Publications by authors named "Xavier Clemente-Casares"

27 Publications

  • Page 1 of 1

Mechanical Stiffness Controls Dendritic Cell Metabolism and Function.

Cell Rep 2021 Jan;34(2):108609

Division of Cellular and Molecular Biology, Diabetes Research Group, Toronto General Hospital Research Institute (TGHRI), University Health Network, Toronto, ON M5G 1L7, Canada; Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, ON M5S 1A8, Canada; Department of Immunology, University of Toronto, Toronto, ON M5S 1A8, Canada; Department of Pathology, University Health Network, Toronto, ON M5G 2C4, Canada; Buck Institute for Research on Aging, 8001 Redwood Boulevard, Novato, CA 94945, USA. Electronic address:

Stiffness in the tissue microenvironment changes in most diseases and immunological conditions, but its direct influence on the immune system is poorly understood. Here, we show that static tension impacts immune cell function, maturation, and metabolism. Bone-marrow-derived and/or splenic dendritic cells (DCs) grown in vitro at physiological resting stiffness have reduced proliferation, activation, and cytokine production compared with cells grown under higher stiffness, mimicking fibro-inflammatory disease. Consistently, DCs grown under higher stiffness show increased activation and flux of major glucose metabolic pathways. In DC models of autoimmune diabetes and tumor immunotherapy, tension primes DCs to elicit an adaptive immune response. Mechanistic workup identifies the Hippo-signaling molecule, TAZ, as well as Ca-related ion channels, including potentially PIEZO1, as important effectors impacting DC metabolism and function under tension. Tension also directs the phenotypes of monocyte-derived DCs in humans. Thus, mechanical stiffness is a critical environmental cue of DCs and innate immunity.
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http://dx.doi.org/10.1016/j.celrep.2020.108609DOI Listing
January 2021

Gut-associated IgA immune cells regulate obesity-related insulin resistance.

Nat Commun 2019 08 13;10(1):3650. Epub 2019 Aug 13.

Division of Cellular and Molecular Biology, Diabetes Research Group, Toronto General Research Institute (TGRI), University Health Network, Toronto, ON, M5G 2C4, Canada.

The intestinal immune system is emerging as an important contributor to obesity-related insulin resistance, but the role of intestinal B cells in this context is unclear. Here, we show that high fat diet (HFD) feeding alters intestinal IgA immune cells and that IgA is a critical immune regulator of glucose homeostasis. Obese mice have fewer IgA immune cells and less secretory IgA and IgA-promoting immune mediators. HFD-fed IgA-deficient mice have dysfunctional glucose metabolism, a phenotype that can be recapitulated by adoptive transfer of intestinal-associated pan-B cells. Mechanistically, IgA is a crucial link that controls intestinal and adipose tissue inflammation, intestinal permeability, microbial encroachment and the composition of the intestinal microbiome during HFD. Current glucose-lowering therapies, including metformin, affect intestinal-related IgA B cell populations in mice, while bariatric surgery regimen alters the level of fecal secretory IgA in humans. These findings identify intestinal IgA immune cells as mucosal mediators of whole-body glucose regulation in diet-induced metabolic disease.
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http://dx.doi.org/10.1038/s41467-019-11370-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6692361PMC
August 2019

Aryl hydrocarbon receptor agonist indigo protects against obesity-related insulin resistance through modulation of intestinal and metabolic tissue immunity.

Int J Obes (Lond) 2019 12 3;43(12):2407-2421. Epub 2019 Apr 3.

Division of Cellular & Molecular Biology, Diabetes Research Group, Toronto General Research Institute (TGRI), University Health Network, 101 College Street, Toronto, ON, M5G 1L7, Canada.

Background/objectives: Low-grade chronic inflammation in visceral adipose tissue and the intestines are important drivers of obesity associated insulin resistance. Bioactive compounds derived from plants are an important source of potential novel therapies for the treatment of chronic diseases. In search for new immune based treatments of obesity associated insulin resistance, we screened for tissue relevant anti-inflammatory properties in 20 plant-based extracts.

Methods: We screened 20 plant-based extracts to assess for preferential production of IL-10 compared to TNFα, specifically targetting metabolic tissues, including the visceral adipose tissue. We assessed the therapeutic potential of the strongest anti-inflammatory compound, indigo, in the C57BL/6J diet-induced obesity mouse model with supplementation for up to 16 weeks by measuring changes in body weight, glucose and insulin tolerance, and gut barrier function. We also utilized flow cytometry, quantitative PCR, enzyme-linked immunosorbent assay (ELISA), and histology to measure changes to immune cells populations and cytokine profiles in the intestine, visceral adipose tissue (VAT), and liver. 16SrRNA sequencing was performed to examine gut microbial differences induced by indigo supplementation.

Results: We identifed indigo, an aryl hydrocarbon receptor (AhR) ligand agonist, as a potent inducer of IL-10 and IL-22, which protects against high-fat diet (HFD)-induced insulin resistance and fatty liver disease in the diet-induced obesity model. Therapeutic actions were mechanistically linked to decreased inflammatory immune cell tone in the intestine, VAT and liver. Specifically, indigo increased Lactobacillus bacteria and elicited IL-22 production in the gut, which improved intestinal barrier permeability and reduced endotoxemia. These changes were associated with increased IL-10 production by immune cells residing in liver and VAT.

Conclusions: Indigo is a naturally occurring AhR ligand with anti-inflammatory properties that effectively protects against HFD-induced glucose dysregulation. Compounds derived from indigo or those with similar properties could represent novel therapies for diseases associated with obesity-related metabolic tissue inflammation.
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http://dx.doi.org/10.1038/s41366-019-0340-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6892742PMC
December 2019

Publisher Correction: Self-renewing resident cardiac macrophages limit adverse remodeling following myocardial infarction.

