FMLP-induced formation of F-actin in HL60 cells is dependent on PI3-K but not on intracellular Ca2+, PKC, ERK or p38 MAPK

Objective and Design: To further understand the mechanisms of signal transd uction pathways for the formation of F-actin (polymerization of actin) and the activation of NADPH oxidase in phagocytic cells, the effects of various inhibitors on them were studied. Materials and Methods: Differentiated HL60 cells were studied to examine th eir N-formyl-methionyl-leucyl-phenyl-alanine (fMLP)-stimulated formation of F-actin and activation of NADPH oxidase following treatment with various i nhibitors. These included a protein kinase C (PKC) inhibitor (GF 109203X), a phosphatidylinositide 3 kinase (PI3-K) inhibitor (wortmannin), an extrace llular response kinase (ERK) inhibitor (PD 98059), a p38 mitogen-activated protein kinase (MAPK) inhibitor (SB 203580) and an intracellular Ca2+-chela tor (BAPTA-AM). Results: The treatment with wortmannin suppressed the formation of F-actin, with less suppression of the activation of NADPH oxidase. BAPTA-AM and GF 109203X did not attenuate the formation of F-actin but completely inhibited the activation of NADPH oxidase. PD 98059 and SE 203580 partially inhibite d the activation of NADPH oxidase without influence on the formation of F-a ctin. Furthermore, wortmannin but not BAPTA-AM and GF 109203X inhibited the fMLP-induced activation of Akt, which is known to regulate NADPH oxidase. Conclusions: These results suggest that the formation of F-actin is depende nt on PI3-K and independent of PKC, ERK and p38 MAPK as well as the increas e in intracellular Ca2+, whereas the activation of NADPH oxidase is partly dependent on ERK, p38 MAPK, Akt regulated by PI3-K, and strongly dependent on the activation of PKC and the increase in intracellular Ca2+.