Publications by authors named "Vinay S Mahajan"

38 Publications

Multisystem inflammatory syndrome in children is driven by zonulin-dependent loss of gut mucosal barrier.

J Clin Invest 2021 07;131(14)

Department of Pediatrics, Division of Infectious Diseases and Immunology, Infectious and Immunologic Diseases Research Center (IIDRC) and Department of Biomedical Sciences, Cedars-Sinai Medical Center, Los Angeles, California, USA.

BACKGROUNDWeeks after SARS-CoV-2 infection or exposure, some children develop a severe, life-threatening illness called multisystem inflammatory syndrome in children (MIS-C). Gastrointestinal (GI) symptoms are common in patients with MIS-C, and a severe hyperinflammatory response ensues with potential for cardiac complications. The cause of MIS-C has not been identified to date.METHODSHere, we analyzed biospecimens from 100 children: 19 with MIS-C, 26 with acute COVID-19, and 55 controls. Stools were assessed for SARS-CoV-2 by reverse transcription PCR (RT-PCR), and plasma was examined for markers of breakdown of mucosal barrier integrity, including zonulin. Ultrasensitive antigen detection was used to probe for SARS-CoV-2 antigenemia in plasma, and immune responses were characterized. As a proof of concept, we treated a patient with MIS-C with larazotide, a zonulin antagonist, and monitored the effect on antigenemia and the patient's clinical response.RESULTSWe showed that in children with MIS-C, a prolonged presence of SARS-CoV-2 in the GI tract led to the release of zonulin, a biomarker of intestinal permeability, with subsequent trafficking of SARS-CoV-2 antigens into the bloodstream, leading to hyperinflammation. The patient with MIS-C treated with larazotide had a coinciding decrease in plasma SARS-CoV-2 spike antigen levels and inflammatory markers and a resultant clinical improvement above that achieved with currently available treatments.CONCLUSIONThese mechanistic data on MIS-C pathogenesis provide insight into targets for diagnosing, treating, and preventing MIS-C, which are urgently needed for this increasingly common severe COVID-19-related disease in children.
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http://dx.doi.org/10.1172/JCI149633DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8279585PMC
July 2021

B1a and B2 cells are characterized by distinct CpG modification states at DNMT3A-maintained enhancers.

Nat Commun 2021 04 13;12(1):2208. Epub 2021 Apr 13.

Ragon Institute of MGH, MIT, and Harvard, Cambridge, MA, USA.

The B1 and B2 lineages of B cells contribute to protection from pathogens in distinct ways. The role of the DNA CpG methylome in specifying these two B-cell fates is still unclear. Here we profile the CpG modifications and transcriptomes of peritoneal B1a and follicular B2 cells, as well as their respective proB cell precursors in the fetal liver and adult bone marrow from wild-type and CD19-Cre Dnmt3a floxed mice lacking DNMT3A in the B lineage. We show that an underlying foundational CpG methylome is stably established during B lineage commitment and is overlaid with a DNMT3A-maintained dynamic methylome that is sculpted in distinct ways in B1a and B2 cells. This dynamic DNMT3A-maintained methylome is composed of novel enhancers that are closely linked to lineage-specific genes. While DNMT3A maintains the methylation state of these enhancers in both B1a and B2 cells, the dynamic methylome undergoes a prominent programmed demethylation event during B1a but not B2 cell development. We propose that the methylation pattern of DNMT3A-maintained enhancers is determined by the coincident recruitment of DNMT3A and TET enzymes, which regulate the developmental expression of B1a and B2 lineage-specific genes.
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http://dx.doi.org/10.1038/s41467-021-22458-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8044213PMC
April 2021

Expansion of Cytotoxic CD4+ T cells in the lungs in severe COVID-19.

medRxiv 2021 Mar 26. Epub 2021 Mar 26.

The contributions of T cells infiltrating the lungs to SARS-CoV-2 clearance and disease progression are poorly understood. Although studies of CD8+ T cells in bronchoalveolar lavage and blood have suggested that these cells are exhausted in severe COVID-19, CD4+ T cells have not been systematically interrogated within the lung parenchyma. We establish here that cytotoxic CD4+ T cells (CD4+CTLs) are prominently expanded in the COVID-19 lung infiltrate. CD4+CTL numbers in the lung increase with disease severity and progression is accompanied by widespread HLA-DR expression on lung epithelial and endothelial cells, increased apoptosis of epithelial cells and tissue remodeling. Based on quantitative evidence for re-activation in the lung milieu, CD4+ CTLs are as likely to drive viral clearance as CD8+ T cells and may also be contributors to lung inflammation and eventually to fibrosis in severe COVID-19.

In Brief: In severe COVID-19 cytotoxic CD4+ T cells accumulate in draining lymph nodes and in the lungs during the resolving phase of the disease. Re-activated cytotoxic CD4+ T cells and cytotoxic CD8+ T cells are present in roughly equivalent numbers in the lungs at this stage and these cells likely collaborate to eliminate virally infected cells and potentially induce fibrosis. A large fraction of epithelial and endothelial cells in the lung express HLA class II in COVID-19 and there is temporal convergence between CD4+CTL accumulation and apoptosis in the lung.

Highlights: In severe COVID-19, activated CD4+ CTLs accumulate in the lungs late in diseaseThese cells likely participate in SARS-CoV-2 clearance, collaborating with CD8+ T cells many of which exhibit an exhausted phenotypeT cells likely contribute to the late exacerbation of inflammationCD4+CTLs have been linked to fibrosis in many disorders and could also be responsible for the eventual induction of fibrosis in a subset of COVID-19 patients.

