Abstract
Adaptive immunity in homeotherms depends greatly on CD4+ Th cells which release cytokines in response to specific antigen stimulation. Whilst bony fish and poikilothermic tetrapods possess cells that express TcR and CD4-related genes (that exist in two forms in teleost fish; termed CD4-1 and CD4-2), to date there is no unequivocal demonstration that cells equivalent to Th exist. Thus, in this study we determined whether CD4-1+ lymphocytes can express cytokines typical of Th cells following antigen specific stimulation, using the zebrafish (Danio rerio). Initially, we analyzed the CD4 locus in zebrafish and found three CD4 homologues, a CD4-1 molecule and two CD4-2 molecules. The zfCD4-1 and zfCD4-2 transcripts were detected in immune organs and were most highly expressed in lymphocytes. A polyclonal antibody to zfCD4-1 was developed and used with an antibody to ZAP70 and revealed double positive cells by immunohistochemistry, and in the Mycobacterium marinum disease model CD4-1+ cells were apparent surrounding the granulomas typical of the infection. Next a prime-boost experiment, using human gamma globulin as antigen, was performed and revealed for the first time in fish that zfCD4-1+ lymphocytes increase the expression of cytokines and master transcription factors relevant to Th1/Th2-type responses as a consequence of boosting with specific antigen.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Adaptive Immunity
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Amino Acid Sequence
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Animals
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Antibodies / chemistry
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CD4-Positive T-Lymphocytes / classification
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CD4-Positive T-Lymphocytes / immunology*
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CD4-Positive T-Lymphocytes / microbiology
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CD4-Positive T-Lymphocytes / pathology
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Cytokines / biosynthesis*
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Cytokines / immunology
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Genetic Loci / immunology
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Histocompatibility Antigens Class II / genetics
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Histocompatibility Antigens Class II / immunology*
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Humans
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Models, Molecular
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Molecular Sequence Data
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Mycobacterium Infections, Nontuberculous / genetics
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Mycobacterium Infections, Nontuberculous / immunology*
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Mycobacterium Infections, Nontuberculous / microbiology
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Mycobacterium Infections, Nontuberculous / veterinary*
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Mycobacterium marinum / immunology
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Phylogeny
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RNA, Messenger / genetics
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RNA, Messenger / immunology*
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Sequence Alignment
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Th1-Th2 Balance
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ZAP-70 Protein-Tyrosine Kinase / genetics
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ZAP-70 Protein-Tyrosine Kinase / immunology
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Zebrafish / classification
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Zebrafish / genetics
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Zebrafish / immunology
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Zebrafish Proteins / genetics
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Zebrafish Proteins / immunology
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gamma-Globulins / administration & dosage
Substances
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Antibodies
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Cytokines
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Histocompatibility Antigens Class II
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RNA, Messenger
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Zebrafish Proteins
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gamma-Globulins
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ZAP-70 Protein-Tyrosine Kinase
Associated data
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GENBANK/HE983357
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GENBANK/HE983358
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GENBANK/HE983359
Grants and funding
Both first authors were supported by the Scottish Overseas Research Scholarship Awards Scheme (SORSAS) to undertake a PhD program at the University of Aberdeen. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.