Philipp Lutz

Impact of Rifaximin on the Frequency and Characteristics of Spontaneous Bacterial Peritonitis in Patients with Liver Cirrhosis and Ascites

Background Rifaximin is a non-absorbable antibiotic used to prevent relapses of hepatic encephalopathy which may also be a candidate for prophylaxis of spontaneous bacterial peritonitis (SBP). Aim To detect the impact of rifaximin on the occurrence and characteristics of SBP. Methods We prospectively studied all hospitalized patients that underwent a diagnostic paracentesis in our department from March 2012 to April 2013 for SBP and recorded all clinical data including type of SBP prophylaxis, prior use of rifaximin, concomitant complications of cirrhosis, as well as laboratory results and bacteriological findings. Patients were divided into the following three groups: no antibiotic prophylaxis, prophylaxis with rifaximin or with systemically absorbed antibiotic prophylaxis. Results Our study cohort comprised 152 patients with advanced liver cirrhosis, 32 of whom developed SBP during the study period. As expected, our study groups differed regarding a history of hepatic encephalopathy and SBP before inclusion into the study. None of the 17 patients on systemic antibiotic prophylaxis developed SBP while 8/27 patients on rifaximin and 24/108 without prophylaxis had SBP (p = 0.02 and p = 0.04 versus systemic antibiotics, respectively). In general, episodes of SBP were similar for patients treated with rifaximin and those without any prophylaxis. However, Escherichia coli and enterococci were dominant in the ascites of patients without any prophylaxis, while mostly klebsiella species were recovered from the ascites samples in the rifaximin group. Conclusion Rifaximin pretreatment did not lead to a reduction of SBP occurrence in hospitalized patients with advanced liver disease. However, the bacterial species causing SBP were changed by rifaximin.

Spontaneous bacterial peritonitis: The clinical challenge of a leaky gut and a cirrhotic liver.

Spontaneous bacterial peritonitis (SBP) is a frequent, life-threatening bacterial infection in patients with liver cirrhosis and ascites. Portal hypertension leads to increased bacterial translocation from the intestine. Failure to eliminate invading pathogens due to immune defects associated with advanced liver disease on the background of genetic predisposition may result in SBP. The efficacy of antibiotic treatment and prophylaxis has declined due to the spread of multi-resistant bacteria. Patients with nosocomial SBP and with prior antibiotic treatment are at a particularly high risk for infection with resistant bacteria. Therefore, it is important to adapt empirical treatment to these risk factors and to the local resistance profile. Rifaximin, an oral, non-absorbable antibiotic, has been proposed to prevent SBP, but may be useful only in a subset of patients. Since novel antibiotic classes are lacking, we have to develop prophylactic strategies which do not induce bacterial resistance. Farnesoid X receptor agonists may be a candidate, but so far, clinical studies are not available. New diagnostic tests which can be carried out quickly at the patients site and provide additional prognostic information would be helpful. Furthermore, we need tools to predict antibiotic resistance in order to tailor first-line antibiotic treatment of spontaneous bacterial peritonitis to the individual patient and to reduce mortality.

Regulatory CD4+ T cells modulate the interaction between NK cells and hepatic stellate cells by acting on either cell type.

