Publications by authors named "Eleanna Kaffe"

27 Publications

  • Page 1 of 1

A humanized mouse model of chronic COVID-19 to evaluate disease mechanisms and treatment options.

Res Sq 2021 Mar 17. Epub 2021 Mar 17.

Coronavirus-associated acute respiratory disease, called coronavirus disease 2019 (COVID-19) is an infectious disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). More than 90 million people have been infected with SARS-CoV-2 and more than 2 million people have died of complications due to COVID-19 worldwide. COVID-19, in its severe form, presents with an uncontrolled, hyperactive immune response and severe immunological injury or organ damage that accounts for morbidity and mortality. Even in the absence of complications, COVID-19 can last for several months with lingering effects of an overactive immune system. Dysregulated myeloid and lymphocyte compartments have been implicated in lung immunopathology. Currently, there are limited clinically-tested treatments of COVID-19 with disparities in the apparent efficacy in patients. Accurate model systems are essential to rapidly evaluate promising discoveries but most currently available in mice, ferrets and hamsters do not recapitulate sustained immunopathology described in COVID19 patients. Here, we present a comprehensively humanized mouse COVID-19 model that faithfully recapitulates the innate and adaptive human immune responses during infection with SARS-CoV-2 by adapting recombinant adeno-associated virus (AAV)-driven gene therapy to deliver human ACE2 to the lungs of MISTRG6 mice. Our unique model allows for the first time the study of chronic disease due to infection with SARS-CoV-2 in the context of patient-derived antibodies to characterize in real time the potential culprits of the observed human driving immunopathology; most importantly this model provides a live view into the aberrant macrophage response that is thought to be the effector of disease morbidity and ARDS in patients. Application of therapeutics such as patient-derived antibodies and steroids to our model allowed separation of the two aspects of the immune response, infectious viral clearance and immunopathology. Inflammatory cells seeded early in infection drove immune-patholgy later, but this very same early anti-viral response was also crucial to contain infection.
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http://dx.doi.org/10.21203/rs.3.rs-279341/v1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7987100PMC
March 2021

The fermented soy beverage Q-CAN® plus induces beneficial changes in the oral and intestinal microbiome.

BMC Nutr 2021 Mar 4;7(1). Epub 2021 Mar 4.

Department of Pediatrics (General Pediatrics), Yale University School of Medicine, New Haven, USA.

Background: Soy products are associated with many beneficial health consequences, but their effects on the human intestinal microbiome are poorly characterized.

Objectives: To identify the changes in the oral and fecal microbiome in lean and obese participants due to consumption of Q-CAN®, and to assess the expected consequences of these changes based on the published literature.

Methods: Prospective study of lean (10) and obese (9) participants consuming Q-CAN® twice daily for 4 weeks with 8 weeks follow-up. Microbial DNA was extracted from saliva and stool samples, amplified against the V4 region of the 16S ribosomal RNA gene and data analyzed using QIIME 1.9.1 bioinformatics. Four hundred forty-four samples were collected in total, 424 of which were productive and yielded good quality data.

Results: STOOL. In the lean population Bifidobacteria and Blautia show a significant increase while taking Q-CAN®, and there was a trend for this in the obese population. ORAL. There were relatively fewer major changes in the oral microbiome with an increase in the family Veillonellaceae in the lean population while on Q-CAN®.

Conclusion: Q-CAN® consumption induced a number of significant changes in the fecal and oral microbiome. Most notably an increase in the stool microbiome of Bifidobacteria and Blautia, both of which are associated with positive health benefits, and in the saliva an increase in Veillonellaceae.

Trial Registration: This trial was registered with Clinicaltrials.gov on January 14th 2016. ClinicalTrials.gov Identifier: NCT02656056.
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http://dx.doi.org/10.1186/s40795-021-00408-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7931600PMC
March 2021

MAP3K8 Regulates Cox-2-Mediated Prostaglandin E Production in the Lung and Suppresses Pulmonary Inflammation and Fibrosis.

J Immunol 2021 02 18;206(3):607-620. Epub 2020 Dec 18.

Institute of Bio-Innovation, Biomedical Sciences Research Center Alexander Fleming, 16672 Athens, Greece;

Idiopathic pulmonary fibrosis (IPF) is characterized by exuberant deposition of extracellular matrix components, leading to the deterioration of lung architecture and respiratory functions. Profibrotic mechanisms are controlled by multiple regulatory molecules, including MAPKs, in turn regulated by multiple phosphorylation cascades. MAP3K8 is an MAPK kinase kinase suggested to pleiotropically regulate multiple pathogenic pathways in the context of inflammation and cancer; however, a possible role in the pathogenesis of IPF has not been investigated. In this report, mRNA levels were found decreased in the lungs of IPF patients and of mice upon bleomycin-induced pulmonary fibrosis. Ubiquitous genetic deletion of in mice exacerbated the modeled disease, whereas bone marrow transfer experiments indicated that although MAP3K8 regulatory functions are active in both hematopoietic and nonhematopoietic cells, Map3k8 in hematopoietic cells has a more dominant role. Macrophage-specific deletion of was further found to be sufficient for disease exacerbation thus confirming a major role for macrophages in pulmonary fibrotic responses and suggesting a main role for Map3k8 in the homeostasis of their effector functions in the lung. deficiency was further shown to be associated with decreased expression, followed by a decrease in PGE production in the lung; accordingly, exogenous administration of PGE reduced inflammation and reversed the exacerbated fibrotic profile of mice. Therefore, MAP3K8 has a central role in the regulation of inflammatory responses and Cox-2-mediated PGE production in the lung, and the attenuation of its expression is integral to pulmonary fibrosis development.
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http://dx.doi.org/10.4049/jimmunol.2000862DOI Listing
February 2021

Structure-Based Discovery of Novel Chemical Classes of Autotaxin Inhibitors.

