New technologies for monitoring human antigen-specific T cells and regulatory T cells by flow-cytometry

Bächer, Petra; Scheffold, Alexander

doi:10.1016/j.coph.2015.04.005

Cited by 36 publications

(37 citation statements)

References 61 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The detection of CD154 as an early activation marker for visualizing antigen-reactive T cells by flow cytometry has been established in human and mouse studies and refined during the last years to properly identify Th cell responses to pathogens, environmental-, tumor-, neo-, or auto-antigens ranging from complex antigenic mixtures to proteins, peptide pools, or even single peptides (24, 45). To establish CD154 marker detection for antigen-reactive T cells in swine, we analyzed the Th cell response toward a complex antigenic mixture of C. albicans (Cand) and SEB, a bacterial superantigen known to cross-link MHC class II α-chain and TCRβ-chain.…”

Section: Resultsmentioning

confidence: 99%

“…For human studies, several ways exist to identify antigen-specific T lymphocytes directly or indirectly for further characterization. These include the use of MHC–peptide multimers and antibody-based methods such as ELISPOT, intracellular cytokine staining (ICS), or antigen-specific cultivation followed by the analysis of defined activation markers (23, 24). One activation marker is CD154 (CD40L), a member of the TNF superfamily and upregulated following short-term (5–7 h) ex vivo stimulation (25–27).…”

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Pathogen-Reactive T Helper Cell Analysis in the Pig

et al. 2017

View full text Add to dashboard Cite

There is growing interest in studying host–pathogen interactions in human-relevant large animal models such as the pig. Despite the progress in developing immunological reagents for porcine T cell research, there is an urgent need to directly assess pathogen-specific T cells—an extremely rare population of cells, but of upmost importance in orchestrating the host immune response to a given pathogen. Here, we established that the activation marker CD154 (CD40L), known from human and mouse studies, identifies also porcine antigen-reactive CD4+ T lymphocytes. CD154 expression was upregulated early after antigen encounter and CD4+CD154+ antigen-reactive T cells coexpressed cytokines. Antigen-induced expansion and autologous restimulation enabled a time- and dose-resolved analysis of CD154 regulation and a significantly increased resolution in phenotypic profiling of antigen-responsive cells. CD154 expression identified T cells responding to staphylococcal Enterotoxin B superantigen stimulation as well as T cells responding to the fungus Candida albicans and T cells specific for a highly prevalent intestinal parasite, the nematode Ascaris suum during acute and trickle infection. Antigen-reactive T cells were further detected after immunization of pigs with a single recombinant bacterial antigen of Streptococcus suis only. Thus, our study offers new ways to study antigen-specific T lymphocytes in the pig and their contribution to host–pathogen interactions.

show abstract

Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Pathogen-Reactive T Helper Cell Analysis in the Pig

et al. 2017

View full text Add to dashboard Cite

show abstract

“…In humans, a functional assay of target-specific polyclonal Treg cells has been developed (27). However, a direct functional assay of TCRs obtained from Treg cells is still not available, reflecting the limited knowledge about epitopes from physiological targets of Treg cells (27, 36). TCRs from tumor specific conventional CD4 + T cells and CD8 + T cells were identified using single-cell RT-PCR methods (23, 24, 37, 38).…”

Section: Discussionmentioning

confidence: 99%

Clonally Expanded Decidual Effector Regulatory T Cells Increase in Late Gestation of Normal Pregnancy, but Not in Preeclampsia, in Humans

