title: Identification of canine natural CD3-positive T cells expressing an invariant T-cell receptor alpha chain.
abstract: A representative T-cell subset exclusively using an invariant TCRalpha chain (iTCRalpha) is natural killer T (NKT) cells which are becoming an emerging topic for cancer and immune disorder in humans and mice. However, NKT cells in dogs have not yet been identified. In this study, CD3(+) T-lymphocyte population reactive to alpha-galactosylceramide-loaded mouse CD1d (alpha-GalCer/CD1d) were identified with flow cytometric analysis in mononuclear cells from spleen, liver, and peripheral blood of a dog with percentages of 0.028%, 0.045%, and 0.004%, respectively. Using cDNA library synthesized from mRNAs of the alpha-GalCer/CD1d reactive CD3(+) lymphocytes in the spleen cells, molecular analysis of canine iTCRalpha was carried out. Consequently, Variable (Valpha) and Joining (Jalpha) regions of iTCRalpha cDNA were found to be homogeneous to both mouse Valpha14-Jalpha281 and human Valpha24-JalphaQ. Characteristic features of iTCRalpha of NKT cells, such as the amino acid sequence of complementarity-determining region (CDR) 3 and extra cysteine residue, were well conserved among dogs, mice, and humans. In quantitative real-time PCR analysis, relative expression of the canine iTCRalpha mRNA in alpha-GalCer/CD1d reactive CD3(+) lymphocytes was 271-fold higher than that in CD3(+) lymphocytes unbound to alpha-GalCer/CD1d, indicating that the iTCRalpha mRNA was preferentially expressed in alpha-GalCer/CD1d-reactive CD3(+) lymphocytes in the dog. Together, the results strongly suggested that alpha-GalCer/CD1d-binding CD3(+) T lymphocytes identified in this study could be considered to be canine NKT cells.
authors.affiliation: Animal Allergy Clinical Laboratories Inc., SIC-1#206 5-4-21 Nishihashimoto, Sagamihara, Kanagawa 229-1131, Japan.
authors.forename: Nobutaka
authors.initials: N
authors.lastname: Yasuda
authors.forename: Kenichi
authors.initials: K
authors.lastname: Masuda
authors.forename: Tohishiro
authors.initials: T
authors.lastname: Tsukui
authors.forename: Annabelle
authors.initials: A
authors.lastname: Teng
authors.forename: Yasuyuki
authors.initials: Y
authors.lastname: Ishii
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D055610:Antigens, CD1d / Q000276:immunology; D001483:Base Sequence; D017252:CD3 Complex / Q000378:metabolism; D017124:Conserved Sequence; D018076:DNA, Complementary / Q000235:genetics; D004285:Dogs / Q000235:genetics* / Q000276:immunology*; D005699:Galactosylceramides / Q000276:immunology; D015870:Gene Expression; D019673:Genes, T-Cell Receptor alpha; D006801:Humans; D066298:In Vitro Techniques; D051379:Mice; D008969:Molecular Sequence Data; D055611:Natural Killer T-Cells / Q000276:immunology*; D012333:RNA, Messenger / Q000235:genetics; D017386:Sequence Homology, Amino Acid; D013045:Species Specificity; D016176:T-Lymphocyte Subsets / Q000276:immunology*
chemical_list: D055610:Antigens, CD1d; D017252:CD3 Complex; D018076:DNA, Complementary; D005699:Galactosylceramides; D012333:RNA, Messenger; C493154:alpha-galactosylceramide