Summary: The contributions of T cells infiltrating the lungs to SARS-CoV-2 clearance and disease progression are poorly understood. Although studies of CD8+ T cells in bronchoalveolar lavage and blood have suggested that these cells are exhausted in severe COVID-19, CD4+ T cells have not been systematically interrogated within the lung parenchyma. We establish here that cytotoxic CD4+ T cells (CD4+CTLs) are prominently expanded in the COVID-19 lung infiltrate. CD4+CTL numbers in the lung increase with disease severity and progression is accompanied by widespread HLA-DR expression on lung epithelial and endothelial cells, increased apoptosis of epithelial cells and tissue remodeling. Based on quantitative evidence for re-activation in the lung milieu, CD4+ CTLs are as likely to drive viral clearance as CD8+ T cells and may also be contributors to lung inflammation and eventually to fibrosis in severe COVID-19.
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http://dx.doi.org/10.1101/2021.03.23.21253885DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8010762PMC
March 2021

CD4CTLs in Fibrosing Mediastinitis Linked to .

J Immunol 2021 02 16;206(3):524-530. Epub 2020 Dec 16.

Ragon Institute of MGH, MIT and Harvard, Cambridge, MA 02139;

Although fibrotic disorders are frequently assumed to be linked to T cells, quantitative tissue interrogation studies have rarely been performed to establish this link and certainly many fibrotic diseases do not fall within the type 2/allergic disease spectrum. We have previously linked two human autoimmune fibrotic diseases, IgG4-related disease and systemic sclerosis, to the clonal expansion and lesional accumulation of CD4CTLs. In both these diseases T cell accumulation was found to be sparse. Fibrosing mediastinitis linked to infection histologically resembles IgG4-related disease in terms of the inflammatory infiltrate and fibrosis, and it provides an example of a fibrotic disease of infectious origin in which the potentially profibrotic T cells may be induced and reactivated by fungal Ags. We show in this study that, in this human disease, CD4CTLs accumulate in the blood, are clonally expanded, infiltrate into disease lesions, and can be reactivated in vitro by Ags. T cells are relatively sparse at lesional sites. These studies support a general role for CD4CTLs in inflammatory fibrosis and suggest that fibrosing mediastinitis is an Ag-driven disease that may provide important mechanistic insights into the pathogenesis of idiopathic fibrotic diseases.
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http://dx.doi.org/10.4049/jimmunol.2000433DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7978153PMC
February 2021

Loss of Bcl-6-Expressing T Follicular Helper Cells and Germinal Centers in COVID-19.

Cell 2020 10 19;183(1):143-157.e13. Epub 2020 Aug 19.

Ragon Institute of MGH, MIT and Harvard, Cambridge, MA 02139, USA.

Humoral responses in coronavirus disease 2019 (COVID-19) are often of limited durability, as seen with other human coronavirus epidemics. To address the underlying etiology, we examined post mortem thoracic lymph nodes and spleens in acute SARS-CoV-2 infection and observed the absence of germinal centers and a striking reduction in Bcl-6 germinal center B cells but preservation of AID B cells. Absence of germinal centers correlated with an early specific block in Bcl-6 T cell differentiation together with an increase in T-bet T cells and aberrant extra-follicular TNF-α accumulation. Parallel peripheral blood studies revealed loss of transitional and follicular B cells in severe disease and accumulation of SARS-CoV-2-specific "disease-related" B cell populations. These data identify defective Bcl-6 T cell generation and dysregulated humoral immune induction early in COVID-19 disease, providing a mechanistic explanation for the limited durability of antibody responses in coronavirus infections, and suggest that achieving herd immunity through natural infection may be difficult.
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http://dx.doi.org/10.1016/j.cell.2020.08.025DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7437499PMC
October 2020

The Loss of Bcl-6 Expressing T Follicular Helper Cells and the Absence of Germinal Centers in COVID-19.

SSRN 2020 Jul 16:3652322. Epub 2020 Jul 16.

Ragon Institute of MGH, MIT and Harvard, Cambridge, MA 02139, USA.

Humoral responses in COVID-19 disease are often of limited durability, as seen with other human coronavirus epidemics. To address the underlying etiology, we examined postmortem thoracic lymph nodes and spleens in acute SARS-CoV-2 infection and observed the absence of germinal centers, a striking reduction in Bcl-6+ germinal center B cells but preservation of AID+ B cells. Absence of germinal centers correlated with an early specific block in Bcl-6+TFH cell differentiation together with an increase in T-bet+TH1 cells and aberrant extra-follicular TNF-a accumulation.  Parallel peripheral blood studies revealed loss of transitional and follicular B cells in severe disease and accumulation of SARS-CoV-2-specific "disease-related" B cell populations. These data identify defective Bcl-6+TFH cell generation and dysregulated humoral immune induction early in COVID-19 disease, providing a mechanistic explanation for the limited durability of antibody responses in coronavirus infections and suggest that achieving herd immunity through natural infection may be difficult. Funding: This work was supported by NIH U19 AI110495 to SP, NIH R01 AI146779 to AGS, NIH R01AI137057 and DP2DA042422 to DL, BMH was supported by NIGMS T32 GM007753, TMC was supported by T32 AI007245. Funding for these studies from the Massachusetts Consortium of Pathogen Readiness, the Mark and Lisa Schwartz Foundation and Enid Schwartz is also acknowledged. Conflict of Interest: None. Ethical Approval: This study was performed with the approval of the Institutional Review Boards at the Massachusetts General Hospital and the Brigham and Women's Hospital.
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http://dx.doi.org/10.2139/ssrn.3652322DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7385482PMC
July 2020

CD4 and CD8 cytotoxic T lymphocytes may induce mesenchymal cell apoptosis in IgG-related disease.

J Allergy Clin Immunol 2021 01 30;147(1):368-382. Epub 2020 May 30.

