ROLE OF ACID BASE HOMEOSTASIS IN THE SUPPRESSION OF APOPTOSIS IN HEMATOPOIETIC-CELLS BY V-ABL PROTEIN-TYROSINE KINASE/

Removal of interleukin-3 from murine IC.DP pre-mast cells results in i rreversible commitment to apoptosis within 18 hours. To identify early events necessary for the engagement of apoptosis we examined the regu lation of intracellular pH (pH(i)). IC,DP cells acidified 2 hours afte r removal of interleukin-3 (before discernible signs of apoptosis) and by 18 hours pH(i) had decreased by 0.15 units. The acidification was due to both an increase in an acid-loading process which only occurs w hen intracellular pH is above 6.8 and a slight reduction in H+ efflux via Na+/H+ exchange. Activation of a temperature sensitive mutant of v -Abl protein tyrosine kinase suppressed apoptosis of IC.DP cells in th e absence of interleukin-3 but did not stimulate proliferation, and mo reover prevented cellular acidification. Acidification of the cells by 0.2 units to pH 6.86 by complete inhibition of Na+/H+ exchange by 10 mu M 5'-(N-methyl-N-isobutyl)-amiloride prevented the suppression of a poptosis by v-abl protein tyrosine kinase following IL 3 withdrawal. H owever in the presence of interleukin-3, addition of 10 mu M 5'-(N-met hyl-N-isobutyl)-amiloride only resulted in a fall of pH(i) to 7.17. Ap optosis did not occur and the cells continued to proliferate. Thus, in this model intracellular pH must fall below a critical value for apop tosis to occur. Together these data point to a step in cytokine depriv ation induced apoptosis (at least in some haemopoietic cell types) whi ch is either enhanced by or dependent upon an acidic intracellular env ironment which is the result of an increase in acid loading and inhibi tion of Na+/H+ exchange activity, One of the mechanisms by which activ ation of v-Abl protein tyrosine kinase suppresses apoptosis is by prev ention of intracellular acidification.