BACKGROUND & AIMS NK cells regulate liver fibrosis by killing activated hepatic stellate cells (HSCs) and are controlled themselves by immune cells and/or soluble factors. Here, we analysed if CD4(+) regulatory T cells (Tregs) modify the interaction between NK cells and HSCs. METHODS The modification of NK cell activity against HSCs was studied in CD56(high)CD16(-) NK cells, using a flow cytometric CD107a degranulation assay and co-cultures with Tregs from healthy donors and patients with hepatitis C, respectively. We studied the underlying mechanisms in detail, applying Treg supernatants, Treg pretreated HSCs, and recombinant IL-8, TGF-ß1, and IL-10 as well as blocking experiments with neutralizing antibodies and analysed Treg-associated changes in the expression of NK cell receptor ligands on HSCs. RESULTS Tregs suppressed NK cell activation during HSC co-culture in a cell-contact-dependent manner involving the cytotoxic T-lymphocyte antigen 4 (CTLA-4). NK cell degranulation was further reduced, when HSCs had been pretreated with Tregs (p=0.043), Treg supernatants (p=0.001) or recombinant IL-8 (R=0.630, p=0.001) and TGF-ß1 (R=0.608, p=0.002), respectively. This additional inhibitory effect corresponded to the IL-8/TGF-ß1-mediated downregulation of MIC-A/B and HLA class-I on HSCs. Tregs from hepatitis C likewise inhibited NK cell activity, which was reversed significantly in specific blocking experiments. CONCLUSIONS Our data indicate that Tregs interfere with NK cell regulation of fibrogenesis via both direct cell-contact-dependent inhibition of NK cells and release of soluble factors, downregulating activating NK cell receptor ligands on HSCs. Our data may be particularly relevant for the intrahepatic accumulation of Tregs in chronic hepatitis C because downregulated NK cell activity against HSCs may blunt their control of fibrogenesis.

A farnesoid X receptor polymorphism predisposes to spontaneous bacterial peritonitis

BACKGROUND In mice, the farnesoid X receptor is involved in bacterial translocation, which can result in spontaneous bacterial peritonitis in patients with cirrhosis. We investigated if polymorphisms in the farnesoid X receptor gene influence the risk for spontaneous bacterial peritonitis. METHODS Laboratory and clinical data of 293 cirrhotic patients with ascites and 226 healthy controls were prospectively collected. The rs56163822, rs11110390 and rs12313471 polymorphisms of the farnesoid X receptor were determined. RESULTS 115 (39%) patients had spontaneous bacterial peritonitis. Distribution of all farnesoid X receptor genotypes matched the Hardy-Weinberg equilibrium. Patients with spontaneous bacterial peritonitis had a higher frequency of the rs56163822 GT genotype (7.0%) than patients without (1.7%, OR=4.4, p=0.02). This genotype was confirmed as predictor of spontaneous bacterial peritonitis by binary logistic regression analysis (OR=6.8, p=0.018). CONCLUSION The farnesoid X receptor rs56163822 GT genotype increases the risk for spontaneous bacterial peritonitis in cirrhotic patients with ascites.

The ratio of calprotectin to total protein as a diagnostic and prognostic marker for spontaneous bacterial peritonitis in patients with liver cirrhosis and ascites.

Abstract Background: Diagnosis of spontaneous bacterial peritonitis (SBP) is based on a differential ascites leukocyte count which does not provide prognostic information. We performed a pilot study to assess calprotectin in ascites as an alternative diagnostic and prognostic marker. Methods: We collected ascites from patients with liver cirrhosis from March 2012 to July 2013. Routine clinical and laboratory data of the patients were recorded. Ascites calprotectin levels were determined by ELISA. Results: Overall, we collected 120 ascites samples from 100 patients with liver cirrhosis and from eight patients with malignant peritoneal effusion as disease control. Samples without infection had significantly lower calprotectin levels (median 34 ng/mL, range 5–795) than SBP samples (median 928 ng/mL, range 21–110,480; p<0.001) and malignant effusions (median 401, range 47–2596 ng/mL; p<0.001). In non-infected ascites, calprotectin levels were higher in Child-Pugh stage B versus C (median 57 ng/mL vs. 17 ng/mL; p<0.001) and in alcoholic versus viral cirrhosis (median 37 ng/mL vs. 10 ng/mL; p=0.015). The ratio of ascites calprotectin to total protein was a better marker for SBP than calprotectin alone (AUROC=0.93; p<0.001; sensitivity 93%, specificity 79%; positive predictive value 60%; negative predictive value 97%). In addition, high levels of the ratio to total protein were associated with poor 30-day survival (p=0.042). Conclusions: The ratio of ascites calprotectin to total protein may be a promising new diagnostic and prognostic marker in patients with liver cirrhosis and SBP and should be evaluated further.

