infections. RESULTS Prevalence of ANCA IgA was significantly higher in cirrhosis (52.2%) compared to chronic liver diseases (18.6%) or healthy controls (0%, p<0.001 for both). ANCA IgA subtyping assays revealed marked increase in the proportion of IgA2 subtype (46% of total ANCA IgA) and presence of the secretory component concurrently. Presence of ANCA IgA was associated with disease-specific clinical characteristics (Child-Pugh stage and presence of ascites, p<0.001). During a 2-year follow-up period, risk of infections was higher among patients with ANCA IgA compared to those without (41.8% vs. 23.4%, p<0.001). ANCA IgA positivity was associated with a shorter time to the first infectious complication (pLogRank <0.001) in Kaplan-Meier analysis and was identified as an independent predictor in multivariate Cox-regression analysis (HR:1.74, 95% CI: 1.18-2.56, p=0.006). CONCLUSIONS Presence of IgA type ANCA is common in cirrhosis. Involvement of gut mucosal immune system is in center of their formation and probably reflects sustained exposure to bacterial constituents.

Rediscovery of the Anti-Pancreatic Antibodies and Evaluation of their Prognostic Value in a Prospective Clinical Cohort of Crohn’s Patients: The Importance of Specific Target Antigens [GP2 and CUZD1]

BACKGROUNDS Glycoprotein 2[GP2] and CUB zona pellucida-like domain 1[CUZD1] belong to protein families involved in gut innate immunity processes and have recently been identified as specific targets of anti-pancreatic autoantibodies [PAbs] in Crohns disease[CD]. We aimed to determine the prognostic potential of novel target-specific PAbs regarding long-term disease course of an adult CD patient cohort. METHODS Sera of 458 consecutive well-characterised IBD patients from a single referral IBD centre were tested by enzyme-linked immunosorbent assay [ELISA] with isoform 4 of recombinant GP2 [anti-MZGP2 and anti-GP2 IgA/IgG] and indirect immunofluorescence test [IIFT] system with GP2 and CUZD1 expressing transfected HEK 293 cells [anti-rPAg2 and rPAg1 IgA/IgG]. Clinical data were available on complicated disease or surgical interventions as well as disease activity and medical treatment during the prospective follow-up [median, 108 months]. RESULTS Totals of 12.4% and 20.8% of CD patients were positive for IgA/IgG type of anti-GP2 and anti-CUZD1, respectively, with a significant difference compared with UC [p < 0.01]. Antibody status was stable over time. Agreement among three different anti-GP2 assays was good. Positivity for PAbs, mainly IgA subtypes, predicted a faster progression towards complicated disease course. In Kaplan-Meier analysis, time to surgery or development of perianal disease was associated with anti-GP2 IgA [pLogRank < 0.01] or anti-CUZD1 IgA [pLogRank < 0.001] positivity, respectively. Anti-CUZD1 IgA remained an independent predictor in the multivariate Cox-regression model (hazard ratio [HR]: 3.43, 95% confidence interval [CI]: 1.68-7.02, p < 0.001). CONCLUSIONS The present study has shown that specific PAbs [especially IgA subtype] predict complicated disease course including the development of perianal disease in CD.

Macrophage activation marker, soluble CD163 is an independent predictor of short-term mortality in patients with cirrhosis and bacterial infection

Innate immune system dysfunction is common in advanced cirrhosis, with a central role of the monocyte/macrophage system. Monocytes and macrophages express the scavenger receptor CD163, which is regulated by inflammatory mediators. Cleavage of the receptor leads to the formation of soluble (s)CD163 that represents an anti‐inflammatory response. We aimed to study the clinical importance of sCD163 in cirrhosis.

Lectin‐complement pathway molecules are decreased in patients with cirrhosis and constitute the risk of bacterial infections

Lectin pathway molecules of the complement system are synthesized by hepatocytes and have pivotal role in innate host defence against infectious organisms. Ficolins (FCNs) act as soluble pattern recognition molecules, while mannan‐binding lectin serine proteases(MASPs) do as effector molecules in elimination of pathogens. We aimed to study the significance of low level of these molecules in the development of cirrhosis‐associated bacterial infections, which has not been elucidated so far.