Nat Immunol 2019 May;20(5):664

Toronto General Hospital Research Institute, University Health Network (UHN), Toronto, Canada.

In the version of this article initially published, the equal contribution of the third author was omitted. The footnote links for that author should be "Sara Nejat" and the correct statement is as follows: "These authors contributed equally: Sarah A. Dick, Jillian A. Macklin, Sara Nejat." The error has been corrected in the HTML and PDF versions of the article.
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http://dx.doi.org/10.1038/s41590-019-0363-8DOI Listing
May 2019

Self-renewing resident cardiac macrophages limit adverse remodeling following myocardial infarction.

Nat Immunol 2019 01 11;20(1):29-39. Epub 2018 Dec 11.

Toronto General Hospital Research Institute, University Health Network (UHN), Toronto, Canada.

Macrophages promote both injury and repair after myocardial infarction, but discriminating functions within mixed populations remains challenging. Here we used fate mapping, parabiosis and single-cell transcriptomics to demonstrate that at steady state, TIMD4LYVE1MHC-IICCR2 resident cardiac macrophages self-renew with negligible blood monocyte input. Monocytes partially replaced resident TIMD4LYVE1MHC-IICCR2 macrophages and fully replaced TIMD4LYVE1MHC-IICCR2 macrophages, revealing a hierarchy of monocyte contribution to functionally distinct macrophage subsets. Ischemic injury reduced TIMD4 and TIMD4 resident macrophage abundance, whereas CCR2 monocyte-derived macrophages adopted multiple cell fates within infarcted tissue, including those nearly indistinguishable from resident macrophages. Recruited macrophages did not express TIMD4, highlighting the ability of TIMD4 to track a subset of resident macrophages in the absence of fate mapping. Despite this similarity, inducible depletion of resident macrophages using a Cx3cr1-based system led to impaired cardiac function and promoted adverse remodeling primarily within the peri-infarct zone, revealing a nonredundant, cardioprotective role of resident cardiac macrophages.
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http://dx.doi.org/10.1038/s41590-018-0272-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6565365PMC
January 2019

The Macrophage in Cardiac Homeostasis and Disease: JACC Macrophage in CVD Series (Part 4).

J Am Coll Cardiol 2018 10;72(18):2213-2230

Zena and Michael A. Wiener Cardiovascular Institute, Icahn School of Medicine at Mount Sinai, New York, New York. Electronic address:

Macrophages are integral components of cardiac tissue and exert profound effects on the healthy and diseased heart. Paradigm shifting studies using advanced molecular techniques have revealed significant complexity within these macrophage populations that reside in the heart. In this final of a 4-part review series covering the macrophage in cardiovascular disease, the authors review the origins, dynamics, cell surface markers, and respective functions of each cardiac macrophage subset identified to date, including in the specific scenarios of myocarditis and after myocardial infarction. Looking ahead, a deeper understanding of the diverse and often dichotomous functions of cardiac macrophages will be essential for the development of targeted therapies to mitigate injury and orchestrate recovery of the diseased heart. Moreover, as macrophages are critical for cardiac healing, they are an emerging focus for therapeutic strategies aimed at minimizing cardiomyocyte death, ameliorating pathological cardiac remodeling, and for treating heart failure and after myocardial infarction.
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http://dx.doi.org/10.1016/j.jacc.2018.08.2149DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6209119PMC
October 2018

Isolation and Identification of Extravascular Immune Cells of the Heart.

J Vis Exp 2018 08 23(138). Epub 2018 Aug 23.

Toronto General Hospital Research Institute, University Health Network (UHN); Dept of Laboratory Medicine and Pathobiology, University of Toronto;

The immune system is an essential component of a healthy heart. The myocardium is home to a rich population of different immune cell subsets with functional compartmentalization both during steady state and during different forms of inflammation. Until recently, the study of immune cells in the heart required the use of microscopy or poorly developed digestion protocols, which provided enough sensitivity during severe inflammation but were unable to confidently identify small - but key - populations of cells during steady state. Here, we discuss a simple method combining enzymatic (collagenase, hyaluronidase and DNAse) and mechanical digestion of murine hearts preceded by intravascular administration of fluorescently-labelled antibodies to differentiate small but unavoidable intravascular cell contaminants. This method generates a suspension of isolated viable cells that can be analyzed by flow cytometry for identification, phenotyping and quantification, or further purified with fluorescence-activated cell sorting or magnetic bead separation for transcriptional analysis or in vitro studies. We include an example of a step-by-step flow cytometric analysis to differentiate the key macrophage and dendritic cell populations of the heart. For a medium sized experiment (10 hearts) the completion of the procedure requires 2-3 h.
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http://dx.doi.org/10.3791/58114DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6231712PMC
August 2018

Insulin Receptor-Mediated Stimulation Boosts T Cell Immunity during Inflammation and Infection.

Cell Metab 2018 12 30;28(6):922-934.e4. Epub 2018 Aug 30.

