title: Characterization of ζ-associated protein, 70 kd (ZAP70)-deficient human lymphocytes.
abstract: BACKGROUND
  ζ-associated protein, 70 kd (ZAP70), deficiency in human subjects results in a combined immunodeficiency characterized by normal numbers of circulating CD4 T cells and CD8 lymphocytopenia. Patients who live beyond infancy can also experience autoimmune manifestations.
  
  
  OBJECTIVE
  We sought to further characterize the nature of the T-cell populations found in ZAP70-deficient patients and explored the mechanisms that might predispose them to autoimmunity.
  
  
  METHODS
  T-cell development was assessed by examining T-cell receptor (TCR) gene rearrangements and thymopoiesis by measuring TCR exclusion circle levels. TCR repertoire on CD4 and CD8 T-cell populations was assessed by means of flow cytometry. T-cell gene expression patterns were examined by means of exonic microarray analysis and apoptotic responses by means of Annexin V binding and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling.
  
  
  RESULTS
  Cells displaying recombination events from all stages of TCR gene rearrangement were present in the peripheral blood of ZAP70-deficient patients; however, the late TCRD-deleting rearrangement was significantly reduced. TCR exclusion circle levels were also found to be low. Surprisingly, all Vβ families were detected in both CD4(+) and CD8(+) circulating T cells. Several Vβ families were significantly overrepresented, which is reminiscent of autoimmune disorders. Levels of mRNA for cytotoxic T lymphocyte-associated antigen 4, TGF-β, and IL-10 were found to be low, a signature of autoimmunity. Finally, Fas-mediated CD4 T-cell apoptosis was found to be reduced in vitro, and staining of thymus biopsy specimens revealed reduced thymocyte apoptosis.
  
  
  CONCLUSIONS
  We show that in the absence of ZAP70, thymopoiesis is altered and differentiation to double-positive cells is hampered. Circulating T cells appear poorly regulated, do not differentiate into T(H)2 T cells, lack a number of inhibitory growth controls, and display reduced apoptosis, all predisposing patients to exaggerated inflammation and autoimmunity.
authors.affiliation: Division of Immunology and Allergy, Canadian Centre for Primary Immunodeficiency, Jeffrey Modell Research Laboratory for the diagnosis of Primary Immunodeficiency, Hospital for Sick Children, Toronto, Ontario, Canada. chaim.roifman@sickkids.ca
authors.forename: Chaim M
authors.initials: CM
authors.lastname: Roifman
authors.forename: Harjit
authors.initials: H
authors.lastname: Dadi
authors.forename: Raz
authors.initials: R
authors.lastname: Somech
authors.forename: Amit
authors.initials: A
authors.lastname: Nahum
authors.forename: Nigel
authors.initials: N
authors.lastname: Sharfe
mesh_terms: D017209:Apoptosis / Q000235:genetics; D015551:Autoimmunity; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology / Q000378:metabolism* / Q000473:pathology; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology / Q000378:metabolism* / Q000473:pathology; D002454:Cell Differentiation / Q000235:genetics / Q000276:immunology; D048708:Cell Growth Processes / Q000235:genetics / Q000276:immunology; D019672:Genes, T-Cell Receptor / Q000276:immunology; D006801:Humans; D007249:Inflammation; D008231:Lymphopenia; D046228:Microarray Analysis; D016511:Severe Combined Immunodeficiency / Q000276:immunology / Q000473:pathology / Q000503:physiopathology; D013950:Thymus Gland / Q000276:immunology / Q000378:metabolism* / Q000473:pathology; D051746:ZAP-70 Protein-Tyrosine Kinase / Q000172:deficiency / Q000235:genetics / Q000276:immunology / Q000378:metabolism*
chemical_list: D051746:ZAP-70 Protein-Tyrosine Kinase; C497261:ZAP70 protein, human