Quantitative high content imaging of cellular adaptive stress response pathways in toxicity for chemical safety assessment.

Over the past decade, major leaps forward have been made on the mechanistic understanding and identification of adaptive stress response landscapes underlying toxic insult using transcriptomics approaches. However, for predictive purposes of adverse outcome several major limitations in these approaches exist. First, the limited number of samples that can be analyzed reduces the in depth analysis of concentration-time course relationships for toxic stress responses. Second these transcriptomics analysis have been based on the whole cell population, thereby inevitably preventing single cell analysis. Third, transcriptomics is based on the transcript level, totally ignoring (post)translational regulation. We believe these limitations are circumvented with the application of high content analysis of relevant toxicant-induced adaptive stress signaling pathways using bacterial artificial chromosome (BAC) green fluorescent protein (GFP) reporter cell-based assays. The goal is to establish a platform that incorporates all adaptive stress pathways that are relevant for toxicity, with a focus on drug-induced liver injury. In addition, cellular stress responses typically follow cell perturbations at the subcellular organelle level. Therefore, we complement our reporter line panel with reporters for specific organelle morphometry and function. Here, we review the approaches of high content imaging of cellular adaptive stress responses to chemicals and the application in the mechanistic understanding and prediction of chemical toxicity at a systems toxicology level.

[1]  A. Mally,et al.  Role of drug-independent stress factors in liver injury associated with diclofenac intake. , 2013, Toxicology.

[2]  A. Levine,et al.  Nucleocytoplasmic shuttling of oncoprotein Hdm2 is required for Hdm2-mediated degradation of p53. , 1999, Proceedings of the National Academy of Sciences of the United States of America.

[3]  M. Sporn,et al.  Pharmacodynamic characterization of chemopreventive triterpenoids as exceptionally potent inducers of Nrf2-regulated genes , 2006, Molecular Cancer Therapeutics.

[4]  Y. Shiloh ATM and related protein kinases: safeguarding genome integrity , 2003, Nature Reviews Cancer.

[5]  Y. Shiloh,et al.  Rapid ATM-dependent phosphorylation of MDM2 precedes p53 accumulation in response to DNA damage. , 1999, Proceedings of the National Academy of Sciences of the United States of America.

[6]  O. Shirihai,et al.  Mitochondrial morphology transitions and functions: implications for retrograde signaling? , 2013, American journal of physiology. Regulatory, integrative and comparative physiology.

[7]  S. Nagini,et al.  Chemoprevention of rat mammary carcinogenesis by black tea polyphenols: Modulation of xenobiotic‐metabolizing enzymes, oxidative stress, cell proliferation, apoptosis, and angiogenesis , 2007, Molecular carcinogenesis.

[8]  P. Bernardi,et al.  High concordance of drug-induced human hepatotoxicity with in vitro cytotoxicity measured in a novel cell-based model using high content screening , 2006, Archives of Toxicology.

[9]  Y. Sakai,et al.  Induced pluripotent stem cell-derived hepatocytes as an alternative to human adult hepatocytes. , 2012, Journal of stem cells.

[10]  Fred G. Silva Chemical-Induced Nephropathy: A Review of the Renal Tubulointerstitial Lesions in Humans , 2004, Toxicologic pathology.

[11]  E. Mariussen Neurotoxic effects of perfluoroalkylated compounds: mechanisms of action and environmental relevance , 2012, Archives of Toxicology.

[12]  S. Veeriah,et al.  Modulation of xenobiotic metabolising enzymes by anticarcinogens -- focus on glutathione S-transferases and their role as targets of dietary chemoprevention in colorectal carcinogenesis. , 2005, Mutation research.

[13]  Paul Jennings,et al.  An overview of transcriptional regulation in response to toxicological insult , 2012, Archives of Toxicology.

[14]  S. Nair,et al.  γ‐Tocopherol‐enriched mixed tocopherol diet inhibits prostate carcinogenesis in TRAMP mice , 2009, International journal of cancer.

[15]  Lei Li,et al.  Sensing, signaling, and responding to DNA damage: Organization of the checkpoint pathways in mammalian cells , 2005, Journal of cellular biochemistry.

