Classification of common variable immunodeficiencies using flow cytometry and a memory B-cell functionality assay
The population of patients with common variable immunodeficiency (CVID) comprises a heterogeneous group of patients with different causes of hypogammaglobulinemia predisposing to recurrent infections, higher incidence of autoimmunity, and malignancy. Although memory B cells (memBcs) are key players...
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| Published in | Journal of allergy and clinical immunology Vol. 135; no. 1; pp. 198 - 208.e5 |
|---|---|
| Main Authors | , , , , , , , , , , |
| Format | Journal Article |
| Language | English |
| Published |
United States
Elsevier Inc
01.01.2015
Elsevier Limited |
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| Online Access | Get full text |
| ISSN | 0091-6749 1097-6825 1097-6825 |
| DOI | 10.1016/j.jaci.2014.06.022 |
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| Abstract | The population of patients with common variable immunodeficiency (CVID) comprises a heterogeneous group of patients with different causes of hypogammaglobulinemia predisposing to recurrent infections, higher incidence of autoimmunity, and malignancy. Although memory B cells (memBcs) are key players in humoral defense and their numbers are commonly reduced in these patients, their functionality is not part of any current classification.
We established and validated a memBc enzyme-linked immunosorbent spot (ELISpot) assay that reveals the capacity of memBcs to develop into antibody-secreting cells and present an idea for a new classification based on this functional capacity.
The memBc ELISpot assay, combined with flow cytometry, was applied to patients with confirmed CVID in comparison with age-matched healthy control subjects.
Ex vivo frequency of IgG-, IgM-, and IgA-secreting plasmablasts was significantly diminished by 27.2-, 2.4-, and 23.3-fold, respectively, compared with that seen in healthy control subjects. Moreover, in vitro differentiation of memBcs into antibody-secreting cells was 6.1-, 2.6-, and 3.7-fold significantly reduced for IgG-, IgM-, and IgA-secreting cells, respectively. Proliferation of memBcs correlates inversely to immunoglobulin-secreting capacity, suggesting compensatory hyperproliferation. Furthermore, patients with no serum IgA can still have a detectable IgA ELISpot assay result in vitro. Most importantly, the large heterogeneity of memBc function in patients with CVID homogenously grouped by means of fluorescence-activated cell sorting allowed additional subclassification based on memBc/plasmablast function.
These data suggest almost normal memBc/immunoglobulin-secreting plasmablast functionality in some patients if sufficient stimulatory signals are delivered, which might open up opportunities for new therapeutic approaches. |
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| AbstractList | Background The population of patients with common variable immunodeficiency (CVID) comprises a heterogeneous group of patients with different causes of hypogammaglobulinemia predisposing to recurrent infections, higher incidence of autoimmunity, and malignancy. Although memory B cells (memBcs) are key players in humoral defense and their numbers are commonly reduced in these patients, their functionality is not part of any current classification. Objective We established and validated a memBc enzyme-linked immunosorbent spot (ELISpot) assay that reveals the capacity of memBcs to develop into antibody-secreting cells and present an idea for a new classification based on this functional capacity. Methods The memBc ELISpot assay, combined with flow cytometry, was applied to patients with confirmed CVID in comparison with age-matched healthy control subjects. Results Ex vivo frequency of IgG-, IgM-, and IgA-secreting plasmablasts was significantly diminished by 27.2-, 2.4-, and 23.3-fold, respectively, compared with that seen in healthy control subjects. Moreover, in vitro differentiation of memBcs into antibody-secreting cells was 6.1-, 2.6-, and 3.7-fold significantly reduced for IgG-, IgM-, and IgA-secreting cells, respectively. Proliferation of memBcs correlates inversely to immunoglobulin-secreting capacity, suggesting compensatory hyperproliferation. Furthermore, patients with no serum IgA can still have a detectable IgA ELISpot assay result in vitro . Most importantly, the large heterogeneity of memBc function in patients with CVID homogenously grouped by means of fluorescence-activated cell sorting allowed additional subclassification based on memBc/plasmablast function. Conclusion These data suggest almost normal memBc/immunoglobulin-secreting plasmablast functionality in some patients if sufficient stimulatory signals are delivered, which might open up opportunities for new therapeutic approaches. The population of patients with common variable immunodeficiency (CVID) comprises a heterogeneous group of patients with