Ragon Institute of MGH, MIT, and Harvard, Cambridge, Mass. Electronic address:

Background: IgG-related disease (IgG-RD) is an immune-mediated fibrotic disorder that has been linked to CD4 cytotoxic T lymphocytes (CD4CTLs). The effector phenotype of CD4CTLs and the relevance of both CD8 cytotoxic T lymphocytes (CD8CTLs) and apoptotic cell death remain undefined in IgG-RD.

Objective: We sought to define CD4CTL heterogeneity, characterize the CD8CTL response in the blood and in lesions, and determine whether enhanced apoptosis may contribute to the pathogenesis of IgG-RD.

Methods: Blood analyses were undertaken using flow cytometry, cell sorting, transcriptomic analyses at the population and single-cell levels, and next-generation sequencing for the TCR repertoire. Tissues were interrogated using multicolor immunofluorescence. Results were correlated with clinical data.

Results: We establish that among circulating CD4CTLs in IgG-RD, CD27CD28CD57 cells are the dominant effector subset, exhibit marked clonal expansion, and differentially express genes relevant to cytotoxicity, activation, and enhanced metabolism. We also observed prominent infiltration of granzyme A-expressing CD8CTLs in disease tissues and clonal expansion in the blood of effector/memory CD8 T cells with an activated and cytotoxic phenotype. Tissue studies revealed an abundance of cells undergoing apoptotic cell death disproportionately involving nonimmune, nonendothelial cells of mesenchymal origin. Apoptotic cells showed significant upregulation of HLA-DR.

Conclusions: CD4CTLs and CD8CTLs may induce apoptotic cell death in tissues of patients with IgG-RD with preferential targeting of nonendothelial, nonimmune cells of mesenchymal origin.
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http://dx.doi.org/10.1016/j.jaci.2020.05.022DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7704943PMC
January 2021

Cytotoxic CD4+ T lymphocytes may induce endothelial cell apoptosis in systemic sclerosis.

J Clin Invest 2020 05;130(5):2451-2464

Ragon Institute, Massachusetts General Hospital, Massachusetts Institute of Technology and Harvard University, Boston, Massachusetts, USA.

Systemic sclerosis (SSc) is an autoimmune fibrotic disease whose pathogenesis is poorly understood and lacks effective therapies. We undertook quantitative analyses of T cell infiltrates in the skin of 35 untreated patients with early diffuse SSc and here show that CD4+ cytotoxic T cells and CD8+ T cells contribute prominently to these infiltrates. We also observed an accumulation of apoptotic cells in SSc tissues, suggesting that recurring cell death may contribute to tissue damage and remodeling in this fibrotic disease. HLA-DR-expressing endothelial cells were frequent targets of apoptosis in SSc, consistent with the prominent vasculopathy seen in patients with this disease. A circulating effector population of cytotoxic CD4+ T cells, which exhibited signatures of enhanced metabolic activity, was clonally expanded in patients with systemic sclerosis. These data suggest that cytotoxic T cells may induce the apoptotic death of endothelial and other cells in systemic sclerosis. Cell loss driven by immune cells may be followed by overly exuberant tissue repair processes that lead to fibrosis and tissue dysfunction.
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http://dx.doi.org/10.1172/JCI131700DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7190971PMC
May 2020

DOCK2 Sets the Threshold for Entry into the Virtual Memory CD8 T Cell Compartment by Negatively Regulating Tonic TCR Triggering.

J Immunol 2020 01 18;204(1):49-57. Epub 2019 Nov 18.

Ragon Institute of MGH, MIT and Harvard, Cambridge, MA 02139;

The control of cytoskeletal dynamics by dedicator of cytokinesis 2 (DOCK2), a hematopoietic cell-specific actin effector protein, has been implicated in TCR signaling and T cell migration. Biallelic mutations in have been identified in patients with a recessive form of combined immunodeficiency with defects in T, B, and NK cell activation. Surprisingly, we show in this study that certain immune functions of CD8 T cells are enhanced in the absence of DOCK2. -deficient mice have a pronounced expansion of their memory T cell compartment. Bone marrow chimera and adoptive transfer studies indicate that these memory T cells develop in a cell-intrinsic manner following thymic egress. Transcriptional profiling, TCR repertoire analyses, and cell surface marker expression indicate that -deficient naive CD8 T cells directly convert into virtual memory cells without clonal effector T cell expansion. This direct conversion to memory is associated with a selective increase in TCR sensitivity to self-peptide MHC in vivo and an enhanced response to weak agonist peptides ex vivo. In contrast, the response to strong agonist peptides remains unaltered in -deficient T cells. Collectively, these findings suggest that the regulation of the actin dynamics by DOCK2 enhances the threshold for entry into the virtual memory compartment by negatively regulating tonic TCR triggering in response to weak agonists.
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http://dx.doi.org/10.4049/jimmunol.1900440DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6920543PMC
January 2020

Update on Clinical Immunology Laboratory Testing.

Authors:
Vinay S Mahajan

Clin Lab Med 2019 12 3;39(4):xi-xii. Epub 2019 Oct 3.

Ragon Institute of MGH, MIT and Harvard, 400 Technology Square, Room 8-890, Cambridge, MA 02139, USA. Electronic address:

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http://dx.doi.org/10.1016/j.cll.2019.09.001DOI Listing
December 2019

The Future of Clinical Immunology Laboratory Testing.

Clin Lab Med 2019 12 3;39(4):699-708. Epub 2019 Oct 3.