Division of Cellular & Molecular Biology, Diabetes Research Group, Toronto General Research Institute (TGRI), University Health Network, 101 College Street, Toronto, ON M5G 1L7, Canada; Department of Immunology, University of Toronto, 1 King's College Circle, Toronto, ON M5S 1A8, Canada; Department of Pathology, University Health Network, 200 Elizabeth Street, Toronto, ON M5G 2C4, Canada; Department of Laboratory Medicine and Pathobiology, University of Toronto, 1 King's College Circle, Toronto, ON M5S 1A8, Canada; Toronto General Research Institute (TGRI), Toronto, ON M5G 2C4, Canada. Electronic address:

T cells represent a critical effector of cell-mediated immunity. Activated T cells engage in metabolic reprogramming during effector differentiation to accommodate dynamic changes in energy demands. Here, we show that the hormone, insulin, and downstream signaling through its insulin receptor shape adaptive immune function through modulating T cell metabolism. T cells lacking insulin receptor expression (LckCre+ Insr) show reduced antigen-specific proliferation and compromised production of pro-inflammatory cytokines. In vivo, T cell-specific insulin receptor deficiency reduces T cell-driven colonic inflammation. In a model of severe influenza infection with A/PR8 (H1N1), lack of insulin receptor on T cells curtails antigen-specific immunity to influenza viral antigens. Mechanistically, insulin receptor signaling reinforces a metabolic program that supports T cell nutrient uptake and associated glycolytic and respiratory capacities. These data highlight insulin receptor signaling as an important node integrating immunometabolic pathways to drive optimal T cell effector function in health and disease.
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http://dx.doi.org/10.1016/j.cmet.2018.08.003DOI Listing
December 2018

A CD103 Conventional Dendritic Cell Surveillance System Prevents Development of Overt Heart Failure during Subclinical Viral Myocarditis.

Immunity 2017 11;47(5):974-989.e8

Toronto General Hospital Research Institute, University Health Network (UHN), Toronto ON, M5G 1L7, Canada; Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto ON, M5S 1A1, Canada; Department of Immunology, University of Toronto, Toronto ON, M5S 1A1, Canada; Peter Munk Cardiac Centre, Toronto ON, M5G 1L7, Canada; Ted Rogers Centre for Heart Research, Toronto ON, M5G 1L7, Canada. Electronic address:

Innate and adaptive immune cells modulate heart failure pathogenesis during viral myocarditis, yet their identities and functions remain poorly defined. We utilized a combination of genetic fate mapping, parabiotic, transcriptional, and functional analyses and demonstrated that the heart contained two major conventional dendritic cell (cDC) subsets, CD103 and CD11b, which differentially relied on local proliferation and precursor recruitment to maintain their tissue residency. Following viral infection of the myocardium, cDCs accumulated in the heart coincident with monocyte infiltration and loss of resident reparative embryonic-derived cardiac macrophages. cDC depletion abrogated antigen-specific CD8 T cell proliferative expansion, transforming subclinical cardiac injury to overt heart failure. These effects were mediated by CD103 cDCs, which are dependent on the transcription factor BATF3 for their development. Collectively, our findings identified resident cardiac cDC subsets, defined their origins, and revealed an essential role for CD103 cDCs in antigen-specific T cell responses during subclinical viral myocarditis.
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http://dx.doi.org/10.1016/j.immuni.2017.10.011DOI Listing
November 2017

A Gut Microbial Mimic that Hijacks Diabetogenic Autoreactivity to Suppress Colitis.

Cell 2017 Oct;171(3):655-667.e17

Julia McFarlane Diabetes Research Centre (JMDRC), University of Calgary, Calgary AB T2N 4N1, Canada; Autoimmunity Research Group, Institut D'Investigacions Biomèdiques August Pi i Sunyer, 08036 Barcelona, Spain; Department of Microbiology, Immunology, and Infectious Diseases, Snyder Institute for Chronic Diseases, University of Calgary, Calgary AB T2N 4N1, Canada. Electronic address:

The gut microbiota contributes to the development of normal immunity but, when dysregulated, can promote autoimmunity through various non-antigen-specific effects on pathogenic and regulatory lymphocytes. Here, we show that an integrase expressed by several species of the gut microbial genus Bacteroides encodes a low-avidity mimotope of the pancreatic β cell autoantigen islet-specific glucose-6-phosphatase-catalytic-subunit-related protein (IGRP). Studies in germ-free mice monocolonized with integrase-competent, integrase-deficient, and integrase-transgenic Bacteroides demonstrate that the microbial epitope promotes the recruitment of diabetogenic CD8+ T cells to the gut. There, these effectors suppress colitis by targeting microbial antigen-loaded, antigen-presenting cells in an integrin β7-, perforin-, and major histocompatibility complex class I-dependent manner. Like their murine counterparts, human peripheral blood T cells also recognize Bacteroides integrase. These data suggest that gut microbial antigen-specific cytotoxic T cells may have therapeutic value in inflammatory bowel disease and unearth molecular mimicry as a novel mechanism by which the gut microbiota can regulate normal immune homeostasis. PAPERCLIP.
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http://dx.doi.org/10.1016/j.cell.2017.09.022DOI Listing
October 2017

Peptide-MHC-based nanomedicines for autoimmunity function as T-cell receptor microclustering devices.

Nat Nanotechnol 2017 07 24;12(7):701-710. Epub 2017 Apr 24.

Julia McFarlane Diabetes Research Centre (JMDRC) and Department of Microbiology, Immunology and Infectious Diseases, Snyder Institute for Chronic Diseases and Hotchkiss Brain Institute, Cumming School of Medicine, University of Calgary, Calgary, Alberta T2N 4N1, Canada.

We have shown that nanoparticles (NPs) can be used as ligand-multimerization platforms to activate specific cellular receptors in vivo. Nanoparticles coated with autoimmune disease-relevant peptide-major histocompatibility complexes (pMHC) blunted autoimmune responses by triggering the differentiation and expansion of antigen-specific regulatory T cells in vivo. Here, we define the engineering principles impacting biological activity, detail a synthesis process yielding safe and stable compounds, and visualize how these nanomedicines interact with cognate T cells. We find that the triggering properties of pMHC-NPs are a function of pMHC intermolecular distance and involve the sustained assembly of large antigen receptor microclusters on murine and human cognate T cells. These compounds show no off-target toxicity in zebrafish embryos, do not cause haematological, biochemical or histological abnormalities, and are rapidly captured by phagocytes or processed by the hepatobiliary system. This work lays the groundwork for the design of ligand-based NP formulations to re-program in vivo cellular responses using nanotechnology.
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http://dx.doi.org/10.1038/nnano.2017.56DOI Listing
July 2017

Expanding antigen-specific regulatory networks to treat autoimmunity.

