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CD27 expression discriminates porcine T helper cells with functionally distinct properties.

ABSTRACT: Differentiation of porcine T helper cells is still poorly investigated, partly due to a lack of monoclonal antibodies (mAbs) specific for molecules involved in this process. Recently, we identified a mAb specific for porcine CD27 and showed that CD27 is expressed by all naïve CD8?- T helper cells but divides CD8?+ T helper cells into a CD27+ and a CD27- subset. In the present study, detailed phenotypical and functional analyses of these T-helper cell subpopulations were performed. Naïve CD8?-CD27+ T helper cells predominantly resided in various lymph nodes, whereas higher proportions of CD8?+CD27+ and CD8?+CD27- T helper cells were found in blood, spleen and liver. Both CD8?+CD27+ and CD8?+CD27- T helper cells were capable of producing IFN-? upon in vitro polyclonal stimulation and antigen-specific restimulation. Experiments with sorted CD8?-CD27+, CD8?+CD27+ and CD8?+CD27- T-helper cell subsets following polyclonal stimulation revealed the lowest proliferative response but the highest ability for IFN-? and TNF-? production in the CD8?+CD27- subset. Therefore, these cells resembled terminally differentiated effector memory cells as described in human. This was supported by analyses of CCR7 and CD62L expression. CD8?+CD27- T helper cells were mostly CCR7- and had considerably reduced CD62L mRNA levels. In contrast, expression of both homing-receptors was increased on CD8?+CD27+ T helper cells, which also had a proliferation rate similar to naïve CD8?-CD27+ T helper cells and showed intermediate levels of cytokine production. Therefore, similar to human, CD8?+CD27+ T helper cells displayed a phenotype and functional properties of central memory cells.

SUBMITTER: Reutner K

PROVIDER: S-EPMC3610194 | biostudies-literature | 2013

REPOSITORIES: biostudies-literature

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CD27 expression discriminates porcine T helper cells with functionally distinct properties.

Reutner Katharina K Leitner Judith J Müllebner Andrea A Ladinig Andrea A Essler Sabine E SE Duvigneau J Catharina JC Ritzmann Mathias M Steinberger Peter P Saalmüller Armin A Gerner Wilhelm W

Veterinary research 20130311

Differentiation of porcine T helper cells is still poorly investigated, partly due to a lack of monoclonal antibodies (mAbs) specific for molecules involved in this process. Recently, we identified a mAb specific for porcine CD27 and showed that CD27 is expressed by all naïve CD8α- T helper cells but divides CD8α+ T helper cells into a CD27+ and a CD27- subset. In the present study, detailed phenotypical and functional analyses of these T-helper cell subpopulations were performed. Naïve CD8α-CD2 ...[more]

PMID: 23497134

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Project description:The origin and function of blood IgM+IgD+CD27+ B cells is controversial, and they are considered a heterogeneous population. Previous staining of circulating B cells of healthy donors with rotavirus fluorescent virus-like particles allowed us to differentiate two subsets of IgM+IgD+CD27+: IgMhi and IgMlo B cells. Here, we confirmed this finding and compared the phenotype, transcriptome, in vitro function, and Ig gene repertoire of these two subsets. Eleven markers phenotypically discriminated both subsets (CD1c, CD69, IL21R, CD27, MTG, CD45RB, CD5, CD184, CD23, BAFFR, and CD38) with the IgMhi phenotypically resembling previously reported marginal zone B cells and the IgMlo resembling both naïve and memory B cells. Transcriptomic analysis showed that both subpopulations clustered close to germinal center-experienced IgM only B cells with a Principal Component Analysis, but differed in expression of 78 genes. Moreover, IgMhi B cells expressed genes characteristic of previously reported marginal zone B cells. After stimulation with CpG and cytokines, significantly (p < 0.05) higher frequencies (62.5%) of IgMhi B cells proliferated, compared with IgMlo B cells (35.37%), and differentiated to antibody secreting cells (14.22% for IgMhi and 7.19% for IgMlo). IgMhi B cells had significantly (p < 0.0007) higher frequencies of mutations in IGHV and IGKV regions, IgMlo B cells had higher usage of IGHJ6 genes (p < 0.0001), and both subsets differed in their HCDR3 properties. IgMhi B cells shared most of their shared IGH clonotypes with IgM only memory B cells, and IgMlo B cells with IgMhi B cells. These results support the notion that differential expression of IgM and IgD discriminates two subpopulations of human circulating IgM+IgD+CD27+ B cells, with the IgMhi B cells having similarities with previously described marginal zone B cells that passed through germinal centers, and the IgMlo B cells being the least differentiated amongst the IgM+CD27+ subsets.

Project description:Unlike mice and humans, porcine γδ T cells represent a prominent subset of T cells in blood and secondary lymphatic organs. GATA-3, T-bet and Eomesodermin (Eomes) are transcription factors with crucial functions in T-cell development and functional differentiation, but their expression has not been investigated in porcine γδ T cells so far. We analyzed the expression of these transcription factors in γδ thymocytes, mature γδ T cells from blood, spleen, lymph nodes, and lung tissue as well as in vitro stimulated γδ T cells on the protein level by flow cytometry. GATA-3 was present in more than 80% of all γδ-thymocytes. Extra-thymic CD2- γδ T cells expressed high levels of GATA-3 in all investigated organs and had a CD8α-/dimCD27+perforin- phenotype. T-bet expression was mainly found in a subset of CD2+ γδ T cells with an opposing CD8αhighCD27dim/-perforin+ phenotype. Eomes+ γδ T cells were also found within CD2+ γδ T cells but were heterogeneous in regard to expression of CD8α, CD27, and perforin. Eomes+ γδ T cells frequently co-expressed T-bet and dominated in the spleen. During aging, CD2-GATA-3+ γδ T cells strongly prevailed in young pigs up to an age of about 2 years but declined in older animals where CD2+T-bet+ γδ T cells became more prominent. Despite high GATA-3 expression levels, IL-4 production could not be found in γδ T cells by intracellular cytokine staining. Experiments with sorted and ConA + IL-2 + IL-12 + IL-18-stimulated CD2- γδ T cells showed that proliferating cells start expressing CD2 and T-bet, produce IFN-γ, but retain GATA-3 expression. In summary, our data suggest a role for GATA-3 in the development of γδ-thymocytes and in the function of peripheral CD2-CD8α-/dimCD27+perforin- γδ T cells. In contrast, T-bet expression appears to be restricted to terminal differentiation stages of CD2+ γδ T cells, frequently coinciding with perforin expression. The functional relevance of high GATA-3 expression levels in extra-thymic CD2- γδ T cells awaits further clarification. However, their unique phenotype suggests that they represent a thymus-derived separate lineage of γδ T cells in the pig for which currently no direct counterpart in rodents or humans has been described.

Dataset Information

CD27 expression discriminates porcine T helper cells with functionally distinct properties.

Publications

CD27 expression discriminates porcine T helper cells with functionally distinct properties.

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