Cluster formation of canine dendritic cells and lymphocytes is calcium dependent and not inhibited by cyclosporine.

Canine peripheral blood mononuclear cells (PBMC), dendritic cells (DC), and lymphocytes either alone or in combination were cultured with concanavalin A (ConA), calcium ionophore, and phorobol-12-myristate-13-acetate (PMA), and examined for lymphocyte/DC cluster formation and lymphocyte proliferation as determined by thymidine uptake. ConA- or calcium ionophore-stimulated proliferation of PBMC required the presence of normal DC, and was preceded by cluster formation of DC and lymphocytes. PMA-triggered proliferation was not preceded by cluster formation but also required the presence of normal DC. The presence of ultraviolet (UV)-irradiated rather than normal DC did not permit ConA-, calcium ionophore-, or PMA-triggered lymphocyte proliferation. Addition of interleukin 2 (IL-2) to cultures of lymphocytes and UV-irradiated DC restored responsiveness to PMA, suggesting that a decrease in cytokine production was the central event in UV-induced accessory cell inhibition. Cyclosporine, known to interfere with IL-2 release and responses, completely blocked both ConA- and PMA-induced lymphocyte proliferation but did not interfere with ConA-triggered cluster formation. Verapamil blocked both cluster formation and proliferation. These data show that DC/lymphocyte cluster formation is Ca2+ dependent and cannot be inhibited by cyclosporine. The data show, furthermore, that in agreement with findings in other species, triggering of canine lymphocytes by lectins and phorbol esters follows distinct pathways.

Canine lymphocyte subpopulations.

Ten to 23% of cells in blood, lymph node and bone marrow from normal dogs formed rosettes with human erythrocytes, and 12-27% formed rosettes with erythrocyte-antibody-complement (EAC) complexes. In contrast, only 3% of thymocytes, and 1% of thoracic duct cells formed rosettes with human erythroyctes, and 0 and 15% respectively formed EAC rosettes. When peripheral blood mononuclear cells were separated by rosette sedimentation into populations depleted of, or enriched for, cells forming rosettes with human erythrocytes (H-RFC), the population depleted of H-RFC responded more vigorously to alloantigens in mixed leukocyte culture (MLC) (P < 0.01) and to the mitogens phytohemagglutinin (PHA) (P = 0.01) and concanavalin A (P = 0.01) than did the population enriched for H-RFC. Passage of peripheral blood mononuclear cells over nylon wool columns produced a nonadherent population depleted of H-RFC, EAC rosette-forming cells and cells binding surface immunoglobulin (SIg), while the adherent population was enriched for each of these markers. In 3 dogs 36%, 44% and 64% of adherent cells that formed rosettes with human erythrocytes also possessed SIg, suggesting that canine B cells form rosettes with human red cells. The nonadherent population showed a more vigorous response to alloantigens in MLC (P < 0.01) and to PHA (P < 0.05) than the adherent population, and also stimulated the growth of autologous erythroid colonies better than the adherent population (P = 0.02). A T cell rich population can thus be obtained from canine peripheral blood, but no specific marker for T cells has been identified. Specifically, the capacity to form rosettes with human red cells is not a marker for the canine T cell.

Prevention of transfusion-induced graft-versus-host disease in dogs by ultraviolet irradiation.

Ten dogs were given 9.2 Gy of total body irradiation and autologous bone marrow infusion followed by ten daily transfusions of leukocytes for a total of 11.5 to 36.2 (median, 18.8) x 10(8)/kg obtained via leukapheresis from histoincompatible unrelated donors. Four dogs were given unirradiated leukocytes, and all developed graft-versus-host disease (GVHD). In contrast, only two of three dogs given leukocytes irradiated with 20 mJ/cm2 of ultraviolet (UV) light (200 to 300 nm), and none of three dogs given leukocytes irradiated with 1,000 mJ/cm2 developed GVHD. These data indicate that UV irradiation abrogates the alloreactive potential of transfused leukocytes, and suggest that UV irradiation can be used to prevent the development of transfusion-induced GVHD.