Nature 2016 Feb 17;530(7591):434-40. Epub 2016 Feb 17.

Julia McFarlane Diabetes Research Centre (JMDRC), and Department of Microbiology, Immunology and Infectious Diseases, Snyder Institute for Chronic Diseases and Hotchkiss Brain Institute, Cumming School of Medicine, University of Calgary, Calgary, Alberta T2N 4N1, Canada.

Regulatory T cells hold promise as targets for therapeutic intervention in autoimmunity, but approaches capable of expanding antigen-specific regulatory T cells in vivo are currently not available. Here we show that systemic delivery of nanoparticles coated with autoimmune-disease-relevant peptides bound to major histocompatibility complex class II (pMHCII) molecules triggers the generation and expansion of antigen-specific regulatory CD4(+) T cell type 1 (TR1)-like cells in different mouse models, including mice humanized with lymphocytes from patients, leading to resolution of established autoimmune phenomena. Ten pMHCII-based nanomedicines show similar biological effects, regardless of genetic background, prevalence of the cognate T-cell population or MHC restriction. These nanomedicines promote the differentiation of disease-primed autoreactive T cells into TR1-like cells, which in turn suppress autoantigen-loaded antigen-presenting cells and drive the differentiation of cognate B cells into disease-suppressing regulatory B cells, without compromising systemic immunity. pMHCII-based nanomedicines thus represent a new class of drugs, potentially useful for treating a broad spectrum of autoimmune conditions in a disease-specific manner.
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http://dx.doi.org/10.1038/nature16962DOI Listing
February 2016

Are obesity-related insulin resistance and type 2 diabetes autoimmune diseases?

Diabetes 2015 Jun;64(6):1886-97

Division of Cellular and Molecular Biology, Diabetes Research Group, Toronto General Research Institute, University Health Network, Toronto, Ontario, Canada Department of Pathology, University Health Network, Toronto, Ontario, Canada Division of Endocrinology and Metabolism, Department of Medicine, University Health Network, Toronto, Ontario, Canada Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, Ontario, Canada Department of Immunology, University of Toronto, Toronto, Ontario, Canada

Obesity and associated insulin resistance predispose individuals to develop chronic metabolic diseases, such as type 2 diabetes and cardiovascular disease. Although these disorders affect a significant proportion of the global population, the underlying mechanisms of disease remain poorly understood. The discovery of elevated tumor necrosis factor-α in adipose tissue as an inducer of obesity-associated insulin resistance marked a new era of understanding that a subclinical inflammatory process underlies the insulin resistance and metabolic dysfunction that precedes type 2 diabetes. Advances in the field identified components of both the innate and adaptive immune response as key players in regulating such inflammatory processes. As antigen specificity is a hallmark of an adaptive immune response, its role in modulating the chronic inflammation that accompanies obesity and type 2 diabetes begs the question of whether insulin resistance and type 2 diabetes can have autoimmune components. In this Perspective, we summarize current data that pertain to the activation and perpetuation of adaptive immune responses during obesity and discuss key missing links and potential mechanisms for obesity-related insulin resistance and type 2 diabetes to be considered as potential autoimmune diseases.
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http://dx.doi.org/10.2337/db14-1488DOI Listing
June 2015

Regulation of obesity-related insulin resistance with gut anti-inflammatory agents.

Cell Metab 2015 Apr;21(4):527-42

Division of Cellular & Molecular Biology, Diabetes Research Group, Toronto General Research Institute (TGRI), University Health Network, 101 College Street, Toronto, ON M5G 1L7, Canada; Department of Immunology, University of Toronto, 1 King's College Circle, Toronto, ON M5S 1A8, Canada; Department of Laboratory Medicine and Pathobiology, University of Toronto, 1 King's College Circle, Toronto, ON M5S 1A8, Canada; Department of Pathology, University Health Network, 200 Elizabeth Street, Toronto, ON M5G 2C4, Canada. Electronic address:

Obesity has reached epidemic proportions, but little is known about its influence on the intestinal immune system. Here we show that the gut immune system is altered during high-fat diet (HFD) feeding and is a functional regulator of obesity-related insulin resistance (IR) that can be exploited therapeutically. Obesity induces a chronic phenotypic pro-inflammatory shift in bowel lamina propria immune cell populations. Reduction of the gut immune system, using beta7 integrin-deficient mice (Beta7(null)), decreases HFD-induced IR. Treatment of wild-type HFD C57BL/6 mice with the local gut anti-inflammatory, 5-aminosalicyclic acid (5-ASA), reverses bowel inflammation and improves metabolic parameters. These beneficial effects are dependent on adaptive and gut immunity and are associated with reduced gut permeability and endotoxemia, decreased visceral adipose tissue inflammation, and improved antigen-specific tolerance to luminal antigens. Thus, the mucosal immune system affects multiple pathways associated with systemic IR and represents a novel therapeutic target in this disease.
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http://dx.doi.org/10.1016/j.cmet.2015.03.001DOI Listing
April 2015

Nanoparticle-based immunotherapy for cancer.

ACS Nano 2015 Jan 23;9(1):16-30. Epub 2014 Dec 23.

Julia McFarlane Diabetes Research Centre (JMDRC) and Department of Microbiology, Immunology and Infectious Diseases, Snyder Institute for Chronic Diseases and Hotchkiss Brain Institute, Cummings School of Medicine, University of Calgary , Calgary, Alberta T2N 4N1 Canada.