Ragon Institute of MGH, MIT and Harvard, 400 Technology Square, Cambridge, MA 02139, USA; Department of Pathology, Brigham and Women's Hospital, 75 Francis Street, Armory 3, Boston, MA 02115, USA. Electronic address:

For decades, autoantibody detection has comprised the bulk of clinical laboratory immunology. However, most immune disorders are caused by imbalances in both humoral and cellular immunity. Our knowledge of the immune system has grown exponentially, resulting in new treatment paradigms in immunology. Extensive functional characterization of lymphocyte subsets is routinely carried out in a research laboratories, facilitated by the emergence of high-dimensional analysis technologies for low cell numbers. It will not be long before these approaches enter the diagnostic realm. This chapter outlines emerging trends in laboratory immunology testing with a focus on deep immune profiling or high-dimensional testing modalities.
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http://dx.doi.org/10.1016/j.cll.2019.07.014DOI Listing
December 2019

Molecular Diagnosis of Inherited Immune Disorders.

Clin Lab Med 2019 12 26;39(4):685-697. Epub 2019 Sep 26.

Ragon Institute of MGH, MIT and Harvard, Cambridge, MA, USA; Center for Advanced Molecular Diagnostics, Brigham and Women's Hospital, Boston, MA, USA.

Primary immunodeficiency diseases are a heterogeneous group of rare inherited disorders of innate or adaptive immune system function. Patients with primary immunodeficiencies typically present with recurrent and severe infections in infancy or young adulthood. More recently, the co-occurrence of autoimmune, benign lymphoproliferative, atopic, and malignant complications has been described. The diagnosis of a primary immunodeficiency disorder requires a thorough assessment of a patient's underlying immune system function. Historically, this has been accomplished at the time of symptomatic presentation by measuring immunoglobulins, complement components, protective antibody titers, or immune cell counts in the peripheral blood. Although these data can be used to critically assess the degree of immune dysregulation in the patient, this approach fall short in at least 2 regards. First, this assessment often occurs after the patient has suffered life-threatening infectious or autoinflammatory complications. Second, these data fail to uncover an underlying molecular cause of the patient's primary immune dysfunction, prohibiting the use of molecularly targeted therapeutic interventions. Within the last decade, the field of primary immunodeficiency diagnostics has been revolutionized by 2 major molecular advancements: (1) the onset of newborn screening in 2008, and (2) the onset of next-generation sequencing in 2010. In this article, the techniques of newborn screening and next-generation sequencing are reviewed and their respective impacts on the field of primary immunodeficiency disorders are discussed with a specific emphasis on severe combined immune deficiency and common variable immune deficiency.
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http://dx.doi.org/10.1016/j.cll.2019.07.013DOI Listing
December 2019

Induction of metabolic quiescence defines the transitional to follicular B cell switch.

Sci Signal 2019 10 22;12(604). Epub 2019 Oct 22.

Ragon Institute of Massachusetts General Hospital, Massachusetts Institute of Technology, and Department of Medicine, Harvard University, Cambridge, MA 02139, USA.

Transitional B cells must actively undergo selection for self-tolerance before maturing into their resting follicular B cell successors. We found that metabolic quiescence was acquired at the follicular B cell stage in both humans and mice. In follicular B cells, the expression of genes involved in ribosome biogenesis, aerobic respiration, and mammalian target of rapamycin complex 1 (mTORC1) signaling was reduced when compared to that in transitional B cells. Functional metabolism studies, profiling of whole-cell metabolites, and analysis of cell surface proteins in human B cells suggested that this transition was also associated with increased extracellular adenosine salvage. Follicular B cells increased the abundance of the cell surface ectonucleotidase CD73, which coincided with adenosine 5'-monophosphate-activated protein kinase (AMPK) activation. Differentiation to the follicular B cell stage in vitro correlated with surface acquisition of CD73 on human transitional B cells and was augmented with the AMPK agonist, AICAR. Last, individuals with gain-of-function (PI3Kδ) mutations and increased pS6 activation exhibited a near absence of circulating follicular B cells. Together, our data suggest that mTORC1 attenuation may be necessary for human follicular B cell development. These data identify a distinct metabolic switch during human B cell development at the transitional to follicular stages, which is characterized by an induction of extracellular adenosine salvage, AMPK activation, and the acquisition of metabolic quiescence.
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http://dx.doi.org/10.1126/scisignal.aaw5573DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7301641PMC
October 2019

IgG4-related disease: Association with a rare gene variant expressed in cytotoxic T cells.

Mol Genet Genomic Med 2019 06 16;7(6):e686. Epub 2019 Apr 16.

Vanderbilt Center for Undiagnosed Disease, Vanderbilt University, Nashville, Tennessee.

Background: Family screening of a 48-year-old male with recently diagnosed IgG4-related disease (IgG4-RD) revealed unanticipated elevations in plasma IgG4 in his two healthy teenaged sons.

Methods: We performed gene sequencing, immune cell studies, HLA typing, and analyses of circulating cytotoxic CD4+ T lymphocytes and plasmablasts to seek clues to pathogenesis. DNA from a separate cohort of 99 patients with known IgG4-RD was also sequenced for the presence of genetic variants in a specific gene, FGFBP2.

Results: The three share a previously unreported heterozygous single base deletion in fibroblast growth factor binding protein type 2 (FGFBP2), which causes a frameshift in the coding sequence. The FGFBP2 protein is secreted by cytotoxic T-lymphocytes and binds fibroblast growth factor. The variant sequence in the FGFBP2 protein is predicted to form a disordered random coil rather than a helical-turn-helix structure, unable to adopt a stable conformation. The proband and the two sons had 5-10-fold higher numbers of circulating cytotoxic CD4 + T cells and plasmablasts compared to matched controls. The three members also share a homozygous missense common variant in FGFBP2 found in heterozygous form in ~40% of the population. This common variant was found in 73% of an independent, well characterized IgG4-RD cohort, showing enrichment in idiopathic IgG4-RD.

