Hirsova, Petra

The search for non-invasive, fast, and low-cost diagnostic tools has gained significant traction among many researchers worldwide. Dielectric properties calculated from microwave signals offer unique insights into biological tissue. Material properties, such as relative permittivity (εr) and conductivity (σ), can vary significantly between healthy and unhealthy tissue types at a given frequency. Understanding this difference in properties is key for identifying the disease state. The frequency-dependent nature of the dielectric measurements results in large datasets, which can be postprocessed using artificial intelligence (AI) methods. In this work, the dielectric properties of liver tissues in three mouse models of liver disease are characterized using dielectric spectroscopy. The measurements are grouped into four categories based on the diets or disease state of the mice, i.e., healthy mice, mice with non-alcoholic steatohepatitis (NASH) induced by choline-deficient high-fat diet, mice with NASH induced by western diet, and mice with liver fibrosis. Multi-class classification machine learning (ML) models are then explored to differentiate the liver tissue groups based on dielectric measurements. The results show that the support vector machine (SVM) model was able to differentiate the tissue groups with an accuracy up to 90%. This technology pipeline, thus, shows great potential for developing the next generation non-invasive diagnostic tools.

Nonalcoholic steatohepatitis (NASH) has become a growing public health problem worldwide, yet its pathophysiology remains unclear. Liver sinusoidal endothelial cells (LSEC) have unique morphology and function, and play a critical role in liver homeostasis. Emerging literature implicates LSEC in many pathological processes in the liver, including metabolic dysregulation, inflammation, angiogenesis, and carcinogenesis. In this review, we highlight the current knowledge of the role of LSEC in each of the progressive phases of NASH pathophysiology (steatosis, inflammation, fibrosis, and the development of hepatocellular carcinoma). We discuss processes that have important roles in NASH progression including the detrimental transformation of LSEC called “capillarization”, production of inflammatory and profibrogenic mediators by LSEC as well as LSEC-mediated angiogenesis. The current review has a special emphasis on LSEC adhesion molecules, and their key role in the inflammatory response in NASH. Moreover, we discuss the pathogenic role of extracellular vesicles and their bioactive cargos in liver intercellular communication, inflammation, and fibrosis. Finally, we highlight LSEC-adhesion molecules and derived bioactive product as potential therapeutic targets for human NASH.

Background and Aims During liver fibrosis, liver sinusoidal endothelial cells (LSECs) release angiocrine signals to recruit inflammatory cells into the liver. p300, a master regulator of gene transcription, is associated with pathological inflammatory response. Therefore, we examined how endothelial p300 regulates angiocrine signaling and inflammation related to portal hypertension and fibrogenesis. Approach and Results CCl4 or partial inferior vena cava ligation (pIVCL) was used to induce liver injury. Mice with LSEC‐specific p300 deletion (p300 LSECΔ/Δ) or C‐C motif chemokine ligand 2 (Ccl2) deficiency, nuclear factor kappa B (NFκB)–p50 knockout mice, and bromodomain containing 4 (BRD4) inhibitors in wild‐type mice were used to investigate mechanisms of inflammation regulation. Leukocytes were analyzed by mass cytometry by time‐of‐flight. Epigenetic histone marks were modified by CRISPR endonuclease‐deficient CRISPR‐associated 9‐fused with the Krüppel associated box domain (CRISPR‐dCas9‐KRAB)–mediated epigenome editing. Portal pressure and liver fibrosis were reduced in p300 LSECΔ/Δ mice compared to p300 fl/fl mice following liver injury. Accumulation of macrophages was also reduced in p300 LSECΔ/Δ mouse livers. Ccl2 was the most up‐regulated chemokine in injured LSECs, but its increase was abrogated in p300 LSECΔ/Δ mice. While the macrophage accumulation was increased in NFκB‐p50 knockout mice with enhanced NFκB activity, it was reduced in mice with LSEC‐specific Ccl2 deficiency and mice treated with specific BRD4 inhibitors. In vitro, epigenome editing of CCL2 enhancer and promoter regions by CRISPR‐dCas9‐KRAB technology repressed TNFα‐induced CCL2 transcription through H3K9 trimethylation. In contrast, TNFα activated CCL2 transcription by promoting p300 interaction with NFκB and BRD4, leading to histone H3 lysine 27 acetylation at CCL2 enhancer and promoter regions. Conclusions In summary, endothelial p300 interaction with NFκB and BRD4 increases CCL2 expression, leading to macrophage accumulation, portal hypertension, and liver fibrosis. Inhibition of p300 and its binding partners might serve as therapy in the treatment of liver diseases.

