Recent advances in 2D and 3D in vitro systems using primary hepatocytes, alternative hepatocyte sources and non-parenchymal liver cells and their use in investigating mechanisms of hepatotoxicity, cell signaling and ADME

Archives of Toxicology, Journal Year: 2013, Volume and Issue: 87(8), P. 1315 - 1530

Published: Aug. 1, 2013

This review encompasses the most important advances in liver functions and hepatotoxicity analyzes which mechanisms can be studied vitro. In a complex architecture of nested, zonated lobules, consists approximately 80 % hepatocytes 20 non-parenchymal cells, latter being involved secondary phase that may dramatically aggravate initial damage. Hepatotoxicity, as well hepatic metabolism, is controlled by set nuclear receptors (including PXR, CAR, HNF-4α, FXR, LXR, SHP, VDR PPAR) signaling pathways. When isolating some pathways are activated, e.g., RAS/MEK/ERK pathway, whereas others silenced (e.g. HNF-4α), resulting up- downregulation hundreds genes. An understanding these changes crucial for correct interpretation vitro data. The possibilities limitations useful systems summarized, including three-dimensional culture techniques, co-cultures with hepatospheres, precision cut slices isolated perfused liver. Also discussed how closely hepatoma, stem cell iPS cell-derived hepatocyte-like-cells resemble real hepatocytes. Finally, summary given state art mathematical modeling currently used pharmaceutical industry an emphasis on drug prediction clearance, interaction, transporter studies hepatotoxicity. One key message despite our enthusiasm systems, we must never lose sight vivo situation. Although have been decades, hunt relevant alternative has only just begun.

Language: Английский

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Liver inflammation and fibrosis

Journal of Clinical Investigation, Journal Year: 2017, Volume and Issue: 127(1), P. 55 - 64

Published: Jan. 2, 2017

Chronic liver inflammation leads to fibrosis and cirrhosis, which is the 12th leading cause of death in United States. Hepatocyte steatosis a component metabolic syndrome insulin resistance. Hepatic may be benign or progress hepatocyte injury initiation inflammation, activates immune cells. While Kupffer cells are resident macrophage liver, inflammatory such as infiltrating macrophages, T lymphocytes, neutrophils, DCs all contribute inflammation. The activate hepatic stellate cells, major source myofibroblasts liver. Here we review their crosstalk with myofibroblasts.

Language: Английский

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Hepatic lipotoxicity and the pathogenesis of nonalcoholic steatohepatitis: The central role of nontriglyceride fatty acid metabolites