Int J Mol Sci 2020 Sep 23;21(19). Epub 2020 Sep 23.

Institute for Bioinnovation, Biomedical Sciences Research Center Alexander Fleming, Fleming 34, 16672 Athens, Greece.

Autotaxin (ATX) is a secreted glycoprotein, widely present in biological fluids, largely responsible for extracellular lysophosphatidic acid (LPA) production. LPA is a bioactive growth-factor-like lysophospholipid that exerts pleiotropic effects in almost all cell types, exerted through at least six G-protein-coupled receptors (LPAR1-6). Increased ATX expression has been detected in different chronic inflammatory diseases, while genetic or pharmacological studies have established ATX as a promising therapeutic target, exemplified by the ongoing phase III clinical trial for idiopathic pulmonary fibrosis. In this report, we employed an in silico drug discovery workflow, aiming at the identification of structurally novel series of ATX inhibitors that would be amenable to further optimization. Towards this end, a virtual screening protocol was applied involving the search into molecular databases for new small molecules potentially binding to ATX. The crystal structure of ATX in complex with a known inhibitor (HA-155) was used as a molecular model docking reference, yielding a priority list of 30 small molecule ATX inhibitors, validated by a well-established enzymatic assay of ATX activity. The two most potent, novel and structurally different compounds were further structurally optimized by deploying further in silico tools, resulting to the overall identification of six new ATX inhibitors that belong to distinct chemical classes than existing inhibitors, expanding the arsenal of chemical scaffolds and allowing further rational design.
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http://dx.doi.org/10.3390/ijms21197002DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7582705PMC
September 2020

Paracrine orchestration of intestinal tumorigenesis by a mesenchymal niche.

Nature 2020 04 1;580(7804):524-529. Epub 2020 Apr 1.

Department of Immunobiology, Yale University School of Medicine, New Haven, CT, USA.

The initiation of an intestinal tumour is a probabilistic process that depends on the competition between mutant and normal epithelial stem cells in crypts. Intestinal stem cells are closely associated with a diverse but poorly characterized network of mesenchymal cell types. However, whether the physiological mesenchymal microenvironment of mutant stem cells affects tumour initiation remains unknown. Here we provide in vivo evidence that the mesenchymal niche controls tumour initiation in trans. By characterizing the heterogeneity of the intestinal mesenchyme using single-cell RNA-sequencing analysis, we identified a population of rare pericryptal Ptgs2-expressing fibroblasts that constitutively process arachidonic acid into highly labile prostaglandin E (PGE). Specific ablation of Ptgs2 in fibroblasts was sufficient to prevent tumour initiation in two different models of sporadic, autochthonous tumorigenesis. Mechanistically, single-cell RNA-sequencing analyses of a mesenchymal niche model showed that fibroblast-derived PGE drives the expansion οf a population of Sca-1 reserve-like stem cells. These express a strong regenerative/tumorigenic program, driven by the Hippo pathway effector Yap. In vivo, Yap is indispensable for Sca-1 cell expansion and early tumour initiation and displays a nuclear localization in both mouse and human adenomas. Using organoid experiments, we identified a molecular mechanism whereby PGE promotes Yap dephosphorylation, nuclear translocation and transcriptional activity by signalling through the receptor Ptger4. Epithelial-specific ablation of Ptger4 misdirected the regenerative reprogramming of stem cells and prevented Sca-1 cell expansion and sporadic tumour initiation in mutant mice, thereby demonstrating the robust paracrine control of tumour-initiating stem cells by PGE-Ptger4. Analyses of patient-derived organoids established that PGE-PTGER4 also regulates stem-cell function in humans. Our study demonstrates that initiation of colorectal cancer is orchestrated by the mesenchymal niche and reveals a mechanism by which rare pericryptal Ptgs2-expressing fibroblasts exert paracrine control over tumour-initiating stem cells via the druggable PGE-Ptger4-Yap signalling axis.
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http://dx.doi.org/10.1038/s41586-020-2166-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7490650PMC
April 2020

Genetic deletion of Autotaxin from CD11b+ cells decreases the severity of experimental autoimmune encephalomyelitis.

PLoS One 2020 2;15(4):e0226050. Epub 2020 Apr 2.

Biomedical Sciences Research Center Alexander Fleming, Athens, Greece.

Autotaxin (ATX) is a secreted lysophospholipase D catalyzing the extracellular production of lysophosphatidic acid (LPA), a growth factor-like signaling lysophospholipid. ATX and LPA signaling have been incriminated in the pathogenesis of different chronic inflammatory diseases and various types of cancer. In this report, deregulated ATX and LPA levels were detected in the spinal cord and plasma of mice during the development of experimental autoimmune encephalomyelitis (EAE). Among the different sources of ATX expression in the inflamed spinal cord, F4/80+ CD11b+ cells, mostly activated macrophages and microglia, were found to express ATX, further suggesting an autocrine role for ATX/LPA in their activation, an EAE hallmark. Accordingly, ATX genetic deletion from CD11b+ cells attenuated the severity of EAE, thus proposing a pathogenic role for the ATX/LPA axis in neuroinflammatory disorders.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0226050PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7117669PMC
July 2020

Deregulated Lysophosphatidic Acid Metabolism and Signaling in Liver Cancer.

Cancers (Basel) 2019 Oct 23;11(11). Epub 2019 Oct 23.

Biomedical Sciences Research Center "Alexander Fleming", 16672 Athens, Greece.