Gut barrier failure biomarkers are associated with poor disease outcome in patients with primary sclerosing cholangitis

AIM To assess the prevalence of a panel of serologic markers that reflect gut barrier dysfunction in a mixed cohort of pediatric and adult primary sclerosing cholangitis (PSC) patients. METHODS Sera of 67 PSC patients [median age (range): 32 (5-79) years, concomitant IBD: 67% and cirrhosis: 20%] were assayed for the presence of antibodies against to F-actin (AAA IgA/IgG) and gliadin (AGA IgA/IgG)] and for serum level of intestinal fatty acid-binding protein (I-FABP) by ELISA. Markers of lipopolysaccharide (LPS) exposure [LPS binding protein (LBP)] and various anti-microbial antibodies [anti-OMP Plus IgA and endotoxin core IgA antibody (EndoCAb)] were also determined. Poor disease outcome was defined as orthotopic liver transplantation and/or liver-related death during the follow-up [median: 99 (14-106) mo]. One hundred and fifty-three healthy subjects (HCONT) and 172 ulcerative colitis (UC) patients were the controls. RESULTS A total of 28.4%, 28.0%, 9% and 20.9% of PSC patients were positive for AAA IgA, AAA IgG, AGA IgA and AGA IgG, respectively. Frequencies of AAA IgA and AAA IgG (P < 0.001, for both) and AGA IgG (P = 0.01, for both) but not AGA IgA were significantly higher compared to both of the HCONT and the UC groups. In survival analysis, AAA IgA-positivity was revealed as an independent predictor of poor disease outcome after adjusting either for the presence of cirrhosis [HR = 5.15 (1.27-20.86), P = 0.022 or for the Mayo risk score (HR = 4.24 (0.99-18.21), P = 0.052]. AAA IgA-positivity was significantly associated with higher frequency of anti-microbial antibodies (P < 0.001 for EndoCab IgA and P = 0.012 for anti-OMP Plus IgA) and higher level of the enterocyte damage marker (median I-FABPAAA IgA pos vs neg: 365 vs 166 pg/mL, P = 0.011), but not with serum LBP level. CONCLUSION Presence of IgA type AAA identified PSC patients with progressive disease. Moreover, it is associated with enhanced mucosal immune response to various microbial antigens and enterocyte damage further highlighting the importance of the gut-liver interaction in PSC.

Presepsin teardown - Pitfalls of biomarkers in the diagnosis and prognosis of bacterial infection in cirrhosis

AIM To evaluate the diagnostic and prognostic value of presepsin in cirrhosis-associated bacterial infections. METHODS Two hundred and sixteen patients with cirrhosis were enrolled. At admission, the presence of bacterial infections and level of plasma presepsin, serum C-reactive protein (CRP) and procalcitonin (PCT) were evaluated. Patients were followed for three months to assess the possible association between presepsin level and short-term mortality. RESULTS Present 34.7 of patients had bacterial infection. Presepsin levels were significantly higher in patients with infection than without (median, 1002 pg/mL vs 477 pg/mL, P < 0.001), increasing with the severity of infection [organ failure (OF): Yes vs No, 2358 pg/mL vs 710 pg/mL, P < 0.001]. Diagnostic accuracy of presepsin for severe infections was similar to PCT and superior to CRP (AUC-ROC: 0.85, 0.85 and 0.66, respectively, P = NS for presepsin vs PCT and P < 0.01 for presepsin vs CRP). At the optimal cut-off value of presepsin > 1206 pg/mL sensitivity, specificity, positive predictive values and negative predictive values were as follows: 87.5%, 74.5%, 61.8% and 92.7%. The accuracy of presepsin, however, decreased in advanced stage of the disease or in the presence of renal failure, most probably because of the significantly elevated presepsin levels in non-infected patients. 28-d mortality rate was higher among patients with > 1277 pg/mL compared to those with ≤ 1277 pg/mL (46.9% vs 11.6%, P < 0.001). In a binary logistic regression analysis, however, only PCT (OR = 1.81, 95%CI: 1.09-3.01, P = 0.022) but neither presepsin nor CRP were independent risk factor for 28-d mortality after adjusting with MELD score and leukocyte count. CONCLUSION Presepsin is a valuable new biomarker for defining severe infections in cirrhosis, proving same efficacy as PCT. However, it is not a useful marker of short-term mortality.

