Anti-CD6 monoclonal antibody, clone PY-63 [FITC]

We sought to determine whether osteoblasts (OBs) can serve as accessory cells (ACs) for T-cell activation and whether T cells directly activate OB production of IL-6, using primary human OBs (NHOst), the transformed fetal osteoblast line hFOB1.19, and an osteosarcoma line SaOS-2. Robust, bidirectional activating interactions were shown using each of these three human ostoblast lines. Introduction: Osteoblasts (OBs) could come into contact with lymphocytes during inflammatory joint destruction and fracture repair. Materials and Methods: We used several in vitro assays to assess the ability of T cells and OBs to interact in the generation of immune and inflammatory responses. Results: By flow cytometry, three OB cell lines all were found to express ligands for T-cell co-stimulation. The integrin ligand CD54/1CAM-1 was constitutively expressed by hFOB1.19 and NHOst and was upregulated on SaOS-2 by IFN-gamma. MHC Class 11 was upregulated on all three lines by IFN-gamma. CD166/ALCAM, a ligand of the T-cell molecule CD6, was constitutively expressed on all three lines. A second putative CD6 ligand designated 3A11 was expressed on hFOB1.19 and NHOst, but not consistently on SaOS-2. The ectoenzyme CD26 (dipeptidyl peptidase IV) was expressed on hFOB1.19 and NHOst, but not on SaOS-2. All three cell lines presented superantigen to T cells, especially after treatment with IFN-gamma. Superantigen presentation was inhibited by antibodies to the leukocyte integrin CD11a/CD18 (LFA-1), MHC Class 11, and CD54/ICAM-1. T cells, particularly when cytokine activated for 7 days before co-culture, stimulated all three osteoblast lines to produce interleukin (IL)-6, and this effect was boosted when IL-17 was added to the co-cultures with either resting T cells or cytokine-activated T cells. Conclusions: Bidirectional activating interactions are readily shown between human T cells and several types of human OBs. The expression by OBs of ligands for the T cell-specific molecule CD6, as well as other molecules involved in immune interactions, strongly suggests that such in vitro interactions are representative of physiologic or pathologic events that occur in vivo.

The CD3(-)NKp46(+) phenotype is frequently used for the identification of natural killer (NK) cells in various mammalian species. Recently, NKp46 expression was analyzed in more detail in swine. It could be shown that besides CD3(-)NKp46(+) lymphocytes, a small but distinct population of CD3(+)NKp46(+) cells exists. In this study, we report low frequencies of CD3(+)NKp46(+) lymphocytes in blood, lymph nodes, and spleen, but increased frequencies in non-lymphatic organs, like liver and lung. Phenotypic analyses showed that the majority of CD3(+)NKp46(+) cells coexpressed the CD8 alpha beta heterodimer, while a minor subset expressed the TCR-gamma delta, which was associated with a CD8 alpha alpha(+) phenotype. Despite these T-cell associated receptors, the majority of CD3(+)NKp46(+) lymphocytes displayed a NK-related phenotype (CD2(+)CD5(-)CD6(-)CD16(+)perforin(+)) and expressed mRNA of NKp30, NKp44, and NKG2D at similar levels as NK cells. Functional tests showed that CD3(+)NKp46(+) lymphocytes produced IFN-gamma and proliferated upon cytokine stimulation to a similar extent as NK cells, but did not respond to the T-cell mitogen, ConA. Likewise, CD3(+)NKp46(+) cells killed K562 cells with an efficiency comparable to NK cells. Cross-linking of NKp46 and CD3 led to degranulation of CD3(+)NKp46(+) cells, indicating functional signaling pathways for both receptors. Additionally, influenza A(H1N1) pdm09-infected pigs had reduced frequencies of CD3(+)NKp46(+) lymphocytes in blood, but increased frequencies in the lung in the early phase of infection. Thus, CD3(+)NKp46(+) cells appear to be involved in the early phase of influenza infections. In summary, we describe a lymphocyte population in swine with a mixed phenotype of NK and T cells, with results so far indicating that this cell population functionally resembles NK cells.