A transient early HBV-DNA increase during PEG-IFNα therapy of hepatitis D indicates loss of infected cells and is associated with HDV-RNA and HBsAg reduction.

HBV-DNA levels are low or even undetectable in the majority HDV-infected patients. The impact of PEG-IFNα on HBV-DNA kinetics in HDV-infected patients has not been studied in detail. We analysed data of a prospective treatment trial where 120 HDV-RNA-positive patients were randomized to receive PEG-IFNα-2a plus tenofovir-disoproxil-fumarate (PEG-IFNα/TDF, n = 59) or placebo (PEG-IFNα/PBO; n = 61) for 96 weeks. At week 96, HBV-DNA was still quantifiable in 71% of PEG-IFNα/PBO-treated patients but also in 76% of PEG-IFNα/TDF-treated patients, despite low HBV-DNA baseline values. Surprisingly, a transient HBV-DNA increase between weeks 12 and 36 was observed in 12 in PEG-IFNα/TDF-treated and 12 PEG-IFNα/PBO-treated patients. This increase was positively associated with HBsAg loss [(P = 0.049, odds ratio (OR) 5.1] and HDV-RNA suppression (P = 0.007, OR 4.1) at week 96. Biochemical markers of cell death (M30 and ALT) were higher during the HBV-DNA peak but no distinct systemic immune pattern could be observed by screening 91 soluble inflammatory markers. In conclusion, an early increase in HBV-DNA during PEG-IFNα-2a therapy occurred in more than 20% of patients, even in TDF-treated patients. This transient HBV-DNA rise may indicate PEG-IFNα-induced cell death and lead to long-term HDV-RNA suppression and HBsAg loss.

In Severe Alcoholic Hepatitis, Serum Keratin-18 Fragments Are Diagnostic, Prognostic, and Theragnostic Biomarkers.

INTRODUCTION: Up to 40% of patients with severe alcoholic hepatitis (AH) die within 6 months of presentation, making prompt diagnosis and appropriate treatment essential. We determined the associations between serum keratin-18 (K18) and histological features, prognosis, and differential response to prednisolone in patients with severe AH. METHODS: Total (K18-M65) and caspase-cleaved K18 (K18-M30) were quantified in pretreatment sera from 824 patients enrolled in the Steroids or Pentoxifylline for Alcoholic Hepatitis trial (87 with suitable histological samples) and disease controls. RESULTS: K18 fragments were markedly elevated in severe AH and strongly predicted steatohepatitis (alcoholic steatohepatitis) on biopsy (area under receiver operating characteristics: 0.787 and 0.807). Application of published thresholds to predict alcoholic steatohepatitis would have rendered biopsy unnecessary in 84% of all AH cases. K18-M30 and M65 were associated with 90-day mortality, independent of age and Model for End-stage Liver Disease score in untreated patients. The association for K18-M65 was independent of both age and Model for End-stage Liver Disease in prednisolone-treated patients. Modelling of the effect of prednisolone on 90-day mortality as a function of pretreatment serum K18 levels indicated benefit in those with high serum levels of K18-M30. At low pretreatment serum K18 levels, prednisolone was potentially harmful. A threshold of K18-M30 5 kIU/L predicted therapeutic benefit from prednisolone above this level (odds ratio: 0.433, 95% confidence interval: 0.19-0.95, P = 0.0398), but not below (odds ratio: 1.271, 95% confidence interval: 0.88-1.84, P = 0.199). Restricting prednisolone usage to the former group would have reduced exposure by 87%. DISCUSSION: In a large cohort of patients with severe AH, serum K18 strongly correlated with histological severity, independently associated with 90-day mortality, and predicted response to prednisolone therapy. Quantification of serum K18 levels could assist in clinical decision-making.

Autophagy alleviates amiodarone-induced hepatotoxicity.

Amiodarone is a widely used antiarrhythmic drug that can cause the development of steatohepatitis as well as liver fibrosis and cirrhosis. The molecular mechanisms of amiodarone-mediated liver injury remain largely unknown. We therefore analyzed amiodarone-mediated hepatocellular injury in patients with chronic heart failure, in primary hepatocytes and HepG2 cells. We found that amiodarone-treated patients with chronic heart failure revealed significantly higher serum levels of caspase-cleaved keratin-18, an apoptosis biomarker, compared to healthy individuals or patients not receiving amiodarone. Furthermore, amiodarone treatment of hepatocytes resulted in apoptosis associated with lipid accumulation and ER-stress induction. Liver cell steatosis was accompanied by enhanced de novo lipogenesis which, after reaching peak levels, declined together with decreased activation of ER stress. The decline of amiodarone-mediated lipotoxicity was associated with protective autophagy induction. In contrast, in hepatocytes treated with the autophagy inhibitor chloroquine as well as in autophagy gene (ATG5 or ATG7)-deficient hepatocytes, amiodarone-triggered toxicity was increased. In conclusion, we demonstrate that amiodarone induces lipid accumulation associated with ER stress and apoptosis in hepatocytes, which is mirrored by increased keratin-18 fragment serum levels in amiodarone-treated patients. Autophagy reduces amiodarone-mediated lipotoxicity and could provide a therapeutic strategy for protection from drug-induced liver injury.