Loss of tolerance to gut immunity protein, glycoprotein 2 (GP2) is associated with progressive disease course in primary sclerosing cholangitis

Glycoprotein 2[GP2] is a specific target of pancreatic autoantibodies[PAbs] in Crohn’s disease(CD) and is involved in gut innate immunity processes. Our aim was to evaluate the prevalence and prognostic potential of PAbs in primary sclerosing cholangitis(PSC). Sixty-five PSC patients were tested for PAbs by indirect immunofluorescence and compared with healthy (n = 100) and chronic liver disease controls(CLD, n = 488). Additionally, a panel of anti-microbial antibodies and secretory (s)IgA levels were measured, as markers of bacterial translocation and immune dysregulation. PAbs were more frequent in PSC(46.2%) compared to controls(healthy:0% and CLD:4.5%), [P < 0.001, for each]. Occurrence of anti-GP2 antibody was 30.8% (20/65) and was exclusively of IgA isotype. Anti-GP2 IgA positive patients had higher sIgA levels (P = 0.021). With flow-cytometry, 68.4% (13/19) of anti-GP2 IgA antibodies were bound with secretory component, suggesting an active retro-transportation of anti-GP2 from the gut lumen to the mucosa. Anti-GP2 IgA was associated with shorter transplant-free survival [PLogRank < 0.01] during the prospective follow-up (median, IQR: 87 [9–99] months) and remained an independent predictor after adjusting for Mayo risk score(HR: 4.69 [1.05–21.04], P = 0.043). These results highlight the significance of gut-liver interactions in PSC. Anti-GP2 IgA might be a valuable tool for risk stratification in PSC and considered as a potential therapeutic target.

High versus low energy administration in the early phase of acute pancreatitis (GOULASH trial): protocol of a multicentre randomised double-blind clinical trial

Introduction Acute pancreatitis (AP) is an inflammatory disease with no specific treatment. Mitochondrial injury followed by ATP depletion in both acinar and ductal cells is a recently discovered early event in its pathogenesis. Importantly, preclinical research has shown that intracellular ATP delivery restores the physiological function of the cells and protects from cell injury, suggesting that restoration of energy levels in the pancreas is therapeutically beneficial. Despite several high quality experimental observations in this area, no randomised trials have been conducted to date to address the requirements for energy intake in the early phase of AP. Methods/design This is a randomised controlled two-arm double-blind multicentre trial. Patients with AP will be randomly assigned to groups A (30 kcal/kg/day energy administration starting within 24 hours of hospital admission) or B (low energy administration during the first 72 hours of hospital admission). Energy will be delivered by nasoenteric tube feeding with additional intravenous glucose supplementation or total parenteral nutrition if necessary. A combination of multiorgan failure for more than 48 hours and mortality is defined as the primary endpoint, whereas several secondary endpoints such as length of hospitalisation or pain will be determined to elucidate more detailed differences between the groups. The general feasibility, safety and quality checks required for high quality evidence will be adhered to. Ethics and dissemination The study has been approved by the relevant organisation, the Scientific and Research Ethics Committee of the Hungarian Medical Research Council (55961-2/2016/EKU). This study will provide evidence as to whether early high energy nutritional support is beneficial in the clinical management of AP. The results of this trial will be published in an open access way and disseminated among medical doctors. Trial registration The trial has been registered at the ISRCTN (ISRTCN 63827758).