Conclusions: The presence of a shared deleterious variant and homozygous common variant in FGFBP2 in the proband and sons strongly implicates this cytotoxic T cell product in the pathophysiology of IgG4-RD. The high prevalence of a common FGFBP2 variant in sporadic IgG4-RD supports the likelihood of participation in disease.
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http://dx.doi.org/10.1002/mgg3.686DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6565556PMC
June 2019

Alterations in sialic-acid O-acetylation glycoforms during murine erythrocyte development.

Glycobiology 2019 03;29(3):222-228

Ragon Institute of MGH, MIT and Harvard, Cambridge, MA, USA.

We used Casd1-deficient mice to confirm that this enzyme is responsible for 9-O-acetylation of sialic acids in vivo. We observed a complete loss of 9-O-acetylation of sialic acid on the surface of myeloid, erythroid and CD4+ T cells in Casd1-deficient mice. Although 9-O-acetylation of sialic acids on multiple hematopoietic lineages was lost, there were no obvious defects in hematopoiesis. Interestingly, erythrocytes from Casd1-deficient mice also lost reactivity to TER-119, a rat monoclonal antibody that is widely used to mark the murine erythroid lineage. The sialic acid glyco-epitope recognized by TER-119 on erythrocytes was sensitive to the sialic acid O-acetyl esterase activity of the hemagglutinin-esterase from bovine coronavirus but not to the corresponding enzyme from the influenza C virus. During erythrocyte development, TER-119+ Ery-A and Ery-B cells could be stained by catalytically inactive bovine coronavirus hemagglutinin-esterase but not by the inactive influenza C hemagglutinin-esterase, while TER-119+ Ery-C cells and mature erythrocytes were recognized by both virolectins. Although the structure of the sialoglycoconjugate recognized by TER-119 was not chemically demonstrated, its selective binding to virolectins suggests that it may be comprised of a 7,9-di-O-acetyl form of sialic acid. As erythrocytes mature, the surfaces of Ery-C cells and mature erythrocytes also acquire an additional distinct CASD1-dependent 9-O-acetyl sialic acid moiety that can be recognized by virolectins from both influenza C and bovine coronavirus.
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http://dx.doi.org/10.1093/glycob/cwy110DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6381321PMC
March 2019

The expansion in lymphoid organs of IL-4 BATF T follicular helper cells is linked to IgG4 class switching in vivo.

Life Sci Alliance 2018 Jan 5;1(1). Epub 2018 Apr 5.

Ragon Institute of MGH, MIT, and Harvard, Massachusetts General Hospital, Harvard Medical School, Boston, MA, USA.

Distinct T follicular helper (T) subsets that influence specific class-switching events are assumed to exist, but the accumulation of isotype-specific T subsets in secondary lymphoid organs (SLOs) and tertiary lymphoid organs has not been hitherto demonstrated. IL-4-expressing T cells are surprisingly sparse in human SLOs. In contrast, in IgG4-related disease (IgG4-RD), a disorder characterized by polarized Ig class switching, most T cells in tertiary and SLOs make IL-4. Human IL-4 T cells do not express GATA-3 but express nuclear BATF, and the transcriptomes of IL-4-secreting T cells differ from both PD1 T cells that do not secrete IL-4 and IL-4-secreting non-T cells. Unlike IgG4-RD, IL-4 T cells are rarely found in tertiary lymphoid organs in Sjögren's syndrome, a disorder in which IgG4 is not elevated. The proportion of CD4IL-4BATF T cells and CD4IL-4CXCR5 T cells in IgG4-RD tissues correlates tightly with tissue IgG4 plasma cell numbers and plasma IgG4 levels in patients but not with the total plasma levels of other isotypes. These data describe a disease-related T subpopulation in human tertiary lymphoid organs and SLOs that is linked to IgG4 class switching.
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http://dx.doi.org/10.26508/lsa.201800050DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6034714PMC
January 2018

Lesional CD4+ IFN-γ+ cytotoxic T lymphocytes in IgG4-related dacryoadenitis and sialoadenitis.

Ann Rheum Dis 2017 Feb 29;76(2):377-385. Epub 2016 Jun 29.

Massachusetts General Hospital, Harvard Medical School, Boston, Massachusetts, USA.

Objectives: IgG4-related disease (IgG4-RD) is a chronic, systemic, inflammatory condition of unknown aetiology. We have recently described clonally expanded circulating CD4 cytotoxic T lymphocytes (CTLs) in IgG4-RD that infiltrate affected tissues where they secrete interleukin (IL)-1β and transforming growth factor -β1 (TGF-β1). In this study, we sought to examine the role of CD4 CTLs in the pathogenesis of IgG4-related dacryoadenitis and sialoadenitis (IgG4-DS) and to determine whether these cells secrete interferon-gamma (IFN-γ) at lesional sites.

Methods: Salivary glands of 25 patients with IgG4-DS, 22 patients with Sjögren's syndrome (SS), 12 patients with chronic sialoadenitis (CS) and 12 healthy controls were analysed in this study. Gene expression analysis was performed on submandibular glands (SMGs) from five patients with IgG4-DS, three with CS and three healthy controls. Infiltrating CD4 CTLs were examined by quantitative multicolour imaging in tissue samples from 20 patients with IgG4-DS, 22 patients with SS, 9 patients with CS and 9 healthy controls.

Results: In IgG4-DS tissues, nine genes associated with CD4 CTLs were overexpressed. The expression of granzyme A (GZMA) mRNA was significantly higher in samples from patients with IgG4-RD compared with corresponding tissues from SS and healthy controls. Quantitative imaging showed that infiltrating CD4 GZMA CTLs were more abundant in patients with IgG4-DS than in the other groups. The ratio of CD4GZMA CTLs in SMGs from patients with IgG4-DS correlated with serum IgG4 concentrations and the number of affected organs. A large fraction of CD4GZMA CTLs in SMGs from patients with IgG4-DS secreted IFN-γ.