TNF-Receptor-1 inhibition reduces liver steatosis, hepatocellular injury and fibrosis in NAFLD mice.

Non-alcoholic fatty liver disease (NAFLD) shows an increasing prevalence and is associated with the development of liver fibrosis and cirrhosis as the major risk factors of liver-related mortality in this disease. The therapeutic possibilities are limited and restricted to life style intervention, since specific drugs for NAFLD are unavailable so far. TNFα has been implicated as a major pathogenic driver of NAFLD. TNFα-mediated liver injury occurs mainly via TNF-receptor-1 (TNFR1) signaling, whereas TNFR2 mediates protective pathways. In this study, we analyzed the therapeutic effects of a novel antibody, which selectively inhibits TNFR1 while retaining protective TNFR2 signaling in a high-fat diet (HFD) mouse model of NAFLD. Mice were fed with HFD for 32 weeks and treated with anti-TNFR1-antibody or control-antibody for the last 8 weeks. We then investigated the mechanisms of TNFR1 inhibition on liver steatosis, inflammatory liver injury, insulin resistance and fibrosis. Compared to control-antibody treatment, TNFR1 inhibition significantly reduced liver steatosis and triglyceride content, which was accompanied by reduced expression and activation of the transcription factor SREBP1 and downstream target genes of lipogenesis. Furthermore, inhibition of TNFR1 resulted in reduced activation of the MAP kinase MKK7 and its downstream target JNK, which was associated with significant improvement of insulin resistance. Apoptotic liver injury, NAFLD activity and alanine aminotransferase (ALT) levels, as well as liver fibrosis significantly decreased by anti-TNFR1 compared to control-antibody treatment. Thus, our results suggest selective TNFR1 inhibition as a promising approach for NAFLD treatment.

Multicenter Validation Study of a Diagnostic Algorithm to Detect NASH and Fibrosis in NAFLD Patients With Low NAFLD Fibrosis Score or Liver Stiffness.

OBJECTIVES: Nonalcoholic steatohepatitis (NASH) and fibrosis play critical roles for the prognosis of patients with nonalcoholic fatty liver disease (NAFLD). Identification of patients at risk of NASH and fibrosis is therefore critical for disease management. NAFLD Fibrosis Score (NFS) and transient elastography (TE) have been suggested to exclude advanced fibrosis. However, there is increasing evidence that also patients with NASH and early fibrosis are at risk of disease progression and complications, emphasizing the need for improved noninvasive risk stratification in NAFLD. METHODS: Because hepatocyte apoptosis plays an early role in NASH pathogenesis, we evaluated whether the apoptosis biomarker M30 might identify NAFLD patients who are at risk of NASH and fibrosis despite low NFS or TE values. Serum M30 levels were assessed by enzyme-linked immunosorbent assay in combination with NFS and/or TE in an exploration (n = 103) and validation (n = 100) cohort of patients with biopsy-proven NAFLD. RESULTS: Most patients with low NFS (cutoff value < -1.455) revealed increased M30 levels (>200 U/L) in the exploration (62%) and validation (67%) cohort, and more than 70% of them had NASH, mostly with histological fibrosis. Vice versa, most patients with NFS < -1.455 but nonelevated M30 levels showed no NASH. NASH was also detected in most patients with indeterminate NFS (-1.455 to 0.676) but elevated M30 levels, from which ∼90% showed fibrosis. Similar results were obtained when using TE instead of NFS. DISCUSSION: The combination of the M30 biomarker with NFS or TE enables a more reliable identification of patients with an increased risk of progressed NAFLD and improves patient stratification.

More than just one: multiplicity of Hirudins and Hirudin-like Factors in the Medicinal Leech, Hirudo medicinalis.

Blood-sucking leeches like the medicinal leech, Hirudo medicinalis, have been used for medical purposes since ancient times. During feeding, medicinal leeches transfer a broad range of bioactive substances into the host's wound to prevent premature hemostasis and blood coagulation. Hirudin is probably the best known of these substances. Despite its long history of investigation, recombinant production and clinical use, there still exist conflicting data regarding the primary structure of hirudin. Entirely unclear is the potential biological significance of three different subtypes and many isoforms of hirudins that have been characterized so far. Furthermore, there is only incomplete information on their cDNA sequences and no information at all on gene structures and DNA sequences are available in the databases. Our efforts to fill these gaps revealed the presence of multiple hirudin-encoding genes in the genome of Hirudo medicinalis. We have strong evidence for the expression of all three subtypes of hirudin within individual leeches and for the expression of additional hirudins or hirudin-like factors that may have different biological functions and may be promising candidates for new drugs.