Liver cancer is one of the leading causes of death worldwide due to late diagnosis and scarcity of treatment options. The major risk factor for liver cancer is cirrhosis with the underlying causes of cirrhosis being viral infection (hepatitis B or C), metabolic deregulation (Non-alcoholic fatty liver disease (NAFLD) in the presence of obesity and diabetes), alcohol or cholestatic disorders. Lysophosphatidic acid (LPA) is a bioactive phospholipid with numerous effects, most of them compatible with the hallmarks of cancer (proliferation, migration, invasion, survival, evasion of apoptosis, deregulated metabolism, neoangiogenesis, etc.). Autotaxin (ATX) is the enzyme responsible for the bulk of extracellular LPA production, and together with LPA signaling is involved in chronic inflammatory diseases, fibrosis and cancer. This review discusses the most important findings and the mechanisms related to ATX/LPA/LPAR involvement on metabolic, viral and cholestatic liver disorders and their progression to liver cancer in the context of human patients and mouse models. It focuses on the role of ATX/LPA in NAFLD development and its progression to liver cancer as NAFLD has an increasing incidence which is associated with the increasing incidence of liver cancer. Bearing in mind that adipose tissue accounts for the largest amount of LPA production, many studies have implicated LPA in adipose tissue metabolism and inflammation, liver steatosis, insulin resistance, glucose intolerance and lipogenesis. At the same time, LPA and ATX play crucial roles in fibrotic diseases. Given that hepatocellular carcinoma (HCC) is usually developed on the background of liver fibrosis, therapies that both delay the progression of fibrosis and prevent its development to malignancy would be very promising. Therefore, ATX/LPA signaling appears as an attractive therapeutic target as evidenced by the fact that it is involved in both liver fibrosis progression and liver cancer development.
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http://dx.doi.org/10.3390/cancers11111626DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6893780PMC
October 2019

Autotaxin and chronic inflammatory diseases.

J Autoimmun 2019 11 28;104:102327. Epub 2019 Aug 28.

Biomedical Sciences Research Center Alexander Fleming, 16672, Athens, Greece. Electronic address:

Autotaxin (ATX) is a secreted glycoprotein, widely present in biological fluids including blood. ATX catalyzes the hydrolysis of lysophosphatidylcholine (LPC) to lysophosphatidic acid (LPA), a growth factor-like, signaling phospholipid. LPA exerts pleiotropic effects mediated by its G-protein-coupled receptors that are widely expressed and exhibit overlapping specificities. Although ATX also possesses matricellular properties, the majority of ATX reported functions in adulthood are thought to be mediated through the extracellular production of LPA. ATX-mediated LPA synthesis is likely localized at the cell surface through the possible interaction of ATX with integrins or other molecules, while LPA levels are further controlled by a group of membrane-associated lipid-phosphate phosphatases. ATX expression was shown to be necessary for embryonic development, and ATX deficient embryos exhibit defective vascular homeostasis and aberrant neuronal system development. In adult life, ATX is highly expressed in the adipose tissue and has been implicated in diet-induced obesity and glucose homeostasis with multiple implications in metabolic disorders. Additionally, LPA has been shown to affect multiple cell types, including stromal and immune cells in various ways. Therefore, LPA participates in many processes that are intricately involved in the pathogenesis of different chronic inflammatory diseases such as vascular homeostasis, skeletal and stromal remodeling, lymphocyte trafficking and immune regulation. Accordingly, increased ATX and LPA levels have been detected, locally and/or systemically, in patients with chronic inflammatory diseases, most notably idiopathic pulmonary fibrosis (IPF), chronic liver diseases, and rheumatoid arthritis. Genetic and pharmacological studies in mice have confirmed a pathogenetic role for ATX expression and LPA signaling in chronic inflammatory diseases, and provided the proof of principle for therapeutic interventions, as exemplified by the ongoing clinical trials for IPF.
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http://dx.doi.org/10.1016/j.jaut.2019.102327DOI Listing
November 2019

Pharmacologic targeting of the ATX/LPA axis attenuates bleomycin-induced pulmonary fibrosis.

Pulm Pharmacol Ther 2018 10 7;52:32-40. Epub 2018 Sep 7.

Division of Immunology, Biomedical Sciences Research Center Alexander Fleming, Athens, Greece. Electronic address:

Idiopathic pulmonary fibrosis (IPF) is a chronic fibrosing lung disease with a dismal prognosis and a largely unknown etiology. Autotaxin (ATX) is a secreted lysophospholipase D, largely responsible for extracellular production of lysophosphatidic acid (LPA), a bioactive phospholipid. LPA has numerous effects in most cell types, signaling through at least 6 receptors (LPAR) exhibiting wide spread distribution and overlapping specificities. The ATX/LPA axis has been suggested as a therapeutic target in different chronic inflammatory and fibroproliferative disorders, including pulmonary fibrosis. In this report, we examined head-to-head the efficacy of a potent inhibitor of ATX (PF-8380), that has not been tested in pulmonary fibrosis models, and an antagonist of LPAR1 (AM095) in bleomycin (BLM)-induced pulmonary fibrosis. Both compounds abrogated the development of pulmonary fibrosis and prevented the distortion of lung architecture, exhibiting qualitative and quantitative differences in different manifestations of the modeled disease.
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http://dx.doi.org/10.1016/j.pupt.2018.08.003DOI Listing
October 2018

Animal models of cholangiocarcinoma: What they teach us about the human disease.

Clin Res Hepatol Gastroenterol 2018 10 9;42(5):403-415. Epub 2018 May 9.