AbstractNonalcoholic hepatitis (NASH) is the progressive inflammatory form of nonalcoholic fatty liver disease. Although the mechanisms of hepatic inflammation in NASH remain incompletely understood, emerging literature implicates the proinflammatory environment created by toxic lipid-induced hepatocyte injury, termed lipotoxicity. Interestingly, numerous NASH-promoting kinases in hepatocytes, immune cells, and adipocytes are activated by the lipotoxic insult associated with obesity. In the current review, we discuss recent advances in NASH-promoting kinases as disease mediators and therapeutic targets. The focus of the review is mainly on the mitogen-activated protein kinases including mixed lineage kinase 3, apoptosis signal-regulating kinase 1, c-Jun N-terminal kinase, and p38 MAPK; the endoplasmic reticulum (ER) stress kinases protein kinase RNA-like ER kinase and inositol-requiring protein-1α; as well as the Rho-associated protein kinase 1. We also discuss various pharmacological agents targeting these stress kinases in NASH that are under different phases of development.

Fatty acid‐induced upregulation of death receptor 5 (DR5) and its cognate ligand, tumor necrosis factor‐related apoptosis‐inducing ligand (TRAIL), promotes hepatocyte lipoapoptosis, which is a key mechanism in the progression of fatty liver disease. Accordingly, inhibition of DR5 signaling represents an attractive strategy for treating fatty liver disease. Ligand competition is prevalent in tumor necrosis factor receptor antagonism, but recent studies suggest a compelling alternative: non‐competitive inhibition through perturbation of receptor conformation. We used a yeast‐displayed combinatorial library to identify a synthetic affibody that specifically binds DR5. Biophysical and biochemical studies show that the affibody neither blocks TRAIL binding nor prevents the receptor‐receptor interaction. Live‐cell fluorescence lifetime measurements indicate that the affibody induces a conformational change in transmembrane dimers of DR5 and favors an inactive state of the receptor. The affibody inhibits apoptosis in TRAIL‐ or palmitate‐treated Huh‐7 cells, an in vitro model of fatty liver disease. Thus, this lead affibody serves as a potential drug candidate, with a unique mechanism of action, for fatty liver disease. Support or Funding Information National Institutes of Health Grants R01EB023339, R01DK41876, R35GM131814 and R01EB028274.

With the epidemic of obesity, nonalcoholic fatty liver disease (NAFLD) has become the most common pediatric liver disease. The influence of a perinatal obesity‐inducing diet (OID) on the development and progression of NAFLD in offspring is important but incompletely studied. Hence, we fed breeding pairs of C57BL/6J mice during gestation and lactation (perinatally) either chow or an OID rich in fat, fructose, and cholesterol (FFC). The offspring were weaned to either chow or an FFC diet, generating four groups: perinatal (p)Chow‐Chow, pChow‐FFC, pFFC‐Chow, and pFFC‐FFC. Mice were sacrificed at 10 weeks of age. We examined the whole‐liver transcriptome by RNA sequencing (RNA‐seq) and whole‐liver genome methylation by reduced representation bisulfite sequencing (RRBS). Our results indicated that the pFFC‐FFC mice had a significant increase in hepatic steatosis, injury, inflammation, and fibrosis, as assessed histologically and biochemically. We identified 189 genes that were differentially expressed and methylated in the pFFC‐FFC mice versus the pChow‐FFC mice. Gene set enrichment analysis identified hepatic fibrosis/hepatic stellate cell activation as the top canonical pathway, suggesting that the differential DNA methylation events in the mice exposed to the FFC diet perinatally were associated with a profibrogenic transcriptome. To verify that this finding was consistent with perinatal nutritional reprogramming of the methylome, we exposed pFFC‐Chow mice to an FFC diet in adulthood. These mice developed significant hepatic steatosis, injury, inflammation, and more importantly fibrosis when compared to the appropriate controls.Conclusion: Perinatal exposure to an OID primes the immature liver for an accentuated fibrosing nonalcoholic steatohepatitis (NASH) phenotype, likely through nutritional reprogramming of the offspring methylome. These data have potential clinical implications for monitoring children of obese mothers and risk stratification of children with NAFLD.