CD7 and CD28 are Ig superfamily molecules expressed on thymocytes and mature T cells that share common signaling 0mechanisms and are co-mitogens for T cell activation. CD7-deficient mice are resistant to lipopolysaccharide (LPS)-induced shock syndrome, and have diminished in vivo LPS-triggered IFN-gamma and tumor necrosis factor (TNF)-alpha production. CD28-deficient mice have decreased serum Ig levels, defective IgG isotype switching, decreased T cell IL-2 production and are resistant to Staphylococcus aureus enterotoxin-induced shock. To determine synergistic roles CD7 and CD28 might play in thymocyte development and function, we have generated and characterized CD7/CD28 double-deficient mice. CD7/CD28-deficient mice were healthy, reproduced normally, had normal numbers of thymocyte subsets and had normal thymus histology. Anti-CD3 mAb induced similar levels of apoptosis in CD7-deficient, CD28-deficient and CD7/CD28 double-deficient thymocytes as in control C57BL/6 mice (P = NS). Similarly, thymocyte viability, apoptosis and necrosis following ionomycin or dexamethasone treatment were the same in control, CD7-deficient, CD28-deficient and CD7/CD28-deficient mice. CD28-deficient and CD7/CD28-deficient thymocytes had decreased [3H]thymidine incorporation responses to concanavalin A (Con A) stimulation compared to control mice (P < or = 0.01 and P < or = 0.05 respectively). CD7/CD28 double-deficient mice had significantly reduced numbers of B7-1/B7-2 double-positive cells compared to freshly isolated wild-type, CD7-deficient and CD28-deficient thymocytes. Con A-stimulated CD4/CD8 double-negative (DN) thymocytes from CD7/CD28 double-deficient mice expressed significantly lower levels of CD25 when compared to CD4/CD8 DN thymocytes from wild-type, CD7-deficient and CD28-deficient mice (P < 0.05). Anti-CD3-triggered CD7/CD28-deficient thymocytes also had decreased IFN-gamma and TNF-alpha production compared to C57BL/6 control, CD7-deficient and CD28-deficient mice (P < or = 0.05). Thus, CD7 and CD28 deficiencies combined to produce abnormalities in the absolute number of B7-1/B7-2-expressing cells in the thymus, thymocyte IL-2 receptor expression and CD3-triggered cytokine production.