different causes of hypogammaglobulinemia predisposing to recurrent infections, higher incidence of autoimmunity, and malignancy. Although memory B cells (memBcs) are key players in humoral defense and their numbers are commonly reduced in these patients, their functionality is not part of any current classification. We established and validated a memBc enzyme-linked immunosorbent spot (ELISpot) assay that reveals the capacity of memBcs to develop into antibody-secreting cells and present an idea for a new classification based on this functional capacity. The memBc ELISpot assay, combined with flow cytometry, was applied to patients with confirmed CVID in comparison with age-matched healthy control subjects. Ex vivo frequency of IgG-, IgM-, and IgA-secreting plasmablasts was significantly diminished by 27.2-, 2.4-, and 23.3-fold, respectively, compared with that seen in healthy control subjects. Moreover, in vitro differentiation of memBcs into antibody-secreting cells was 6.1-, 2.6-, and 3.7-fold significantly reduced for IgG-, IgM-, and IgA-secreting cells, respectively. Proliferation of memBcs correlates inversely to immunoglobulin-secreting capacity, suggesting compensatory hyperproliferation. Furthermore, patients with no serum IgA can still have a detectable IgA ELISpot assay result in vitro. Most importantly, the large heterogeneity of memBc function in patients with CVID homogenously grouped by means of fluorescence-activated cell sorting allowed additional subclassification based on memBc/plasmablast function. These data suggest almost normal memBc/immunoglobulin-secreting plasmablast functionality in some patients if sufficient stimulatory signals are delivered, which might open up opportunities for new therapeutic approaches. The population of patients with common variable immunodeficiency (CVID) comprises a heterogeneous group of patients with different causes of hypogammaglobulinemia predisposing to recurrent infections, higher incidence of autoimmunity, and malignancy. Although memory B cells (memBcs) are key players in humoral defense and their numbers are commonly reduced in these patients, their functionality is not part of any current classification.BACKGROUNDThe population of patients with common variable immunodeficiency (CVID) comprises a heterogeneous group of patients with different causes of hypogammaglobulinemia predisposing to recurrent infections, higher incidence of autoimmunity, and malignancy. Although memory B cells (memBcs) are key players in humoral defense and their numbers are commonly reduced in these patients, their functionality is not part of any current classification.We established and validated a memBc enzyme-linked immunosorbent spot (ELISpot) assay that reveals the capacity of memBcs to develop into antibody-secreting cells and present an idea for a new classification based on this functional capacity.OBJECTIVEWe established and validated a memBc enzyme-linked immunosorbent spot (ELISpot) assay that reveals the capacity of memBcs to develop into antibody-secreting cells and present an idea for a new classification based on this functional capacity.The memBc ELISpot assay, combined with flow cytometry, was applied to patients with confirmed CVID in comparison with age-matched healthy control subjects.METHODSThe memBc ELISpot assay, combined with flow cytometry, was applied to patients with confirmed CVID in comparison with age-matched healthy control subjects.Ex vivo frequency of IgG-, IgM-, and IgA-secreting plasmablasts was significantly diminished by 27.2-, 2.4-, and 23.3-fold, respectively, compared with that seen in healthy control subjects. Moreover, in vitro differentiation of memBcs into antibody-secreting cells was 6.1-, 2.6-, and 3.7-fold significantly reduced for IgG-, IgM-, and IgA-secreting cells, respectively. Proliferation of memBcs correlates inversely to immunoglobulin-secreting capacity, suggesting compensatory hyperproliferation. Furthermore, patients with no serum IgA can still have a detectable IgA ELISpot assay result in vitro. Most importantly, the large heterogeneity of memBc function in patients with CVID homogenously grouped by means of fluorescence-activated cell sorting allowed additional subclassification based on memBc/plasmablast function.RESULTSEx vivo frequency of IgG-, IgM-, and IgA-secreting plasmablasts was significantly diminished by 27.2-, 2.4-, and 23.3-fold, respectively, compared with that seen in healthy control subjects. Moreover, in vitro differentiation of memBcs into antibody-secreting cells was 6.1-, 2.6-, and 3.7-fold significantly reduced for IgG-, IgM-, and IgA-secreting cells, respectively. Proliferation of memBcs correlates inversely to immunoglobulin-secreting capacity, suggesting compensatory hyperproliferation. Furthermore, patients with no serum IgA can still have a detectable IgA ELISpot assay result in vitro. Most importantly, the large heterogeneity of memBc function in patients with CVID