Phenotype and adhesion characteristics of rat thymic macrophages cultivated in serum-free medium

Rat thymic macrophages (TMF) were cultivated in serum-free medium (Biorich 2) using an explant technique. This medium favored the growth of macrophages and inhibited the proliferation of other non-lymphoid cells. Macrophage characteristics of cultivated cells were confirmed by their morphology, NSE activity and positivity (approx. 95%) with ED1 and Mar 1 mAbs (pan-macrophage markers in the rat). The majority of non-adherent cells (NAC) released in culture medium during the proliferation of TMF also possessed macrophage characteristics. Both adherent TMF and NAC were heterogeneous cells expressing cortical/CMZ phenotype (ED2+, R-MC 41+, R-MC 42+) and CMZ/medullary phenotype (R-MC 43+, R-MC 44+). NAC population contained higher proportion of cells expressing class II MHC molecules, IL-2R and 1F119 antigen compared to adherent TMF. The TMF phenotype changed during cultivation and upon stimulation with inflammatory cytokines (IL-1, IFN-gamma, TNF-alpha). TMF formed rosettes with syngeneic or allogeneic thymocytes both at 37 and 4 degrees C. The percentages of rosettes was dependent on the incubation time and TMF/thymocyte ratio. A higher percentage of immature thymocytes (CD4-CD8- or CD4+CD8+) bound to TMF compared to mature, cortisone-resistant thymocytes. The involvement of TMF membrane molecules in TMF/thymocyte adhesion was tested by preincubation of TMF with specific mAbs. It was found that Mar 1 mAb was partially inhibitory, OX-42 (anti CD11b) was slightly stimulatory, whereas other mAbs had no significant effect.