Department of Molecular Medicine, University of Padua, Viale G. Colombo 3, 35131 Padua, Italy; International Center for Digestive Health (ICDH), University of Milan-Bicocca, Milan, Italy; Section of Digestive Diseases, Department of Internal Medicine, Yale University, New Haven, CT, USA. Electronic address:

Despite recent advances, pathogenesis of cholangiocarcinoma, a highly lethal cancer, remains enigmatic. Furthermore, treatment options are still limited and often disappointing. For this reason, in the last few years there has been a mounting interest towards the generation of experimental models able to reproduce the main features associated with this aggressive behavior. Toxic and infestation-induced, genetically engineered and cell implantation rodent models have been generated, contributing to a deeper understanding of the complex cell biology of the tumor, sustained by multiple cell interactions and driven by a huge variety of molecular perturbations. Herein, we will overview the most relevant animal models of biliary carcinogenesis, highlighting the methodological strategy, the molecular, histological and clinical phenotypes consistent with the human condition, their particular strengths and weaknesses and the novel therapeutic approaches that have been developed.
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http://dx.doi.org/10.1016/j.clinre.2018.04.008DOI Listing
October 2018

The Autotaxin-Lysophosphatidic Acid Axis Promotes Lung Carcinogenesis.

Cancer Res 2018 07 3;78(13):3634-3644. Epub 2018 May 3.

Division of Immunology, Biomedical Sciences Research Center "Alexander Fleming," Greece.

Pathogenesis and progression of lung cancer are governed by complex interactions between the environment and host genetic susceptibility, which is further modulated by genetic and epigenetic changes. Autotaxin (ATX, ENPP2) is a secreted glycoprotein that catalyzes the extracellular production of lysophosphatidic acid (LPA), a growth-factor-like phospholipid that is further regulated by phospholipid phosphatases (PLPP). LPA's pleiotropic effects in almost all cell types are mediated through at least six G-protein coupled LPA receptors (LPAR) that exhibit overlapping specificities, widespread distribution, and differential expression profiles. Here we use both preclinical models of lung cancer and clinical samples (from patients and healthy controls) to investigate the expression levels, activity, and biological role of the above components of the ATX/LPA axis in lung cancer. was genetically altered in 8% of patients with lung cancer, whereas increased ATX staining and activity were detected in patient biopsies and sera, respectively. Moreover, expression was consistently downregulated in patients with lung cancer. Comparable observations were made in the two most widely used animal models of lung cancer, the carcinogen urethane-induced and the genetically engineered -driven models, where genetic deletion of or resulted in disease attenuation, thus confirming a procarcinogenic role of LPA signaling in the lung. Expression profiling data analysis suggested that metabolic rewiring may be implicated in the procarcinogenic effects of the ATX/LPA axis in - -driven lung cancer pathogenesis. These findings establish the role of ATX/LPA in lung carcinogenesis, thus expanding the mechanistic links between pulmonary fibrosis and cancer. .
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http://dx.doi.org/10.1158/0008-5472.CAN-17-3797DOI Listing
July 2018

Hydroxamic Acids Constitute a Novel Class of Autotaxin Inhibitors that Exhibit in Vivo Efficacy in a Pulmonary Fibrosis Model.

J Med Chem 2018 04 18;61(8):3697-3711. Epub 2018 Apr 18.

Laboratory of Organic Chemistry, Department of Chemistry , National and Kapodistrian University of Athens , Panepistimiopolis , Athens 15771 , Greece.

Autotaxin (ATX) catalyzes the hydrolysis of lysophosphatidylcholine (LPC) generating the lipid mediator lysophosphatidic acid (LPA). Both ATX and LPA are involved in various pathological inflammatory conditions, including fibrosis and cancer, and have attracted great interest as medicinal targets over the past decade. Thus, the development of novel potent ATX inhibitors is of great importance. We have developed a novel class of ATX inhibitors containing the zinc binding functionality of hydroxamic acid. Such novel hydroxamic acids that incorporate a non-natural δ-amino acid residue exhibit high in vitro inhibitory potency over ATX (IC values 50-60 nM). Inhibitor 32, based on δ-norleucine, was tested for its efficacy in a mouse model of pulmonary inflammation and fibrosis induced by bleomycin and exhibited promising efficacy. The novel hydroxamic ATX inhibitors provide excellent tools for the study of the role of the enzyme and could contribute to the development of novel therapeutic agents for the treatment of fibrosis and other chronic inflammatory diseases.
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http://dx.doi.org/10.1021/acs.jmedchem.8b00232DOI Listing
April 2018

Notch signaling and progenitor/ductular reaction in steatohepatitis.

PLoS One 2017 15;12(11):e0187384. Epub 2017 Nov 15.

International Center for Digestive Health, Department of Medicine and Surgery, University of Milan-Bicocca, Monza, Italy.

Background And Objective: Persistent hepatic progenitor cells (HPC) activation resulting in ductular reaction (DR) is responsible for pathologic liver repair in cholangiopathies. Also, HPC/DR expansion correlates with fibrosis in several chronic liver diseases, including steatohepatitis. Increasing evidence indicates Notch signaling as a key regulator of HPC/DR response in biliary and more in general liver injuries. Therefore, we aimed to investigate the role of Notch during HPC/DR activation in a mouse model of steatohepatitis.

Methods: Steatohepatitis was generated using methionine-choline deficient (MCD) diet. For hepatocyte lineage tracing, R26R-YFP mice were infected with AAV8-TBG-Cre.

Results: MCD diet promoted a strong HPC/DR response that progressively diffused in the lobule, and correlated with increased fibrosis and TGF-β1 expression. Notch signaling was unchanged in laser-capture microdissected HPC/DR, whereas Notch receptors were down regulated in hepatocytes. However, in-vivo lineage tracing experiments identified discrete hepatocytes showing Notch-1 activation and expressing (the Notch-dependent) Sox9. Stimulation of AML-12 hepatocyte-cell line with immobilized Jag1 induced Sox9 and down-regulated albumin and BSEP expression. TGF-β1 treatment in primary hepatic stellate cells (HSC) induced Jag1 expression. In MCD diet-fed mice, αSMA-positive HSC were localized around Sox9 expressing hepatocytes, suggesting that Notch activation in hepatocytes was promoted by TGF-β1 stimulated HSC. In-vivo Notch inhibition reduced HPC response and fibrosis progression.

