Journal of Clinical Immunology

, Volume 31, Issue 4, pp 540–549 | Cite as

Thymic and Bone Marrow Output in Patients with Common Variable Immunodeficiency

  • Federico Serana
  • Paolo Airò
  • Marco Chiarini
  • Cinzia Zanotti
  • Mirko Scarsi
  • Micol Frassi
  • Vassilios Lougaris
  • Alessandro Plebani
  • Luigi Caimi
  • Luisa Imberti



The study aims to obtain more information about the immune deficit of common variable immunodeficiency (CVID) patients.

Materials and Methods

A new real-time PCR assay was used to quantify T and B lymphocyte mobilization from the production and maturation sites through the detection of T cell receptor excision circles (TRECs) and kappa-deleting recombination circles (KRECs) and to allow the estimation of the average number of B cell divisions. T and B lymphocyte subsets were analyzed by flow cytometry.


The number of TREC+ lymphocytes, which depends on age and gender, was significantly reduced in CVID patients. Similarly, KREC concentration was lower than in controls. Classification of patients according to the percentage of memory switched B cells showed that patients belonging to MB2 group and therefore with conserved B cell maturation have the lowest new B cell output but increased average peripheral divisions, leading to the highest B cell number.


TREC and KREC quantification can be helpful for a more complete and informative understanding of a heterogeneous disease such as CVID.


Common variable immunodeficiency kappa-deleting recombination circles T cell receptor excision circles 

Supplementary material

10875_2011_9526_MOESM1_ESM.ppt (70 kb)
Supplementary Figure 1Correlation between the average number of divisions and KRECs per milliliter in CVID patients. The number of KRECs per milliliter of each patient is plotted against the respective average number of B cell divisions. The line was obtained by simple linear regression calculated after log transformation of KRECs per milliliter values. (PPT 70 kb)
10875_2011_9526_MOESM2_ESM.xls (30 kb)
Supplementary Table 1Percentage and number of T and B lymphocyte subpopulations (XLS 30 kb)