[16]  L. Hendershot,et al.  Herp Is Dually Regulated by Both the Endoplasmic Reticulum Stress-specific Branch of the Unfolded Protein Response and a Branch That Is Shared with Other Cellular Stress Pathways* , 2004, Journal of Biological Chemistry.

[17]  Laura P James,et al.  Acetaminophen-induced hepatotoxicity. , 2003, Drug metabolism and disposition: the biological fate of chemicals.

[18]  Q. Ma Transcriptional responses to oxidative stress: pathological and toxicological implications. , 2010, Pharmacology & therapeutics.

[19]  H. Yamada,et al.  The Japanese toxicogenomics project: application of toxicogenomics. , 2010, Molecular nutrition & food research.

[20]  M. McMahon,et al.  p62/SQSTM1 Is a Target Gene for Transcription Factor NRF2 and Creates a Positive Feedback Loop by Inducing Antioxidant Response Element-driven Gene Transcription* , 2010, The Journal of Biological Chemistry.

[21]  Min Zhang,et al.  A decade of toxicogenomic research and its contribution to toxicological science. , 2012, Toxicological sciences : an official journal of the Society of Toxicology.

[22]  A. Dinkova-Kostova,et al.  Importance of phase 2 gene regulation in protection against electrophile and reactive oxygen toxicity and carcinogenesis. , 2003, Advances in enzyme regulation.

[23]  Takeharu Nagai,et al.  Visualization of ATP levels inside single living cells with fluorescence resonance energy transfer-based genetically encoded indicators , 2009, Proceedings of the National Academy of Sciences.

[24]  P. Bastiaens,et al.  Imaging Activation of Two Ras Isoforms Simultaneously in a Single Cell , 2005, Chembiochem : a European journal of chemical biology.

[25]  T. Rushmore,et al.  The antioxidant responsive element. Activation by oxidative stress and identification of the DNA consensus sequence required for functional activity. , 1991, The Journal of biological chemistry.

[26]  S. Lindquist,et al.  Heat Shock , 1991, Springer Berlin Heidelberg.

[27]  L. Hartwell,et al.  Checkpoints: controls that ensure the order of cell cycle events. , 1989, Science.

[28]  L. Smirnova,et al.  Metabolomics in toxicology and preclinical research. , 2013, ALTEX.

[29]  J Wade Harper,et al.  The DNA damage response: ten years after. , 2007, Molecular cell.

[30]  P. Talalay,et al.  Identification of a common chemical signal regulating the induction of enzymes that protect against chemical carcinogenesis. , 1988, Proceedings of the National Academy of Sciences of the United States of America.

[31]  Peter Walter,et al.  Functional and Genomic Analyses Reveal an Essential Coordination between the Unfolded Protein Response and ER-Associated Degradation , 2000, Cell.

[32]  Eduardo Sontag,et al.  Transcriptional control of human p53-regulated genes , 2008, Nature Reviews Molecular Cell Biology.

[33]  Terry S Peters,et al.  Do Preclinical Testing Strategies Help Predict Human Hepatotoxic Potentials? , 2005, Toxicologic pathology.

[34]  Hiroshi Yamada,et al.  Gene expression profiling in rat liver treated with compounds inducing elevation of bilirubin , 2009, Human & experimental toxicology.

[35]  D. Scheuner,et al.  Translation attenuation through eIF2alpha phosphorylation prevents oxidative stress and maintains the differentiated state in beta cells. , 2009, Cell metabolism.

[36]  Galit Lahav,et al.  Oscillations by the p53-Mdm2 feedback loop. , 2008, Advances in experimental medicine and biology.

[37]  M. Massiah,et al.  Potency of Michael reaction acceptors as inducers of enzymes that protect against carcinogenesis depends on their reactivity with sulfhydryl groups , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[38]  C. Prives,et al.  Blinded by the Light: The Growing Complexity of p53 , 2009, Cell.

[39]  Q. Ma,et al.  Molecular Basis of Electrophilic and Oxidative Defense: Promises and Perils of Nrf2 , 2012, Pharmacological Reviews.

[40]  Min Zhang,et al.  Is toxicogenomics a more reliable and sensitive biomarker than conventional indicators from rats to predict drug-induced liver injury in humans? , 2012, Chemical research in toxicology.