Conclusion: Our data suggest that Notch signaling is an important regulator of DR and that in steatohepatitis, hepatocytes exposed to Jag1-positive HSC, contribute to pathologic DR by undergoing Notch-mediated differentiation towards an HPC-like phenotype. Given the roles of Notch in fibrosis and liver cancer, these data suggest mesenchymal expression of Jag1 as an alternative therapeutic target.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0187384PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5687773PMC
December 2017

β-Catenin and interleukin-1β-dependent chemokine (C-X-C motif) ligand 10 production drives progression of disease in a mouse model of congenital hepatic fibrosis.

Hepatology 2018 05 25;67(5):1903-1919. Epub 2018 Mar 25.

Section of Digestive Diseases, Liver Center, Yale University, New Haven, CT.

Congenital hepatic fibrosis (CHF), a genetic disease caused by mutations in the polycystic kidney and hepatic disease 1 (PKHD1) gene, encoding for the protein fibrocystin/polyductin complex, is characterized by biliary dysgenesis, progressive portal fibrosis, and a protein kinase A-mediated activating phosphorylation of β-catenin at Ser675. Biliary structures of Pkhd1 mice, a mouse model of CHF, secrete chemokine (C-X-C motif) ligand 10 (CXCL10), a chemokine able to recruit macrophages. The aim of this study was to clarify whether CXCL10 plays a pathogenetic role in disease progression in CHF/Caroli disease and to understand the mechanisms leading to increased CXCL10 secretion. We demonstrate that treatment of Pkhd1 mice for 3 months with AMG-487, an inhibitor of CXC chemokine receptor family 3, the cognate receptor of CXCL10, reduces the peribiliary recruitment of alternative activated macrophages (cluster of differentiation 45 F4/80 cells), spleen size, liver fibrosis (sirius red), and cyst growth (cytokeratin 19-positive area), consistent with a pathogenetic role of CXCL10. Furthermore, we show that in fibrocystin/polyductin complex-defective cholangiocytes, isolated from Pkhd1 mice, CXCL10 production is mediated by Janus kinase/signal transducer and activator of transcription 3 in response to interleukin 1beta (IL-1β) and β-catenin. Specifically, IL-1β promotes signal transducer and activator of transcription 3 phosphorylation, whereas β-catenin promotes its nuclear translocation. Increased pro-IL-1β was regulated by nuclear factor kappa-light-chain-enhancer of activated B cells, and increased secretion of active IL-1β was mediated by the activation of Nod-like receptors, pyrin domain containing 3 inflammasome (increased expression of caspase 1 and Nod-like receptors, pyrin domain containing 3).

Conclusion: In fibrocystin/polyductin complex-defective cholangiocytes, β-catenin and IL-1β are responsible for signal transducer and activator of transcription 3-dependent secretion of CXCL10; in vivo experiments show that the CXCL10/CXC chemokine receptor family 3 axis prevents the recruitment of macrophages, reduces inflammation, and halts the progression of the disease; the increased production of IL-1β highlights the autoinflammatory nature of CHF and may open novel therapeutic avenues. (Hepatology 2018;67:1903-1919).
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http://dx.doi.org/10.1002/hep.29652DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5906178PMC
May 2018

Pathophysiologic implications of innate immunity and autoinflammation in the biliary epithelium.

Biochim Biophys Acta Mol Basis Dis 2018 04 25;1864(4 Pt B):1374-1379. Epub 2017 Jul 25.

Digestive Disease Section, Yale University School of Medicine, New Haven, CT, USA; International Center for Digestive Health, Department of Surgery and Translational Medicine, University of Milan-Bicocca, Milan, Italy; Department of Molecular Medicine, University of Padova School of Medicine, Padova, Italy.

The most studied physiological function of biliary epithelial cells (cholangiocytes) is to regulate bile flow and composition, in particular the hydration and alkalinity of the primary bile secreted by hepatocytes. After almost three decades of studies it is now become clear that cholangiocytes are also involved in epithelial innate immunity, in inflammation, and in the reparative processes in response to liver damage. An increasing number of evidence highlights the ability of cholangiocyte to undergo changes in phenotype and function in response to liver damage. By participating actively to the immune and inflammatory responses, cholangiocytes represent a first defense line against liver injury from different causes. Indeed, cholangiocytes express a number of receptors able to recognize pathogen- or damage-associated molecular patterns (PAMPs/DAMPs), such as Toll-like receptors (TLR), which modulate their pro-inflammatory behavior. Cholangiocytes can be both the targets and the initiators of the inflammatory process. Derangements of the signals controlling these mechanisms are at the basis of the pathogenesis of different cholangiopathies, both hereditary and acquired, such as cystic fibrosis-related liver disease and sclerosing cholangitis. This article is part of a Special Issue entitled: Cholangiocytes in Health and Diseaseedited by Jesus Banales, Marco Marzioni, Nicholas LaRusso and Peter Jansen.
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http://dx.doi.org/10.1016/j.bbadis.2017.07.023DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5785585PMC
April 2018

Hepatocyte autotaxin expression promotes liver fibrosis and cancer.

Hepatology 2017 04 7;65(4):1369-1383. Epub 2017 Feb 7.

Division of Immunology, Biomedical Sciences Research Center Alexander Fleming, Athens, Greece.