  1. 1.
    Cunningham-Rundles C, Bodian C. Common variable immunodeficiency: clinical and immunological features of 248 patients. Clin Immunol. 1999;92:34–48.PubMedCrossRefGoogle Scholar
  2. 2.
    Conley ME, Notarangelo LD, Etzioni A. Diagnostic criteria for primary immunodeficiencies. Representing PAGID (Pan-American Group for Immunodeficiency) and ESID (European Society for Immunodeficiencies). Clin Immunol. 1999;93:190–7.PubMedCrossRefGoogle Scholar
  3. 3.
    Grimbacher B, Hutloff A, Schlesier M, Glocker E, Warnatz K, Drager R, et al. Homozygous loss of ICOS is associated with adult-onset common variable immunodeficiency. Nat Immunol. 2003;4:261–8.PubMedCrossRefGoogle Scholar
  4. 4.
    Castigli E, Wilson SA, Garibyan L, Rachid R, Bonilla F, Schneider L, et al. TACI is mutant in common variable immunodeficiency and IgA deficiency. Nat Genet. 2005;37:829–34.PubMedCrossRefGoogle Scholar
  5. 5.
    Losi CG, Silini A, Fiorini C, Soresina A, Meini A, Ferrari S, et al. Mutational analysis of human BAFF receptor TNFRSF13C (BAFF-R) in patients with common variable immunodeficiency. J Clin Immunol. 2005;25:496–502.PubMedCrossRefGoogle Scholar
  6. 6.
    Salzer U, Chapel HM, Webster AD, Pan-Hammarström Q, Schmitt-Graeff A, Schlesier M, et al. Mutations in TNFRSF13B encoding TACI are associated with common variable immunodeficiency in humans. Nat Genet. 2005;37:820–8.PubMedCrossRefGoogle Scholar
  7. 7.
    van Zelm MC, Reisli I, van der Burg M, Castano D, van Noesel CJ, van Tol MJ, et al. An antibody-deficiency syndrome due to mutations in the CD19 gene. N Engl J Med. 2006;354:1901–12.PubMedCrossRefGoogle Scholar
  8. 8.
    Vorechovsky I, Zetterquist H, Paganelli R, Koskinen S, Webster AD, Bjorkander J, et al. Family and linkage study of selective IgA deficiency and common variable immunodeficiency. Clin Immunol Immunopathol. 1995;77:185–92.PubMedCrossRefGoogle Scholar
  9. 9.
    Agematsu K, Nagumo H, Yang FC, Nakazawa T, Fukushima K, Ito S, et al. B cell subpopulations separated by CD27 and crucial collaboration of CD27+ B cells and helper T cells in immunoglobulin production. Eur J Immunol. 1997;27:2073–9.PubMedCrossRefGoogle Scholar
  10. 10.
    Brouet JC, Chedeville A, Fermand JP, Royer B. Study of the B cell memory compartment in common variable immunodeficiency. Eur J Immunol. 2000;30:2516–20.PubMedCrossRefGoogle Scholar
  11. 11.
    Jacquot S, Macon-Lemaitre L, Paris E, Kobata T, Tanaka Y, Morimoto C, et al. B cell co-receptors regulating T cell-dependent antibody production in common variable immunodeficiency: CD27 pathway defects identify subsets of severely immuno-compromised patients. Int Immunol. 2001;13:871–6.PubMedCrossRefGoogle Scholar
  12. 12.
    Agematsu K, Futatani T, Hokibara S, Kobayashi N, Takamoto M, Tsukada S, et al. Absence of memory B cells in patients with common variable immunodeficiency. Clin Immunol. 2002;103:34–42.PubMedCrossRefGoogle Scholar
  13. 13.
    Piqueras B, Lavenu-Bombled C, Galicier L, Bergeron-van der Cruyssen F, Mouthon L, Chevret S, et al. Common variable immunodeficiency patient classification based on impaired B cell memory differentiation correlates with clinical aspects. J Clin Immunol. 2003;23:385–400.PubMedCrossRefGoogle Scholar
  14. 14.
    Warnatz K, Denz A, Dräger R, Braun M, Groth C, Wolff-Vorbeck G, et al. Severe deficiency of switched memory B cells (CD27(+)IgM(−)IgD(−)) in subgroups of patients with common variable immunodeficiency: a new approach to classify a heterogeneous disease. Blood. 2002;99:1544–51.PubMedCrossRefGoogle Scholar
  15. 15.
    Wehr C, Kivioja T, Schmitt C, Ferry B, Witte T, Eren E, et al. The EUROclass trial: defining subgroups in common variable immunodeficiency. Blood. 2008;111:77–85.PubMedCrossRefGoogle Scholar
  16. 16.
    Bryant A, Calver NC, Toubi E, Webster AD, Farrant J. Classification of patients with common variable immunodeficiency by B cell secretion of IgM and IgG in response to anti-IgM and interleukin-2. Clin Immunol Immunopathol. 1990;56:239–48.PubMedCrossRefGoogle Scholar
  17. 17.
    Eisenstein EM, Jaffe JS, Strober W. Reduced interleukin-2 (IL-2) production in common variable immunodeficiency is due to a primary abnormality of CD4+ T cell differentiation. J Clin Immunol. 1993;13:247–58.PubMedCrossRefGoogle Scholar
  18. 18.
    Stagg AJ, Funauchi M, Knight SC, Webster AD, Farrant J. Failure in antigen responses by T cells from patients with common variable immunodeficiency (CVID). Clin Exp Immunol. 1994;96:48–53.PubMedCrossRefGoogle Scholar
