Gastroenterology

Gastroenterology

Volume 123, Issue 4, October 2002, Pages 1031-1043
Gastroenterology

Clinical–Liver, Pancreas, and Biliary Tract
Quantitation and phenotypic analysis of natural killer T cells in primary biliary cirrhosis using a human CD1d tetramer,☆☆

https://doi.org/10.1053/gast.2002.36020Get rights and content

Abstract

Background & Aims: Natural killer T (NKT) cells are a subset of lymphocytes incriminated in playing an important role in the modulation of the innate immune response and the development of autoimmunity. However, there have been only limited studies attempting to quantitate the number of NKT cells in autoimmune disease, particularly because of difficulties associated with definition of this subpopulation. Methods: We used a human CD1d (hCD1d) tetramer produced by a baculovirus expressing recombinant CD1d protein complexed with α-galactosylceramide (α-GalCer) and quantitated hCD1d tetramer reactive cells in blood and liver from controls and patients with primary biliary cirrhosis (PBC). Results: The majority of CD1d-αGalCer-restricted NKT cells were positive for TCR Vα24 and Vβ11. There was a distinct CD4 CD8+ population within the CD1d-αGalCer-restricted NKT cells in addition to the CD4 CD8 and CD4+ CD8 population. The frequency of CD1d-αGalCer-restricted NKT cells was similar between blood and liver in healthy individuals. In contrast, the frequency of CD1d-αGalCer-restricted NKT cells in the liver was significantly higher than in the blood of PBC patients. The frequency of CD1d-α-GalCer-restricted NKT cells in the liver was also significantly higher in PBC patients than in healthy individuals. Conclusions: The frequency and function of such cells should be studied not only in blood but also in the target organ of the autoimmune disease. Selective enrichment of CD1d-αGalCer-restricted NKT cells at the site of inflammation is observed in PBC, suggesting a role of these cells in the development of PBC.

GASTROENTEROLOGY 2002;123:1031-1043

Section snippets

Tetrameric human CD1d-α-GalCer complex

α-GalCer was synthesized and kindly provided by the Pharmaceutical Research Laboratory of Kirin Brewery Co.18 Then, tetrameric human CD1d complexes loaded with α-GalCer and control unloaded hCD1d tetramers were prepared. First, a human CD1d heavy chain was PCR amplified and cloned into the SalI and BamHI sites of the previously described mouse CD1/β2-microglobulin (β2m) expression vector pBacP10pH (gift of J. Kappler; National Jewish Center, Denver, CO), thereby replacing the mouse CD1d heavy

Detection of hCD1d tetramer-binding cells in PBMCs

Our initial attempts were focused on defining the specific peripheral lymphocytes stained with hCD1d tetramer loaded with α-GalCer. Thus, PBMCs from 28 healthy individuals were stained with FITC-labeled anti-CD3 antibody and PE-labeled hCD1d tetramer loaded with α-GalCer (hCD1d tetramer) or an “empty” hCD1d tetramer not complexed with α-GalCer. A distinct population of CD3+ lymphocytes positively stained with α-GalCer loaded hCD1d tetramer was detected in all 28 samples, at a frequency of 0.07%

Discussion

Several lines of evidence suggest that NKT cells are implicated in human as well as murine models of autoimmune diseases. Nonobese diabetic (NOD) mice are numerically and functionally deficient in NKT cells in the thymus24, 25 and, to a lesser extent, in the periphery26 well before the onset of diabetes.27 A quantitative defect in NKT cells has also been reported in other autoimmune mouse models such as lpr/lpr and NZB and NZB/NZW F1 mice.28, 29

Transfer of double-negative NKT thymocytes

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    Address requests for reprints to: M. Eric Gershwin, M.D., Division of Rheumatology, Allergy, and Clinical Immunology, University of California at Davis School of Medicine, TB 192, One Shields Avenue, Davis, California 95616. e-mail: [email protected]; fax: (530) 752-4669.

    ☆☆

    Supported by grants DK39588 (to M. E. G.) and AI45053 (to M. K.) from the National Institutes of Health.

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