Hepatology, Journal Year: 2010, Volume and Issue: 52(2), P. 774 - 788

Published: April 19, 2010

"…the cause of steatosis, and not the fat accumulation by itself, produces cirrhosis"—Heribert Thaler, 19751 W ith nonalcoholic steatohepatitis (NASH), NASH cirrhosis, NASH-related hepatocellular carcinoma becoming increasingly prevalent, need for effective therapies has never been greater. Unfortunately, our understanding what causes at molecular level remains mostly speculative, thus ability to design clinical trials that test rationally designed is limited. More than a decade passed since Christopher Day Oliver James first proposed oft-cited two-hit hypothesis fatty liver disease (NAFLD) explain pathogenesis NASH.2, 3 According this appealing hypothesis, lipid in form triglyceride needed development constitutes "hit" disease. The injury setting lipid-loaded hepatocytes was be oxidant stress leading peroxidation milieu ample substrate. This second then triggers necroinflammatory changes we recognize histopathologically as NASH. Although intuitive appeal, emerging data now suggests liver, other organs, droplets truly just an "innocent bystander" processes cellular inflammation,4, 5 explanation presciently suggested eminent Austrian pathologist Heribert Thaler,1 considered others,6 acknowledged viable alternative when they hypothesis.3 Convincing evidence central role causing also established. A research confirmed these occur,7 but studies have unable prove or are necessary humans. As points metabolites acids real culprits NASH, target organs lipotoxicity (Fig. 1).8-12 Because often accumulates parallel process during lipotoxic injury, it understandably difficult separate respective roles acid cell death. with all good hypotheses, nontriglyceride generates more questions can currently answer. Fortunately, provides new avenues investigators pursue search treatments common model Emerging indicate free manifested ER stress, inflammation, apoptosis, necrosis, dysmorphic features such ballooning Mallory-Denk body formation. generation typically occurs (steatosis), resulting phenotype recognized where steatosis present together. feature distinguishes from earlier models and, fact, may protective. caveat does "progress" Metabolic abnormalities predisposing include increased supply impaired disposal acids. Insulin resistance plays allowing excessive flow adipose tissue impairing peripheral glucose disposal. Fatty through oxidative pathways formation which stored temporarily secreted VLDL. Supply-side salutary reduction carbohydrate precursors de novo lipogenesis prevention inappropriate lipolysis improving adipocyte insulin responsiveness. Favorable on side increasing (with intracellular antioxidant defenses must adequate handle ROS produced) triglyceride. Red arrows identify contribute flux hepatocytes, promoting injury; green eliminate reduce injury. Not shown mechanistic aspects repair, fibrogenesis ultimately determine whether leads cirrhosis. ER, endoplasmic reticulum; IR, resistance; P450, cytochrome P450 mixed function oxidases; ROS, reactive oxygen species; SER, smooth VLCFA, very long chain acids; VLDL, low density lipoprotein. CoA, coenzyme A; CYP, P450; DAG, diacylglycerol; DGAT2, diacylglycerol acyltransferase 2; DNL, lipogenesis; FABP, binding proteins; JNK, c-Jun N-terminal kinase; LPC, lysophosphatidyl choline; NAFLD, disease; steatohepatitis; PPAR, peroxisome proliferator-activated receptor; SCD-1, stearoyl-CoA desaturase-1; SREBP-1c, sterol regulatory element protein 1c; Lipotoxicity term coined Unger 15 years ago describe toxic effects pancreatic beta survival.13 formally defined used variably death caused acids, their metabolites, even triglyceride, although generally found relatively inert.8, 14 wide variety derived having varying lengths degrees locations desaturation, mention cis versus trans configuration double bonds, number permutations enormous, making identification species challenging.15 Identifying important derivatives area intense lipidomic study understand diseases heart, islets, vasculature, well liver.10, 12, 16-21 Shown Fig. 2 few many putative mediators lipotoxicity. We should anticipate advance discovered, list will appear quite naïve retrospect. Possible acid–derived Currently available suggest participate