Andreas Wannhoff

Reduction in alkaline phosphatase is associated with longer survival in primary sclerosing cholangitis, independent of dominant stenosis

Alkaline phosphatase (ALP) is an important serum marker in primary sclerosing cholangitis (PSC). Patients with obstruction of the large bile ducts due to dominant strictures (DS) are a special, clinically important phenotype.

FUT2 and FUT3 genotype determines CA19-9 cut-off values for detection of cholangiocarcinoma in patients with primary sclerosing cholangitis.

BACKGROUND & AIMS Allelic variants of fucosyltransferases 2 and 3 (FUT2/3) influence serum levels of CA19-9, a screening parameter commonly used for detection of biliary malignancy in PSC. We aimed at improving diagnostic accuracy of CA19-9 by determining the impact of FUT2/3 genotypes. METHODS CA19-9 levels were measured in 433 PSC patients, 41 of whom had biliary malignancy. Genotypes for FUT3 and FUT2 were used to assign patients to one of three groups: A, no FUT3 activity regardless of FUT2 activity; B, both FUT2 and FUT3 activity and C, no FUT2 activity without loss of FUT3 activity. Group-specific cut-off values were determined by Youdens index. RESULTS The median CA19-9 values of cancer-free patients were significantly different (p<0.001) in Groups A (2.0U/ml), B (17.0U/ml), and C (37.0U/ml). Biliary malignancy patients in Groups B and C had significantly higher CA19-9 values than cancer-free patients (p<0.001). The optimal cut-off, as determined by ROC analysis, for all patients was 88.5U/ml. Optimal cut-off values in Groups A, B, and C were 4.0U/ml, 74.5U/ml, and 106.8U/ml, respectively. Use of these values improved sensitivity of CA19-9 in Groups B and C. Further, use of group-dependent cut-off values with 90% sensitivity resulted in a 42.9% reduction of false positive results. CONCLUSIONS Use of FUT2/3 genotype-dependent cut-off values for CA19-9 improved sensitivity and reduced the number of false positive results.

PNPLA3 in end-stage liver disease: Alcohol consumption, hepatocellular carcinoma development, and transplantation-free survival

The rs738409 variant (I148M) of the patatin‐like phospholipase domain‐containing protein 3 (PNPLA3) gene is associated with several liver malfunctions. Its impact on end‐stage liver disease has not been addressed yet.

Fut2 genotype is a risk factor for dominant stenosis and biliary candida infections in primary sclerosing cholangitis

A recent genome‐wide association study identified the FUT2 secretor status and genotype defined by the single‐nucleotide polymorphism rs601338 as potential genetic risk factor in primary sclerosing cholangitis (PSC), which significantly influences biliary bacterial composition.

Increased levels of rivaroxaban in patients after liver transplantation treated with cyclosporine A.

Calcineurin inhibitors (CNIs) form the backbone of immunosuppression in solid organ transplantation, but are also frequently used in patients with autoimmune diseases. It is important to understand drug-drug interactions in transplant patients treated with cyclosporine A (CsA) or tacrolimus (Tac), but no data on the interactions between these CNIs and new oral anticoagulants are available. We investigated differences in rivaroxaban blood levels in liver transplant patients treated with CsA or Tac. Trough levels of rivaroxaban (determined by a chromogenic anti-Xa activity assay; Haemochrom Diagnostica, Germany) were evaluated in patients after liver transplantation treated with CNIs, and results were compared between the CsA and Tac groups. Bleeding events and thrombembolic events were retrospectively evaluated. The Mann-Whitney U test and Fisher’s exact test were used for statistical analysis. Statistical analyses were performed using IBM Statistical Package for the Social Sciences version 21 (IBM Corp, Armonk, NY). All patients provided written informed consent, and the study was previously approved by the local ethics committee. Overall, nine patients were included, five in the CsA group and four in the Tac group. Reasons for anticoagulant therapy included atrial fibrillation (n=2), pulmonary embolism (n=2), peripheral vein thrombosis (n=1), portal vein thrombosis (n=1), liver vein thrombosis (n=2), and hepatic artery thrombosis (n=1). Demographic and medical characteristics of the patients are summarized in Table 1. Mean (T standard deviation) for rivaroxaban trough level was 131.7 ng/mL (T119.5) in patients treated with CsA compared to 20.3 ng/mL (T14.4) in the Tac group (P=0.014), even though one patient in the CsA group received a lower rivaroxaban dose than recommended. Furthermore, in the CsA group, in three of five patients with initially increased rivaroxaban level (therapeutic range, 7Y65 ng/mL; internal reference range, adopted from (1)), the dose was reduced by 5 mg, resulting in trough levels within the therapeutic range. No dose reductions were necessary in the Tac group (P=0.164). Median time between