Autotaxin (ATX) is a secreted lysophospholipase D that catalyzes the production of lysophosphatidic acid (LPA), a pleiotropic growth-factor-like lysophospholipid. Increased ATX expression has been detected in various chronic inflammatory disorders and different types of cancer; however, little is known about its role and mode of action in liver fibrosis and cancer. Here, increased ATX expression was detected in chronic liver disease (CLD) patients of different etiologies, associated with shorter overall survival. In mice, different hepatotoxic stimuli linked with the development of different forms of CLDs were shown to stimulate hepatocyte ATX expression, leading to increased LPA levels, activation of hepatic stellate cells (HSCs), and amplification of profibrotic signals. Hepatocyte-specific, conditional genetic deletion and/or transgenic overexpression of ATX established a liver profibrotic role for ATX/LPA, whereas pharmacological ATX inhibition studies suggested ATX as a possible therapeutic target in CLDs. In addition, hepatocyte ATX ablation and the consequent deregulation of lipid homeostasis was also shown to attenuate hepatocellular carcinoma (HCC) development, thus implicating ATX/LPA in the causative link of cirrhosis and HCC.

Conclusion: ATX is a novel player in the pathogenesis of liver fibrosis and cancer and a promising therapeutic target. (Hepatology 2017;65:1369-1383).
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http://dx.doi.org/10.1002/hep.28973DOI Listing
April 2017

Computer Aided Drug Design Approaches for Identification of Novel Autotaxin (ATX) Inhibitors.

Curr Med Chem 2016 ;23(17):1708-24

Autotaxin (ATX) has become an attractive target with a huge pharmacological and pharmacochemical interest in LPA-related diseases and to date many small organic molecules have been explored as potential ATX inhibitors. As a useful aid in the various efforts of identifying novel effective ATX inhibitors, in silico methods can serve as an important and valuable tool. Especially, Virtual Screening (VS) has recently received increased attention due to the large datasets made available, the development of advanced VS techniques and the encouraging fact that VS has contributed to the discovery of several compounds that have either reached the market or entered clinical trials. Different techniques and workflows have been reported in literature with the goal to prioritize possible potent hits. In this review article several deployed virtual screening strategies for the identification of novel potent ATX inhibitors are described.
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http://dx.doi.org/10.2174/0929867323666160321122228DOI Listing
February 2017

The Bulk of Autotaxin Activity Is Dispensable for Adult Mouse Life.

PLoS One 2015 16;10(11):e0143083. Epub 2015 Nov 16.

Division of Immunology, Biomedical Sciences Research Center Alexander Fleming, Athens, Greece.

Autotaxin (ATX, Enpp2) is a secreted lysophospholipase D catalysing the production of lysophosphatidic acid, a pleiotropic growth factor-like lysophospholipid. Increased ATX expression has been detected in a number of chronic inflammatory diseases and different types of cancer, while genetic interventions have proven a role for ATX in disease pathogenesis. Therefore, ATX has emerged as a potential drug target and a large number of ATX inhibitors have been developed exhibiting promising therapeutic potential. However, the embryonic lethality of ATX null mice and the ubiquitous expression of ATX and LPA receptors in adult life question the suitability of ATX as a drug target. Here we show that inducible, ubiquitous genetic deletion of ATX in adult mice, as well as long-term potent pharmacologic inhibition, are well tolerated, alleviating potential toxicity concerns of ATX therapeutic targeting.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0143083PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4646642PMC
June 2016

Autotaxin and Endotoxin-Induced Acute Lung Injury.

PLoS One 2015 21;10(7):e0133619. Epub 2015 Jul 21.

Division of Immunology, Biomedical Sciences Research Center "Alexander Fleming", Athens, Greece.

Acute Lung Injury (ALI) is a life-threatening, diffuse heterogeneous lung injury characterized by acute onset, pulmonary edema and respiratory failure. Lipopolysaccharide (LPS) is a common cause of both direct and indirect lung injury and when administered to a mouse induces a lung phenotype exhibiting some of the clinical characteristics of human ALI. Here, we report that LPS inhalation in mice results in increased bronchoalveolar lavage fluid (BALF) levels of Autotaxin (ATX, Enpp2), a lysophospholipase D largely responsible for the conversion of lysophosphatidylcholine (LPC) to lysophosphatidic acid (LPA) in biological fluids and chronically inflamed sites. In agreement, gradual increases were also detected in BALF LPA levels, following inflammation and pulmonary edema. However, genetic or pharmacologic targeting of ATX had minor effects in ALI severity, suggesting no major involvement of the ATX/LPA axis in acute inflammation. Moreover, systemic, chronic exposure to increased ATX/LPA levels was shown to predispose to and/or to promote acute inflammation and ALI unlike chronic inflammatory pathophysiological situations, further suggesting a differential involvement of the ATX/LPA axis in acute versus chronic pulmonary inflammation.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0133619PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4509763PMC
May 2016

Autotaxin, a secreted lysophospholipase D, as a promising therapeutic target in chronic inflammation and cancer.

Prog Lipid Res 2015 Apr 20;58:76-96. Epub 2015 Feb 20.

Laboratory of Organic Chemistry, Department of Chemistry, University of Athens, Panepistimiopolis, Athens 15771, Greece.

Autotaxin (ATX) is a member of the nucleotide pyrophosphatase/phosphodiesterase family of ectoenzymes that hydrolyzes phosphodiester bonds of various nucleotides. It possesses lysophospholipase D activity, catalyzing the hydrolysis of lysophosphatidylcholine into lysophosphatidic acid (LPA), and it is considered the major LPA-producing enzyme in the circulation. LPA is a bioactive phospholipid with diverse functions in almost every mammalian cell type, which exerts its action through binding to specific G protein-coupled receptors and stimulates various cellular functions, including migration, proliferation and survival. As a consequence, both ATX and LPA have attracted the interest of researchers, in an effort to understand their roles in physiology and pathophysiology. The present review article aims to summarize the existing knowledge as to the implications of ATX in chronic inflammatory diseases and cancer and to highlight the low molecular weight compounds, which have been developed as leads for the discovery of novel medicines to treat inflammatory diseases and cancer.
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http://dx.doi.org/10.1016/j.plipres.2015.02.001DOI Listing
April 2015

Intestinal myofibroblast-specific Tpl2-Cox-2-PGE2 pathway links innate sensing to epithelial homeostasis.

