HSP27 PHOSPHORYLATION IS INDUCED IN ALVEOLAR MACROPHAGES EXPOSED TO CDO-COATED SILICA PARTICLES

Protein phosphorylation in bovine alveolar macrophages (BAM:) activate d by quartz dusts and metal oxide-coated silica particles was investig ated by means of two-dimensional electrophoresis (BD-PAGE) and densito metric (32)[P]-phosphate image analysis. BAM activity was monitored by determining generated superoxide anions and hydrogen peroxide, In vit ro stimulation of BAM with cadmium oxide-coated silica particles (LiC- CdO) resulted in characteristic time-dependent changes in ED-PAGE spot patterns, that were similar to the effects induced by 4 beta-phorbol- 12-myristate-13-acetate (PMA). Phosphorylation of two proteins with ap parent molecular masses of 29 and 42 kDa appeared as main signals in b oth LiC-CdO and in PMA treated BAM but with different kinetics. Phosph oprotein spot pp29 was identified as an isoelectric form of Hsp27 by m icrosequence and Western blot analysis. In contrast to PMA stimulation , LiC-CdO-induced Hsp27 and p42 phosphorylation did not correlate with the amount of generated reactive oxygen intermediates. Other potent B AM activators like quartz dust SIKRON F600 or VO-coated silica particl es did not show Hsp27 and p42 phosphorylation. LiC-CdO-mediated Hsp27 phosphorylation was inhibited by SE 203580 indicating that p38 MAP kin ase is the upstream mediator of the activated signaling pathway(s), wh ile MEK inhibitor PD 98059 had no effect. (C) 1998 Academic Press.