Conclusions: The pathogenesis of IgG4-DS is associated with tissue infiltration by CD4GZMA CTLs that secrete IFN-γ.
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http://dx.doi.org/10.1136/annrheumdis-2016-209139DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5435236PMC
February 2017

Striking Immune Phenotypes in Gene-Targeted Mice Are Driven by a Copy-Number Variant Originating from a Commercially Available C57BL/6 Strain.

Cell Rep 2016 05 19;15(9):1901-9. Epub 2016 May 19.

Ragon Institute of MGH, MIT and Harvard, 400 Technology Square, Cambridge, MA 02139, USA. Electronic address:

We describe a homozygous copy-number variant that disrupts the function of Dock2 in a commercially available C57BL/6 mouse strain that is widely used for backcrossing. This Dock2 allele was presumed to have spontaneously arisen in a colony of Irf5 knockout mice. We discovered that this allele has actually been inadvertently backcrossed into multiple mutant mouse lines, including two engineered to be deficient in Siae and Cmah. This particular commercially obtained subline of C57BL/6 mice also exhibits several striking immune phenotypes that have been previously described in the context of Dock2 deficiency. Inadvertent backcrossing of a number of gene-targeted mice into this background has complicated the interpretation of several immunological studies. In light of these findings, published studies involving immune or hematopoietic phenotypes in which these C57BL/6 mice have been used as controls, as experimental animals, or for backcrossing will need to be reinterpreted.
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http://dx.doi.org/10.1016/j.celrep.2016.04.080DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4892502PMC
May 2016

Clonal expansion of CD4(+) cytotoxic T lymphocytes in patients with IgG4-related disease.

J Allergy Clin Immunol 2016 09 11;138(3):825-838. Epub 2016 Mar 11.

Massachusetts General Hospital, Harvard Medical School, Boston, Mass. Electronic address:

Background: IgG4-related disease (IgG4-RD) is a systemic condition of unknown cause characterized by highly fibrotic lesions with dense lymphoplasmacytic infiltrates. CD4(+) T cells constitute the major inflammatory cell population in IgG4-RD lesions.

Objective: We used an unbiased approach to characterize CD4(+) T-cell subsets in patients with IgG4-RD based on their clonal expansion and ability to infiltrate affected tissue sites.

Methods: We used flow cytometry to identify CD4(+) effector/memory T cells in a cohort of 101 patients with IgG4-RD. These expanded cells were characterized by means of gene expression analysis and flow cytometry. Next-generation sequencing of the T-cell receptor β chain gene was performed on CD4(+)SLAMF7(+) cytotoxic T lymphocytes (CTLs) and CD4(+)GATA3(+) TH2 cells in a subset of patients to identify their clonality. Tissue infiltration by specific T cells was examined by using quantitative multicolor imaging.

Results: CD4(+) effector/memory T cells with a cytolytic phenotype were expanded in patients with IgG4-RD. Next-generation sequencing revealed prominent clonal expansions of these CD4(+) CTLs but not CD4(+)GATA3(+) memory TH2 cells in patients with IgG4-RD. The dominant T cells infiltrating a range of inflamed IgG4-RD tissue sites were clonally expanded CD4(+) CTLs that expressed SLAMF7, granzyme A, IL-1β, and TGF-β1. Clinical remission induced by rituximab-mediated B-cell depletion was associated with a reduction in numbers of disease-associated CD4(+) CTLs.

Conclusions: IgG4-RD is prominently linked to clonally expanded IL-1β- and TGF-β1-secreting CD4(+) CTLs in both peripheral blood and inflammatory tissue lesions. These active, terminally differentiated, cytokine-secreting effector CD4(+) T cells are now linked to a human disease characterized by chronic inflammation and fibrosis.
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http://dx.doi.org/10.1016/j.jaci.2015.12.1330DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5014627PMC
September 2016

Predictors of disease relapse in IgG4-related disease following rituximab.

Rheumatology (Oxford) 2016 Jun 16;55(6):1000-8. Epub 2016 Feb 16.

Division of Rheumatology, Allergy & Immunology, Massachusetts General Hospital, Harvard Medical School, Boston, MA, USA

Objective: IgG4-related disease (IgG4-RD) is a relapsing-remitting condition responsible for fibroinflammatory lesions that can lead to organ damage and life-threatening complications at nearly any anatomical site. The duration of remission following treatment varies and predictors of relapse are unclear. The objectives of this study were to review our experience with rituximab as remission induction in IgG4-RD, to clarify the duration of efficacy and to identify predictors of flare following treatment.

Methods: In this retrospective cohort study, all patients were treated with two doses of rituximab (1 g) separated by 15 days. Clinical, radiographic and laboratory data pertaining to rituximab response and disease relapse were collected from the electronic medical record. Kaplan-Meier curves were constructed to estimate the time to disease relapse. Log-rank analyses were performed to compare times to relapse among subgroups. Potential relapse predictors were evaluated with Cox regression analysis.

Results: Fifty-seven of 60 patients (95%) had clinical responses to rituximab. Forty-one patients (68%) were treated without glucocorticoids. Twenty-one patients (37%) experienced relapses following treatment at a median time from the first infusion of 244 days. Baseline concentrations of serum IgG4, IgE and circulating eosinophils predicted subsequent relapses, with hazard ratios of 6.2 (95% CI: 1.2, 32.0), 8.2 (95% CI: 1.4, 50.0) and 7.9 (95% CI: 1.8, 34.7), respectively. The higher the baseline values, the greater the risk of relapse and the shorter the time to relapse. Only 10% of the patients had elevations of all three major risk factors, underscoring the importance of measuring all three at baseline.