[41]  A. Sonnenberg,et al.  Integrins Uncouple Src-induced Morphological and Oncogenic Transformation* , 2008, Journal of Biological Chemistry.

[42]  Tomomi Gotoh,et al.  ER Stress Triggers Apoptosis by Activating BH3-Only Protein Bim , 2007, Cell.

[43]  M. Black,et al.  Drug induced liver disease. , 1983, Postgraduate medical journal.

[44]  K. Iwaisako,et al.  CHOP deficiency attenuates cholestasis-induced liver fibrosis by reduction of hepatocyte injury. , 2008, American journal of physiology. Gastrointestinal and liver physiology.

[45]  H. Macfarland,et al.  Inhalation toxicology. , 1975, Journal - Association of Official Analytical Chemists.

[46]  V. Carraro,et al.  Induction of CHOP Expression by Amino Acid Limitation Requires Both ATF4 Expression and ATF2 Phosphorylation* , 2004, Journal of Biological Chemistry.

[47]  M. Schapira,et al.  Regulated translation initiation controls stress-induced gene expression in mammalian cells. , 2000, Molecular cell.

[48]  Sean C. Warren,et al.  Rapid Global Fitting of Large Fluorescence Lifetime Imaging Microscopy Datasets , 2013, PloS one.

[49]  N. Thomas Review Article: High-Content Screening: A Decade of Evolution , 2010, Journal of biomolecular screening.

[50]  Y Taya,et al.  A role for ATR in the DNA damage-induced phosphorylation of p53. , 1999, Genes & development.

[51]  Wolfgang Link,et al.  High content screening: seeing is believing. , 2010, Trends in biotechnology.

[52]  L. Attardi,et al.  p53 at a glance , 2010, Journal of Cell Science.

[53]  J. Bartek,et al.  DNA damage signalling guards against activated oncogenes and tumour progression , 2007, Oncogene.

[54]  Z. Tian,et al.  Natural killer cells in liver disease , 2013, Hepatology.

[55]  P. Ganey,et al.  Tumor Necrosis Factorα Is a Proximal Mediator of Synergistic Hepatotoxicity from Trovafloxacin/Lipopolysaccharide Coexposure , 2009, Journal of Pharmacology and Experimental Therapeutics.

[56]  Xi Chen,et al.  ER stress regulation of ATF6 localization by dissociation of BiP/GRP78 binding and unmasking of Golgi localization signals. , 2002, Developmental cell.

[57]  A. Jaiswal,et al.  The transcription factor NF‐E2‐related Factor 2 (Nrf2): a protooncogene? , 2013, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[58]  J. Valentin,et al.  Safety pharmacology and risk assessment , 2002, Fundamental & clinical pharmacology.

[59]  R. Voellmy,et al.  Chaperone regulation of the heat shock protein response. , 2007, Advances in experimental medicine and biology.

[60]  B. S. Manjunath,et al.  Biological imaging software tools , 2012, Nature Methods.

[61]  Adrian L Harris,et al.  Activating transcription factor 4. , 2008, The international journal of biochemistry & cell biology.

[62]  Erez Zadok,et al.  Unifying biological image formats with HDF5 , 2009, CACM.

[63]  E. Danen,et al.  Identification of Cisplatin-Regulated Metabolic Pathways in Pluripotent Stem Cells , 2013, PloS one.

[64]  T. Uehara,et al.  Toxicogenomics discrimination of potential hepatocarcinogenicity of non‐genotoxic compounds in rat liver , 2012, Journal of applied toxicology : JAT.

[65]  P. Walter,et al.  Signal integration in the endoplasmic reticulum unfolded protein response , 2007, Nature Reviews Molecular Cell Biology.

[66]  B. van de Water,et al.  Cleavage of the Actin-capping Protein α-Adducin at Asp-Asp-Ser-Asp633-Ala by Caspase-3 Is Preceded by Its Phosphorylation on Serine 726 in Cisplatin-induced Apoptosis of Renal Epithelial Cells* , 2000, The Journal of Biological Chemistry.

