The p38 Mitogen-Activated Protein Kinase Pathway Is Involved in the Regulation of Heme Oxygenase-1 by Acidic Extracellular pH in Aortic Smooth Muscle Cells

Extracellular acidosis (EA) regulates Heme Oxygenase-1 (HO-1) expression in vascular smooth muscle cells via transcriptional and posttranscriptional mechanisms but the signaling pathways involved are not known. We examined the role of Mitogen-Activated Protein Kinase (MAPK) pathways in HO-1 regulation by EA. Primary rat aortic smooth muscle cells were exposed to EA or physiologic pH. Levels of the total and phosphorylated forms of p38, extracellular signal-regulated protein kinases 1/2 (ERK 1/2), c-Jun N-terminal kinases/stress-activated protein kinases (JNK 1/2), and HO-1 protein were assessed by Western analysis and HO-1 mRNA levels were assessed by quantitative PCR. Inhibition of p38 MAPK was achieved with the chemical inhibitor SB203580, or adenoviral infection of a dominant-negative form of p38 alpha. Phospho p38 MAPK activity was evaluated with an in vitro kinase activity assay. Binding of Activator Protein-1 (AP-1), a known target of MAPK pathways, was assessed by Electromobility shift assay (EMSA). EA induced phosphorylation of p38 MAPK in a biphasic manner while total p38 was unchanged. EA did not alter levels of phospho ERK 1/2 and phospho JNK 1/2. There was increased phospho p38 MAPK activity in the setting of EA which preceded the induction of HO-1. Inhibition of phospho p38 activity with either SB20358 or a dominant negative p38 alpha oligonucleotide abrogated the induction of HO-1 by EA. Increased specific binding of AP-1 in the setting of EA was shown by EMSA. Increased phospho p38 activity precedes and likely mediates HO-1 induction by EA. Increased AP-1 binding may underlie the transcriptional regulation of HO-1 by EA. J. Cell. Biochem. 105: 1298-1306, 2008. (c) 2008 Wiley-Liss, Inc.