Phytohemagglutinin stimulation of guanylate cyclase in human lymphocytes.

Preincubation of human peripheral blood lymphocytes with optimum mitogenic concentrations of phytohemagglutinin (PHA) produced increased activities of both membrane and soluble forms of guanylate cyclase, measured with MnZ+, Mg"', or Ca2+.No significant effects of a 5-min PHA pretreatment on lymphocyte basal or NaF-stimulated adenylate cyclase or on CAMP or cGMP phosphodiesterases were found.It is concluded that the early effects of mitogens to elevate lymphocyte cGMP levels are mediated by stimulation of activities of membrane-bound and soluble guanylate cyclase.Guanylate cyclase increases varied with the time of preexposure of cells to PHA, with peak Ca2+-or Mg"+supported activities occurring at 10 min and peak Mn2+supported activities at times up to 40 min.Guanylate cyclase stimulation was dependent on Ca2+ in the preincubation medium, was associated with increased Vma, and was not prevented by actinomycin D. A requirement for an oxidation step in the mechanism of PHA activation of guanylate cyclase is indicated by the observation that preincubation of lymphocytes with dithioerythritol prevented PHA stimulation.It was also prevented by 4 inhibitors of phospholipase Az: N6, 02'dibutyryl cyclic AMP; dimethyl-~L-2,3-distearoyloxypropyl-2-hydroxyethylammonium acetate, p-bromophenacyl bromide; and quinacrine.5,8,11,14-Eicosatetraynoic acid and nordihydroguaiaretic acid markedly inhibited activation of guanylate cyclase, while indomethacin did not.These results indicate that inhibition of the lipoxygenase ,rather than the cyclooxygenase pathway of arachidonate metabolism interferes with the effects of PHA.We suggest that hydroperoxides and hydroxy fatty acid derivatives of arachidonate, formed by the lipoxygenase reaction, represent the proximal stimulants of guanylate cyclase.Human peripheral blood lymphocytes are induced by plant lectin mitogens such as PHA' and Con A to undergo blast