Visceral obesity is intimately associated with metabolic disease and adverse health outcomes. However, a direct association between increasing amounts of visceral fat and end-organ inflammation and scarring has not been demonstrated. We examined the association between visceral fat and liver inflammation in patients with nonalcoholic fatty liver disease (NAFLD) to delineate the importance of visceral fat to progressive steatohepatitis and hence the inflammatory pathogenesis of the metabolic syndrome. We undertook a cross-sectional, proof of concept study in 38 consecutive adults with NAFLD at a tertiary liver clinic. All subjects had a complete physical examination, anthropometric assessment, and fasting blood tests on the day of liver biopsy. Abdominal fat volumes were assessed by magnetic resonance imaging within 2 weeks of liver biopsy. The extent of hepatic inflammation and fibrosis augmented incrementally with increases in visceral fat (P < 0.01). For each 1% increase in visceral fat, the odds ratio for increasing liver inflammation and fibrosis was 2.4 (confidence interval [CI]: 1.3-4.2) and 3.5 (CI: 1.7-7.1), respectively. Visceral fat remained an independent predictor of advanced steatohepatitis (odds ratio [OR] 2.1, CI: 1.1-4.2, P ‫؍‬ 0.05) and fibrosis (OR 2.9, CI: 1.4-6.3, P ‫؍‬ 0.006) even when controlled for insulin resistance and hepatic steatosis. Interleukin-6 (IL-6) levels, which correlated with visceral fat, also independently predicted increasing liver inflammation. Visceral fat was associated with all components of the metabolic syndrome. Conclusion: Visceral fat is directly associated with liver inflammation and fibrosis independent of insulin resistance and hepatic steatosis. Visceral fat should therefore be a central target for future interventions in nonalcoholic steatohepatitis and indeed all metabolic disease. (HEPATOLOGY 2008;48:449-457.)

BACKGROUNDManagement of nonalcoholic steatohepatitis (NASH) is an unmet clinical need. Lanifibranor is a pan-PPAR (peroxisome proliferator-activated receptor) agonist that modulates key metabolic, inflammatory, and fibrogenic pathways in the pathogenesis of NASH. METHODSIn this phase 2b, double-blind, randomized, placebo-controlled trial, patients with noncirrhotic, highly active NASH were randomly assigned in a 1:1:1 ratio to receive 1200 mg or 800 mg of lanifibranor or placebo once daily for 24 weeks. The primary end point was a decrease of at least 2 points in the SAF-A score (the activity part of the Steatosis, Activity, Fibrosis [SAF] scoring system that incorporates scores for ballooning and inflammation) without worsening of fibrosis; SAF-A scores range from 0 to 4, with higher scores indicating more-severe disease activity. Secondary end points included resolution of NASH and regression of fibrosis. RESULTSA total of 247 patients underwent randomization, of whom 103 (42%) had type 2 diabetes mellitus and 188 (76%) had significant (moderate) or advanced fibrosis. The percentage of patients who had a decrease of at least 2 points in the SAF-A score without worsening of fibrosis was significantly higher among those who received the 1200-mg dose, but not among those who received the 800-mg dose, of lanifibranor than among those who received placebo (1200-mg dose vs. placebo, 55% vs. 33%, P = 0.007; 800-mg dose vs. placebo, 48% vs. 33%, P = 0.07). The results favored both the 1200-mg and 800-mg doses of lanifibranor over placebo for resolution of NASH without worsening of fibrosis (49% and 39%, respectively, vs. 22%), improvement in fibrosis stage of at least 1 without worsening of NASH (48% and 34%, respectively, vs. 29%), and resolution of NASH plus improvement in fibrosis stage of at least 1 (35% and 25%, respectively, vs. 9%). Liver enzyme levels decreased and the levels of the majority of lipid, inflammatory, and fibrosis biomarkers improved in the lanifibranor groups. The dropout rate for adverse events was less than 5% and was similar across the trial groups. Diarrhea, nausea, peripheral edema, anemia, and weight gain occurred more frequently with lanifibranor than with placebo. CONCLUSIONSIn this phase 2b trial involving patients with active NASH, the percentage of patients who had a decrease of at least 2 points in the SAF-A score without worsening of fibrosis was significantly higher with the 1200-mg dose of lanifibranor than with placebo. These findings support further assessment of lanifibranor in phase 3 trials. (Funded by Inventiva Pharma; NATIVE ClinicalTrials.gov number, NCT03008070.

