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Invariant Natural Killer T Cells are Reduced in Hereditary Hemochromatosis Patients



Invariant natural killer T (iNKT) cells are CD1d restricted-T cells that react to lipid antigens. iNKT cells were shown to be important in infection, autoimmunity and tumor surveillance. Alterations in the number and function of these cells were described in several pathological conditions including autoimmune and/or liver diseases. CD1d is critical for antigen presentation to iNKT cells, and its expression is increased in liver diseases. The liver is the major organ affected in Hereditary Hemochromatosis (HH), an autosomal recessive disorder caused by excessive iron absorption. Herein, we describe the study of iNKT cells of HH patients.


Twenty-eight HH patients and 24 control subjects from Santo António Hospital, Porto, were included in this study. Patient’s iron biochemical parameters (serum transferrin saturation and ferritin levels) and the liver function marker alanine transaminase (ALT) were determined at the time of study. Peripheral blood iNKT cells were analyzed by flow cytometry using an anti-CD3 antibody and the CD1d tetramer loaded with PBS57.


We found a decrease in the percentage and number of circulating iNKT cells from HH patients when compared with control population independently of age. iNKT cell defects were more pronounced in untreated patients, relating with serum ferritin and transferrin saturation levels. No correlation was found with ALT, a marker of active liver dysfunction.


Altogether, our results demonstrate that HH patients have reduced numbers of iNKT cells and that these are influenced by iron overload.

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  1. Godfrey DI, Stankovic S, Baxter AG. Raising the NKT cell family. Nat Immunol. 2010;11:197–206.

    Article  CAS  PubMed  Google Scholar 

  2. Berzins SP, Smyth MJ, Baxter AG. Presumed guilty: natural killer T cell defects and human disease. Nat Rev Immunol. 2011;11:131–42.

    Article  CAS  PubMed  Google Scholar 

  3. De Libero G, Mori L. How the immune system detects lipid antigens. Prog Lipid Res. 2010;49:120–7.

    Article  PubMed  Google Scholar 

  4. Lynch L, O’Shea D, Winter DC, Geoghegan J, Doherty DG, O’Farrelly C. Invariant NKT cells and CD1d(+) cells amass in human omentum and are depleted in patients with cancer and obesity. Eur J Immunol. 2009;39:1893–901.

    Article  CAS  PubMed  Google Scholar 

  5. Olszak T, An D, Zeissig S, Vera MP, Richter J, Franke A, et al. Microbial exposure during early life has persistent effects on natural killer T cell function. Science. 2012;336:489–93.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  6. Montoya CJ, Cataño JC, Ramirez Z, Rugeles MT, Wilson SB, Landay AL. Invariant NKT cells from HIV-1 or Mycobacterium tuberculosis-infected patients express an activated phenotype. Clin Immunol. 2008;127:1–6.

    Article  CAS  PubMed  Google Scholar 

  7. Kita H, Naidenko OV, Kronenberg M, Ansari AA, Rogers P, He X-S, et al. Quantitation and phenotypic analysis of natural killer T cells in primary biliary cirrhosis using a human CD1d tetramer. Gastroenterology. 2002;123:1031–43.

    Article  CAS  PubMed  Google Scholar 

  8. Lucas M, Gadola S, Meier U, Young NT, Harcourt G, Karadimitris A, et al. Frequency and phenotype of circulating Valpha24/Vbeta11 double-positive natural killer T cells during hepatitis C virus infection. J Virol. 2003;77:2251–7.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  9. Exley MA, Koziel MJ. To be or not to be NKT: natural killer T cells in the liver. Hepatology. 2004;40:1033–40.

    Article  PubMed  Google Scholar 

  10. Kukreja A, Cost G, Marker J, Zhang C, Sun Z, Lin-Su K, et al. Multiple immuno-regulatory defects in type-1 diabetes. J Clin Invest. 2002;109:131–40.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  11. Driver JP, Scheuplein F, Chen Y-G, Grier AE, Wilson SB, Serreze DV. Invariant natural killer T-cell control of type 1 diabetes: a dendritic cell genetic decision of a silver bullet or Russian roulette. Diabetes. 2010;59:423–32.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  12. Van der Vliet HJ, von Blomberg BM, Nishi N, Reijm M, Voskuyl AE, van Bodegraven AA, et al. Circulating V(alpha24+) Vbeta11+ NKT cell numbers are decreased in a wide variety of diseases that are characterized by autoreactive tissue damage. Clin Immunol. 2001;100:144–8.

    Article  PubMed  Google Scholar 

  13. Bollyky PL, Wilson SB. CD1d-restricted T-cell subsets and dendritic cell function in autoimmunity. Immunol. Cell Biol. 2004;307–14.

  14. De Libero G, Collmann A, Mori L. The cellular and biochemical rules of lipid antigen presentation. Eur J Immunol. 2009;39:2648–56.

    Article  PubMed  Google Scholar 

  15. Durante-Mangoni E, Wang R, Shaulov A, He Q, Nasser I, Afdhal N, et al. Hepatic CD1d Expression in Hepatitis C Virus Infection and Recognition by Resident Proinflammatory CD1d-Reactive T Cells. J Immunol. 2004;173:2159–66.

    Article  CAS  PubMed  Google Scholar 

  16. Brossay L, Jullien D, Cardell S, Sydora BC, Burdin N, Modlin RL, et al. Mouse CD1 is mainly expressed on hemopoietic-derived cells. J Immunol. 1997;159:1216–24.

    CAS  PubMed  Google Scholar 

  17. Canchis PW, Bhan AK, Landau SB, Yang L, Balk SP, Blumberg RS. Tissue distribution of the non-polymorphic major histocompatibility complex class I-like molecule, CD1d. Immunology. 1993;80:561–5.

