Background: Nonalcoholic fatty liver disease (NAFLD) is increasingly prevalent and represents a growing challenge in terms of prevention and treatment. A minority of affected patients develops inflammation, subsequently fibrosis, cirrhosis and hepatocellular carcinoma (HCC). HCC is a leading cause of cancer-related death. An increased number of senescent cells correlate with age-related tissue degeneration during NAFLD-induced HCC. Senolytics are promising agents that target selectively senescent cells. Previous studies showed that whereas a combination of the senolytic drugs dasatinib and quercetin (D + Q) reduced NAFLD in mice, D + Q lacked efficacy in removing doxorubicin-induced β-gal-positive senescent cells in human HCC xenografted mice. Whether D + Q has an effect on the age-associated spectrum of NAFLD-inflammation-HCC remains unknown. Methods: Here, we utilized an established model of age- and obesity-associated HCC, the low dose diethylnitrosamine (DEN)/high fat diet (HFD), a regimen promoting liver inflammation and tumorigenesis over a long period of 9 months. Four groups of mice each were created: group 1 included control untreated mice; group 2 included mice treated with D + Q; group 3 included mice undergoing the DEN/HFD protocol; group 4 included mice undergoing the DEN/HFD protocol with the administration of D + Q. At the end of the chemical/dietary regimen, we analyzed liver damage and cell senescence by histopathology, qPCR and immunoblotting approaches. Results: Unexpectedly, D + Q worsened liver disease progression in the DEN/HFD mouse model, slightly increasing histological damage and tumorigenesis, while having no effect on senescent cells removal. Conclusions: In summary, using an animal model that fully recapitulates NAFLD, we demonstrate that these compounds are ineffective against age-associated NAFLD-induced HCC.

Type 1 diabetes mellitus (T1D) is a chronic autoimmune disease resulting in the destruction of insulin producing β-cells of the pancreas, with consequent insulin deficiency and excessive glucose production. Hyperglycemia results in increased levels of reactive oxygen species (ROS) and nitrogen species (RNS) with consequent oxidative/nitrosative stress and tissue damage. Oxidative damage of the pancreatic tissue may contribute to endothelial dysfunction associated with diabetes. The aim of the present study was to investigate if the potentially protective effects of phenethyl ester of caffeic acid (CAPE), a natural phenolic compound occurring in a variety of plants and derived from honeybee hive propolis, and of a novel CAPE analogue, as heme oxygenase-1 (HO-1) inducers, could reduce pancreatic oxidative damage induced by excessive amount of glucose, affecting the nitric oxide synthase/dimethylarginine dimethylaminohydrolase (NOS/DDAH) pathway in streptozotocin-induced type 1 diabetic rats. Our data demonstrated that inducible nitric oxide synthase/gamma-Glutamyl-cysteine ligase (iNOS/GGCL) and DDAH dysregulation may play a key role in high glucose mediated oxidative stress, whereas HO-1 inducers such as CAPE or its more potent derivatives may be useful in diabetes and other stress-induced pathological conditions.

Recent studies suggest that PGC1-α plays a crucial role in mitochondrial and vascular function, yet the physiological significance of PGC1α and HO expression in adipose tissues in the context of obesity-linked vascular dysfunction remains unclear. We studied three groups of six-week-old C57BL/6J male mice: (1) mice fed a normal chow diet; (2) mice fed a high-fat diet (H.F.D.) for 28 weeks, and (3) mice fed a high-fat diet (H.F.D.) for 28 weeks, treated with adipose-specific overexpression of PGC-1α (transgenic-adipocyte-PGC-1α) at week 20, and continued on H.F.D. for weeks 20–28. R.N.A. arrays examined 88 genes involved in adipocyte proliferation and maturation. Blood pressure, tissue fibrosis, fasting glucose, and oxygen consumption were measured, as well as liver steatosis, and the expression levels of metabolic and mitochondrial markers. Obese mice exhibited a marked reduction of PGC1α and developed adipocyte hypertrophy, fibrosis, hepatic steatosis, and decreased mitochondrial respiration. Mice with adipose-specific overexpression of PGC1-α exhibited improvement in HO-1, mitochondrial biogenesis and respiration, with a decrease in fasting glucose, reduced blood pressure and fibrosis, and increased oxygen consumption. PGC-1α led to the upregulated expression of processes associated with the browning of fat tissue, including UCP1, FGF21, and pAMPK signaling, with a reduction in inflammatory adipokines, NOV/CCN3 expression, and TGFβ. These changes required HO-1 expression. The R.N.A. array analysis identified subgroups of genes positively correlated with contributions to the browning of adipose tissue, all dependent on HO-1. Our observations reveal a positive impact of adipose-PGC1-α on distal organ systems, with beneficial effects on HO-1 levels, reversing obesity-linked cardiometabolic disturbances.

