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Nonalcoholic fatty liver disease (NAFLD) is a major health problem with complex pathogenesis. Although sex differences in NAFLD pathogenesis have been reported, the mechanisms underlying such differences remain understudied. Interleukin (IL)22 is a pleiotropic cytokine with both protective and/or pathogenic effects during liver injury. IL22 was shown to be hepatoprotective in NAFLD-related liver injury. However, these studies relied primarily on exogenous administration of IL22 and did not examine the sex-dependent effect of IL22. Here, we sought to characterize the role of endogenous IL22-receptor signaling during NAFLD-induced liver injury in males and females.We used immunofluorescence, flow cytometry, histopathologic assessment, and gene expression analysis to examine IL22 production and characterize the intrahepatic immune landscape in human subjects with NAFLD (n = 20; 11 men and 9 women) and in an in vivo Western high-fat diet-induced NAFLD model in IL22RA knock out mice and their wild-type littermates.Examination of publicly available data sets from 2 cohorts with NAFLD showed increased hepatic IL22 gene expression in females compared with males. Furthermore, our immunofluorescence analysis of liver sections from NAFLD subjects (n = 20) showed increased infiltration of IL22-producing cells in females. Similarly, IL22-producing cells were increased in wild-type female mice with NAFLD and the hepatic IL22/IL22 binding protein messenger RNA ratio correlated with expression of anti-apoptosis genes. The lack of endogenous IL22-receptor signaling (IL22RA knockout) led to exacerbated liver damage, inflammation, apoptosis, and liver fibrosis in female, but not male, mice with NAFLD.Our data suggest a sex-dependent hepatoprotective antiapoptotic effect of IL22-receptor signaling during NAFLD-related liver injury in females.

BackgroundInterleukin-6 (IL-6) plays important and dynamic roles in inflammation associated with fatty liver disease over all stages, from simple steatosis to steatohepatitis, cirrhosis and cancer. IL-6 signals locally, but also circulates with multiple co-factors that control paracrine and endocrine signaling. As inflammation is a main driver of liver fibrosis, we investigated relationships between circulating components of the interleukin-6 signaling pathway (IL-6, sIL-6R and sgp130) and liver pathology in subjects with metabolically associated fatty liver disease (MAFLD) or steatohepatitis (MASH).MethodsPredictive performances of plasma IL-6, sIL-6R and sgp130 were investigated in two independent cohorts: 1) patients with biopsy-confirmed MASH (n=49), where magnetic resonance spectroscopy (MRS), imaging (MRI) and elastography (MRE) assessed liver fat, volume and stiffness; and 2) patients with morbid obesity (n=245) undergoing bariatric surgery where histological staging of steatosis, activity, and fibrosis determined MASH severity. Correlations were evaluated between IL-6, sIL-6R and sgp130 and anthropomorphic characteristics, plasma markers of metabolic disease or liver pathology.ResultsIn patients with MASH, plasma IL-6 and sgp130 strongly correlated with liver stiffness, which for sgp130 was independent of age, sex, BMI, diabetes, hyperlipidemia, hypertension or history of HCC. Plasma sgp130 was the strongest predictor of liver stiffness compared to common predictors and risk scores. Plasma sIL-6R correlated with liver volume independent of age, sex, and BMI. In patients with morbid obesity, circulating sgp130 correlated with advanced liver fibrosis.ConclusionLevels of circulating sgp130 can predict progressing MASH and may be used alone or in combination with other predictors as a non-invasive measure of liver disease severity.GRAPHICAL ABSTRACT

