Frontiers in peptide–MHC class II multimer technology

Hackett, Charles J.; Sharma, Opendra K.

doi:10.1038/ni1002-887

Cited by 24 publications

(21 citation statements)

References 19 publications

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“…Although MHC class II tetramers were first used in mice in 1998 [23,24], only a handful of reports utilizing these reagents have been published, suggesting that technical obstacles still prevent their routine use [10]. In addition to these technical difficulties, MHC multimers are also restricted to exactly defined antigenic peptide epitopes, limiting their use to a few well-characterized model antigens and common MHC alleles and confining the analysis to only those T cells that recognize the very same peptide.…”

Section: Discussionmentioning

confidence: 99%

“…Both methods allow the analysis of live cells on the singlecell level but are limited by several restrictions: MHC multimers require defined antigenic peptide epitopes, and for many pathogens and autoimmune diseases no epitopes are known. Moreover, the preparation of functional MHC class II multimers is still a challenge [10], and thus the technology is currently not applicable to the routine analysis of CD4 T cell responses. Functional readouts such as the detection of cytokinesecreting cells do not require defined antigenic epitopes; whole proteins or crude lysates of pathogens can be used for in vitro stimulation [8].…”

Section: Introductionmentioning

confidence: 99%

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Identification and isolation of murine antigen‐reactive T cells according to CD154 expression

et al. 2007

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T helper (Th) cells are central regulators of adaptive immune responses. However, the detection of the small number of Th cells specific for a particular antigen or pathogen is still a major challenge. CD154 was recently introduced as a marker for antigen-specific Th cells. To date, this technology was not applicable for mice -arguably the most important immunological model system. CD154 is difficult to detect due to its rapid removal from the cell surface upon binding to CD40 during antigen-specific activation by APC. We present an efficient strategy to block the degradation of murine CD154 by combined use of antibodies against CD40 and CD154. This strategy makes CD154 easily accessible for surface staining, which allows isolation and expansion of rare antigen specific T cells. Importantly, CD154 identified all specific T cells in strongly Th1-or Th2-polarized immune responses against pathogens like Salmonella typhimurium and Heligmosomoides polygyrus, independent of their potential to produce cytokines. We demonstrate that CD154 can in fact be used as a reliable marker for antigen-specific CD4 T cells in mice, offering a unique option to analyze, isolate and rapidly expand the entire pool of Th-cells generated during a physiological T cell response in vivo.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Identification and isolation of murine antigen‐reactive T cells according to CD154 expression

et al. 2007

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“…Formation of the SMAC on the T cell membrane is induced in T cell hybridoma or naive transgenic T cells by employing a system of planar lipid bilayers in which MHC-peptide complexes and adhesion molecules are uniformly distributed. SMAC formation was found to correlate with cellular activation (12). Accumulation of MHC-peptide complexes at the APC-T cell interface was assumed to occur passively upon the interaction with the TCR.…”

mentioning

confidence: 99%

Clustering of T Cell Ligands on Artificial APC Membranes Influences T Cell Activation and Protein Kinase C θ Translocation to the T Cell Plasma Membrane

Giannoni

Barnett

et al. 2005

The Journal of Immunology

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T cell activation is associated with active clustering of relevant molecules in membrane microdomains defined as the supramolecular activation cluster. The contact area between these regions on the surface of T cells and APC is defined as the immunological synapse. It has been recently shown that preclustering of MHC-peptide complexes in membrane microdomains on the APC surface affects the efficiency of immune synapse formation and the related T cell activation. Disruption of such clusters may reduce the efficiency of stimulation. We describe here an entirely artificial system for Ag-specific, ex vivo stimulation of human polyclonal T cells (artificial APC (aAPC)). aAPC are based on artificial membrane bilayers containing discrete membrane microdomains encompassing T cell ligands (i.e., appropriate MHC-peptide complexes in association with costimulatory molecules). We show here that preclustering of T cell ligands triggered a degree of T cell activation significantly higher than the one achieved when we used either soluble tetramers or aAPC in which MHC-peptide complexes were uniformly distributed within artificial bilayer membranes. This increased efficiency in stimulation was mirrored by increased translocation from the cytoplasm to the membrane of protein kinase θ, a T cell signaling molecule that colocalizes with the TCR within the supramolecular activation cluster, thus indicating efficient engagement of T cell activation pathways. Engineered aAPC may have immediate application for basic and clinical immunology studies pertaining to modulation of T cells ex vivo.

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“…Recently, soluble ligands for TCRs have been developed to target Ag-specific T cells (11)(12)(13). These TCR ligands consist of engineered MHC molecules in multimeric form, some with covalently attached peptide Ags.…”

mentioning

confidence: 99%

Peptide-MHC Class II Dimers as Therapeutics to Modulate Antigen-Specific T Cell Responses in Autoimmune Diabetes

Masteller

Warner

Ferlin

et al. 2003

The Journal of Immunology

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Type 1 diabetes is an autoimmune disorder caused by autoreactive T cells that mediate destruction of insulin-producing β cells of the pancreas. Studies have shown that T cell tolerance can be restored by inducing a partial or altered signal through the TCR. To investigate the potential of bivalent peptide-MHC class II/Ig fusion proteins as therapeutics to restore Ag-specific tolerance, we have developed soluble peptide I-Ag7 dimers for use in the nonobese diabetic mouse model of diabetes. I-Ag7 dimers with a linked peptide specific for islet-reactive BDC2.5 TCR transgenic CD4+ T cells were shown to specifically bind BDC2.5 T cells as well as a small population of Ag-specific T cells in nonobese diabetic mice. In vivo treatment with BDC2.5 peptide I-Ag7 dimers protected mice from diabetes mediated by the adoptive transfer of diabetogenic BDC2.5 CD4+ T cells. The dimer therapy resulted in the activation and increased cell death of transferred BDC2.5 CD4+ T cells. Surviving cells were hypoproliferative to challenge by Ag and produced increased levels of IL-10 and decreased levels of IFN-γ compared with cells from control I-Ag7 dimer-treated mice. Anti-IL-10R therapy reversed the tolerogenic effects of the dimer. Thus, peptide I-Ag7 dimers induce tolerance of BDC2.5 TCR T cells through a combination of the induction of clonal anergy and anti-inflammatory cytokines.

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Frontiers in peptide–MHC class II multimer technology

Abstract: Although tetramer technology has been wildly successful for examination of MHC class I-recognizing T cells, the same hasn't been true for MHC class II reagents.

Cited by 24 publications

References 19 publications

Identification and isolation of murine antigen‐reactive T cells according to CD154 expression

Identification and isolation of murine antigen‐reactive T cells according to CD154 expression

Clustering of T Cell Ligands on Artificial APC Membranes Influences T Cell Activation and Protein Kinase C θ Translocation to the T Cell Plasma Membrane

Peptide-MHC Class II Dimers as Therapeutics to Modulate Antigen-Specific T Cell Responses in Autoimmune Diabetes

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