The aim of this study is to determine whether heat-induced heat shock gene expression in rat pulmonary alveolar macrophages (PAMs) would protect PAMs against hydrogen peroxide (H2O2)-mediated cell killing. In response to sublethal heat shock at 42 degrees C for 2 h, the cells synthesized heat shock protein 70 kD (HSP70) and other different molecular weight heat shock proteins, which were detected with gel electrophoresis after [35S]-methionin labeling cellular protein and Western blotting analysis in PAMs. Northern blot analysis showed the induction of HSP70 mRNA with heat shock treatment. Futhermore, with heat shock, there was a significant increment of survival cells after H2O2 (1,2,3 mmol.L-1, 45 min) exposure. This increase was blocked both by Cycloheximide and by Actinomycin D, and was associated with inhibition of synthesis of HSP70 protein and transcription of HSP70 mRNA. These results strongly suggest that expression of heat shock gene (especially HSP70 gene) plays an important role in the intracelluar mechanism of cytoprotection against H2O2 in rat PAMs.