Proc Natl Acad Sci U S A 2014 Oct 14;111(43):E4658-67. Epub 2014 Oct 14.

Institute of Immunology, Biomedical Sciences Research Center "Alexander Fleming", Vari 16672, Greece;

Tumor progression locus-2 (Tpl2) kinase is a major inflammatory mediator in immune cell types recently found to be genetically associated with inflammatory bowel diseases (IBDs). Here we show that Tpl2 may exert a dominant homeostatic rather than inflammatory function in the intestine mediated specifically by subepithelial intestinal myofibroblasts (IMFs). Mice with complete or IMF-specific Tpl2 ablation are highly susceptible to epithelial injury-induced colitis showing impaired compensatory proliferation in crypts and extensive ulcerations without significant changes in inflammatory responses. Following epithelial injury, IMFs sense innate or inflammatory signals and activate, via Tpl2, the cyclooxygenase-2 (Cox-2)-prostaglandin E2 (PGE2) pathway, which we show here to be essential for the epithelial homeostatic response. Exogenous PGE2 administration rescues mice with complete or IMF-specific Tpl2 ablation from defects in crypt function and susceptibility to colitis. We also show that Tpl2 expression is decreased in IMFs isolated from the inflamed ileum of IBD patients indicating that Tpl2 function in IMFs may be highly relevant to human disease. The IMF-mediated mechanism we propose also involves the IBD-associated genes IL1R1, MAPK1, and the PGE2 receptor-encoding PTGER4. Our results establish a previously unidentified myofibroblast-specific innate pathway that regulates intestinal homeostasis and may underlie IBD susceptibility in humans.
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http://dx.doi.org/10.1073/pnas.1415762111DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4217397PMC
October 2014

Oxidative stress and antioxidant status in patients with autoimmune liver diseases.

Redox Rep 2015 Jan 13;20(1):33-41. Epub 2014 Aug 13.

Objective: To estimate oxidative stress and antioxidant components during different stages of autoimmune liver diseases and assess their possible implication on disease progression.

Methods: We determined several markers of oxidative injury (isoprostane, aldehydes, protein carbonyls, 3-nitrotyrosine, and myeloperoxidase) and antioxidant components (glutathione, glutathione peroxidase, glutathione reductase, superoxide dismutase, and catalase) in whole blood, serum, and urine in 49 patients with autoimmune cholestatic liver diseases (AC) and 36 patients with autoimmune hepatitis (AIH) and healthy subjects matched for sex and age.

Results: Both AC and AIH patients had increased levels of all lipid and protein oxidative injury products and significantly decreased whole blood glutathione levels compared to controls. AIH patients had significantly higher levels of aldehydes and glutathione peroxidase activity and significantly lower protein carbonyl levels compared to AC patients. Protein carbonyl and isoprostane levels increased and glutathione levels decreased gradually with progression from mild fibrosis to severe fibrosis and cirrhosis in both AC and AIH patients. In addition, both cirrhotic AC and AIH patients had significantly higher protein carbonyls compared to non-cirrhotics.

Discussion: We provide novel findings in support of a major contribution of oxidant/antioxidant imbalance in the progression of liver injury in AC and AIH.
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http://dx.doi.org/10.1179/1351000214Y.0000000101DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6837668PMC
January 2015

Autotaxin production of lysophosphatidic acid mediates allergic asthmatic inflammation.

Am J Respir Crit Care Med 2013 Oct;188(8):928-40

1 Section of Pulmonary, Critical Care, Sleep, and Allergy, Department of Medicine.

Rationale: Bioactive lipid mediators, derived from membrane lipid precursors, are released into the airway and airspace where they bind high-affinity cognate receptors and may mediate asthma pathogenesis. Lysophosphatidic acid (LPA), a bioactive lipid mediator generated by the enzymatic activity of extracellular autotaxin (ATX), binds LPA receptors, resulting in an array of biological actions on cell proliferation, migration, survival, differentiation, and motility, and therefore could mediate asthma pathogenesis.

Objectives: To define a role for the ATX-LPA pathway in human asthma pathogenesis and a murine model of allergic lung inflammation.

Methods: We investigated the profiles of LPA molecular species and the level of ATX exoenzyme in bronchoalveolar lavage fluids of human patients with asthma subjected to subsegmental bronchoprovocation with allergen. We interrogated the role of the ATX-LPA pathway in allergic lung inflammation using a murine allergic asthma model in ATX-LPA pathway-specific genetically modified mice.

Measurements And Main Results: Subsegmental bronchoprovocation with allergen in patients with mild asthma resulted in a remarkable increase in bronchoalveolar lavage fluid levels of LPA enriched in polyunsaturated 22:5 and 22:6 fatty acids in association with increased concentrations of ATX protein. Using a triple-allergen mouse asthma model, we showed that ATX-overexpressing transgenic mice had a more severe asthmatic phenotype, whereas blocking ATX activity and knockdown of the LPA2 receptor in mice produced a marked attenuation of Th2 cytokines and allergic lung inflammation.

Conclusions: The ATX-LPA pathway plays a critical role in the pathogenesis of asthma. These preclinical data indicate that targeting the ATX-LPA pathway could be an effective antiasthma treatment strategy.
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http://dx.doi.org/10.1164/rccm.201306-1014OCDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3826286PMC
October 2013

A metabolically-stabilized phosphonate analog of lysophosphatidic acid attenuates collagen-induced arthritis.

