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-CD27+ T helper cells predominantly resided in various lymph nodes, whereas higher proportions of CD8α+CD27+ and CD8α+CD27- T helper cells were found in blood, spleen and liver. Both CD8α+CD27+ and CD8α+CD27- T helper cells were capable of producing IFN-γ upon in vitro polyclonal stimulation and antigen-specific restimulation. Experiments with sorted CD8α-CD27+, CD8α+CD27+ and CD8α+CD27- T-helper cell subsets following polyclonal stimulation revealed the lowest proliferative response but the highest ability for IFN-γ and TNF-α production in the CD8α+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. CD8α+CD27- T helper cells were mostly CCR7- and had considerably reduced CD62L mRNA levels. In contrast, expression of both homing-receptors was increased on CD8α+CD27+ T helper cells, which also had a proliferation rate similar to naïve CD8α-CD27+ T helper cells and showed intermediate levels of cytokine production. Therefore, similar to human, CD8α+CD27+ T helper cells displayed a phenotype and functional properties of central memory cells.

Natural Killer (NK) cells play a crucial role in the early phase of immune responses against various pathogens. In swine so far only little information about this lymphocyte population exists. Phenotypical analyses with newly developed monoclonal antibodies (mAbs) against porcine NKp46 recently revealed that in blood NKp46- and NKp46+ cells with NK phenotype exist with comparable cytotoxic properties. In spleen a third NKp46-defined population with NK phenotype was observed that was characterised by a low to negative CD8α and increased NKp46 expression. In the current study it is shown that this NKp46high phenotype was correlated with an increased expression of CD16 and CD27 compared to the CD8α+NKp46- and NKp46+ NK-cell subsets in spleen and blood. Additionally NKp46high NK cells expressed elevated levels of the chemokine receptor CXCR3 on mRNA level. Functional analyses revealed that splenic NKp46high NK cells produced much higher levels of Interferon-γ and Tumor Necrosis Factor-α upon stimulation with cytokines or phorbol-12-myristate-13-acetate/Ionomycin compared to the other two subsets. Furthermore, cross-linking of NKp46 by NKp46-specific mAbs led to a superior CD107a expression in the NKp46high NK cells, thus indicating a higher cytolytic capacity of this subset. Therefore porcine splenic NKp46high NK cells represent a highly activated subset of NK cells and may play a profound role in the immune surveillance of this organ.

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