Medical care of patients with Wilson disease in Germany: a multidisciplinary survey among university centers.

BACKGROUND: Wilson disease (WD) is a rare, hereditary disorder of copper metabolism. Due to its variable symptoms and manifestations, diagnosis remains challenging. Affected patients must obtain lifelong medical treatment, as the disease is fatal if untreated. Patients require continuous monitoring, but little is known about the care of these patients in Germany. Therefore, we analyzed the medical care of WD patients at German university centers. We sent a questionnaire containing 20 questions to a total of 108 departments of pediatrics, neurology and gastroenterology in 36 university hospitals. Our questions referred to the characteristics of WD patients at the different sites and internal procedures regarding diagnosis, therapy and follow-up. A descriptive statistical analysis was performed. RESULTS: Sixty-three departments (58%) returned our questionnaire. In total, approximately one-third of the estimated WD patients in Germany are seen annually in the outpatient clinics of these departments (approx. 950 patients). There are only a few departments which treat patients in a multidisciplinary setting (12%). Our survey revealed that for diagnosis, 51% of all departments used an algorithm based on the Leipzig score as recommended by international guidelines. Most departments apply essential parameters recommended by WD guidelines. Routine monitoring is performed at least biannually by 84% of the departments, and standard investigations for monitoring are regularly applied. A routine family screening is performed by 84% of all departments. A reduction in medical therapy during pregnancy is recommended by 46% of the departments. Only 14% suggested that WD patients should not breastfeed. Liver transplantation (LT) due to WD is a rare but repeatedly occurring event. Most departments of gastroenterology (72%) reported at least one patient with LT within the last decade. CONCLUSIONS: Medical care of WD patients at German university centers follows the recommendations set forth by international guidelines, but only a few centers treat significant numbers of patients. The surveillance of patients does not follow specified standards, but most departments adhere to the accepted guidelines. The formation of central units and networks in a multidisciplinary setting should be evaluated to improve the care of WD patients.

Hydrophobic Bile Salts Induce Pro-Fibrogenic Proliferation of Hepatic Stellate Cells through PI3K p110 Alpha Signaling.

Bile salts accumulating during cholestatic liver disease are believed to promote liver fibrosis. We have recently shown that chenodeoxycholate (CDC) induces expansion of hepatic stellate cells (HSCs) in vivo, thereby promoting liver fibrosis. Mechanisms underlying bile salt-induced fibrogenesis remain elusive. We aimed to characterize the effects of different bile salts on HSC biology and investigated underlying signaling pathways. Murine HSCs (mHSCs) were stimulated with hydrophilic and hydrophobic bile salts. Proliferation, cell mass, collagen deposition, and activation of signaling pathways were determined. Activation of the human HSC cell line LX 2 was assessed by quantification of α-smooth muscle actin (αSMA) expression. Phosphatidyl-inositol-3-kinase (PI3K)-dependent signaling was inhibited both pharmacologically and by siRNA. CDC, the most abundant bile salt accumulating in human cholestasis, but no other bile salt tested, induced Protein kinase B (PKB) phosphorylation and promoted HSC proliferation and subsequent collagen deposition. Pharmacological inhibition of the upstream target PI3K-inhibited activation of PKB and pro-fibrogenic proliferation of HSCs. The PI3K p110α-specific inhibitor Alpelisib and siRNA-mediated knockdown of p110α ameliorated pro-fibrogenic activation of mHSC and LX 2 cells, respectively. In summary, pro-fibrogenic signaling in mHSCs is selectively induced by CDC. PI3K p110α may be a potential therapeutic target for the inhibition of bile salt-induced fibrogenesis in cholestasis.

Multidisciplinary lifestyle intervention is associated with improvements in liver damage and in surrogate scores of NAFLD and liver fibrosis in morbidly obese patients.