The goal of this study was to evaluate the effect of the Model for End-Stage Liver Disease (MELD)-based allocation system on mortality, bleeding, and transfusion requirement in orthotopic liver transplantation (OLT).OLTs were studied for this observational study (before-and-after observational cohort study).One community hospital.The study comprised 686 patients who underwent 750 consecutive OLTs.None.Patients who underwent OLT in the MELD era had an adjusted lower 1-year mortality (adjusted odds ratio 0.45 [0.24-0.83]) compared with patients who underwent OLT the pre-MELD era. No significant difference in 1-month mortality was observed. Other variables with a significant effect on 1-year mortality in multivariate analysis were preoperative international normalized ratio, intraoperative use of a phlebotomy, total intraoperative volume of crystalloid infused, and retransplantation. Blood loss was greater in the MELD era (median difference 200 mL; p0.001), as were red blood cell, fresh frozen plasma, and cryoprecipitate transfusions. More patients in the MELD era received at least 1 transfusion (27% v 20%; p = 0.024).The MELD allocation system did not affect 1-month mortality, but a decrease in 1-year mortality was demonstrated. Blood loss and transfusions increased during OLTs performed in the MELD era. The role of other variables should be explored further to explain postoperative morbidity and mortality.

Disulfiram (DSF), a sulfur-containing compound, has been used to treat chronic alcoholism and cancer for decades by inactivating aldehyde dehydrogenase (ALDH). Hydrogen sulfide (H2S) is a new gasotransmitter and regulates various cellular functions by S-sulfhydrating cysteine in the target proteins. H2S exhibits similar properties to DSF in the sensitization of cancer cells. The interaction of DSF and H2S on ALDH activity and liver cancer cell survival are not clear. Here it was demonstrated that DSF facilitated H2S release from thiol-containing compounds, and DSF and H2S were both capable of regulating ALDH through inhibition of gene expression and enzymatic activity. The supplement of H2S sensitized human liver cancer cells (HepG2) to DSF-inhibited cell viability. The expression of cystathionine gamma-lyase (a major H2S-generating enzyme) was lower but ALDH was higher in mouse liver cancer stem cells (Dt81Hepa1-6) in comparison with their parental cells (Hepa1-6), and H2S was able to inhibit liver cancer stem cell adhesion. In conclusion, these data point to the potential of combining DSF and H2S for inhibition of cancer cell growth and tumor development by targeting ALDH.

Genetic iron overload, or hemochromatosis, can be caused by mutations in HFE, hemojuvelin, and hepcidin genes. Hepcidin, a negative regulator of intestinal iron absorption, is found to be inappropriately low in both patients and in animal models, indicating that proper control of basal hepcidin levels requires both hemojuvelin and HFE. In mice, repulsive guidance molecule c (Rgmc, the hemojuvelin mouse ortholog) and hepcidin levels are transcriptionally regulated during inflammation. Here, we report that basal Rgmc levels in Hfe-deficient mice are normal and that these mice retain the ability to suppress Rgmc expression after lipopolysaccharide (LPS) challenge. Thus, Rgmc regulation by LPS is Hfe-independent. The response of Rgmc to LPS involves signaling through toll-like receptor 4 (Tlr4), because Tlr4-deficient mice do not show altered Rgmc expression after LPS administration. We further show that tumor necrosis factor-alpha, but not interleukin-6, is sufficient to cause Rgmc down-regulation by LPS. These results contrast with previous data demonstrating that hepcidin levels are directly regulated by interleukin-6 but not by tumor necrosis factor-alpha. The regulation of iron-related genes by different cytokines may allow for time-dependent control of iron metabolism changes during inflammation and may be relevant to chronic inflammation, infections, and cancer settings, leading to the development of anemia of chronic disease.

In some public goods environments it may be advantageous for heterogeneous groups to be coordinated by a single individual. This “volunteer” will bear private costs for acting as the leader while enabling each member of the group to achieve maximum potential gains. This environment is modeled as a War of Attrition game in which everyone can wait for someone else to volunteer. Since these games generally have multiple Nash equilibria but a unique subgameperfect equilibrium, we tested experimentally the predictive power of the subgame-perfection criterion. Our data contradict that subjects saw the subgame-perfect strategy combination as the obvious way to play the game. An alternative behavioral hypothesis – that subjects were unable to predict accurately how their opponents would play and tried to maximize their expected payoff – is proposed. This hypothesis fits the observed data generally well.