    CAS  PubMed Central  PubMed  Google Scholar 

  18. Pietrangelo A. Hemochromatosis: an endocrine liver disease. Hepatology. 2007;46:1291–301.

    Article  CAS  PubMed  Google Scholar 

  19. Santos PCJL, Krieger JE, Pereira AC. Molecular diagnostic and pathogenesis of hereditary hemochromatosis. Int J Mol Sci. 2012;13:1497–511.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  20. Gton CAW, Ley KVK. Review article: haemochromatosis. Aliment Pharmacol Ther. 2002;1963–75.

  21. Porto G, Cardoso CS, Gordeuk V, Cruz E, Fraga J, Areias J, et al. Clinical and genetic heterogeneity in hereditary haemochromatosis: association between lymphocyte counts and expression of iron overload. Eur J Haematol. 2001;67:110–8.

    Article  CAS  PubMed  Google Scholar 

  22. Arosa FA, Oliveira L, Porto G, da Silva BM, Kruijer W, Veltman J, et al. Anomalies of the CD8+ T cell pool in haemochromatosis: HLA-A3-linked expansions of CD8 + CD28- T cells. Clin Exp Immunol. 1997;107:548–54.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  23. Porto B, Vieira R, Porto G. Increased capacity of lymphocytes from hereditary hemochromatosis patients homozygous for the C282Y HFE mutation to respond to the genotoxic effect of diepoxybutane. Mutat Res. 2009;673:37–42.

    Article  CAS  PubMed  Google Scholar 

  24. Cruz E, Vieira J, Almeida S, Lacerda R, Gartner A, Cardoso CS, et al. A study of 82 extended HLA haplotypes in HFE-C282Y homozygous hemochromatosis subjects: relationship to the genetic control of CD8+ T-lymphocyte numbers and severity of iron overload. BMC Med Genet. 2006;7:16.

    Article  PubMed Central  PubMed  Google Scholar 

  25. De Sousa M, Porto G. The immunological system in hemochromatosis. J Hepatol. 1998;28 Suppl 1:1–7.

    Article  PubMed  Google Scholar 

  26. Macedo MF, Porto G, Costa M, Vieira CP, Rocha B, Cruz E. Low numbers of CD8+ T lymphocytes in hereditary haemochromatosis are explained by a decrease of the most mature CD8+ effector memory T cells. Clin Exp Immunol. 2010;159:363–71.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  27. Jing Y, Gravenstein S, Chaganty NR, Chen N, Lyerly KH, Joyce S, et al. Aging is associated with a rapid decline in frequency, alterations in subset composition, and enhanced Th2 response in CD1d-restricted NKT cells from human peripheral blood. Exp Gerontol. 2007;42:719–32.

    Article  CAS  PubMed  Google Scholar 

  28. Peralbo E, DelaRosa O, Gayoso I, Pita ML, Tarazona R, Solana R. Decreased frequency and proliferative response of invariant Valpha24Vbeta11 natural killer T (iNKT) cells in healthy elderly. Biogerontology. 2006;7:483–92.

    Article  CAS  PubMed  Google Scholar 

  29. Reuben A, Phénix M, Santos MM, Lapointe R. The WT hemochromatosis protein HFE inhibits CD8+ T-lymphocyte activation. Eur J Immunol. 2014;44:1604–14.

    Article  CAS  PubMed  Google Scholar 

  30. Niemelä O, Parkkila S, Britton RS, Brunt E, Janney C, Bacon B. Hepatic lipid peroxidation in hereditary hemochromatosis and alcoholic liver injury. J Lab Clin Med. 1999;133:451–60.

    Article  PubMed  Google Scholar 

  31. Salio M, Silk JD, Cerundolo V. Recent advances in processing and presentation of CD1 bound lipid antigens. Curr Opin Immunol. 2010;22:81–8.

    Article  CAS  PubMed  Google Scholar 

  32. Myers BM, Prendergast FG, Holman R, Kuntz SM, LaRusso NF. Alterations in the structure, physicochemical properties, and pH of hepatocyte lysosomes in experimental iron overload. J Clin Invest. 1991;88:1207–15.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  33. Girelli D, Pasino M, Goodnough JB, Nemeth E, Guido M, Castagna A, et al. Reduced serum hepcidin levels in patients with chronic hepatitis C. J Hepatol. 2009;51:845–52.

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  34. Bonkovsky HL. Iron as a comorbid factor in chronic viral hepatitis. Am J Gastroenterol. 2002;97:1–4.

    Article  CAS  PubMed  Google Scholar 

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This work was supported by Fundação para a Ciência e a Tecnologia (PEST-C/SAU/LA0002/2013 - FCOMP-01-0124-FEDER-037277) and by FEDER Funds through the Operational Competitiveness Programme — COMPETE and by National Funds through FCT — Fundação para a Ciência e a Tecnologia under the project FCOMP-01-0124-FEDER-015955 (PTDC/SAU-ORG/110112/2009). MAE was partially supported by NIH grant CA170194. CSP was supported by a fellowship from Fundação para a Ciência e Tecnologia (SFRH/BD/79211/2011). The authors would like to thank the NIH Tetramer Core, Emory University, USA, for providing the CD1d-PBS57 tetramer.

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The authors declare that they have no conflict of interest.

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Correspondence to M. F. Macedo.

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Maia, M.L., Pereira, C.S., Melo, G. et al. Invariant Natural Killer T Cells are Reduced in Hereditary Hemochromatosis Patients. J Clin Immunol 35, 68–74 (2015).

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  • Invariant natural killer T cells
  • hereditary hemochromatosis
  • iron
  • lipid