Chronic hepatic inflammation from multiple etiologies leads to a fibrogenic response that can progress to cirrhosis and liver failure. Transplantation of human amniotic epithelial cells (hAEC) from term delivered placenta has been shown to decrease mild to moderate hepatic fibrosis in a murine model. To model advanced human liver disease and assess the efficacy of hAEC therapy, we transplanted hAEC in mice with advanced hepatic fibrosis. Immunocompetent C57BL/6 mice were administered carbon tetrachloride (CCl 4 ) twice weekly resulting in bridging fibrosis by 12 weeks. hAEC (2×10 6 ) were infused via the tail vein at week 8 or weeks 8 and 10 (single and double dose, respectively). Human cells were detected in mouse liver four weeks after transplantation showing hAEC engraftment. CCl 4 treated mice receiving single or double hAEC doses showed a significant but similar decrease in liver fibrosis area associated with decreased activation of collagen-producing hepatic stellate cells and decreased hepatic protein levels of the pro-fibrogenic cytokine, transforming growth factor-beta1. CCl 4 administration caused hepatic T cell infiltration that decreased significantly following hAEC transplantation. Hepatic macrophages play a crucial role in both fibrogenesis and fibrosis resolution. Mice exposed to CCl 4 demonstrated increased numbers of hepatic macrophages compared to normal mice; the number of macrophages decreased significantly in CCl 4 treated mice given hAEC. These mice had significantly lower hepatic protein levels of the chemokine monocyte chemoattractant protein-1 than mice given CCl 4 alone. Alternatively activated M2 macrophages are associated with fibrosis resolution. CCl 4 treated mice given hAEC showed increased expression of genes associated with M2 macrophages including YM-1, IL-10 and CD206. We provide novel data showing that hAEC transplantation induces a wound healing M2 macrophage phenotype associated with reduction of established hepatic fibrosis that justifies further investigation of this potential cell-based therapy for advanced hepatic fibrosis.

Current conductive materials for use in cardiac regeneration are limited by cytotoxicity or cost in implementation. In this manuscript, we demonstrate for the first time the application of a biocompatible, conductive polypyrrole-polycaprolactone film as a platform for culturing cardiomyocytes for cardiac regeneration. This study shows that the novel conductive film is capable of enhancing cell-cell communication through the formation of connexin-43, leading to higher velocities for calcium wave propagation and reduced calcium transient durations among cultured cardiomyocyte monolayers. Furthermore, it was demonstrated that chemical modification of polycaprolactone through alkaline-mediated hydrolysis increased overall cardiomyocyte adhesion. The results of this study provide insight into how cardiomyocytes interact with conductive substrates and will inform future research efforts to enhance the functional properties of cardiomyocytes, which is critical for their use in pharmaceutical testing and cell therapy.