We examine, both in general games in strategic form and in games of voluntary provision of a public good, some implications of the assumption that individuals may obey ethical codes of conduct. The notion of morality considered captures the intuition (often attributed to Kant) that a moral action leads to the best outcome when it is properly universalized . We propose a formalization of this idea which generalizes earlier attempts made in this direction in the literature by allowing the players to differ both in their strategy sets and their preferences. We show that it is easy to find examples of games in which no moral behavior of this type exists or where the only existing ‘Kantian’ code of conduct leads to a Pareto-inefficient outcome. We then more specifically examine the issues of existence and Pareto-efficiency of Kantian norms of behavior in games of voluntary provision of a public good. We find in this context that there is no conflict between morality and Pareto-efficiency since any Kantian norm of behavior is Pareto-efficient. We also prove the existence of a Kantian norm of individual contribution.

Background/Aims: Total ligation of the left portal vein is thought to induce both hepatocyte apoptosis and necrosis. The pathological impact of partial ligation of a branch of the portal vein has not yet been evaluated. Methods: We studied the degree of hepatocyte injury following 0, 43, 48, 59, 68, 72, 78 and 100% left portal vein stenosis in 200-g Sprague-Dawley male rats. Serum alanine aminotransferase levels, total body weight, and left and right liver lobe weights were measured at 2 and 7 days. Mitosis and 3 H-thymidine labelling indices were measured as markers of proliferation; the apoptotic index and TUNEL stain were used as markers to measure apoptotic cell death. Necrosis was assessed morphologically. All these parameters were evaluated 2 days after ligation. Results: There was a direct relation between the increase in weight of the right lobes and the reduction in weight of the left lobes. The degree of weight change correlated significantly with the degree of stenosis. In the right lobes, mitosis and 3 H-thymidine labelling were increased in proportion to the degree of stenosis. In the left lobes, the decrease in volume of hepatocytes correlated with the degree of ligation, especially in the pericentral areas. Necrosis was identified only when ligation was ≥68%, this being associated with an increase in alanine aminotransferase levels. On the other hand, apoptotic cells were identified in increasing numbers, starting from the lowest degree of ligation to 100% ligation. This was found both morphologically and with TUNEL stain. Conclusions: Partial ligation of the left portal vein induces left liver atrophy through hepatocyte volume loss and apoptosis. Necrosis is found only when the degree of ligation is severe.

Which member of a heterogeneous population will volunteer to provide the entrepreneurial services needed to found a nonprofit firm to privately provide a public good? We develop a model of this decision. The model predicts that nonprofit entrepreneurs will be those who incur relatively low private costs, or get a relatively high private payoff. Less obviously, they will tend to have a high discount rate and relatively long time horizons, either due to their youth, or due to their consideration of generations to come far into the future. If the public good is one which individuals tend to contribute large amounts to, such entrepreneurs will tend to be relatively wealthy. If private contributions to the firm are relatively small, the entrepreneur is more likely to come from the center of the wealth distribution.

Quantitative measurement of hepatic iron by biochemical analysis of liver biopsy samples is required to assess hepatic iron stores accurately. Cirrhotic livers, however, contain variable amounts of fibrous tissue and the distribution of iron within the hepatic parenchyma is not always uniform. The aim of this study was to assess the variability in hepatic iron concentration measurement from needle-biopsy specimens.The livers from eight patients with cirrhosis selected because of elevated serum ferritin were obtained at the time of liver transplantation (n = 6) or at autopsy (n = 2). Multiple needle biopsies were done, and hepatic iron concentration was measured by atomic absorption spectroscopy. The hepatic iron index was calculated as iron concentration divided by age.Four cases had a mean hepatic iron index above 2.0, in the range of that reported in patients with homozygous genetic hemochromatosis, whereas the other four had an hepatic iron index of less than 2.0. The intra-individual coefficient of variation for hepatic iron concentration ranged from 11.3 to 43.7%, averaging 24.9%. The coefficient of variation was smaller in biopsy samples4 mg dry weight than in samples4 mg (19.8% vs 28.6%, p0.05). Histological examination of surgical biopsies from these livers showed large amounts of fibrous tissue, and inhomogeneous distribution or iron in the hepatic parenchyma.This study demonstrates an important variability in the measurement of hepatic iron content from needle biopsy specimens in patients with severe cirrhosis.