  19. 19.
    Thon V, Eggenbauer H, Wolf HM, Fischer MB, Litzman J, Lokaj J, et al. Antigen presentation by common variable immunodeficiency (CVID) B cells and monocytes is unimpaired. Clin Exp Immunol. 1997;108:1–8.PubMedCrossRefGoogle Scholar
  20. 20.
    Takahashi N, Matsumoto K, Saito H, Nanki T, Miyasaka N, Kobata T, et al. Impaired CD4 and CD8 effector function and decreased memory T cell populations in ICOS-deficient patients. J Immunol. 2009;182:5515–27.PubMedCrossRefGoogle Scholar
  21. 21.
    Pandolfi F, Trentin L, San Martin JE, Wong JT, Kurnick JT, Moscicki RA. T cell heterogeneity in patients with common variable immunodeficiency as assessed by abnormalities of T cell subpopulations and T cell receptor gene analysis. Clin Exp Immunol. 1992;89:198–203.PubMedCrossRefGoogle Scholar
  22. 22.
    Farrant J, Spickett G, Matamoros N, Copas D, Hernandez M, North M, et al. Study of B and T cell phenotypes in blood from patients with common variable immunodeficiency (CVID). Immunodeficiency. 1994;5:159–69.PubMedGoogle Scholar
  23. 23.
    Guazzi V, Aiuti F, Mezzaroma I, Mazzetta F, Andolfi G, Mortellaro A, et al. Assessment of thymic output in common variable immunodeficiency patients by evaluation of T cell receptor excision circles. Clin Exp Immunol. 2002;129:346–53.PubMedCrossRefGoogle Scholar
  24. 24.
    Moratto D, Gulino AV, Fontana S, Mori L, Pirovano S, Soresina A, et al. Combined decrease of defined B and T cell subsets in a group of common variable immunodeficiency patients. Clin Immunol. 2006;121:203–14.PubMedCrossRefGoogle Scholar
  25. 25.
    Giovannetti A, Pierdominici M, Mazzetta F, Marziali M, Renzi C, Mileo AM, et al. Unravelling the complexity of T cell abnormalities in common variable immunodeficiency. J Immunol. 2007;178:3932–43.PubMedGoogle Scholar
  26. 26.
    Isgrò A, Marziali M, Mezzaroma I, Luzi G, Mazzone AM, Guazzi V, et al. Bone marrow clonogenic capability, cytokine production, and thymic output in patients with common variable immunodeficiency. J Immunol. 2005;174:5074–81.PubMedGoogle Scholar
  27. 27.
    Giovannetti A, Pierdominici M, Aiuti F. T-cell homeostasis: the dark(ened) side of common variable immunodeficiency. Blood. 2008;112:446–7.PubMedCrossRefGoogle Scholar
  28. 28.
    Sottini A, Ghidini C, Zanotti C, Chiarini M, Caimi L, Lanfranchi A, et al. Simultaneous quantification of recent thymic T-cell and bone marrow B-cell emigrants in patients with primary immunodeficiency undergone to stem cell transplantation. Clin Immunol. 2010;136:217–27.PubMedCrossRefGoogle Scholar
  29. 29.
    van Zelm MC, Szczepanski T, van der Burg M, van Dongen JJM. Replication history of B lymphocytes reveals homeostatic proliferation and extensive antigen-induced B cell expansion. J Exp Med. 2007;204:645–55.PubMedCrossRefGoogle Scholar
  30. 30.
    Hazenberg MD, Verschuren MC, Hamann D, Miedema F, van Dongen JJ. T cell receptor excision circles as markers for recent thymic emigrants: basic aspects, technical approach, and guidelines for interpretation. J Mol Med. 2001;79:631–40.PubMedCrossRefGoogle Scholar
  31. 31.
    Fronkova E, Muzikova K, Mejstrikova E, Kovac M, Formankova R, Sedlacek P, et al. B-cell reconstitution after allogeneic SCT impairs minimal residual disease monitoring in children with ALL. Bone Marrow Transplant. 2008;42:187–96.PubMedCrossRefGoogle Scholar
  32. 32.
    Sallusto F, Lenig D, Förster R, Lipp M, Lanzavecchia A. Two subsets of memory T lymphocytes with distinct homing potentials and effector functions. Nature. 1999;401:708–12.PubMedCrossRefGoogle Scholar
  33. 33.
    Kimmig S, Przybylski GK, Schmidt CA, Laurisch K, Möwes B, Radbruch A, et al. Two subsets of naive T helper cells with distinct T cell receptor excision circle content in human adult peripheral blood. J Exp Med. 2002;195:789–94.PubMedCrossRefGoogle Scholar
  34. 34.
    Chiarini M, Sottini A, Ghidini C, Zanotti C, Serana F, Rottoli M, et al. Renewal of the T-cell compartment in multiple sclerosis patients treated with glatiramer acetate. Mult Scler. 2010;16:218–27.PubMedCrossRefGoogle Scholar
  35. 35.
    Maurer D, Fischer GF, Fae I, Majdic O, Stuhlmeier K, Von Jeney N, et al. IgM and IgG but not cytokine secretion is restricted to the CD27+ B lymphocyte subset. J Immunol. 1992;148:3700–5.PubMedGoogle Scholar
  36. 36.