reintroduction play regulating exposure cells intermediaries highly regulated family proteins.41, 67, 125 specificity regulation reviewed.66, 69, 70, 72 ACSL, acyl-CoA synthase; AGPAT, acyl-glycerolphosphate acyltransferase; ATGL, lipase; CPT, choline phosphotransferase; DAGK, DGATs, acyltransferases; GPAT, glycerol monophosphate HSL, hormone-sensitive LPAAT, lysophosphatidic LPAP, phosphatase; LysoPLD, lysophospholipase D; MAGK, monoacylglycerol MGL, MOGAT, PLA, phospholipase PLD, D. once thought underlying muscle, tissues, phenomenon altered signaling pathways.12, 14, 22-24 Similarly, implicating within hepatocyte human animal demonstrate only association between two phenomena, rather showing causality. However, probably totally benign. When extreme, some might compromise hepatic blood simply compression sinusoids necrosis,25 debated.26 Excessive render donor livers susceptible ischemia reperfusion transplantation. associated markers reticulum response uncertain.27 Mice overexpressing (DGAT2), final enzyme catalyzing formation, phosphorylation PERK (protein kinase R–like kinase), marker stress. mice did exhibit increases proinflammatory (JNK) nuclear factor-κB activation, elevated steatohepatitis.27, 28 Examined opposite way, blocking DGAT2 expression prevented acid–induced prevent both culture mice.22, 29 Overall, experiments manipulate discern consistent demonstrating there conflicting data, possibly because compensatory up-regulation under various experimental conditions burden confound interpretation results.30-32 additional dissociated droplet For example, inhibiting incorporation into nascent lipoprotein (VLDL) microsomal transfer secretion accumulation, yet injury.33 view emerged recent actually protective prevents species.4, 5, 10, 34 Early demonstrated cardiomyocytes,35 subsequently, effect cultured islet cells36 hepatocytes.37 storage temporizing measure; released point, if still them appropriately metabolic pathways, could serve source intermediates.38 Free noxious biological systems. mechanism, circulating bound high affinity albumin, proteins (FABPs) mechanisms uptake (e.g., adipocytes, enterocytes). Measurement concentrations tissues provide information about net system, patients increased.39-41 tightly FABPs, levels FABP reflective how much flowing system. Studies concentration fairly constant liver,17 seen NASH,42 suggesting elevations unlikely lipotoxicity, whereas delivery enzymes organelles markedly increased.14 On hand, overexpression cardiac promotes lipotoxicity,43 deletion lines,44, 45 observations confirm system facilitating FABPs conduit use substrates, bigger (i.e., higher concentrations) delivered. decreased immunostaining compared those no steatohepatitis, significance uncertain.46 shuttled off so responsible lipotoxicity.38 Saturated activating ligands toll-like receptor-4, cascade events precipitating apoptosis.47 One inhibition loss receptor-4 mouse models.48 capable directly destabilizing lysosomal membranes, release cathepsin B activation apoptotic pathways.49, 50 receptor alpha (PPARα), PPARγ, G protein–coupled receptors, mediate diverse indirectly causative lipotoxicity.11 Activation PPARα PPARγ serves adaptive against respectively. Despite acyl-coenzyme (acyl-CoA) requisite step Experiments Chang β-cells synthesis triacsin C saturated acids.51, 52 Therefore, likely Ceramides were initially major injury53 disease.16, 54 Ceramide cells,55 several ceramide types diminish fumonisin B1 palmitate liver-derived lines,51, 56 ceramides seem involved palmitate-induced Chinese hamster ovary despite being elevated.57 Whether results relevant shown, pointing candidates. Diacylglycerol (DAG) well-known most isoforms, group kinases essential pathways. Thus, DAG carefully coordinate functions DAGs signal transduction substrates phospholipids.58 Increases suspected contributor lipotoxicity;34, 59 however, analysis biopsy specimens NAFLD without similar levels, had normal liver.17 reflect trafficking causally related steatohepatitis. Further further clarify Phosphatidyl choline, lecithin, component membrane bilayers envelop monolayers, VLDL required its secretion. Removal one sn-2 position (the middle backbone) (LPC), biologically active molecule implicated direct monocyte activation.60 LPC mediator NASH.51 In study, biopsies