Inhibition of prolyl hydroxylase domain-containing protein suppressed lipopolysaccharide-induced TNF-alpha expression.

Objective: Prolyl hydroxylase domain-containing proteins (PHDs) play pivotal roles in oxygen-sensing system through the regulation of alpha-subunit of hypoxia-inducible factor (HIF), a key transcription factor governing a large set of gene expression to adapt hypoxia. Although tissue hypoxia plays an essential role in maintaining inflammation, the role of PHDs in the inflammatory responses has not been clearly determined. Here, we investigated the role of PHDs in lipopolysaccharide (LPS)-induced tumor necrosis factor alpha (TNF-alpha) induction in macrophages.

Methods And Results: Northern blot analysis and ELISA revealed that LPS-induced TNF-alpha upregulation was strongly suppressed by PHD inhibitors, dimethyloxallyl glycine (DMOG), and TM6008 in RAW264.7 macrophages. DMOG suppressed LPS-induced TNF-alpha upregulation in HIF-1alpha-depleted cells and HIF-1alpha overexpression failed to suppress the induction of TNF-alpha. DMOG rather suppressed LPS-induced NF-kappaB transcriptional activity. Downregulation of Phd1 or Phd2 mRNA by RNA interference partially attenuated LPS-induced TNF-alpha induction. DMOG also inhibited LPS-induced TNF-alpha production in peritoneal macrophages as well as human macrophages.

Conclusions: PHD inhibition by DMOG or RNA interference inhibited LPS-induced TNF-alpha upregulation in macrophages possibly through NF-kappaB inhibition, which is independent of HIF-1alpha accumulation. This study suggests that PHDs are positive regulators of LPS-induced inflammatory process, and therefore inhibition of PHD may be a novel strategy for the treatment of inflammatory diseases.