[67]  S. Westerheide,et al.  Heat Shock Response Modulators as Therapeutic Tools for Diseases of Protein Conformation* , 2005, Journal of Biological Chemistry.

[68]  Bracken M. King,et al.  Synergistic drug-cytokine induction of hepatocellular death as an in vitro approach for the study of inflammation-associated idiosyncratic drug hepatotoxicity. , 2009, Toxicology and applied pharmacology.

[69]  Karl Mechtler,et al.  BAC TransgeneOmics: a high-throughput method for exploration of protein function in mammals , 2008, Nature Methods.

[70]  C. Dinarello,et al.  Immunological and inflammatory functions of the interleukin-1 family. , 2009, Annual review of immunology.

[71]  Roger Y. Tsien,et al.  A genetically encoded fluorescent reporter reveals oscillatory phosphorylation by protein kinase C , 2003, The Journal of cell biology.

[72]  R. Roth,et al.  Hypoxia, drug therapy and toxicity. , 2007, Pharmacology & therapeutics.

[73]  Guido Kroemer,et al.  Autophagy and the integrated stress response. , 2010, Molecular cell.

[74]  Melvin E. Andersen,et al.  The Future of Toxicity Testing , 2010, Journal of toxicology and environmental health. Part B, Critical reviews.

[75]  Erwin van Vliet,et al.  Current standing and future prospects for the technologies proposed to transform toxicity testing in the 21st century. , 2011 .

[76]  Jian Yu,et al.  No PUMA, no death: implications for p53-dependent apoptosis. , 2003, Cancer cell.

[77]  Henry Jay Forman,et al.  Reactive oxygen species and cell signaling: respiratory burst in macrophage signaling. , 2002, American journal of respiratory and critical care medicine.

[78]  P Smith,et al.  Concordance of the toxicity of pharmaceuticals in humans and in animals. , 2000, Regulatory toxicology and pharmacology : RTP.

[79]  C. Winder,et al.  The sensitivity and specificity of the MTS tetrazolium assay for detecting the in vitro cytotoxicity of 20 chemicals using human cell lines. , 1997, Toxicology.

[80]  J. Hayes,et al.  The hepatotoxic metabolite of acetaminophen directly activates the Keap1‐Nrf2 cell defense system , 2008, Hepatology.

[81]  S. Elledge,et al.  The DNA damage response: putting checkpoints in perspective , 2000, Nature.

[82]  A. D. Jones,et al.  Sulindac Metabolism and Synergy with Tumor Necrosis Factor-α in a Drug-Inflammation Interaction Model of Idiosyncratic Liver Injury , 2009, Journal of Pharmacology and Experimental Therapeutics.

[83]  A. Tanoue,et al.  Expression of albumin and cytochrome P450 enzymes in HepG2 cells cultured with a nanotechnology-based culture plate with microfabricated scaffold. , 2011, The Journal of toxicological sciences.

[84]  Agustín Lahoz,et al.  Development of a multiparametric cell-based protocol to screen and classify the hepatotoxicity potential of drugs. , 2012, Toxicological sciences : an official journal of the Society of Toxicology.

[85]  H. Federoff,et al.  HUMMR, a hypoxia- and HIF-1α–inducible protein, alters mitochondrial distribution and transport , 2009, The Journal of cell biology.

[86]  Hiderou Yoshida,et al.  Transcriptional induction of mammalian ER quality control proteins is mediated by single or combined action of ATF6alpha and XBP1. , 2007, Developmental cell.

[87]  W. Schoonen,et al.  High content screening analysis of phospholipidosis: validation of a 96-well assay with CHO-K1 and HepG2 cells for the prediction of in vivo based phospholipidosis. , 2011, Toxicology in vitro : an international journal published in association with BIBRA.

[88]  L. Attardi,et al.  Deconstructing p53 transcriptional networks in tumor suppression. , 2012, Trends in cell biology.

[89]  Fons J. Verbeek,et al.  Automated Analysis of NF-κB Nuclear Translocation Kinetics in High-Throughput Screening , 2012, PloS one.

[90]  G. Isani,et al.  Metallothionein functions and structural characteristics. , 2007, Journal of trace elements in medicine and biology : organ of the Society for Minerals and Trace Elements.