small proportion severity. using lines, primary culture, mice. Mechanistically, LPC-induced appeared depend unidentified induction apoptosis mitochondrial depolarization. generated action A2 inhibitors culture. needs validation, provocative identity potential known unknown, molecules act specific phosphatidic promote bending folding, interact large molecules. homeostatic metabolism running smoothly, termed "liporegulation",53 production minimized. genetic stresses alter homeostasis either precursor Up-regulation enzymatic generate intermediates dispose speculated remain inform us hypothetical Stresses liporegulation characterized primarily models, corroborating findings seminal briefly reviewed below. 4 decades.61 sources blood, (DNL), excess carbohydrates amino 1).62 Adipose delivered DNL normally contributes 5%; up 26% acids.40 Relatively minor contributors pool short gut breakdown remnants autophagosomes.63 Except limited residual remnants, triglycerides liver. designing efforts serum pharmacologically significant benefit liver.31, 33 synthesis, described.62, 64 Enzymatic hydrolysis circulation interplay hormonal, neurologic, pharmacologic stimuli complex simple lipase, (Table 1).65 mediated neutral lipases, proteins, regulators cyclic adenosine levels.66-71 inhibitor lipolytic discussed below, largely diseases. tissue, re-esterified return adipocytes; seemingly futile cycling thermogenesis consumes triphosphate.72 prevailing liberated lipases transported newer adipocytes fate, playing energy wasting thermogenesis,73 continues debated.70 Bile regulate tissue. Acting TGR5 increase bile activate thyroid hormone locally thermogenesis.74, 75 impact modulated muscle uptake. It unbridled lipolysis, prolonged fasting, send handle.76, 77 Older adverse fasting accelerating injury,78, 79 attenuated muscle. Muscle-specific lipase spares uptake, expense deleterious organs.80 Also, strains develop fasting-induced biogenesis particularly prone steatosis,81 SJL/J strain oxidation tolerates developing steatosis.82 Manipulating recognizing defects humans predispose sedentary behaviors certainly areas exploration trials, especially respect exercise capacity assumption would equally. fate presented dietary intake Dietary habits consumption, fructose corn syrup,83, 84 overwhelming liver's DNL. Fructose enters glycolytic pathway downstream phosphofructokinase, rate-limiting pathway, unregulated acetyl-CoA substrate DNL.84, 85 importance sugars steatohepatitis,86 cross-sectional linked consumption advanced NASH.87 To efficiently incorporate newly synthesized fraction undergo desaturation monounsaturated backbone optimally esterified unsaturated acid. desaturase-1 (SCD-1) efficient absence Recent SCD-1 lipotoxicity.23 forcing diet sucrose deficient fats severe lacking SCD-1.88 Such add Previous accompanied pathways,89, 90 indicating available, averted.91, 92 De amplified 1c (SREBP-1c), transcription factor inducing Overexpression SREBP-1c identified injury.93, 94 physiologically down-regulated acting farnesoid X receptor, rationale pursuing means thwarting preventing NASH.95, 96 Uptake chylomicron Recently comparison autophagy releasing droplets. Termed lipophagy, turnover droplets.63, 97 Autophagosome way reducing investigated. complicated difficulties knowing drugs manipulations affecting both. Inhibiting 18 β-glycyrrhetinic lard-fed rats HepG2 cells, latter finding response.98 An problem robust. oxidize carbon dioxide water three different caveat, endogenous able (ROS) superoxide hydrogen peroxide inevitably amounts electrons transferred bonds oxidized. coincidentally, well-endowed antioxidants. Glutathione mitochondria; glutathione eliminates peroxides via peroxidase abundant S-transferases Substantial depletion reliably occur NASH,99, 100 circumstances, cytoprotective mechanism handling load acids.7, 101-104 mitochondria cannot make glutathione, sensitive depleted, steatohepatitis.105, 106 After meals, away β-oxidation By comparison, state, triphosphate ketone bodies. Partitioning controlled. β-cells, shut induced lipotoxicity.52 after feeding insulin-resistant switch puts disadvantage Deleting carboxylase 2, gatekeeper β-oxidation, diet-induced injury.107 Mitochondrial dysfunction, acquired, steatohepatitis.64, 108-111 develops circumstances due solely