homogenously grouped by means of fluorescence-activated cell sorting allowed additional subclassification based on memBc/plasmablast function.These data suggest almost normal memBc/immunoglobulin-secreting plasmablast functionality in some patients if sufficient stimulatory signals are delivered, which might open up opportunities for new therapeutic approaches.CONCLUSIONThese data suggest almost normal memBc/immunoglobulin-secreting plasmablast functionality in some patients if sufficient stimulatory signals are delivered, which might open up opportunities for new therapeutic approaches. The population of patients with common variable immunodeficiency (CVID) comprises a heterogeneous group of patients with different causes of hypogammaglobulinemia predisposing to recurrent infections, higher incidence of autoimmunity, and malignancy. Although memory B cells (memBcs) are key players in humoral defense and their numbers are commonly reduced in these patients, their functionality is not part of any current classification. We established and validated a memBc enzyme-linked immunosorbent spot (ELISpot) assay that reveals the capacity of memBcs to develop into antibody-secreting cells and present an idea for a new classification based on this functional capacity. The memBc ELISpot assay, combined with flow cytometry, was applied to patients with confirmed CVID in comparison with age-matched healthy control subjects. Ex vivo frequency of IgG-, IgM-, and IgA-secreting plasmablasts was significantly diminished by 27.2-, 2.4-, and 23.3-fold, respectively, compared with that seen in healthy control subjects. Moreover, in vitro differentiation of memBcs into antibody-secreting cells was 6.1-, 2.6-, and 3.7-fold significantly reduced for IgG-, IgM-, and IgA-secreting cells, respectively. Proliferation of memBcs correlates inversely to immunoglobulin-secreting capacity, suggesting compensatory hyperproliferation. Furthermore, patients with no serum IgA can still have a detectable IgA ELISpot assay result in vitro. Most importantly, the large heterogeneity of memBc function in patients with CVID homogenously grouped by means of fluorescence-activated cell sorting allowed additional subclassification based on memBc/plasmablast function. These data suggest almost normal memBc/immunoglobulin-secreting plasmablast functionality in some patients if sufficient stimulatory signals are delivered, which might open up opportunities for new therapeutic approaches. Background The population of patients with common variable immunodeficiency (CVID) comprises a heterogeneous group of patients with different causes of hypogammaglobulinemia predisposing to recurrent infections, higher incidence of autoimmunity, and malignancy. Although memory B cells (memBcs) are key players in humoral defense and their numbers are commonly reduced in these patients, their functionality is not part of any current classification. Objective We established and validated a memBc enzyme-linked immunosorbent spot (ELISpot) assay that reveals the capacity of memBcs to develop into antibody-secreting cells and present an idea for a new classification based on this functional capacity. Methods The memBc ELISpot assay, combined with flow cytometry, was applied to patients with confirmed CVID in comparison with age-matched healthy control subjects. Results Ex vivofrequency of IgG-, IgM-, and IgA-secreting plasmablasts was significantly diminished by 27.2-, 2.4-, and 23.3-fold, respectively, compared with that seen in healthy control subjects. Moreover,in vitrodifferentiation of memBcs into antibody-secreting cells was 6.1-, 2.6-, and 3.7-fold significantly reduced for IgG-, IgM-, and IgA-secreting cells, respectively. Proliferation of memBcs correlates inversely to immunoglobulin-secreting capacity, suggesting compensatory hyperproliferation. Furthermore, patients with no serum IgA can still have a detectable IgA ELISpot assay resultin vitro. Most importantly, the large heterogeneity of memBc function in patients with CVID homogenously grouped by means of fluorescence-activated cell sorting allowed additional subclassification based on memBc/plasmablast function. Conclusion These data suggest almost normal memBc/immunoglobulin-secreting plasmablast functionality in some patients if sufficient stimulatory signals are delivered, which might open up opportunities for new therapeutic approaches. Background The population of patients with common variable immunodeficiency (CVID) comprises a heterogeneous group of patients with different causes of hypogammaglobulinemia predisposing to recurrent infections, higher incidence of autoimmunity, and malignancy. Although memory B cells (memBcs) are key players in humoral defense and their numbers are commonly reduced in these patients, their functionality is not part of any current classification. Objective We established and validated a memBc enzyme-linked immunosorbent spot (ELISpot) assay that reveals the capacity