et al. 2018

View full text Add to dashboard Cite

Background: Regulatory T (Treg) cells are necessary for the maintenance of allogenic pregnancy. However, the repertoire of effector Treg cells at the feto-maternal interface in human pregnancy remains unknown. Our objective was to study T cell receptor (TCR) repertoires of Treg cells during pregnancy compared to normal and complicated pregnancies.Methods:Paired samples of peripheral blood and decidua in induced abortion and miscarriage cases were obtained from consenting patients. CD4+CD25+CD127low/−CD45RA− effector Treg cells were single-cell sorted from mononuclear cells. cDNAs of complementarity determining region 3 (CDR3) in TCRβ were amplified from the single cells by RT-PCR and the sequences were analyzed. The TCRβ repertoires were determined by amino acid and nucleotide sequences. Treg cells were classified into clonally expanded and non-expanded populations by CDR3 sequences.Results: We enrolled nine induced abortion cases in the 1st trimester, 12 cases delivered without complications in the 3rd trimester, 11 miscarriages with abnormal chromosomal karyotyped embryo, seven miscarriages with normal chromosomal karyotyped embryo, and seven cases of preeclampsia [median gestational week (interquartile range): 7 (7–9), 39 (38–40), 9 (8–10), 8 (8–10), and 34 (32–37), respectively]. The frequency of clonally expanded populations of effector Treg cells increased in decidua of 3rd trimester cases compared to 1st trimester cases [4.5% (1.4–10.8%) vs. 20.9% (15.4–28.1%), p < 0.001]. Clonally expanded Treg cells were rarely seen in peripheral blood. The ratio of clonally expanded populations of decidual effector Treg cells in miscarriages with abnormal and normal embryos was not significantly different compared with that in 1st trimester normal pregnancy. Interestingly, clonally expanded populations of effector Treg cells decreased in preeclampsia compared with that in 3rd trimester normal pregnancy [9.3% (4.4–14.5%) vs. 20.9% (15.4–28.1%), p = 0.003]. When repertoires in previous pregnancy and subsequent pregnancy were compared, some portions of the repertoire were shared.Conclusion: TCR repertoires of decidual effector Treg cells are skewed in the 3rd trimester of normal pregnancy. Failure of clonal expansion of populations of decidual effector Treg cells might be related to the development of preeclampsia.

show abstract

“…The detection of PAD4-specific T cells in patients with RA is significant given the substantial challenges to studying human CD4+ T cell responses in autoimmune diseases including the marked heterogeneity in the human population, the low precursor frequency of autoantigen-specific T cells in the circulation relative to the target tissue, 46 and the immunomodulatory effects of disease-modifying antirheumatic drugs (DMARDs). 47 The immunologic studies shown here were performed using circulating PBMCs from RA patients with long-standing disease that was well controlled with one or more DMARDs, including targeted biological therapies (Table 2).…”

Section: Discussionmentioning

confidence: 99%

Proteolysis by Granzyme B Enhances Presentation of Autoantigenic Peptidylarginine Deiminase 4 Epitopes in Rheumatoid Arthritis

et al. 2016

View full text Add to dashboard Cite

Proteolysis of autoantigens can alter normal MHC class II antigen processing and has been implicated in the induction of autoimmune diseases. Many autoantigens are substrates for the protease granzyme B (GrB), but the mechanistic significance of this association is unknown. Peptidylarginine deiminase 4 (PAD4) is a frequent target of autoantibodies in patients with rheumatoid arthritis (RA) and a substrate for GrB. RA is strongly associated with specific MHC class II alleles, and elevated levels of GrB and PAD4 are found in the joints of RA patients, suggesting that GrB may alter the presentation of PAD4 by RA-associated class II alleles. In this study, complementary proteomic and immunologic approaches were utilized to define the effects of GrB cleavage on the structure, processing, and immunogenicity of PAD4. Hydrogen–deuterium exchange and a cell-free MHC class II antigen processing system revealed that proteolysis of PAD4 by GrB induced discrete structural changes in PAD4 that promoted enhanced presentation of several immunogenic peptides capable of stimulating PAD4-specific CD4+ T cells from patients with RA. This work demonstrates the existence of PAD4-specific T cells in patients with RA and supports a mechanistic role for GrB in enhancing the presentation of autoantigenic CD4+ T cell epitopes.

show abstract

New technologies for monitoring human antigen-specific T cells and regulatory T cells by flow-cytometry

Cited by 36 publications

References 61 publications

Pathogen-Reactive T Helper Cell Analysis in the Pig

Pathogen-Reactive T Helper Cell Analysis in the Pig

Clonally Expanded Decidual Effector Regulatory T Cells Increase in Late Gestation of Normal Pregnancy, but Not in Preeclampsia, in Humans

Proteolysis by Granzyme B Enhances Presentation of Autoantigenic Peptidylarginine Deiminase 4 Epitopes in Rheumatoid Arthritis

Contact Info

Product

Resources

About