Background: Hepatic stellate cells (HSCs) are the primary collagen-secreting cells in the liver. While HSCs are the major cell type involved in the pathogenesis of liver fibrosis, hepatic macrophages also play an important role in mediating fibrogenesis and fibrosis resolution. Previously, we observed a reduction in HSC activation, proliferation, and collagen synthesis following exposure to human amnion epithelial cells (hAEC) and hAEC-conditioned media (hAEC-CM). This suggested that specific factors secreted by hAEC might be effective in ameliorating liver fibrosis. hAEC-derived extracellular vesicles (hAEC-EVs), which are nanosized (40–100 nm) membrane bound vesicles, may act as novel cell–cell communicators. Accordingly, we evaluated the efficacy of hAEC-EV in modulating liver fibrosis in a mouse model of chronic liver fibrosis and in human HSC.Methods: The hAEC-EVs were isolated and characterized. C57BL/6 mice with CCl4-induced liver fibrosis were administered hAEC-EV, hAEC-CM, or hAEC-EV depleted medium (hAEC-EVDM). LX2 cells, a human HSC line, and bone marrow-derived mouse macrophages were exposed to hAEC-EV, hAEC-CM, and hAEC-EVDM. Mass spectrometry was used to examine the proteome profile of each preparation.Results: The extent of liver fibrosis and number of activated HSCs were reduced significantly in CCl4-treated mice given hAEC-EVs, hAEC-CM, and hAEC EVDM compared to untreated controls. Hepatic macrophages were significantly decreased in all treatment groups, where a predominant M2 phenotype was observed. Human HSCs cultured with hAEC-EV and hAEC-CM displayed a significant reduction in collagen synthesis and hAEC-EV, hAEC-CM, and hAEC-EVDM altered macrophage polarization in bone marrow-derived mouse macrophages. Proteome analysis showed that 164 proteins were unique to hAEC-EV in comparison to hAEC-CM and hAEC-EVDM, and 51 proteins were co-identified components with the hAEC-EV fraction.Conclusion: This study provides novel data showing that hAEC-derived EVs significantly reduced liver fibrosis and macrophage infiltration to an extent similar to hAEC-EVDM and hAEC-CM. hAEC-EV-based therapy may be a potential therapeutic option for liver fibrosis.

Human engineered heart tissues have potential to revolutionize cardiac development research, drug-testing, and treatment of heart disease; however, implementation is limited by the need to use pre-differentiated cardiomyocytes (CMs). Here we show that by providing a 3D poly(ethylene glycol)-fibrinogen hydrogel microenvironment, we can directly differentiate human pluripotent stem cells (hPSCs) into contracting heart tissues. Our straight-forward, ontomimetic approach, imitating the process of development, requires only a single cell-handling step, provides reproducible results for a range of tested geometries and size scales, and overcomes inherent limitations in cell maintenance and maturation, while achieving high yields of CMs with developmentally appropriate temporal changes in gene expression. Here we demonstrate that hPSCs encapsulated within this biomimetic 3D hydrogel microenvironment develop into functional cardiac tissues composed of self-aligned CMs with evidence of ultrastructural maturation, mimicking heart development, and enabling investigation of disease mechanisms and screening of compounds on developing human heart tissue.

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Liver cirrhosis is the 6th leading cause of death in adults aged 15–59 years in high-income countries. For many who progress to cirrhosis, the only prospect for survival is liver transplantation. While there is some indication that mesenchymal stem cells may be useful in reversing established liver fibrosis, there are limitations to their widespread use – namely their rarity, the need for extensive serial passaging and the associated potential for genomic instability and cellular senescence. To this end, we propose the use of allogeneic amnion epithelial cells. This clinical trial will assess the safety of intravenously delivered allogeneic human amnion epithelial cells (hAECs) in patients with compensated liver cirrhosis. This will also provide clinical data that will inform phases 2 and 3 clinical trials with the ultimate goal of developing hAECs as a therapeutic option for patients with cirrhosis who are at significant risk of disease progression. We will recruit 12 patients with compensated cirrhosis, based on their hepatic venous pressure gradient, for a dose escalation study. Patients will be closely monitored in the first 24 h post-infusion, then via daily telephone interviews until clinical assessment on day 5. Long term follow up will include standard liver tests, transient elastography and hepatic ultrasound. Ethics approval was obtained from Monash Health for this trial 16052A, “A Pilot Study Evaluating the Safety of Intravenously Administered Human Amnion Epithelial Cells for the Treatment of Liver Fibrosis, A First in Adult Human Study.” The trial will be conducted in accordance to Monash Health Human Ethics guidelines. Outcomes from this study will be disseminated in the form of conference presentations and submission to a peer reviewed journal. This trial has been registered on the Australian and New Zealand Clinical Trials Registry ACTRN12616000437460.