Previously unreported CD8(+) CD28(-) and CD8(+) CD28(+) T-cell subsets occur in healthy individuals and expand in patients suffering from autoimmune disease. Here we studied, for the first time, the expression of CD8(+) CD28(+) , CD8(+) CD28(-) , and CD8(+) CD56(+) subpopulations in induced sputum from asthmatics. Using sputum samples, purified CD8(+) T cells were stained for surface antigen CD28, CD56, FITC-conjugated anti-perforin, and anti-IFN-gamma. Cytotoxic activity was evaluated in a chromium releasing test. Induced sputum CD8(+) CD28(-) T cells were found to be more expanded and expressed low levels of IFN-gamma in severe asthmatics than mild asthma and age-matched healthy controls. The predominance of CD8(+) CD28(-) T cells can be in part explained by the expansion of CD8(+) CD56(+). CD8(+) CD28(-) T cells from severe asthmatics produced high intracytoplasmic perforin and exerted a potent cytotoxic activity. Considering their phenotyping and functional properties, the CD8(+) CD28(-) T-cell subset may constitute an intermediate phenotype in the process of CD8(+) T-cell differentiation of effector-type cells in severe asthmatics. Functional studies showed that CD8(+) CD28(-) T cells had cytotoxic function.