Language: Английский

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Mitochondrial Glutathione, a Key Survival Antioxidant

Antioxidants and Redox Signaling, Journal Year: 2009, Volume and Issue: 11(11), P. 2685 - 2700

Published: June 26, 2009

Mitochondria are the primary intracellular site of oxygen consumption and major source reactive species (ROS), most them originating from mitochondrial respiratory chain. Among arsenal antioxidants detoxifying enzymes existing in mitochondria, glutathione (mGSH) emerges as main line defense for maintenance appropriate redox environment to avoid or repair oxidative modifications leading dysfunction cell death. mGSH importance is based not only on its abundance, but also versatility counteract hydrogen peroxide, lipid hydroperoxides, xenobiotics, mainly a cofactor such peroxidase glutathione-S-transferase (GST). Many death-inducing stimuli interact with causing stress; addition, numerous pathologies characterized by consistent decrease levels, which may sensitize additional insults. From evaluation influence different pathologic settings hypoxia, ischemia/reperfusion injury, aging, liver diseases, neurologic disorders, it becoming evident that has an important role pathophysiology biomedical strategies aimed boost levels.

Language: Английский

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The interaction of hepatic lipid and glucose metabolism in liver diseases

Journal of Hepatology, Journal Year: 2011, Volume and Issue: 56(4), P. 952 - 964

Published: Dec. 13, 2011

It is widely known that the liver a central organ in lipogenesis, gluconeogenesis and cholesterol metabolism. However, over last decades, variety of pathological conditions highlighted importance metabolic functions within diseased liver. As observed Western societies, an increase prevalence obesity syndrome promotes pathophysiological changes cause non-alcoholic fatty disease (NAFLD). NAFLD increases susceptibility to acute injury may lead cirrhosis hepatocellular cancer. Alterations insulin response, β-oxidation, lipid storage transport, autophagy imbalance chemokines nuclear receptor signaling are held accountable for these changes. Furthermore, recent studies revealed role accumulation inflammation ER stress clinical context regeneration hepatic carcinogenesis. This review focuses on novel findings related - including vitamin D homolog 1 glucose uptake, metabolism NAFLD, regeneration,

Language: Английский

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Liver Fibrosis: Mechanistic Concepts and Therapeutic Perspectives

Cells, Journal Year: 2020, Volume and Issue: 9(4), P. 875 - 875

Published: April 3, 2020

Liver fibrosis due to viral or metabolic chronic liver diseases is a major challenge of global health. Correlating with disease progression, key factor for outcome and risk hepatocellular carcinoma (HCC). Despite different mechanism primary injury disease-specific cell responses, the progression fibrotic follows shared patterns across main etiologies. Scientific discoveries within last decade have transformed understanding mechanisms fibrosis. Removal elimination causative agent such as control cure infection has shown that reversible. However, reversal often occurs too slowly infrequent avoid life-threatening complications particularly in advanced Thus, there huge unmet medical need anti-fibrotic therapies prevent HCC development. while many candidate agents robust effects experimental animal models, their clinical trials been limited absent. no approved therapy exists In this review we summarize cellular drivers molecular fibrogenesis discuss impact development urgently needed therapies.

Language: Английский

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Free fatty acids promote hepatic lipotoxicity by stimulating TNF-α expression via a lysosomal pathway

Hepatology, Journal Year: 2004, Volume and Issue: 40(1), P. 185 - 194

Published: July 1, 2004

Nonalcoholic fatty liver disease (NAFLD) is a serious health problem. Although NAFLD represents form of lipotoxicity, its pathogenesis remains poorly understood. The aim this study was to examine the cellular mechanisms involved in free acid (FFA)-mediated hepatic lipotoxicity. FFA treatment cells resulted Bax translocation lysosomes and lysosomal destabilization with release cathepsin B (ctsb), cysteine protease, into cytosol. This process also partially dependent on ctsb. Lysosomal nuclear factor kappa B-dependent tumor necrosis alpha expression. Release ctsb cytoplasm observed humans correlated severity. In dietary murine model NAFLD, either genetic or pharmacological inactivation protected against development steatosis, injury, insulin resistance associated "dysmetabolic syndrome." conclusion, these data support lipotoxic FFA-mediated destabilization.

Language: Английский

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Endoplasmic reticulum stress signalling and the pathogenesis of non-alcoholic fatty liver disease

Journal of Hepatology, Journal Year: 2018, Volume and Issue: 69(4), P. 927 - 947

Published: June 27, 2018

Language: Английский

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Triggering and resolution of inflammation in NASH

Nature Reviews Gastroenterology & Hepatology, Journal Year: 2018, Volume and Issue: 15(6), P. 349 - 364

Published: May 8, 2018

Language: Английский

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Nonalcoholic fatty liver disease

Nature Reviews Disease Primers, Journal Year: 2015, Volume and Issue: 1(1)

Published: Dec. 16, 2015

Language: Английский

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