Conclusion: Baseline elevations in serum IgG4, IgE and blood eosinophil concentrations all predict IgG4-RD relapses independently.
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http://dx.doi.org/10.1093/rheumatology/kev438DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4900135PMC
June 2016

Sialic acids and autoimmune disease.

Immunol Rev 2016 Jan;269(1):145-61

Ragon Institute of MGH, MIT and Harvard, Cambridge, MA, USA.

An important underlying mechanism that contributes to autoimmunity is the loss of inhibitory signaling in the immune system. Sialic acid-recognizing Ig superfamily lectins or Siglecs are a family of cell surface proteins largely expressed in hematopoietic cells. The majority of Siglecs are inhibitory receptors expressed in immune cells that bind to sialic acid-containing ligands and recruit SH2-domain-containing tyrosine phosphatases to their cytoplasmic tails. They deliver inhibitory signals that can contribute to the constraining of immune cells, and thus protect the host from autoimmunity. The inhibitory functions of CD22/Siglec-2 and Siglec-G and their contributions to tolerance and autoimmunity, primarily in the B lymphocyte context, are considered in some detail in this review. The relevance to autoimmunity and unregulated inflammation of modified sialic acids, enzymes that modify sialic acid, and other sialic acid-binding proteins are also reviewed.
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http://dx.doi.org/10.1111/imr.12344DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4769436PMC
January 2016

IgG4-Related Disease: Clinical and Laboratory Features in One Hundred Twenty-Five Patients.

Arthritis Rheumatol 2015 Sep;67(9):2466-75

Massachusetts General Hospital and Harvard Medical School, Boston.

Objective: IgG4-related disease (IgG4-RD) is an immune-mediated fibroinflammatory condition that can affect nearly any organ. Prior studies have focused on individual cases of IgG4-RD or small case series. This study was undertaken to report detailed clinical and laboratory findings in a larger group of patients with IgG4-RD whose diagnosis was established by strict clinicopathologic correlation.

Methods: The baseline features of 125 patients with biopsy-proven IgG4-RD were reviewed. The diagnosis was confirmed by pathologists' review, based on consensus diagnostic criteria and correlation with clinicopathologic features. Disease activity and damage were assessed using the IgG4-RD Responder Index (RI). Flow cytometry was used to assess levels of circulating plasmablasts.

Results: Of the 125 patients, 107 had active disease and 86 were not receiving treatment for IgG4-RD. Only 51% of the patients with active disease had elevated serum IgG4 concentrations. However, patients with active disease and elevated serum IgG4 concentrations were older, had a higher IgG4-RD RI score, a greater number of organs involved, lower complement levels, higher absolute eosinophil counts, and higher IgE levels compared to those with active disease but normal serum IgG4 concentrations (P < 0.01 for all comparisons). The correlation between IgG4+ plasmablast levels and the IgG4-RD RI of disease activity (Spearman's ρ = 0.45, P = 0.003) was stronger than the correlation between total plasmablast levels and the IgG4-RD RI. Seventy-six (61%) of the patients were male, but no significant differences according to sex were observed with regard to disease severity, organ involvement, or serum IgG4 concentrations. Treatment with glucocorticoids failed to produce sustained remission in 77% of patients.

Conclusion: Nearly 50% of this patient cohort with biopsy-proven, clinically active IgG4-RD had normal serum IgG4 concentrations. Elevations in the serum IgG4 concentration appeared to identify a subset of patients with a more severe disease phenotype. In addition, the levels of IgG4+ plasmablasts correlated well with the extent of disease activity.
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http://dx.doi.org/10.1002/art.39205DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4621270PMC
September 2015

Plasmablasts as a biomarker for IgG4-related disease, independent of serum IgG4 concentrations.

Ann Rheum Dis 2015 Jan 9;74(1):190-5. Epub 2014 May 9.

Division of Rheumatology, Allergy, and Immunology, Massachusetts General Hospital, Boston, Massachusetts, USA.

Objectives: We examined the utility of circulating total and IgG4+ plasmablasts as biomarkers of diagnosis and disease activity in IgG4-related disease (IgG4-RD).

Materials Methods: We evaluated patients with active, untreated, biopsy-proven IgG4-RD affecting various organs. Flow cytometry was used to measure total plasmablast and IgG4+ plasmablast counts by gating peripheral blood for CD19lowCD38+CD20-CD27+ cells and CD19lowCD38+CD20-CD27+IgG4+ cells. Serum IgG4 concentrations were measured by nephelometry. We compared 37 IgG4-RD patients to 35 controls, including healthy individuals (n=14) and patients with other inflammatory diseases before treatment (n=21).

Results: The IgG4-RD patients' mean age was 59, and 68% were male. Fourteen patients (38%) had three or more organs involved. The IgG4-RD patients had substantially elevated total plasmablast counts (median 4698/mL, range 610-79524/mL) compared to both untreated disease controls (median 592/mL, range 19-4294/mL; p < 0.001) and healthy controls (median 94/mL, range 1-653/mL; p < 0.001). Thirteen IgG4-RD patients (36%) had normal serum IgG4 concentrations (mean 60 mg/dL, range 5-123 mg/dL, normal <135 mg/dL). However, the median plasmablast count was not significantly lower in that subset with normal serum IgG4 concentrations (3784/mL) compared to those with elevated serum IgG4 (5155/mL) (p = 0.242). Among the 12 rituximab (RTX)-treated patients, the median plasmablast level during disease flare was 6356/mL (range 1123-41589/mL), declining to 1419/mL (range 386/mL-4150/mL) during remission (p < 0.01).

