Renal mesangial cells are regarded as main players in glomerular inflammatory diseases. To investigate a possible crosstalk between inflammatory and hypoxia-driven signaling processes, we stimulated cultured mouse mesangial cells with different inflammatory agents and analyzed the expression of prolyl hydroxylase domain containing proteins (PHDs), the main regulators of hypoxia-inducible factor (HIF) stability. Administration of IL-1β (1 nM) and TNF-α (1 nM), a combination further referred to as cytokine mix (CM), resulted in a fivefold increase in PHD3 but not PHD1 and PHD2 mRNA expression compared to untreated controls. In contrast, a combination of IL-1β, TNF-α with lipopolysaccharide (10 μg/ml), and interferon-γ (20 ng/ml) designated as CM+ showed a high (60-fold) induction of PHD3 and a moderate (twofold) induction of PHD2 mRNA expression. Interestingly, CM+ but not CM induced the expression of inducible NO synthase and endogenously produced NO was responsible for the immense induction of PHD3 in mesangial cells treated with CM+. We found that CM+ affected PHD3 expression mainly via the NO/HIF axis, whereas PHD3 regulation by CM occurred in a NF-κB-dependent manner. In turn, silencing of PHD3 expression resulted in a decrease in the mRNA expression of ICAM-1, MIP-2, MCP-1, and CXCL-10, which are under control of NF-κB. In a rat model of mesangio-proliferative glomerulonephritis, PHD3 mRNA and protein expression was markedly induced and this effect was nearly abolished when rats were treated with the iNOS-specific inhibitor L-NIL, thus confirming our findings also in vivo.
Key message: PHD3 expression induced by cytokines is NF-κB dependent in mesangial cells. Endogenously produced NO further augments PHD3 expression via HIF-1α. PHD3 expression is induced by NO in anti-Thy-1 glomerulonephritis.
Keywords: Mesangial cells; Nitric oxide; Prolyl hydroxylase domain containing protein 3.