Influence of the CXCL1 rs4074 A Allele on Alcohol Induced Cirrhosis and HCC in Patients of European Descent

Background and Aims CXCL1 (CXC chemokine-ligand-1) is a ligand for CXC chemokine receptor 2 expressed on hepatic stellate cells (HSC). Thus, CXCL1 might contribute to HSC activation and fibrogenesis. In the present study, we investigated the influence of the CXCL1 rs4074 polymorphism on the occurrence of alcohol induced liver cirrhosis and hepatocellular carcinoma (HCC). Methods The study involved 458 patients with alcoholic cirrhosis (170 with HCC), 115 alcoholics without liver disease and 342 healthy controls. All subjects were genotyped for the CXCL1 rs4074 polymorphism and CXCL1 serum levels of 132 patients were measured. In vitro CXCL1 secretion in TLR-transfected cell lines were studied by ELISA. Results Distribution of the CXCL1 genotypes (GG/GA/AA) was 159/219/80 in patients with alcoholic cirrhosis, 52/44/19 in alcoholic controls and 158/140/44 in healthy controls. Patients with alcohol-induced cirrhosis were significantly more often carriers of the CXCL1 rs4074 A allele (65.3%) than alcoholics without liver disease (54.8%, OR=1.55; 95%CI=1.025-2.350; p=0.04) and healthy controls (53.8%, OR=1.62; 95%CI=1.212-2.151; p=0.001). Accordingly, the frequency of the CXCL1 rs4074 A allele was significantly higher in the cirrhotic patients than in the subjects without cirrhosis (41.4% vs. 33.9%, OR=1.38, 95% CI:1.14–1.66, p=0.001). Furthermore cirrhotic carriers of the CXCL1 rs4074 A allele had significantly higher CXCL1 serum levels than carriers of the GG genotype. In contrast to sera from healthy controls, sera from patients with alcoholic cirrhosis induced CXCL1 secretion in TLR2- (p=0.016) and TLR4- (p=0.008) transfected HEK293 cells. This finding indicates that sera from patients with alcoholic cirrhosis contain soluble ligands that can induce CXCL1 production via stimulation of TLRs. Conclusion The enhanced CXCL1 serum levels in carriers of the rs4074 A allele together with their increased frequency in patients with alcohol induced cirrhosis suggest the CXCL1 rs4074 A allele as a genetic risk factor for alcoholic cirrhosis.

Compartment-specific distribution of human intestinal innate lymphoid cells is altered in HIV patients under effective therapy

Innate lymphocyte cells (ILCs), a novel family of innate immune cells are considered to function as key orchestrators of immune defences at mucosal surfaces and to be crucial for maintaining an intact intestinal barrier. Accordingly, first data suggest depletion of ILCs to be involved in human immunodeficiency virus (HIV)-associated damage of the intestinal mucosa and subsequent microbial translocation. However, although ILCs are preferentially localized at mucosal surfaces, only little is known regarding distribution and function of ILCs in the human gastrointestinal tract. Here, we show that in HIV(-) individuals composition and functional capacity of intestinal ILCs is compartment-specific with group 1 ILCs representing the major fraction in the upper gastrointestinal (GI) tract, whereas ILC3 are the predominant population in ileum and colon, respectively. In addition, we present first data indicating that local cytokine concentrations, especially that of IL-7, might modulate composition of gut ILCs. Distribution of intestinal ILCs was significantly altered in HIV patients, who displayed decreased frequency of total ILCs in ileum and colon owing to reduced numbers of both CD127(+)ILC1 and ILC3. Of note, frequency of colonic ILC3 was inversely correlated with serum levels of I-FABP and sCD14, surrogate markers for loss of gut barrier integrity and microbial translocation, respectively. Both expression of the IL-7 receptor CD127 on ILCs as well as mucosal IL-7 mRNA levels were decreased in HIV(+) patients, especially in those parts of the GI tract with reduced ILC frequencies, suggesting that impaired IL-7 responses of ILCs might contribute to incomplete reconstitution of ILCs under effective anti-retroviral therapy. This is the first report comparing distribution and function of ILCs along the intestinal mucosa of the entire human gastrointestinal tract in HIV(+) and HIV(-) individuals.