Conclusions: Circulating plasmablasts are elevated in active IgG4-RD, even in patients with normal serum IgG4 concentrations. Plasmablast counts are a potentially useful biomarker for diagnosis, assessing response to treatment, and determining the appropriate time for re-treatment.
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http://dx.doi.org/10.1136/annrheumdis-2014-205233DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4656194PMC
January 2015

De novo oligoclonal expansions of circulating plasmablasts in active and relapsing IgG4-related disease.

J Allergy Clin Immunol 2014 Sep 6;134(3):679-87. Epub 2014 May 6.

Massachusetts General Hospital and Harvard Medical School, Boston, Mass. Electronic address:

Background: IgG4-related disease (IgG4-RD) is a poorly understood, multiorgan, chronic inflammatory disease characterized by tumefactive lesions, storiform fibrosis, obliterative phlebitis, and accumulation of IgG4-expressing plasma cells at disease sites.

Objective: The role of B cells and IgG4 antibodies in IgG4-RD pathogenesis is not well defined. We evaluated patients with IgG4-RD for activated B cells in both disease lesions and peripheral blood and investigated their role in disease pathogenesis.

Methods: B-cell populations from the peripheral blood of 84 patients with active IgG4-RD were analyzed by using flow cytometry. The repertoire of B-cell populations was analyzed in a subset of patients by using next-generation sequencing. Fourteen of these patients were longitudinally followed for 9 to 15 months after rituximab therapy.

Results: Numbers of CD19(+)CD27(+)CD20(-)CD38(hi) plasmablasts, which are largely IgG4(+), are increased in patients with active IgG4-RD. These expanded plasmablasts are oligoclonal and exhibit extensive somatic hypermutation, and their numbers decrease after rituximab-mediated B-cell depletion therapy; this loss correlates with disease remission. A subset of patients relapse after rituximab therapy, and circulating plasmablasts that re-emerge in these subjects are clonally distinct and exhibit enhanced somatic hypermutation. Cloning and expression of immunoglobulin heavy and light chain genes from expanded plasmablasts at the peak of disease reveals that disease-associated IgG4 antibodies are self-reactive.

Conclusions: Clonally expanded CD19(+)CD27(+)CD20(-)CD38(hi) plasmablasts are a hallmark of active IgG4-RD. Enhanced somatic mutation in activated B cells and plasmablasts and emergence of distinct plasmablast clones on relapse indicate that the disease pathogenesis is linked to de novo recruitment of naive B cells into T cell-dependent responses by CD4(+) T cells, likely driving a self-reactive disease process.
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http://dx.doi.org/10.1016/j.jaci.2014.03.034DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4149918PMC
September 2014

IgG4-related midline destructive lesion.

Ann Rheum Dis 2014 Jul 20;73(7):1434-6. Epub 2014 Mar 20.

Division of Rheumatology, Allergy, & Immunology, Massachusetts General Hospital, Boston, Massachusetts, USA.

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http://dx.doi.org/10.1136/annrheumdis-2014-205187DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4625399PMC
July 2014

Prevalence of atopy, eosinophilia, and IgE elevation in IgG4-related disease.

Allergy 2014 02 25;69(2):269-272. Epub 2013 Nov 25.

Rheumatology, Allergy and Immunology, Massachusetts General Hospital, Boston MA.

IgG4-related disease (IgG4-RD) is a fibroinflammatory disorder that can affect virtually every organ system. T-helper type 2 responses have been presumed to be pathogenic in this disease, and a high proportion of patients with IgG4-RD are reported to have longstanding allergies, peripheral blood eosinophilia, and serum IgE elevation. It has therefore been proposed that allergic mechanisms drive IgG4-RD. However, no epidemiological assessment of atopy, peripheral blood eosinophilia, and serum IgE concentrations has ever been undertaken in patients with IgG4-RD. In this study, we evaluated these parameters in a large cohort of patients with IgG4-RD in whom a wide range of organs were affected by disease. Our results demonstrate that the majority of patients with IgG4-RD are nonatopic. Nevertheless, a subset of nonatopic subjects exhibit peripheral blood eosinophilia and elevated IgE, suggesting that processes inherent to IgG4-RD itself rather than atopy per se contribute to the eosinophilia and IgE elevation observed in the absence of atopy.
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http://dx.doi.org/10.1111/all.12320DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4638385PMC
February 2014

IgG4-related disease.

Annu Rev Pathol 2014 2;9:315-47. Epub 2013 Oct 2.

Cancer Center.

Immunoglobulin G4 (IgG4)-related disease (IgG4-RD) is an immune-mediated condition that can affect almost any organ and is now being recognized with increasing frequency. IgG4-RD is characterized by a lymphoplasmacytic infiltrate composed of IgG4(+) plasma cells, storiform fibrosis, obliterative phlebitis, and mild to moderate eosinophilia. The diagnosis of IgG4-RD unifies many eponymous fibroinflammatory conditions that had previously been thought to be confined to single organs. IgG4-RD lesions are infiltrated by T helper cells, which likely cause progressive fibrosis and organ damage. IgG4 antibodies are generally regarded as noninflammatory. Although autoreactive IgG4 antibodies are observed in IgG4-RD, there is no evidence that they are directly pathogenic. Rituximab-induced B cell depletion in IgG4-RD leads to rapid clinical and histological improvement accompanied by swift declines in serum IgG4 concentrations. Although IgG autoantibodies against various exocrine gland antigens have been described in IgG4-RD, whether they are members of the IgG4 subclass is unknown. The contribution of autoantibodies to IgG4-RD remains unclear.
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http://dx.doi.org/10.1146/annurev-pathol-012513-104708DOI Listing
March 2014

How to interpret elevated cardiac troponin levels.

Circulation 2011 Nov;124(21):2350-4

Brigham and Women's Hospital, 75 Francis St, Boston, MA 02115, USA.

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http://dx.doi.org/10.1161/CIRCULATIONAHA.111.023697DOI Listing
November 2011
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