Risk factors and outcome in patients with primary sclerosing cholangitis with persistent biliary candidiasis

BackgroundCandidiasis is commonly observed in patients with primary sclerosing cholangitis (PSC), but the clinical risk factors associated with its presence have not been fully investigated. In this study, we aimed to analyse the incidence, risk factors, and transplantation-free survival in primary sclerosing cholangitis (PSC) patients with persistent biliary candidiasis.MethodsWe retrospectively analysed patients diagnosed with PSC who were admitted to our department during 2002 to 2012. One-hundred fifty patients whose bile cultures were tested for fungal species were selected, and their clinical and laboratory parameters were investigated. The results of endoscopic retrograde cholangiography (ERC) and bile cultures were analysed using chart reviews. The cases of biliary candidiasis were sub-classified as transient or persistent.ResultsThirty out of 150 (20.0%) patients had biliary candidiasis. Although all patients demonstrated comparable baseline characteristics, those with biliary candidiasis showed significantly reduced transplantation-free survival (p < 0.0001) along with a markedly elevated frequency of cholangiocarcinoma (CCA) (p = 0.04). The patients were further sub-classified according to the transient (15/30) or persistent (15/30) nature of their biliary candidiasis. A subgroup analysis showed reduced survival with a greater necessity for orthotopic liver transplantation (OLT) only in patients with persistence of Candida (p = 0.007). The survival in the patients with transient biliary candidiasis was comparable to that in candidiasis-free patients. In a multivariate regression analysis that included Mayo risk score (MRS), sex, age, dominant stenosis, inflammatory bowel disease, autoimmune hepatitis overlap syndrome, and number of times ERC was performed, biliary candidiasis was an independent risk factor for reduced survival (p = 0.008). Risk factors associated with acquisition of biliary candidiasis were age at PSC diagnosis and number of ERCs.ConclusionsThe persistence of biliary candidiasis is associated with markedly reduced transplantation-free survival in PSC patients. By contrast, actuarial survival in patients with transient biliary candidiasis approaches that for patients without any evidence of biliary candidiasis. Further studies on the treatment of persistent biliary candidiasis in patients with PSC are warranted.

Plasma membrane phospholipase A2 controls hepatocellular fatty acid uptake and is responsive to pharmacological modulation: implications for nonalcoholic steatohepatitis

Excess hepatic fat accumulation leads to nonalcoholic steatohepatitis (NASH), a serious threat to health for which no effective treatment is available. However, the mechanism responsible for fatty acid uptake by hepatocytes remains unclear. Using the human hepatocyte‐derived tumor cell line HepG2, we found that fatty acid influx is mediated by a heterotetrameric plasma membrane protein complex consisting of plasma membrane fatty acid‐binding protein, caveolin‐1, CD36, and calcium‐independent membrane phospholipase A2 (iPLA2β). Blocking iPLA2β with the bile acid‐phospholipid conjugate ursodeoxycholate‐lysophosphatidylethanolamide (UDCA‐LPE) caused the dissociation of the complex, thereby inhibiting fatty acid influx (IC50 47 μM), and suppressed the synthesis of all subunits through a reduction in lysophosphatidylcholine from 8.0 to 3.5 μmol/mg of protein and corresponding depletion of phosphorylated c‐Jun N‐terminal kinase. These findings were substantiated by an observed 56.5% decrease in fatty acid influx in isolated hepatocytes derived from iPLA2β‐knockout mice. Moreover, steatosis and inflammation were abrogated by UDCA‐LPE treatment in a cellular model of NASH. Thus, iPLA2β acts as an upstream checkpoint for mechanisms that regulate fatty acid uptake, and its inhibition by UDCA‐LPE qualifies this nontoxic compound as a therapeutic candidate for the treatment of NASH.—Stremmel, W., Staffer, S., Wannhoff, A., Pathil, A., Chamulitrat, W. Plasma membrane phospholipase A2 controls hepatocellular fatty acid uptake and is responsive to pharmacological modulation: implications for nonalcoholic steatohepatitis. FASEB J. 28, 3159–3170 (2014). www.fasebj.org

Cardiac volume overload and pulmonary hypertension in long‐term follow‐up of patients with a transjugular intrahepatic portosystemic shunt

Transjugular intrahepatic portosystemic shunt (TIPSS) cause haemodynamic changes in patients with cirrhosis, yet little is known about long‐term cardiopulmonary outcomes.