PURPOSE: Non-alcoholic fatty liver disease (NAFLD) is the hepatic manifestation of the metabolic syndrome. Particularly morbidly obese patients are at risk of developing progressive liver disease. Nutritional and lifestyle intervention is recommended as the standard of care in NAFLD. However, there is a striking lack of evidence to support the efficacy of lifestyle intervention to treat NAFLD in morbidly obese patients. Here, we aimed to assess the impact of lifestyle intervention on NAFLD in the morbidly obese in a real-world setting. METHODS: 136 obese patients were included in an industry-independent, multiprofessional lifestyle intervention program with a lead-in phase of 12 weeks of formula diet and a total of 48 weeks intensive counselling. Body weight and markers of the metabolic syndrome were analyzed. Presence of NAFLD was screened for by use of non-invasive markers of fatty liver, non-alcoholic steatohepatitis and liver fibrosis. RESULTS: Weight loss goals (i.e. > 5% or > 10% of initial body weight, respectively, depending on baseline BMI) were achieved in 89.7% of subjects in the intention-to-treat analysis and 93.9% in the per-protocol analysis. This was associated with a pronounced improvement in serum ALT values. The percentage of subjects who fulfilled non-invasive criteria for fatty liver dropped from 95.2 to 54.8%. Risk of NASH improved and the number of patients at risk of liver fibrosis declined by 54.1%. CONCLUSION: Lifestyle intervention was associated with a marked improvement of serum ALT and an improvement of surrogate scores indicative of NAFLD and, importantly, advanced fibrosis, in a real-world cohort of morbidly obese patients.

The Psychosocial Burden on Liver Transplant Recipients during the COVID-19 Pandemic.

INTRODUCTION: Due to the coronavirus disease 19 (COVID-19) pandemic, multiple measures have been implemented including social distancing and curfews. Both the disease and measures might cause stress, particularly in persons at risk, such as liver transplant (LT) recipients. Here, we evaluated the impact on psychosocial well-being of LT recipients. METHODS: Seventy-nine LT recipients and 83 nontransplanted controls participated in this study. Questionnaires comprising the WHO-five well-being index (WHO-5), the University of California at Los Angeles (UCLA) Loneliness Scale, and the preliminary COVID-19 Pandemic Mental Health Questionnaire (CoPaQ) were distributed among them. For the WHO-5 and UCLA Loneliness Scale, means of sum scores were compared between both groups, while a comparison on item level was conducted for the CoPaQ. RESULTS: The general well-being was similar in LT recipients and controls (WHO-5: 64.0 ± 20.5% vs. 66.4 ± 17.3%), while the UCLA Loneliness Scale indicated a higher level of perceived social isolation (1.90 ± 0.51 vs. 1.65 ± 0.53, p = 0.001). The CoPaQ indicated higher risk perception regarding health issues, in particular concerning the fear of having severe consequences in case of a COVID-19 infection (3.1 ± 1.1 vs. 2.2 ± 1.3, p < 0.001), higher risk-avoiding behavior and stronger adherence to pandemic measures in LT recipients. CONCLUSION: During the COVID-19 pandemic, LT recipients displayed a higher risk perception, a more pronounced risk-avoiding behavior and a higher perception of loneliness, while the overall well-being was comparable to nontransplanted controls.

Chemerin-156 is the Active Isoform in Human Hepatic Stellate Cells.

The chemokine chemerin exists as C-terminally processed isoforms whose biological functions are mostly unknown. A highly active human chemerin variant (huChem-157) was protective in experimental hepatocellular carcinoma (HCC) models. Hepatic stellate cells (HSCs) are central mediators of hepatic fibrogenesis and carcinogenesis and express the chemerin receptors chemokine-like receptor 1 (CMKLR1) and G protein-coupled receptor 1 (GPR1). Here we aimed to analyse the effect of chemerin isoforms on the viability, proliferation and secretome of the human HSC cell line LX-2. Therefore, huChem-157, 156 and 155 were over-expressed in LX-2 cells, which have low endogenous chemerin levels. HuChem-157 produced in LX-2 cells activated CMKLR1 and GPR1, and huChem-156 modestly induced GPR1 signaling. HuChem-155 is an inactive chemerin variant. Chemerin isoforms had no effect on cell viability and proliferation. Cellular expression of the fibrotic proteins galectin-3 and alpha-smooth muscle actin was not regulated by any chemerin isoform. HuChem-156 increased IL-6, IL-8 and galectin-3 in cell media. HuChem-157 was ineffective, and accordingly, did not enhance levels of these proteins in media of primary human hepatic stellate cells when added exogenously. These analyses provide evidence that huChem-156 is the biologic active chemerin variant in hepatic stellate cells and acts as a pro-inflammatory factor.

Chemokine (CC-motif) receptor-like 2 mRNA is expressed in hepatic stellate cells and is positively associated with characteristics of non-alcoholic steatohepatitis in mice and men.