[91]  N. Kaplowitz,et al.  Mechanisms of drug-induced liver injury. , 2013, Clinics in liver disease.

[92]  Daniel L Villeneuve,et al.  Adverse outcome pathways: A conceptual framework to support ecotoxicology research and risk assessment , 2010, Environmental toxicology and chemistry.

[93]  D. Schuppan,et al.  Dendritic cells in liver injury and fibrosis: shortcomings and promises. , 2013, Journal of hepatology.

[94]  Y. Haupt,et al.  Mdm2 in growth signaling and cancer , 2005, Growth factors.

[95]  Jie Lu,et al.  Human PXR modulates hepatotoxicity associated with rifampicin and isoniazid co–therapy , 2013, Nature Medicine.

[96]  Kevin W Eliceiri,et al.  NIH Image to ImageJ: 25 years of image analysis , 2012, Nature Methods.

[97]  D. Gerlier,et al.  Use of MTT colorimetric assay to measure cell activation. , 1986, Journal of immunological methods.

[98]  T. Kuwana,et al.  PUMA Couples the Nuclear and Cytoplasmic Proapoptotic Function of p53 , 2005, Science.

[99]  C. Hetz The unfolded protein response: controlling cell fate decisions under ER stress and beyond , 2012, Nature Reviews Molecular Cell Biology.

[100]  C. Prives,et al.  Covalent and noncovalent modifiers of the p53 protein , 1999, Cellular and Molecular Life Sciences CMLS.

[101]  Melvin E. Andersen,et al.  Organotypic liver culture models: Meeting current challenges in toxicity testing , 2012, Critical reviews in toxicology.

[102]  W. Schaffner,et al.  A conserved cysteine cluster, essential for transcriptional activity, mediates homodimerization of human metal-responsive transcription factor-1 (MTF-1). , 2012, Biochimica et biophysica acta.

[103]  D. Segrè,et al.  Supporting Online Material Materials and Methods Tables S1 and S2 References the Effect of Oxygen on Biochemical Networks and the Evolution of Complex Life , 2022 .

[104]  J. Taunton,et al.  Substrate-Specific Translocational Attenuation during ER Stress Defines a Pre-Emptive Quality Control Pathway , 2006, Cell.

[105]  P. Pinton,et al.  Perturbed mitochondrial Ca2+ signals as causes or consequences of mitophagy induction , 2013, Autophagy.

[106]  Bin Chen,et al.  Guidelines for the nomenclature of the human heat shock proteins , 2008, Cell Stress and Chaperones.

[107]  Adam S. Hayward,et al.  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 , 2013, Archives of Toxicology.

[108]  Jiri Bartek,et al.  Cell-cycle checkpoints and cancer , 2004, Nature.

[109]  Ken Itoh,et al.  Redox-regulated Turnover of Nrf2 Is Determined by at Least Two Separate Protein Domains, the Redox-sensitive Neh2 Degron and the Redox-insensitive Neh6 Degron* , 2004, Journal of Biological Chemistry.

[110]  K. Tsuneyama,et al.  Involvement of immune-related factors in diclofenac-induced acute liver injury in mice. , 2012, Toxicology.

[111]  Mark Hannink,et al.  Keap1 Is a Redox-Regulated Substrate Adaptor Protein for a Cul3-Dependent Ubiquitin Ligase Complex , 2004, Molecular and Cellular Biology.

[112]  J. Kleinjans,et al.  Application of toxicogenomics to study mechanisms of genotoxicity and carcinogenicity. , 2009, Toxicology letters.

[113]  B. Rehermann,et al.  The liver as an immunological organ , 2004 .

[114]  Bob van de Water,et al.  The ToxTracker assay: novel GFP reporter systems that provide mechanistic insight into the genotoxic properties of chemicals. , 2012, Toxicological sciences : an official journal of the Society of Toxicology.

[115]  W. Klaffke,et al.  Comparison of In Vitro Assays of Cellular Toxicity in the Human Hepatic Cell Line HepG2 , 2006, Journal of biomolecular screening.

[116]  N. Lowndes,et al.  53BP1: function and mechanisms of focal recruitment. , 2009, Biochemical Society transactions.

