GAMMA-DELTA(-BETA(+) HUMAN T-CELL SUBSET RESPONSES UPON STIMULATION WITH VARIOUS MYCOBACTERIUM-TUBERCULOSIS SOLUBLE EXTRACTS() AND CD4(+) ALPHA)

By using a flow cytometric technique which allows direct identificatio n of proliferating cells within mixed cell populations, we have previo usly described that soluble extracts obtained from Mycobacterium tuber culosis or M. avium represent strong stimuli for human gamma delta(+) T cells. In the present study, we demonstrate that the protocol used f or the preparation of M. tuberculosis soluble extracts may have an imp act on their gamma delta(+) T cell stimulatory capacity. In agreement with our previous data, soluble extracts prepared from bacteria killed at 85 degrees C and directly disrupted by prolonged sonication (TBe), elicited a strong proliferation of gamma delta(+) T cells after 6-7 d ays of stimulation. In contrast, when soluble extracts were obtained f rom bacteria autoclaved (121 degrees C, 25 min) and then washed by cen trifugation, a predominant proportion of CD4(+) alpha beta(+) T cells was achieved in the responding population. The stimulatory activity fo r gamma delta(+) T cells was recovered in the supernatant of the autoc laved bacteria, indicating that autoclaving of M. tuberculosis bacilli releases an antigen(s) into the supernatant which stimulates human ga mma delta(+) T cells. While protease digestion of TBe only partially r educed its stimulatory capacity on gamma delta(+) T cells, the stimula tory component(s) released into the supernatant after autoclavation of bacilli was found to be sensitive to protease digestion. Interestingl y, in contrast to the preponderant proportion of gamma delta(+) T cell s induced in the responding population by unfractionated TBe, when the extract was fractionated by fast performance liquid chromatography (F PLC), most of the fractions exhibited a strong stimulatory capacity on CD4(+) alpha beta(+) T cells only. The gamma delta(+) T cell stimulat ory activity was confined to the low molecular weight range FPLC fract ions. Such results may suggest a possible regulatory role of gamma del ta(+) T cells on CD4(+) alpha beta(+) T cells.