Antibiotic resistance in healthcare-related and nosocomial spontaneous bacterial peritonitis.

Spontaneous bacterial peritonitis (SBP) can be life threatening in patients with liver cirrhosis. In contrast to community‐acquired SBP, no standard treatment has been established for healthcare‐related and nosocomial SBP.

A variant in the nuclear dot protein 52kDa gene increases the risk for spontaneous bacterial peritonitis in patients with alcoholic liver cirrhosis.

BACKGROUND Spontaneous bacterial peritonitis is frequently a fatal infection in patients with liver cirrhosis. We investigated if nuclear dot protein 52kDa (NDP52), a negative regulator of toll-like receptor (TLR) signalling and autophagy adaptor protein, might be involved. METHODS Two cohorts comprising 152 (derivation cohort) and 198 patients (validation cohort) with decompensated liver cirrhosis and 168 healthy controls were genotyped for the rs2303015 polymorphism in the NDP52 gene and prospectively followed-up for spontaneous bacterial peritonitis. RESULTS Overall, 57 (38%) patients in the derivation cohort and 77 (39%) in the validation cohort had spontaneous bacterial peritonitis. Cirrhosis was due to alcohol abuse in 57% of the derivation and 66% of the validation cohort. In patients with alcoholic cirrhosis, patients with spontaneous bacterial peritonitis had an increased frequency of the NDP52 rs2303015 minor variant in the derivation (p=0.04) and in the validation cohort (p=0.01). Multivariate analysis confirmed this minor variant (odds ratio 4.7, p=0.002) and the TLR2 -16934 TT variant (odds ratio 2.5, p=0.008) as risk factors for spontaneous bacterial peritonitis. In addition, presence of the NDP52 minor variant affected survival negatively. CONCLUSION Presence of the NDP52 rs2303015 minor variant increases the risk for spontaneous bacterial peritonitis in patients with alcoholic cirrhosis.

Hypoxia impairs anti-viral activity of natural killer (NK) cells but has little effect on anti-fibrotic NK cell functions in hepatitis C virus infection

BACKGROUND & AIMS Natural killer (NK) cells have been shown to exert anti-viral as well as anti-fibrotic functions in hepatitis C virus (HCV) infection. Previous studies, however, analyzed NK cell functions exclusively under atmospheric oxygen conditions despite the fact that the liver microenvironment is hypoxic. Here, we analyzed the effects of low oxygen tension on anti-viral and anti-fibrotic NK cell activity. METHODS Peripheral (n=34) and intrahepatic (n=15) NK cells from HCV(+) patients as well as circulating NK cells from healthy donors (n=20) were studied with respect to anti-viral and anti-fibrotic activity via co-culture experiments with HuH7 replicon cells and hepatic stellate cells, respectively. RESULTS Anti-viral activity of resting NK cells from healthy controls was not affected by hypoxia. However, hypoxia significantly reduced the response of healthy NK cells to cytokine stimulation. In contrast to healthy controls, we observed resting and cytokine activated peripheral NK cells from HCV patients to display a significantly decreased anti-viral activity when cultured at 5% or 1% oxygen, suggesting HCV NK cells to be very sensitive to hypoxia. These findings could be confirmed when intrahepatic NK cells were tested. Finally, we show that anti-fibrotic NK cell activity was not affected by low oxygen tension. CONCLUSIONS Our results show that anti-viral function of NK cells from HCV(+) patients is critically affected by a hypoxic microenvironment and, therefore, indicate that in order to obtain an accurate understanding of intrahepatic NK cell anti-HCV activity, the laboratory modelling should take into account the liver specific levels of oxygen.