[117]  Deborah A. Brown,et al.  Fluorescent Detection of Lipid Droplets and Associated Proteins , 2007, Current protocols in cell biology.

[118]  Steven O Simmons,et al.  Cellular stress response pathway system as a sentinel ensemble in toxicological screening. , 2009, Toxicological sciences : an official journal of the Society of Toxicology.

[119]  Yuesheng Zhang,et al.  Mechanism of Chemical Activation of Nrf2 , 2012, PloS one.

[120]  N. Kiyosawa,et al.  Gene expression profiling in rat liver treated with various hepatotoxic-compounds inducing coagulopathy. , 2009, The Journal of toxicological sciences.

[121]  K. Mori,et al.  Mammalian transcription factor ATF6 is synthesized as a transmembrane protein and activated by proteolysis in response to endoplasmic reticulum stress. , 1999, Molecular biology of the cell.

[122]  L. Mullenders,et al.  DNA damage response and transcription. , 2011, DNA repair.

[123]  Fei Li,et al.  Disruption of thioredoxin reductase 1 protects mice from acute acetaminophen-induced hepatotoxicity through enhanced NRF2 activity. , 2013, Chemical research in toxicology.

[124]  Anne Bertolotti,et al.  Dynamic interaction of BiP and ER stress transducers in the unfolded-protein response , 2000, Nature Cell Biology.

[125]  L. Laine,et al.  How Common Is Diclofenac-Associated Liver Injury? Analysis of 17,289 Arthritis Patients in a Long-Term Prospective Clinical Trial , 2009, The American Journal of Gastroenterology.

[126]  J. Stevens,et al.  Strategic applications of toxicogenomics in early drug discovery. , 2008, Current opinion in pharmacology.

[127]  A. Levonen,et al.  The Keap1-Nrf2 pathway: Mechanisms of activation and dysregulation in cancer☆ , 2013, Redox biology.

[128]  S. Kain,et al.  Early detection of apoptosis using a fluorescent conjugate of annexin V. , 1997, BioTechniques.

[129]  Ruili Huang,et al.  Chemical Genomics Profiling of Environmental Chemical Modulation of Human Nuclear Receptors , 2011, Environmental health perspectives.

[130]  J. Olsen,et al.  Global Phosphoproteome Profiling Reveals Unanticipated Networks Responsive to Cisplatin Treatment of Embryonic Stem Cells , 2011, Molecular and Cellular Biology.

[131]  H. Yamaguchi,et al.  CHOP Is Involved in Endoplasmic Reticulum Stress-induced Apoptosis by Enhancing DR5 Expression in Human Carcinoma Cells* , 2004, Journal of Biological Chemistry.

[132]  W. Schoonen,et al.  Phase II enzyme levels in HepG2 cells and cryopreserved primary human hepatocytes and their induction in HepG2 cells. , 2007, Toxicology in vitro : an international journal published in association with BIBRA.

[133]  K Botzenhart,et al.  Reactive Oxygen Species , 2014 .

[134]  M. Clarke,et al.  Regulation of p53 localization. , 2001, European journal of biochemistry.

[135]  T. Rülicke,et al.  Metal‐responsive transcription factor‐1 (MTF‐1) is essential for embryonic liver development and heavy metal detoxification in the adult liver , 2004, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[136]  S. Lenna,et al.  The role of endoplasmic reticulum stress and the unfolded protein response in fibrosis , 2012, Current opinion in rheumatology.

[137]  Jonathan M. Austyn,et al.  Dendritic cells , 1998, Current opinion in hematology.

[138]  H. Arnqvist,et al.  Mitogenic Effect of the Insulin Analogue Glargine in Malignant Cells in Comparison with Insulin and IGF-I , 2008, Hormone and metabolic research = Hormon- und Stoffwechselforschung = Hormones et metabolisme.

[139]  P. Escartín,et al.  Hepatocyte damage induced by lymphocytes from patients with chronic liver diseases, as detected by LDH release. , 1978, Clinical and experimental immunology.

[140]  S. Nakajima,et al.  Selective, potent blockade of the IRE1 and ATF6 pathways by 4‐phenylbutyric acid analogues , 2013, British journal of pharmacology.