Question:

The metabolic microenvironment in various diseases (tumor, inflammation, ischemia) often display pronounced acidosis that may influence signaling and gene expression. Hence the impact of metabolic acidosis on activation of MAP kinases ERK1/2 and p38 as well as on the expression of TNF-α (tumor necrosis factor), MCP-1 (monocyte chemotactic protein), COX-2 (cyclooxygenase), iNOS (inducible nitric oxide synthase) and osteopontin in rat fibroblasts (NRKF) was studied.

Methods:

Cells were incubated for 24 h with medium lacking FCS supplementation and thereafter for 3 h at pH 7.4 or pH 6.6 in bicarbonate-buffered Ringer solution. Activation of MAP kinases was analyzed using phosphorylation specific antibodies. Alterations in transcription were determined using qPCR (Platinum SYBR Green qPCR Supermix, Invitrogen) according to the manufacturer's instructions.

Results:

Extracellular acidosis led to an activation of ERK1/2 and p38. The expression of TNF-α, COX-2 and iNOS was elevated after 3 h at pH 6.6, while MCP-1 and osteopontin were not affected by acidosis. Increased COX-2 and iNOS expression depended on p38. ERK1/2 inhibition did not prevent transcriptional changes induced by acidosis, but resulted in a decrease in basal TNF-α and MCP-1 expression.

Conclusion:

The acidic microenvironment induces defined changes in cellular signaling and in the transcriptional program of disease-relevant genes. Part of the transcriptional effects of acidosis depends on p38, but not on ERK1/2 signaling.