Chemokine (CC-motif) receptor-like 2 (CCRL2) is a decoy receptor and regulates the local responses of the chemokine chemerin. Recently our group has shown that the functional chemerin receptor, chemokine-like receptor 1 (CMKLR1), correlates with fibrosis and non-alcoholic steatohepatitis (NASH) score in males only. In our current study, we wanted to know whether CCRL2 shows similar correlations as CMKLR1. Therefore, we analyzed the hepatic expression of CCRL2 in murine NASH and in liver tissues obtained from 85 patients with non-alcoholic fatty liver disease (NAFLD) and 33 controls. CCRL2 mRNA was not significantly changed in murine and human NASH liver. CCRL2 mRNA levels were positively correlated with inflammation, fibrosis and NASH scores in the patients. Concordantly, CCRL2 was related to the mRNA levels of F4/80, transforming growth factor beta and alpha smooth muscle actin in murine NASH. In the human cohort, CCRL2 mRNA correlated with fibrosis score and CMKLR1 mRNA in both gender. CCRL2 mRNA was induced in the liver of type 2 diabetes and hypercholesterolemic patients, but still positively correlated with fibrosis score when these patients were excluded from calculations. Human hepatic stellate cells (HSC), hepatic sinusoidal endothelial cells and Kupffer cells (KC) express CCRL2 mRNA. TNF induces CCRL2 expression in HSC and lipopolysaccharide in KC suggesting that correlations identified in NAFLD patients are partly related to the activation of these cells.

Tubulin alpha 8 is expressed in hepatic stellate cells and is induced in transformed hepatocytes.

Tubulin alpha 8 (TUBA8) is highly abundant in murine liver tumors suggesting a role in hepatocellular carcinoma (HCC). Non-alcoholic steatohepatitis (NASH) is a risk factor for HCC. In mice that are fed with a methionine-choline deficient diet for two weeks to induce advanced murine NASH, we do see increased hepatic levels of TUBA8 protein. In animals given a high-fat diet for 14 weeks or an atherogenic diet for 12 weeks, hepatic TUBA8 is unchanged. TUBA8 is highly expressed in human hepatic stellate cells (HSC) and co-localizes with the HSC marker desmin in the murine liver. Inflammatory (TNF, LPS, IL-6) and profibrotic mediators (TGF-beta) do not regulate TUBA8 in HepG2 cells, primary HSC and the HSC cell line LX-2, when stimulated for 24 h. Agonists of the farnesoid X receptor and peroxisome proliferator activated receptor gamma, which are nuclear receptors involved in NASH and HCC pathophysiology, have no effect on TUBA8 in HepG2 and LX-2 cells. In human HCC tissues of 18 patients TUBA8 is significantly upregulated when compared to the corresponding non-tumorous tissues. Compared to non-transformed hepatocytes, TUBA8 protein is strongly expressed in transformed cells. Thus, TUBA8 is a marker of HSC whose cell number is increased in NASH, while higher levels in HCC may be related to induction of TUBA8 in parenchymal cells.

Hepatic chemerin mRNA expression is reduced in human nonalcoholic steatohepatitis.

BACKGROUND: Chemerin is associated with insulin resistance and is expressed in the liver. Nonalcoholic fatty liver disease (NAFLD) is related to impaired insulin sensitivity, but studies evaluating hepatic and serum chemerin in NAFLD resulted in discordant data. MATERIALS AND METHODS: Chemerin mRNA was determined in the liver tissue obtained from 33 controls and 76 NAFLD patients. Chemerin serum levels were measured in a different cohort of patients with ultrasound-diagnosed NAFLD and the respective controls. Hepatic stellate cells and hepatocytes were exposed to selected metabolites and nuclear receptor agonists to study the regulation of chemerin. Effect of recombinant chemerin on hepatocyte released proteins was analysed. RESULTS: Hepatic chemerin expression was not related to BMI, gender, type 2 diabetes and hypertension. Chemerin mRNA did not correlate with steatosis and was negatively associated with inflammation, fibrosis and nonalcoholic steatohepatitis (NASH) score. Patients with NASH had lower chemerin mRNA compared to those with borderline NASH and controls. Factors with a role in NASH mostly did not regulate chemerin in the liver cells. Of note, liver X receptor agonist reduced chemerin protein. Serum chemerin was not changed in NAFLD. Levels positively correlated with age, waist-to-hip ratio, systolic blood pressure, serum FGF21 and lipocalin 2, and negatively with transferrin saturation. Chemerin induced FGF21 in supernatants of primary human hepatocytes. Hepcidin, a major regulator of iron homoeostasis and lipocalin 2, were not regulated by chemerin. CONCLUSION: Chemerin mRNA is reduced in the liver of NASH patients, and liver X receptor seems to have a role herein.