The design of nanovaccines capable of triggering effective antitumor immunity requires an understanding of how the immune system senses and responds to threats, including pathogens and tumors. Equally important is an understanding of the mechanisms employed by tumor cells to evade immunity and an appreciation of the deleterious effects that antitumor immune responses can have on tumor growth, such as by skewing tumor cell composition toward immunologically silent tumor cell variants. The immune system and tumors engage in a tug-of-war driven by competition where promoting antitumor immunity or tumor cell death alone may be therapeutically insufficient. Nanotechnology affords a unique opportunity to develop therapeutic compounds than can simultaneously tackle both aspects, favoring tumor eradication. Here, we review the current status of nanoparticle-based immunotherapeutic strategies for the treatment of cancer, ranging from antigen/adjuvant delivery vehicles (to professional antigen-presenting cell types of the immune system) to direct tumor antigen-specific T-lymphocyte-targeting compounds and their combinations thereof.
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http://dx.doi.org/10.1021/nn5062029DOI Listing
January 2015

The cross-priming capacity and direct presentation potential of an autoantigen are separable and inversely related properties.

J Immunol 2014 Oct 27;193(7):3296-307. Epub 2014 Aug 27.

Julia McFarlane Diabetes Research Centre, Snyder Institute for Chronic Diseases, Faculty of Medicine, University of Calgary, Calgary, Alberta T2N 4N1, Canada; Department of Microbiology, Immunology and Infectious Diseases, Snyder Institute for Chronic Diseases, Faculty of Medicine, University of Calgary, Calgary, Alberta T2N 4N1, Canada; Institut D'Investigacions Biomèdiques August Pi i Sunyer, Barcelona 08036, Spain

We investigated whether a prevalent epitope of the β-cell-specific autoantigen islet-specific glucose-6-phosphatase catalytic subunit-related protein (IGRP206-214) reaches regional Ag-presentation pathways via unprocessed polypeptide chains, as free IGRP206-214 peptide or via preformed IGRP206-214/K(d) complexes. This was accomplished by expressing bacterial artificial chromosome transgenes encoding wild-type (stable) or ubiquitinated (unstable) forms of IGRP in IGRP-deficient NOD mice carrying MHC class I-deficient β-cells, dendritic cells, or B cells. We investigated the ability of the pancreatic lymph nodes of these mice to prime naive IGRP206-214-reactive CD8(+) T cells in vivo, either in response to spontaneous Ag shedding, or to synchronized forms of β-cell necrosis or apoptosis. When IGRP was made unstable by targeting it for proteasomal degradation within β-cells, the cross-priming, autoimmune-initiating potential of this autoantigen (designated autoantigenicity) was impaired. Yet at the same time, the direct presentation, CTL-targeting potential of IGRP (designated pathogenicity) was enhanced. The appearance of IGRP206-214 in regional Ag-presentation pathways was dissociated from transfer of IGRP206-214 or IGRP206-214/K(d) from β cells to dendritic cells. These results indicate that autoantigenicity and pathogenicity are separable and inversely related properties and suggest that pathogenic autoantigens, capable of efficiently priming CTLs while marking target cells for CTL-induced killing, may have a critical balance of these two properties.
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http://dx.doi.org/10.4049/jimmunol.1401001DOI Listing
October 2014

Nanomedicine in autoimmunity.

Immunol Lett 2014 Mar-Apr;158(1-2):167-74. Epub 2014 Jan 6.

Julia McFarlane Diabetes Research Centre (JMDRC) and Department of Microbiology, Immunology and Infectious Diseases, Snyder Institute for Chronic Diseases, Faculty of Medicine, University of Calgary, Calgary, Alberta, Canada; Institut D'Investigacions Biomediques August Pi i Sunyer, Barcelona, Spain. Electronic address:

The application of nanotechnology to the diagnosis and therapy of human diseases is already a reality and is causing a real revolution in how we design new therapies and vaccines. In this review we focus on the applications of nanotechnology in the field of autoimmunity. First, we review scenarios in which iron oxide nanoparticles have been used in the diagnosis of autoimmune diseases, mostly through magnetic resonance imaging (MRI), both in animal models and patients. Second, we discuss the potential of nanoparticles as an immunotherapeutic platform for autoimmune diseases, for now exclusively in pre-clinical models. Finally, we discuss the potential of this field to generate the 'perfect drug' with the capacity to report on its therapeutic efficacy in real time, that is, the birth of theranostics in autoimmunity.
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http://dx.doi.org/10.1016/j.imlet.2013.12.018DOI Listing
November 2014

Dendritic cell-dependent in vivo generation of autoregulatory T cells by antidiabetogenic MHC class II.

J Immunol 2013 Jul 5;191(1):70-82. Epub 2013 Jun 5.

Department of Microbiology, Immunology and Infectious Diseases and Julia McFarlane Diabetes Research Centre, Snyder Institute for Chronic Diseases, Faculty of Medicine, University of Calgary, Calgary, Alberta, Canada.

