The role of dependent pathway in eosinophil apoptosis / 천식및알레르기

ABSTRACT

BACKGROUND: Interleukin-5 (IL-5), IL-3, and GM-CSF are known to prolong the survival of eosinophils, and IL-5 has the most potent effect on eosinophil survivaL It is also known that divergent signals induce apoptosis in different cells. But, There have been few reports on about the intracellular signals that trigger the effectors of apoptosis. Cyclic AMP (cAMP) can modulate apoptosis in many cells. But, the role of intracellular cAMP in the IL-5 induced eosinophil survival is still not completely understood. OBJECTIVES: This study was aimed to elucidate the role of intracellular cAMP in IL-5 induced eosinophil survival. MATERIAL AND METHOD: Eosinophils were isolated from peripheral blood of atopic patients. Eosinophil viability was measured by means of propidium iodine (PI) method and the number of viable cells was counted by FAC scan (Becton Dickinson, USA). Cells were cultured with or without IL-5, and also with various cAMP-elevating agents (dibutyryl cAMP, 8-bromo-cAMP, N6- benzoyl cAMP). The concentrations of cAMP were measured by cAMP enzyme immunoassay system(BiotrakTM, Amersham). Finally cAMP dependent protein kinase A inhibitor (H8) was added to eosinophils to examine the effect of decreased intracellular cAMP activity on the viability of eosinophils stimulated with IL-5. RESULTS: The percentage of viable eosinophils was reduced rapidly from 92.1+/-1.8% to 8.23+/- 3.41% without IL-5 (p<0.05; n=ll, 4-day incubation). Upon addition of IL-5, it was increased to 33.02+7.8% (p<0.05; n=ll). In the absence of IL-5, the addition of cAMP-elevating agent increased eosinophil viability in a dose-dependent manner. Upon addition of H8 (24 uM), the eosinophil viability increased by IL-5 (52.5+/-6.4%) was significantly reduced to 27.2+/-5.4% (p<0.05;n=7). Compared with tissue culture media (TCM) only, IL-5 produced persistent elevation of intracellular cAMP of eosinophils in a time and dose dependent manner.