Abstract:Nonalcoholic steatohepatitis (NASH) is an emerging health crisis with no approved therapies. Obeticholic acid (OCA), a farnesoid X receptor (FXR) agonist, shows promise in NASH trials. However, the precise mechanisms mediating OCA effects and impact on cholesterol metabolism are not fully understood. We explored the pharmaco-toxicological effects of OCA on pathophysiological pathways in hepatocytes using a previously described perfused organotypic liver system that allows culture in near-physiological insulin/… Show more
“…Downregulation of diacylglycerol acyltransferase 2 (DGAT2) by OCA by greater than 4-fold is evident at 4 h and maintained up to 18 h ( Fig. 6 C), in accordance with a previous study in an organotypic liver model ( Dash et al, 2017 ). Fig.…”
Hepatic steatosis, a reversible state of metabolic dysregulation, can promote the onset of nonalcoholic steatohepatitis (NASH), and its transition is thought to be critical in disease evolution. The association between endoplasmic reticulum (ER) stress response and hepatocyte metabolism disorders prompted us to characterize ER stress-induced hepatic metabolic dysfunction in human induced pluripotent stem cell-derived hepatocytes (hiPSC-Hep), to explore regulatory pathways and validate a phenotypic in vitro model for progression of liver steatosis. We treated hiPSC-Hep with a ratio of unsaturated and saturated fatty acids in the presence of an inducer of ER stress to synergistically promote triglyceride accumulation and dysregulate lipid metabolism. We monitored lipid accumulation by high-content imaging and measured gene regulation by RNA sequencing and reverse transcription quantitative PCR analyses. Our results show that ER stress potentiated intracellular lipid accumulation by 5-fold in hiPSC-Hep in the absence of apoptosis. Transcriptome pathway analysis identified ER stress pathways as the most significantly dysregulated of all pathways affected. Obeticholic acid dose dependently inhibited lipid accumulation and modulated gene expression downstream of the farnesoid X receptor. We were able to identify modulation of hepatic markers and gene pathways known to be involved in steatosis and nonalcoholic fatty liver disease (NAFLD), in support of a hiPSC-Hep disease model that is relevant to clinical data for human NASH. Our results show that the model can serve as a translational discovery platform for the understanding of molecular pathways involved in NAFLD, and can facilitate the identification of novel therapeutic molecules based on high-throughput screening strategies.
“…Downregulation of diacylglycerol acyltransferase 2 (DGAT2) by OCA by greater than 4-fold is evident at 4 h and maintained up to 18 h ( Fig. 6 C), in accordance with a previous study in an organotypic liver model ( Dash et al, 2017 ). Fig.…”
Hepatic steatosis, a reversible state of metabolic dysregulation, can promote the onset of nonalcoholic steatohepatitis (NASH), and its transition is thought to be critical in disease evolution. The association between endoplasmic reticulum (ER) stress response and hepatocyte metabolism disorders prompted us to characterize ER stress-induced hepatic metabolic dysfunction in human induced pluripotent stem cell-derived hepatocytes (hiPSC-Hep), to explore regulatory pathways and validate a phenotypic in vitro model for progression of liver steatosis. We treated hiPSC-Hep with a ratio of unsaturated and saturated fatty acids in the presence of an inducer of ER stress to synergistically promote triglyceride accumulation and dysregulate lipid metabolism. We monitored lipid accumulation by high-content imaging and measured gene regulation by RNA sequencing and reverse transcription quantitative PCR analyses. Our results show that ER stress potentiated intracellular lipid accumulation by 5-fold in hiPSC-Hep in the absence of apoptosis. Transcriptome pathway analysis identified ER stress pathways as the most significantly dysregulated of all pathways affected. Obeticholic acid dose dependently inhibited lipid accumulation and modulated gene expression downstream of the farnesoid X receptor. We were able to identify modulation of hepatic markers and gene pathways known to be involved in steatosis and nonalcoholic fatty liver disease (NAFLD), in support of a hiPSC-Hep disease model that is relevant to clinical data for human NASH. Our results show that the model can serve as a translational discovery platform for the understanding of molecular pathways involved in NAFLD, and can facilitate the identification of novel therapeutic molecules based on high-throughput screening strategies.
“…OCA was found in the clinic to raise LDL, a risk not observed in pre-clinical animal studies. Both hPCLS and the lipotoxic system reported transcriptional signatures that may explain the cholesterol endpoints observed in the clinical trials for OCA [26, 35]. Additionally, in the lipotoxic system, OCA treatment induced high secretion of ApoB (the primary apoprotein associated with LDL) and increased intracellular levels of cholesterol, suggesting dysregulation of this pathway, in agreement with what was observed in the clinic [31, 34].…”
Section: Human In Vitro Organotypic Models Of Nafldsupporting
confidence: 66%
“…Ljssennagger et al used hPCLS to evaluate the farnesoid X receptor (FXR)-agonist, obeticholic acid (OCA, 1 µM for 24 h), which is currently in Phase 3 clinical trials for the treatment of NASH, and reported that gene expression profiling revealed on-target engagement of FXR [26]. Similarly, the lipotoxic system developed by Feaver et al observed several clinically-relevant effects with 0.5-µM OCA treatment, including on-target effects and reductions in steatosis, inflammatory cytokines, fibrotic biomarkers, and triglycerides—all of which were observed in the Phase 2b clinical trial [31, 34, 35]. OCA was found in the clinic to raise LDL, a risk not observed in pre-clinical animal studies.…”
Section: Human In Vitro Organotypic Models Of Nafldmentioning
Non-alcoholic fatty liver disease (NAFLD) is the most common cause of chronic liver disease in the Western world, affecting about 1/3 of the US general population and remaining as a significant cause of morbidity and mortality. The hallmark of the disease is the excessive accumulation of fat within the liver cells (hepatocytes), which eventually paves the way to cellular stress, injury and apoptosis. NAFLD is strongly associated with components of the metabolic syndrome and is fast emerging as a leading cause of liver transplant in the USA. Based on clinico-pathologic classification, NAFLD may present as isolated lipid collection (steatosis) within the hepatocytes (referred to as non-alcoholic fatty liver; NAFL); or as the more aggressive phenotype (known as non-alcoholic steatohepatitis; NASH). There are currently no regulatory agency-approved medication for NAFLD, despite the enormous work and resources that have gone into the study of this condition. Therefore, there remains a huge unmet need in developing and utilizing pre-clinical models that will recapitulate the disease condition in humans. In line with progress being made in developing appropriate disease models, this review highlights the cutting-edge preclinical in vitro and animal models that try to recapitulate the human disease pathophysiology and/or clinical manifestations.
“…This feature has key implications for the application of tissue slice cultures in bioscience. Notably, slice cultures are more resistant to the action of some drugs or toxins than dissociated, 2D-cultures (Miller and Miller, 1985;Elkayam and Amitay-Shaprut, 2006;Olinga and Schuppan, 2013), and respond differently to physiological stimuli such as insulin compared to organotypic cultures from liver tissue (Dash and Figler, 2017).…”
Once surgical tissue is available, the current protocol is easy to perform and produces functional slices from adult human brain. These slice cultures may represent a preferred model for translational studies of neurodegenerative disorders when long term culturing in not required, as in investigations on AβO neurotoxicity.
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