Role for cAMP-protein kinase A signalling in augmented neutrophil adhesion and chemotaxis in sickle cell disease

Abstract

The significance of the leukocyte in sickle cell disease (SCD) pathophysiology is becoming increasingly recognised; we sought to examine whether the chemotactic properties of neutrophils of SCD individuals may be altered and, further, to better understand the signalling events that mediate altered SCD neutrophil function. Adhesion to immobilised fibronectin (FN) and chemotaxis of control and SCD neutrophils were assessed using in vitro static adhesion assays and 96-well chemotaxis chamber assays. Adhesion assays confirmed a significantly higher basal adhesion of SCD neutrophils to FN, compared with control neutrophils. Chemotaxis assays established, for the first time, that SCD neutrophils demonstrate greater spontaneous migration and, also, augmented migration in response to IL-8, when compared with control neutrophils. Co-incubation of SCD neutrophils with KT5720 (an inhibitor of PKA) abrogated increased basal SCD neutrophil adhesion, spontaneous chemotaxis and IL-8-stimulated chemotaxis. Stimulation of SCD neutrophils with IL-8 also significantly augmented SCD neutrophil adhesion to FN with a concomitant increase in cAMP levels and this increase in adhesion was abolished by KT5720. Interestingly, the adhesive properties of neutrophils from SCD individuals on hydroxyurea therapy were not significantly altered and results indicate that a reduction in intracellular cAMP may contribute to lower the adhesive properties of these cells. Data indicate that up-regulated cAMP signalling plays a significant role in the altered adhesive and migratory properties in SCD neutrophils. Such alterations may have important implications for the pathophysiology of the disease and the cAMP-PKA pathway may represent a therapeutic target for the abrogation of altered leukocyte function.