Several mechanisms have been proposed to explain how certain MHC class II molecules afford dominant resistance to autoimmune diseases like type 1 diabetes (T1D). However, it remains unclear how protective MHC types can blunt autoreactive T cell responses directed against a diverse repertoire of autoantigenic epitopes presented by disease-promoting MHCs. In this study, we show that expression of I-E on dendritic cells (DCs) of NOD mice promotes the differentiation of MHC promiscuous autoreactive CD4(+) clonotypes into antidiabetogenic autoregulatory T cells. We expressed an I-Eα(kloxP) transgene in NOD mice and used cell type-specific I-E ablation to show that I-E-expressing DCs, but not B cells, promote the generation of autoreactive CD4(+)Foxp3(+) regulatory T cells (Tregs) and their accumulation in the pancreas-draining lymph nodes. There, these Tregs suppress the presentation of β cell Ags to naive autoreactive CD4(+) and CD8(+) T cells restricted by diabetogenic MHC molecules in an I-E-independent manner. Whereas selective removal of I-E on DCs abrogated autoregulatory Treg formation and T1D protection, selective removal of I-E on B cells was inconsequential. These results explain how certain MHC class II molecules can completely suppress antigenically complex autoimmune responses in an Ag-nonspecific manner.
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http://dx.doi.org/10.4049/jimmunol.1300168DOI Listing
July 2013

Antidiabetogenic MHC class II promotes the differentiation of MHC-promiscuous autoreactive T cells into FOXP3+ regulatory T cells.

Proc Natl Acad Sci U S A 2013 Feb 11;110(9):3471-6. Epub 2013 Feb 11.

Department of Microbiology, Immunology and Infectious Diseases, Julia McFarlane Diabetes Research Centre, Snyder Institute for Chronic Diseases, Faculty of Medicine, University of Calgary, Calgary, AB, Canada T2N 4N1.

Polymorphisms in MHC class II molecules, in particular around β-chain position-57 (β57), afford susceptibility/resistance to multiple autoimmune diseases, including type 1 diabetes, through obscure mechanisms. Here, we show that the antidiabetogenic MHC class II molecule I-A(b) affords diabetes resistance by promoting the differentiation of MHC-promiscuous autoreactive CD4(+) T cells into disease-suppressing natural regulatory T cells, in a β56-67-regulated manner. We compared the tolerogenic and antidiabetogenic properties of CD11c promoter-driven transgenes encoding I-A(b) or a form of I-A(b) carrying residues 56-67 of I-Aβ(g7) (I-A(b-g7)) in wild-type nonobese diabetic (NOD) mice, as well as NOD mice coexpressing a diabetogenic and I-A(g7)-restricted, but MHC-promiscuous T-cell receptor (4.1). Both I-A transgenes protected NOD and 4.1-NOD mice from diabetes. However, whereas I-A(b) induced 4.1-CD4(+) thymocyte deletion and 4.1-CD4(+)Foxp3(+) regulatory T-cell development, I-A(b-g7) promoted 4.1-CD4(+)Foxp3(+) Treg development without inducing clonal deletion. Furthermore, non-T-cell receptor transgenic NOD.CD11cP-I-A(b) and NOD.CD11cP-IA(b-g7) mice both exported regulatory T cells with superior antidiabetogenic properties than wild-type NOD mice. We propose that I-A(b), and possibly other protective MHC class II molecules, afford disease resistance by engaging a naturally occurring constellation of MHC-promiscuous autoreactive T-cell clonotypes, promoting their deviation into autoregulatory T cells.
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http://dx.doi.org/10.1073/pnas.1211391110DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3587200PMC
February 2013

IL-2 promotes the function of memory-like autoregulatory CD8+ T cells but suppresses their development via FoxP3+ Treg cells.

Eur J Immunol 2013 Feb 15;43(2):394-403. Epub 2013 Jan 15.

Julia McFarlane Diabetes Research Centre and Departments of Microbiology, Snyder Institute for Chronic Diseases, the University of Calgary, Calgary, AB, Canada.

IL-2 plays a critical role in both effector T-cell development and FoxP3(+) CD4(+) Treg-cell homeostasis. A reduction in Il2 transcription results in impaired FoxP3(+) CD4(+) Treg-cell recruitment and function, and accounts for the association between murine Il2 and type 1 diabetes (T1D). The progression of T1D elicits a disease-countering negative feedback regulatory loop that involves the differentiation of low-avidity autoreactive CD8(+) T cells into memory-like autoregulatory T cells in a CD4(+) Th-dependent manner. Since these auto-regulatory T cells express IL-2Rβ (CD122), we hypothesized that their development might also be regulated by IL-2. Here, we investigate the effects of differences in IL-2 expression on this autoregulatory subset. We show that decreased IL-2 production impairs the regulatory capacity of memory-like autoregulatory CD8(+) CD122(+) T cells. Surprisingly, we also find that a reduction in IL-2 production capacity increases memory autoregulatory CD8(+) T-cell formation indirectly, by decreasing the development and function of FoxP3(+) Treg cells in nonobese diabetic mice. These results illustrate a complex homeostatic interplay between IL-2, CD4(+) Th cells, FoxP3(+) CD4(+) Treg cells and autoregulatory CD8(+) T-cell memory whereby IL-2 controls the function of both Treg-cell subsets, but IL-2-potentiation of FoxP3(+) CD4(+) Treg-cell function results in the suppression of CD4(+) Th-cell activation and autoregulatory memory CD8(+) T-cell formation.
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http://dx.doi.org/10.1002/eji.201242845DOI Listing
February 2013

Local autoantigen expression as essential gatekeeper of memory T-cell recruitment to islet grafts in diabetic hosts.

Diabetes 2013 Mar 16;62(3):905-11. Epub 2012 Nov 16.

Julia McFarlane Diabetes Research Centre and Department of Microbiology, Immunology, and Infectious Diseases, Faculty of Medicine, University of Calgary, Calgary, Alberta, Canada.