Aim: Obesity is associated with metabolic syndrome, hypertension, dyslipidemia, nonalcoholic fatty liver disease (NAFLD), and type 2 diabetes. In this study, we investigated whether the dietary supplementation of pomegranate seed oil (PSO) exerted a protective effect on liver lipid uptake, fibrosis, and mitochondrial function in a mouse model of obesity and insulin resistance. Method: In this in vivo study, eight-week-old C57BL/6J male mice were fed with a high fat diet (HFD) for 24 weeks and then were divided into three groups as follows: group (1) Lean; group (n = 6) (2) HF diet; group (n = 6) (3) HF diet treated with PSO (40 mL/kg food) (n = 6) for eight additional weeks starting at 24 weeks. Physiological parameters, lipid droplet accumulation, inflammatory biomarkers, antioxidant biomarkers, mitochondrial biogenesis, insulin sensitivity, and hepatic fibrosis were determined to examine whether PSO intervention prevents obesity-associated metabolic syndrome. Results: The PSO group displayed an increase in oxygen consumption, as well as a decrease in fasting glucose and blood pressure (p < 0.05) when compared to the HFD-fed mice group. PSO increased both the activity and expression of hepatic HO-1, downregulated inflammatory adipokines, and decreased hepatic fibrosis. PSO increased the levels of thermogenic genes, mitochondrial signaling, and lipid metabolism through increases in Mfn2, OPA-1, PRDM 16, and PGC1α. Furthermore, PSO upregulated obesity-mediated hepatic insulin receptor phosphorylation Tyr-⁹⁷², p-IRB tyr¹¹⁴⁶, and pAMPK, thereby decreasing insulin resistance. Conclusions: These results indicated that PSO decreased obesity-mediated insulin resistance and the progression of hepatic fibrosis through an improved liver signaling, as manifested by increased insulin receptor phosphorylation and thermogenic genes. Furthermore, our findings indicate a potential therapeutic role for PSO in the prevention of obesity-associated NAFLD, NASH, and other metabolic disorders.

Nonalcoholic fatty liver disease (NAFLD) has become the most common chronic liver disease in children and adolescents, increasing the risk of its progression toward nonalcoholic steatohepatitis (NASH), cirrhosis, and cancer. There is an urgent need for noninvasive early diagnostic and prognostic tools such as epigenetic marks (epimarks), which would replace liver biopsy in the future. We used plasma samples from 67 children with biopsy‐proven NAFLD, and as controls we used samples from 20 children negative for steatosis by ultrasound. All patients were genotyped for patatin‐like phospholipase domain containing 3 (PNPLA3), transmembrane 6 superfamily member 2 (TM6SF2), membrane bound O‐acyltransferase domain containing 7 (MBOAT7), and klotho‐β (KLB) gene variants, and data on anthropometric and biochemical parameters were collected. Furthermore, plasma cell‐free DNA (cfDNA) methylation was quantified using a commercially available kit, and ImageStream(X) was used for the detection of free circulating histone complexes and variants. We found a significant enrichment of the levels of histone macroH2A1.2 in the plasma of children with NAFLD compared to controls, and a strong correlation between cfDNA methylation levels and NASH. Receiver operating characteristic curve analysis demonstrated that combination of cfDNA methylation, PNPLA3 rs738409 variant, coupled with either high‐density lipoprotein cholesterol or alanine aminotransferase levels can strongly predict the progression of pediatric NAFLD to NASH with area under the curve >0.87. Conclusion: Our pilot study combined epimarks and genetic and metabolic markers for a robust risk assessment of NAFLD development and progression in children, offering a promising noninvasive tool for the consistent diagnosis and prognosis of pediatric NAFLD. Further studies are necessary to identify their pathogenic origin and function.

The progression of non-alcoholic fatty liver disease (NAFLD) and the development of hepatic fibrosis is caused by changes in redox balance, leading to an increase of reactive oxygen species (ROS) levels. NAFLD patients are at risk of progressing to non-alcoholic steatohepatitis (NASH), associated to cardiovascular diseases (CVD), coronary heart disease and stroke. Heme Oxygenase-1 (HO-1) is a potent endogenous antioxidant gene that plays a key role in decreasing oxidative stress. The present work was directed to determine whether use of an inhibitor of HO-1 activity affects lipid metabolism and fibrosis process in hepatic cells. Oil Red assay and mRNA analysis were used to evaluate the triglycerides content and the lipid metabolism pathway in HepG2 cells. ROS measurement, RT-PCR and Soluble collagen assay were used to assess the intracellular oxidant, the fibrosis pathway and the soluble collagen in LX2 cells. The activity of HO-1 was inhibited using Tin Mesoporphyrin IX (SnMP). Our study demonstrates that a non-functional HO system results in an increased lipid storage and collagen release in hepatocytes. Consequently, an increase of HO-1 levels may provide a therapeutic approach to address the metabolic alterations associated with NAFLD and its progression to NASH.