Effect of a selective protein kinase C inhibitor, Ro 31-8425, on Mac-1 expression and adhesion of human neutrophils

The role of protein kinase C (PKC) in mediating up-regulation of macrophage 1 adhesion protein (Mac-1) and adhesion of neutrophils in response to physiological agonists is not clear. Previous studies have relied on use of phorbol esters to activate PKC directly or on results obtained with non-selective inhibitors of protein kinases. 3-[8-(Aminomethyl)-6,7,8,9-tetrahydropyridol[1,2-a]-indol-10-yl]-4 -(1- methyl-3-indolyl)-1H-pyrrole-2,5-dione hydrochloride (Ro 31-8425) is a potent and highly selective inhibitor of PKC (Bit et al. J. Med. Chem. 1993. 36: 21). In these studies Ro 31-8425 has been used to define, more definitively, the role of PKC in mediating complement fragment C5a (C5a)-stimulated up-regulation of Mac-1 and adhesion of neutrophils to endothelial cells and to bovine serum albumin (BSA)-coated plastic. Phorbol 12, 13 dibutyrate (PBu2) increased surface expression of Mac-1 and stimulated adhesion of neutrophils to endothelial cells and to BSA-coated plastic. This confirms previous reports that activation of PKC can stimulate these responses. The PKC inhibitor, Ro 31-8425, inhibited the PBu2-stimulated responses, which confirms that Ro 31-8425 was effective in inhibiting PKC in these neutrophils. A more physiological agonist, C5a, also increased surface expression of Mac-1 and adhesion of neutrophils to endothelial cells and BSA-coated plastic. However, the responses to C5a were unaffected by Ro 31-8425. These results suggest that, although activation of PKC can promote up-regulation of Mac-1 and adhesion of neutrophils, this does not appear to be the physiological pathway. A non-selective protein kinase inhibitor, staurosporine, inhibited both PBu2 and C5a-stimulated adhesion. This suggests that a protein kinase other than PKC, possibly a tyrosine protein kinase, is likely to be involved in mediating C5a-stimulated Mac-1 up-regulation and adhesion. These results emphasise the need for caution in interpreting experiments and assuming a role for PKC. Use of a potent and selective inhibitor of PKC, Ro 31-8425, provides more definitive information.