PLoS One 2013 29;8(7):e70941. Epub 2013 Jul 29.

Institute of Immunology, Biomedical Sciences Research Center Alexander Fleming, Athens, Greece.

Rheumatoid arthritis (RA) is a destructive arthropathy with systemic manifestations, characterized by chronic synovial inflammation. Under the influence of the pro-inflammatory milieu synovial fibroblasts (SFs), the main effector cells in disease pathogenesis become activated and hyperplastic while releasing a number of signals that include pro-inflammatory factors and tissue remodeling enzymes. Activated RA SFs in mouse or human arthritic joints express significant quantities of autotaxin (ATX), a lysophospholipase D responsible for the majority of lysophosphatidic acid (LPA) production in the serum and inflamed sites. Conditional genetic ablation of ATX from SFs resulted in attenuation of disease symptoms in animal models, an effect attributed to diminished LPA signaling in the synovium, shown to activate SF effector functions. Here we show that administration of 1-bromo-3(S)-hydroxy-4-(palmitoyloxy)butyl-phosphonate (BrP-LPA), a metabolically stabilized analog of LPA and a dual function inhibitor of ATX and pan-antagonist of LPA receptors, attenuates collagen induced arthritis (CIA) development, thus validating the ATX/LPA axis as a novel therapeutic target in RA.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0070941PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3726599PMC
August 2014

Lysoglycerophospholipids in chronic inflammatory disorders: the PLA(2)/LPC and ATX/LPA axes.

Biochim Biophys Acta 2013 Jan 31;1831(1):42-60. Epub 2012 Jul 31.

Institute of Immunology, Biomedical Sciences Research Center, Athens, Greece.

Lysophosphatidylcholine (LPC) and lysophosphatidic acid (LPA), the most prominent lysoglycerophospholipids, are emerging as a novel class of inflammatory lipids, joining thromboxanes, leukotrienes and prostaglandins with which they share metabolic pathways and regulatory mechanisms. Enzymes that participate in LPC and LPA metabolism, such as the phospholipase A(2) superfamily (PLA(2)) and autotaxin (ATX, ENPP2), play central roles in regulating LPC and LPA levels and consequently their actions. LPC/LPA biosynthetic pathways will be briefly presented and LPC/LPA signaling properties and their possible functions in the regulation of the immune system and chronic inflammation will be reviewed. Furthermore, implications of exacerbated LPC and/or LPA signaling in the context of chronic inflammatory diseases, namely rheumatoid arthritis, multiple sclerosis, pulmonary fibrosis and hepatitis, will be discussed. This article is part of a Special Issue entitled Advances in Lysophospholipid Research.
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http://dx.doi.org/10.1016/j.bbalip.2012.07.019DOI Listing
January 2013

Pulmonary autotaxin expression contributes to the pathogenesis of pulmonary fibrosis.

Am J Respir Cell Mol Biol 2012 Nov 28;47(5):566-74. Epub 2012 Jun 28.

Institute of Immunology, Biomedical Sciences Research Center Alexander Fleming, Athens, Greece.

Idiopathic pulmonary fibrosis (IPF) is a chronic, progressive, fibrotic form of diffuse lung disease occurring mainly in older adults. Increased lysophosphatidic acid (LPA) concentrations have been reported in the alveolar space of both idiopathic pulmonary fibrosis patients and a corresponding animal model, whereas the genetic deletion or pharmacological inhibition of LPA receptor 1 attenuated the development of the modeled disease, suggesting a direct involvement of LPA in disease pathogenesis. In this report, increased concentrations of autotaxin (ATX; ENPP2), the enzyme largely responsible for extracellular LPA production, were detected in both murine and human fibrotic lungs. The genetic deletion of ATX from bronchial epithelial cells or macrophages attenuated disease severity, establishing ATX as a novel player in IPF pathogenesis. Furthermore, the pharmacological inhibition of ATX attenuated the development of the modeled disease, suggesting that ATX is a possible therapeutic target in IPF.
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http://dx.doi.org/10.1165/rcmb.2012-0004OCDOI Listing
November 2012

The metabotropic glutamate 2/3 receptor antagonist LY341495 differentially affects recognition memory in rats.

Behav Brain Res 2012 May;230(2):374-9

Department of Pharmacology, School of Medicine, University of Thessaly, Biopolis, 411-10 Larissa, Greece.

Experimental evidence suggests that metabotropic glutamate 2/3 (mGlu2/3) receptor antagonists affect cognitive function, although contradictory findings have been reported. To clarify the role of mGlu2/3 receptor antagonists in one aspect of cognition, the present study investigated the effects of a broad range of doses of the mGlu2/3 receptor antagonist LY341495 on post-training recognition memory components (storage and/or retrieval) in rats. The efficacy of LY341495 in antagonizing the extinction of recognition memory was also investigated. The novel object recognition test was used as the memory test. The highest LY341495 doses administered (0.3, 1, and 3 mg/kg) disrupted performance in this recognition memory procedure in rats at all delay conditions tested, whereas administration of lower doses (0.05 and 0.1 mg/kg) did not impair recognition memory. Moreover, administration of the low LY341495 doses (0.05 and 0.1 mg/kg) counteracted the extinction of recognition memory. The present results indicate that administration of the mGlu2/3 receptor antagonist LY341495 can either impair or enhance recognition memory in rats, depending on the dose of the compound and delay period used. Thus, together with previously reported findings, the present data suggest complex effects of this compound on cognitive function, particularly recognition memory.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3705739PMC
http://dx.doi.org/10.1016/j.bbr.2012.02.027DOI Listing
May 2012
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