Modification of tumor necrosis factor-α (TNF-α) production by the Na+-dependent HCO3− cotransport in lipopolysaccharide-activated human monocytes

Abstract

Tumor necrosis factor-α (TNF-α) is produced and secreted from monocytes in response to activation with lipopolysaccharide (LPS). The role of Na+ and HCO3− in the production of TNF-α by monocytes was investigated; it was observed that replacement of Na+ in the culture medium with sucrose or choline chloride inhibited TNF-α production completely. The addition of Na+ to Na+ -free culture medium restored TNF-α production with an EC50 value of 35 mmol/l. The amiloride analog 5-(N-ethyl-N-isopropyl) amiloride (EIPA), an inhibitor of the Na+/H+ antiporter, inhibited TNF-α production with an EC50 of 3.3 μM. Without HCO3− in the culture medium TNF-α production was inhibited by 92%. Total protein synthesis was inhibited by 85% in the absence of Na+ but did not change in the absence of bicarbonate in the culture medium. Intracellular pH (pHi) which increased from 6.90 in control monocyte to 7.40 in response to activation with LPS was abrogated to pHi of 6.95 in the absence of Na+ but did not change in the absence of HCO3− in the culture medium. In the presence of 100 μM phloretin or DIDS the pHi of activated monocyte was reduced to control value, TNF-α production was inhibited completely and total protein synthesis was inhibited by 61%. These data suggest that (1) TNF-α production, as other proteins, is dependent on the pHi of monocytes, and (2) TNF-α production, in contrast to total protein, is modulated by Na+-dependent HCO3−