    Chen X, Barfield R, Benaim E, Leung W, Knowles J, Lawrence D, et al. Prediction of T-cell reconstitution by assessment of T-cell receptor excision circle before allogeneic hematopoietic stem cell transplantation in pediatric patients. Blood. 2005;105:886–93.PubMedCrossRefGoogle Scholar
  37. 37.
    Motta M, Chiarini M, Ghidini C, Zanotti C, Lamorgese C, Caimi L, et al. Quantification of newly produced B and T lymphocytes in untreated chronic lymphocytic leukemia patients. J Transl Med. 2010;8:111.PubMedCrossRefGoogle Scholar
  38. 38.
    Lorenzi AR, Patterson AM, Pratt A, Jefferson M, Chapman CE, Ponchel F, et al. Determination of thymic function directly from peripheral blood: a validated modification to an established method. J Immunol Meth. 2008;339:185–94.CrossRefGoogle Scholar
  39. 39.
    Pido-Lopez J, Imami N, Aspinall R. Both age and gender affect thymic output: more recent thymic migrants in females than males as they age. Clin Exp Immunol. 2001;125:409–13.PubMedCrossRefGoogle Scholar
  40. 40.
    Ribeiro RM, Perelson AS. Determining thymic output quantitatively: using models to interpret experimental T-cell receptor excision circle (TREC) data. Immunol Rev. 2007;216:21–34.PubMedGoogle Scholar
  41. 41.
    Vlkova M, Thon V, Sarfyova M, Blaha L, Svobodnik A, Lokaj J, et al. Age dependency and mutual relations in T and B lymphocyte abnormalities in common variable immunodeficiency patients. Clin Exp Immunol. 2006;143:373–9.PubMedCrossRefGoogle Scholar
  42. 42.
    Mouillot G, Carmagnat M, Gérard L, Garnier JL, Fieschi C, Vince N, et al. B-cell and T-cell phenotypes in CVID patients correlate with the clinical phenotype of the disease. J Clin Immunol. 2010;30:746–55.PubMedCrossRefGoogle Scholar
  43. 43.
    De Vera MJ, Al Harthi L, Gewurz AT. Assessing thymopoiesis in patients with common variable immunodeficiency as measured by T-cell receptor excision circles. Ann Allergy Asthma & Immunol. 2004;93:478–84.CrossRefGoogle Scholar
  44. 44.
    Spickett GP, Farrant J, North ME, Zhang JG, Morgan L, Webster AD. Common variable immunodeficiency: how many diseases? Immunol Today. 1997;18:325–8.PubMedCrossRefGoogle Scholar
  45. 45.
    Goldacker S, Warnatz K. Tackling the heterogeneity of CVID. Curr Opin Allergy Clin Immunol. 2005;5:504–9.PubMedCrossRefGoogle Scholar
  46. 46.
    Douek DC, Betts MR, Hill BJ, Little SJ, Lempicki R, Metcalf JA, et al. Evidence for increased T cell turnover and decreased thymic output in HIV infection. J Immunol. 2001;167:6663–8.PubMedGoogle Scholar
  47. 47.
    Vlková M, Fronková E, Kanderová V, Janda A, Ruzicková S, Litzman J, et al. Characterization of lymphocyte subsets in patients with common variable immunodeficiency reveals subsets of naive human B cells marked by CD24 expression. J Immunol. 2010;185:6431–8.PubMedCrossRefGoogle Scholar
  48. 48.
    Banchereau J, Rousset F. Human B lymphocytes: phenotype, proliferation, and differentiation. Adv Immunol. 1992;52:125–262.PubMedCrossRefGoogle Scholar
  49. 49.
    Caraux A, Klein B, Paiva B, Bret C, Schmitz A, Fuhler GM, et al. Circulating human B and plasma cells. Age-associated changes in counts and detailed characterization of circulating normal CD138− and CD138+ plasma cells. Haematologica. 2010;95:1016–20.PubMedCrossRefGoogle Scholar
  50. 50.
    Treml LS, Quinn 3rd WJ, Treml JF, Scholz JL, Cancro MP. Manipulating B cell homeostasis: a key component in the advancement of targeted strategies. Arch Immunol Ther Exp. 2008;56:153–64.CrossRefGoogle Scholar
  51. 51.
    Boileau J, Mouillot G, Gérard L, Carmagnat M, Rabian C, Oksenhendler E, et al. Autoimmunity in common variable immunodeficiency: correlation with lymphocyte phenotype in the French DEFI study. J Autoimmun. 2011;36:25–32.PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2011

Authors and Affiliations

  • Federico Serana
    • 1
  • Paolo Airò
    • 3
  • Marco Chiarini
    • 2
  • Cinzia Zanotti
    • 2
  • Mirko Scarsi
    • 3
  • Micol Frassi
    • 3
  • Vassilios Lougaris
    • 4
  • Alessandro Plebani
    • 4
  • Luigi Caimi
    • 2
  • Luisa Imberti
    • 2
  1. 1.Department of Biomedical Science and BiotechnologyUniversity of BresciaBresciaItaly
  2. 2.Laboratorio di Biotecnologie, Diagnostics DepartmentSpedali Civili di BresciaBresciaItaly
  3. 3.Rheumatology and Clinical Immunology ServiceSpedali Civili and University of BresciaBresciaItaly
  4. 4.Pediatrics and Laboratory of Molecular Medicine “A. Nocivelli”Spedali Civili di BresciaBresciaItaly

Personalised recommendations