CD14 is associated with biliary stricture formation.

The pathogenesis of intrahepatic biliary stricture formation in patients with primary sclerosing cholangitis (PSC) or after liver transplantation (LTx) remains elusive. CD14 receptor signaling is a key mediator of the innate immune system; its common genetic variant is associated with alcoholic liver disease. PSC and LTx cohort patients and primary biliary cirrhosis (PBC) control patients were genotyped for the CD14 ‐260C>T (rs2569190) polymorphism, and genotypes were correlated with long‐term clinical outcome. Biliary tissue, bile, and whole blood of PSC patients and healthy controls were screened for markers of the innate immune system and bacterial infection. In 121 PSC patients, the CD14 ‐260C>T genotype was associated with development of dominant bile duct strictures (P = 0.02). In 365 LTx patients, TT carriers (4.1%) were protected against the formation of nonanastomotic biliary strictures versus CC/CT patients (12.6%; P = 0.01). Chemokine ligand 8 (P = 0.04) and chemokine receptor 6 (P = 0.004) were up‐regulated in biliary tissue of PSC patients with the TT versus the CC/CT genotype. Lipopolysaccharide whole‐blood stimulation resulted in a significant change in interleukin (IL)‐8 (P = 0.05) and IL‐12p40 levels (P = 0.04) in healthy control subjects carrying the TT genotype. TT PSC patients were protected against Gram‐negative bacterial biliary infection (TT: 0% vs. CC/CT: 22.5%; P = 0.02). Serum‐soluble CD14 levels correlated with the CD14 ‐260C>T genotype (P = 0.02), representing an independent risk indicator of survival in PSC patients (hazard ratio, 0.40; 95% confidence interval, 0.19‐0.86; P =0.01). Conclusions: The function of the innate immune response by CD14 is crucial during biliary infection and stricture formation. The benefits of CD14 signaling modification should be addressed in future studies. (Hepatology 2016;64:843‐852)

A common genetic variant of fucosyltransferase 2 correlates with serum carcinoembryonic antigen levels and affects cancer screening in patients with primary sclerosing cholangitis.

Background Primary sclerosing cholangitis (PSC) patients are at increased risk of biliary tract cancer, and carcinoembryonic antigen (CEA) serum levels might be used for screening. Objective To examine cancer screening with CEA in PSC patients and analyse how serum CEA levels are affected by genetic variants of fucosyltransferase (FUT) 2 and 3. Methods In a retrospective cohort analysis we evaluated CEA levels in 226 PSC patients, including 19 with biliary malignancy, and investigated how FUT2 and FUT3 SNPs affected CEA levels. Receiver-operating-characteristic (ROC) analysis was performed and cut-off values were determined based on Youden’s index. A control cohort contained 240 patients, including 28 with biliary malignancy. Results Median CEA concentration was lower in cancer-free patients (1.4 ng/mL) than in cancer patients (2.0 ng/mL, P = 0.014). ROC analysis revealed an area under the curve (AUC) of 0.671, the optimal cut-off was 3.2 ng/mL. The FUT2 variant rs601338 (G428A) correlated with CEA levels, and the effect was most prominent in a subgroup of patients genetically incapable of expressing CA19-9. The AUC improved if ROC analysis was performed separately for wild-type (AUC: 0.731) and homozygous mutant (AUC: 0.816) G428A. The influence of FUT2 on CEA was confirmed in the control cohort. Conclusions CEA is interesting for biliary-malignancy screening in PSC patients, especially in patients who do not express CA19-9. This is the first study to show that the combined use of CEA measurement and FUT genotyping is clinically beneficial and that it might enhance the early detection of biliary malignancy in clinical practice. This approach could also be effective when screening for other common gastrointestinal malignancies.