Several hepatocyte mitogens have been identified, but the signals triggering the G0/G1 transition and cell cycle progression of hepatocytes remain unknown. Using hepatocyte primary cultures, we investigated the role of epidermal growth factor/pyruvate during the entry into and progression through the G1 phase and analyzed the expression of cell cycle markers. We show that the G0/G1 transition occurs during hepatocyte isolation as evidenced by the expression of early genes such as c-fos, c-jun, and c-myc. In culture, hepatocytes progress through G1 regardless of growth factor stimulation until a restriction point (R point) in mid-late G1 beyond which they cannot complete the cell cycle without mitogenic stimulation. Changes in cell cycle gene expression were associated with progression in G1; the cyclin E mRNA level is low early in G1 but increases at the G1/S boundary, while the protein is constantly detected during cell cycle but undergoes a change of electrophoretic mobility in mid-late G1 after the R point. In addition, a drastic induction of cyclin D1 mRNA and protein, and to a lesser extent of cyclin D2 mRNA, takes place in mitogen-stimulated cells after the R point. In contrast, cyclin D3 mRNA appears early in G1, remains constant in stimulated cells, but accumulates in unstimulated arrested cells, paralleling the cyclin-dependent kinase 4 mRNA expression. These results characterize the different steps of G1 phase in hepatocytes.

The performance of tax-earmaking institutions that require individuals to contribute some minimum levels toward public goods but do not constrain the composition of their contributions is examined. It is shown that in a non-negligible class of circumstances, leaving the spending decisions in the hands of individuals would yield a unique non-cooperative equilibrium in the (constrained) core. These results are then applied to the analysis of financing rules for public and separate schools in Canada.

If individuals are free to direct their gifts to any charity, why would they contribute instead to an institution like the United Way that may not disburse their donations as they would have themselves? It is shown that ‘contributing only to the United Fund’ can be a Subgame Perfect Equilibrium, if the Fund plays after everyone else and is able to offset direct contributions. However, donations to a United Fund may be lower than direct contributions would have been, so that an optimal grants policy for the United Fund would involve trading off a less desirable mix of services for higher total contributions.

Background & Aims Development of fibrosis is part of the pathophysiologic process of chronic liver disease. Although it is considered deleterious, it also represents a form of tissue repair. Deposition of extracellular matrix changes the cellular environment of the liver; we investigated whether it increases resistance to noxious stimuli and the role of changes in intracellular signaling to hepatocytes in mediating this effect. Methods Primary cultures of mouse hepatocytes were exposed to type I collagen (COL1); cell injury was assessed by morphologic and biochemical criteria. The expression of Bcl-2 family members was evaluated by immunoblot analyses. Activation of extracellular signal–regulated kinase (ERK) was assessed using phospho-specific antibodies. Liver fibrosis was induced by repeated administration of thioacetamide or carbon tetrachloride to mice; mice were then exposed to Fas antibodies. Results Hepatocytes exposed to COL1 were more resistant to a variety of hepatotoxins, in a dose-dependent manner, and had lower levels of Bad, Bid, and Bax proapoptotic proteins compared with control hepatocytes. Activation of ERK1/2 was stronger and quicker in hepatocytes exposed to COL1. The MEK1/2 inhibitors U0126 and PD98059 reversed the protective effects of COL1 and the decrease in proapoptotic proteins. Hepatocytes isolated from ERK1 −/− mice were insensitive to the protective effect of COL1. Fibrotic livers from wild-type mice had high levels of phospho-ERK1 and were resistant to Fas-induced cell death. ERK1 −/− mice lost this effect. Conclusions Production of COL1 during liver fibrosis induces a hepatoprotective response that is mediated by activation of ERK1 signaling.