It is generally believed that inflammatory cues can attract noncognate, "bystander" T-cell specificities to sites of inflammation. We have shown that recruitment of naive and in vitro activated autoreactive CD8⁺ T cells into endogenous islets requires local autoantigen expression. Here, we demonstrate that absence of an autoantigen in syngeneic extrapancreatic islet grafts in diabetic hosts renders the grafts "invisible" to cognate memory (and naive) T cells. We monitored the recruitment of islet-specific glucose-6-phosphatase catalytic subunit-related protein (IGRP)₂₀₆₋₂₁₄-reactive CD8⁺ T cells into IGRP₂₀₆₋₂₁₄-competent and IGRP₂₀₆₋₂₁₄-deficient islet grafts in diabetic wild-type or IGRP₂₀₆₋₂₁₄(-/-) nonobese diabetic hosts (harboring either naive and memory T cells or only naive IGRP₂₀₆₋₂₁₄-specific T-cells, respectively). All four host-donor combinations had development of recurrent diabetes within 2 weeks. Wild-type hosts recruited IGRP₂₀₆₋₂₁₄-specific T cells into IGRP₂₀₆₋₂₁₄(+/+) but not IGRP₂₀₆₋₂₁₄(-/-) grafts. In IGRP₂₀₆₋₂₁₄(-/-) hosts, there was no recruitment of IGRP₂₀₆₋₂₁₄-specific T cells, regardless of donor type. Graft-derived IGRP₂₀₆₋₂₁₄ activated naive IGRP₂₀₆₋₂₁₄-specific T cells, but graft destruction invariably predated their recruitment. These results indicate that recurrent diabetes is exclusively driven by autoreactive T cells primed during the primary autoimmune response, and demonstrate that local antigen expression is a sine qua non requirement for accumulation of memory T cells into islet grafts. These findings underscore the importance of tackling autoreactive T-cell memory after β-cell replacement therapy.
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http://dx.doi.org/10.2337/db12-0600DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3581210PMC
March 2013

Antigen-specific therapeutic approaches in Type 1 diabetes.

Cold Spring Harb Perspect Med 2012 Feb;2(2):a007773

Julia McFarlane Diabetes Research Centre, University of Calgary, NW Calgary, Alberta T2N 4N1, Canada.

Development of strategies capable of specifically curbing pathogenic autoimmune responses in a disease- and organ-specific manner without impairing foreign or tumor antigen-specific immune responses represents a long sought-after goal in autoimmune disease research. Unfortunately, our current understanding of the intricate details of the different autoimmune diseases that affect mankind, including type 1 diabetes, is rudimentary. As a result, progress in the development of the so-called "antigen-specific" therapies for autoimmunity has been slow and fraught with limitations that interfere with bench-to-bedside translation. Absent or incomplete understanding of mechanisms of action and lack of adequate immunological biomarkers, for example, preclude the rational design of effective drug development programs. Here, we provide an overview of antigen-specific approaches that have been tested in preclinical models of T1D and, in some cases, human subjects. The evidence suggests that effective translation of these approaches through clinical trials and into patients will continue to meet with failure unless detailed mechanisms of action at the level of the organism are defined.
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http://dx.doi.org/10.1101/cshperspect.a007773DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3281592PMC
February 2012

Development of memory-like autoregulatory CD8+ T cells is CD4+ T cell dependent.

J Immunol 2011 Sep 8;187(6):2859-66. Epub 2011 Aug 8.

Julia McFarlane Diabetes Research Centre, Faculty of Medicine, University of Calgary, Calgary, Alberta T2N 4N1, Canada.

Progression of spontaneous autoimmune diabetes is associated with development of a disease-countering negative-feedback regulatory loop that involves differentiation of low-avidity autoreactive CD8(+) cells into memory-like autoregulatory T cells. Such T cells blunt diabetes progression by suppressing the presentation of both cognate and noncognate Ags to pathogenic high-avidity autoreactive CD8(+) T cells in the pancreas-draining lymph nodes. In this study, we show that development of autoregulatory CD8(+) T cell memory is CD4(+) T cell dependent. Transgenic (TG) NOD mice expressing a low-affinity autoreactive TCR were completely resistant to autoimmune diabetes, even after systemic treatment of the mice with agonistic anti-CD40 or anti-4-1BB mAbs or autoantigen-pulsed dendritic cells, strategies that dramatically accelerate diabetes development in TG NOD mice expressing a higher affinity TCR for the same autoantigenic specificity. Furthermore, whereas abrogation of RAG-2 expression, hence endogenous CD4(+) T cell and B cell development, decelerated disease progression in high-affinity TCR-TG NOD mice, it converted the low-affinity TCR into a pathogenic one. In agreement with these data, polyclonal CD4(+) T cells from prediabetic NOD mice promoted disease in high-affinity TCR-TG NOD.Rag2(-/-) mice, but inhibited it in low-affinity TCR-TG NOD.Rag2(-/-) mice. Thus, in chronic autoimmune responses, CD4(+) Th cells contribute to both promoting and suppressing pathogenic autoimmunity.
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http://dx.doi.org/10.4049/jimmunol.1101117DOI Listing
September 2011

CD8(+) Tregs in autoimmunity: learning "self"-control from experience.

Cell Mol Life Sci 2011 Dec 14;68(23):3781-95. Epub 2011 Jun 14.

Julia McFarlane Diabetes Research Centre, The University of Calgary, AB, Canada.

Autoreactive CD8(+) regulatory T cells (Tregs) play important roles as modulators of immune responses against self, and numerical and functional defects in CD8(+) Tregs have been linked to autoimmunity. Several subsets of CD8(+) Tregs have been described. However, the origin of these T cells and how they participate in the natural progression of autoimmunity remain poorly defined. We discuss several lines of evidence suggesting that the autoimmune process itself promotes the development of autoregulatory CD8(+) T cells. We posit that chronic autoantigenic exposure fosters the differentiation of non-pathogenic autoreactive CD8(+) T cells into antigen-experienced, memory-like autoregulatory T cells, to generate a "negative feedback" regulatory loop capable of countering pathogenic autoreactive effectors. This hypothesis predicts that approaches capable of boosting autoregulatory T cell memory will be able to blunt autoimmunity without compromising systemic immunity.
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http://dx.doi.org/10.1007/s00018-011-0738-yDOI Listing
December 2011

Peptide-MHC-based nanovaccines for the treatment of autoimmunity: a "one size fits all" approach?