Major changes of von Willebrand factor multimer distribution in cirrhotic patients with stable disease or acute decompensation

Background  There is an unstable balance between pro- and anti-haemostatic processes in patients with cirrhosis. We hypothesized, that in patients with acute decompensation (AD) the major alterations of von Willebrand factor (VWF) could contribute to the pro-thrombotic situation as compared to patients with stable (ST) cirrhosis. Patients and Methods  We analysed different parameters of VWF, including detailed multimer distribution by densitometry and platelet adhesion, together with a d isintegrin-like a nd m etalloproteinase with t hrombo s pondin type-1 motifs 13 (ADAMTS13) activity and antigen and C-reactive protein (CRP) levels in patients with ST cirrhosis ( n  = 99), with AD ( n  = 54) and controls ( n  = 92). Results  VWF antigen, ristocetin co-factor as well as collagen-binding activities were elevated in both cirrhotic groups in a stepwise manner. There was a decrease in high and an increase in low molecular weight multimer ratios in the majority of ST cirrhosis. However, in 24 out of 54 AD patients, ultra-large VWF multimers (ultra-large molecular weight multimers [ULMWM]) were found. ADAMTS13 activity in ST and AD patients without ULMWM was similar to controls (median [interquartile range; IQR]%: 98 [67–132] and 91 [60–110] vs. 106 [88–117], respectively). The presence of ULMWM in AD patients was associated with low ADAMTS13 activity [33 (24–49)%] and high CRP level [23 (7.1–83.6) mg/L]. Adhesion of normal platelets showed a stepwise increase in the presence of cirrhotic plasmas, reaching the highest level in AD patients with ULMWM. Conclusion  Characteristic changes of VWF parameters are seen in ST cirrhosis. In AD patients, highly increased VWF and reduced ADAMTS13 activity could be found, along with the presence of ULMWM, which are possible markers and contributors of the disease progression.

Significance of serological markers in the disease course of ulcerative colitis in a prospective clinical cohort of patients

Background & aims To determine the prognostic potential of classic and novel serologic antibodies regarding unfavorable disease course in a prospective ulcerative colitis (UC) patient cohort, since few and conflicting data are available in the literature regarding this matter. Methods 187 consecutive patients were studied prospectively (median follow-up: 135 months) from a single referral IBD center in Hungary. Sera were tested for different IgA/IgG type autoantibodies (anti-neutrophil cytoplasmic [ANCA], anti-DNA-bound-lactoferrin [anti-LFS], anti-goblet cell [anti-GAB] and anti-pancreatic [PAB: anti-CUZD1 and anti-GP2)]) by indirect immunofluorescence technique and for anti-microbial (anti-Saccharomyces cerevisiae [ASCA] IgG/IgA and anti-OMP Plus™ IgA) antibodies by enzyme-linked immunosorbent assays. Results A total of 73.6%, 62.4% and 11.2% of UC patients were positive for IgA/IgG type of atypical perinuclear-ANCA, anti-LFS and anti-GAB, respectively. Occurrences of PABs were 9.6%, while ASCA IgA/IgG and anti-OMP IgA were 17.6% and 19.8%, respectively. Antibody status was stable over time. IgA type PABs were more prevalent in patients with primary sclerosing cholangitis (37.5% vs. 4.7% for anti-CUZD1 and 12.5% vs. 0% for anti-GP2, p<0.001 for both). IgA type ASCA and anti-CUZD1 antibodies were associated with higher risk of requirement for long-term immunosuppressant therapy in Kaplan-Meier analysis (pLogRank <0.01 for both). However, in multivariate Cox-regression analysis only ASCA IgA (HR: 2.74, 95%CI: 1.46–5.14, p<0.01) remained independent predictor. UC-related hospitalization due to disease activity was only associated with multiple antibody positivity (for 3 or more; HR 2.03 [95% CI: 1.16–3.56]; p = 0.013). None of the individual antibodies or their combination was associated with the risk of development of extensive disease or colectomy. Conclusion Even with low prevalence rates, present study gives further evidence to the role of certain antibodies as markers for distinct phenotype and disease outcome in UC. Considering the result of the multivariate analysis the novel antibodies investigated do not seem to be associated with poor clinical outcome in UC, only a classic antibody, IgA subtype ASCA remained an independent predictor of long-term immunosuppressive therapy.