of memBcs to develop into antibody-secreting cells and present an idea for a new classification based on this functional capacity. Methods The memBc ELISpot assay, combined with flow cytometry, was applied to patients with confirmed CVID in comparison with age-matched healthy control subjects. Results Ex vivo frequency of IgG-, IgM-, and IgA-secreting plasmablasts was significantly diminished by 27.2-, 2.4-, and 23.3-fold, respectively, compared with that seen in healthy control subjects. Moreover, in vitro differentiation of memBcs into antibody-secreting cells was 6.1-, 2.6-, and 3.7-fold significantly reduced for IgG-, IgM-, and IgA-secreting cells, respectively. Proliferation of memBcs correlates inversely to immunoglobulin-secreting capacity, suggesting compensatory hyperproliferation. Furthermore, patients with no serum IgA can still have a detectable IgA ELISpot assay result in vitro. Most importantly, the large heterogeneity of memBc function in patients with CVID homogenously grouped by means of fluorescence-activated cell sorting allowed additional subclassification based on memBc/plasmablast function. Conclusion These data suggest almost normal memBc/immunoglobulin-secreting plasmablast functionality in some patients if sufficient stimulatory signals are delivered, which might open up opportunities for new therapeutic approaches. |
| Author | Warnatz, Klaus Hanitsch, Leif Rösel, Amelia L. Hoffmeister, Bodo Thomas, Sybill Krüger, Renate von Bernuth, Horst Schliesser, Ulrike Scheibenbogen, Carmen Sollwedel, André Volk, Hans-Dieter |
| Author_xml | – sequence: 1 givenname: Amelia L. surname: Rösel fullname: Rösel, Amelia L. organization: Institute for Medical Immunology, Charité University Medicine Berlin, Berlin, Germany – sequence: 2 givenname: Carmen surname: Scheibenbogen fullname: Scheibenbogen, Carmen organization: Institute for Medical Immunology, Charité University Medicine Berlin, Berlin, Germany – sequence: 3 givenname: Ulrike surname: Schliesser fullname: Schliesser, Ulrike organization: Berlin-Brandenburg Centre for Regenerative Therapies, Berlin, Germany – sequence: 4 givenname: André surname: Sollwedel fullname: Sollwedel, André organization: Institute for Medical Immunology, Charité University Medicine Berlin, Berlin, Germany – sequence: 5 givenname: Bodo surname: Hoffmeister fullname: Hoffmeister, Bodo organization: Out-patients Clinic for Immunodeficiencies, Charité University Medicine Berlin, Berlin, Germany – sequence: 6 givenname: Leif surname: Hanitsch fullname: Hanitsch, Leif organization: Out-patients Clinic for Immunodeficiencies, Charité University Medicine Berlin, Berlin, Germany – sequence: 7 givenname: Horst surname: von Bernuth fullname: von Bernuth, Horst organization: Clinic for Pediatrics, Department of Pneumology and Immunology, Charité University Medicine Berlin, Berlin, Germany – sequence: 8 givenname: Renate surname: Krüger fullname: Krüger, Renate organization: Clinic for Pediatrics, Department of Pneumology and Immunology, Charité University Medicine Berlin, Berlin, Germany – sequence: 9 givenname: Klaus surname: Warnatz fullname: Warnatz, Klaus organization: Centre for Chronic Immunodeficiency, University Clinic and University of Freiburg, Freiburg, Germany – sequence: 10 givenname: Hans-Dieter surname: Volk fullname: Volk, Hans-Dieter email: hans-dieter.volk@charite.de organization: Institute for Medical Immunology, Charité University Medicine Berlin, Berlin, Germany – sequence: 11 givenname: Sybill surname: Thomas fullname: Thomas, Sybill organization: Institute for Medical Immunology, Charité University Medicine Berlin, Berlin, Germany |
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| Keywords | enzyme-linked immunosorbent spot assay Ig-sPb Memory B cell common variable immunodeficiency subtyping FACS ELISpot ASC CVID PB PE memBc flow cytometry HC antibody-secreting cells Healthy control Immunoglobulin-secreting plasmablast Common variable immunodeficiency Enzyme-linked immunosorbent spot Pacific Blue Fluorescence-activated cell sorting Antibody-secreting cell Phycoerythrin |
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| SubjectTerms | Adult Age Allergy and Immunology antibody-secreting cells B-Lymphocytes - cytology B-Lymphocytes - immunology Cell Differentiation Cells, Cultured Classification Common Variable Immunodeficiency - blood Common Variable Immunodeficiency - classification Common Variable Immunodeficiency - immunology common variable immunodeficiency subtyping enzyme-linked immunosorbent spot assay Enzyme-Linked Immunospot Assay Female Flow Cytometry Humans Immunoglobulin A - blood Immunoglobulin G - blood Immunoglobulin M - blood Immunologic Memory Leukocytes, Mononuclear - cytology Male Medical treatment Memory B cell Middle Aged Patients |
| Title | Classification of common variable immunodeficiencies using flow cytometry and a memory B-cell functionality assay |
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