J Mol Med (Berl) 2011 Aug 16;89(8):733-42. Epub 2011 Apr 16.

Julia McFarlane Diabetes Research Centre and Department of Microbiology and Infectious Diseases, Institute of Inflammation, Infection and Immunity, Faculty of Medicine, The University of Calgary, Alberta, Canada.

Nanotechnology offers enormous potential in drug delivery and in vivo imaging. Nanoparticles (NPs), for example, are being extensively tested as scaffolds to deliver anti-cancer therapeutics or imaging tags. Our recent work, discussed herein, indicates that an opportunity exists to use NPs to deliver ligands for, and trigger, cognate receptors on T lymphocytes as a way to induce therapeutic immune responses in vivo. Specifically, systemic delivery of NPs coated with Type 1 diabetes (T1D)-relevant peptide-major histocompatibility complex molecules triggered the expansion of cognate memory autoregulatory (disease-suppressing) T cells, suppressed the progression of autoimmune attack against insulin-producing beta cells, and restored glucose homeostasis. This therapeutic avenue exploits a new paradigm in the progression of chronic autoimmune responses that enables the rational design of disease-specific "nanovaccines" capable of blunting autoimmunity without impairing systemic immunity, a long sought-after goal in the therapy of these disorders. Here, we discuss the research paths that led to the discovery of this therapeutic avenue and highlight the features that make it an attractive approach for the treatment, in an antigen-specific manner, of a whole host of autoimmune diseases.
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http://dx.doi.org/10.1007/s00109-011-0757-zDOI Listing
August 2011

Turning human epidermis into pancreatic endoderm.

Rev Diabet Stud 2010 10;7(2):158-67. Epub 2010 Aug 10.

Center of Regenerative Medicine in Barcelona, Dr. Aiguader 88, 08003 Barcelona, Spain.

Objective: Human embryonic stem (hES) cells can be differentiated into pancreatic endoderm structures in vitro. The study was performed to determine whether induced pluripotent stem (iPS) cells can be differentiated into similar structures with comparable efficiency.

Methods: We compared the ability of hES cells and iPS cells derived from human epidermal keratinocytes to progressively differentiate into pancreatic endoderm. Human foreskin keratinocytes were reprogrammed to pluripotency by transduction with retroviruses encoding Oct4, Sox2, and Klf4. The resulting keratinocyte-derived iPS (KiPS) cell lines and a hES cell line were subjected to a modified pancreatic endoderm differentiation protocol. Cells and embryoid-body structures derived from both hES and KiPS cells were compared at different stages of development for expression of stem cell and differentiation markers, including Sox2, Oct4, Mixl1, Brachyury, Gsc, FoxA2, Sox17, Hnf4α, Hnf1β, Nkx2.2, Nkx6.1, Hex, Isl1, Pdx1, and Slc2A, via Taqman real-time PCR, flow-cytometry, and/or immunocytochemistry.

Results: hES cells and KiPS cells expressed similar levels of the stem cell factors Sox2 and Oct4. Upon differentiation, both cell types underwent remarkably similar changes in gene expression. They acquired the definitive endoderm markers Sox17 and FoxA2. Most Sox17+ and FoxA2+ cells co-expressed Hnf4α and Hnf1β, found in the primitive gut tube, a pancreas precursor. Most FoxA2+ cells were also Pdx1+, and many expressed Nkx2.2, Nkx6.1, and Isl1.

Conclusions: Keratinocyte-derived iPS cells can be differentiated into pancreatic endoderm, and the efficiency of this process is comparable to that seen for hES cells. Thus keratinocytes have the potential to serve as a source of patient-specific pancreatic endoderm for transplantation.
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http://dx.doi.org/10.1900/RDS.2010.7.158DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2989788PMC
February 2011

Reversal of autoimmunity by boosting memory-like autoregulatory T cells.

Immunity 2010 Apr 8;32(4):568-80. Epub 2010 Apr 8.

Julia McFarlane Diabetes Research Centre, Department of Microbiology and Infectious Diseases, The University of Calgary, 3330 Hospital Drive N.W., Calgary, AB T2N 4N1, Canada.

Blunting autoreactivity without compromising immunity remains an elusive goal in the treatment of autoimmunity. We show that progression to autoimmune diabetes results in the conversion of naive low-avidity autoreactive CD8(+) T cells into memory-like autoregulatory cells that can be expanded in vivo with nanoparticles coated with disease-relevant peptide-major histocompatibility complexes (pMHC-NP). Treatment of NOD mice with monospecific pMHC-NPs expanded cognate autoregulatory T cells, suppressed the recruitment of noncognate specificities, prevented disease in prediabetic mice, and restored normoglycemia in diabetic animals. pMHC-NP therapy was inconsequential in mice engineered to bear an immune system unresponsive to the corresponding epitope, owing to absence of epitope-experienced autoregulatory T cells. pMHC-NP-expanded autoregulatory T cells suppressed local presentation of autoantigens in an interferon-gamma-, indoleamine 2,3-dioxygenase-, and perforin-dependent manner. Nanoparticles coated with human diabetes-relevant pHLA complexes restored normoglycemia in a humanized model of diabetes. These observations expose a paradigm in the pathogenesis of autoimmunity amenable for therapeutic intervention.
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http://dx.doi.org/10.1016/j.immuni.2010.03.015DOI Listing
April 2010