Prolyl hydroxylase inhibitors increase the production of vascular endothelial growth factor by periodontal fibroblasts

Agis H, Watzek G, Gruber R. Prolyl hydroxylase inhibitors increase the production of vascular endothelial growth factor by periodontal fibroblasts. J Periodont Res 2012; 47: 165–173. © 2011 John Wiley & Sons A/S Background and Objective: Pharmacological inhibitors of prolyl hydroxylases (PHDs) can induce a proangiogenic response that favors wound healing and bone regeneration. However, the response of periodontal cells to PHD inhibitors is unknown. Material and Methods: To determine the effects of PHD inhibitors on periodontal cells, we exposed human fibroblasts from the gingiva and the periodontal ligament to dimethyloxallyl glycine, desferrioxamine, l ‐mimosine and CoCl₂. Viability, proliferation, and protein synthesis were assessed by 3‐(4,5‐dimethylthiazol‐2‐yl)‐2,5‐diphenyltetrazolium bromide (MTT), ³H]thymidine, and ³H]leucine incorporation, respectively. The levels of Ki67, hypoxia‐inducible factor 1α (HIF‐1α), p27, phosphorylated c‐Jun N‐terminal kinase (JNK) and phosphorylated p38 were determined by immunohistochemistry and western blotting. Vascular endothelial growth factor (VEGF) mRNA levels were measured by quantitative PCR. Protein levels of VEGF and interleukin (IL)‐6 were evaluated by immunoassays. Results: We found that PHD inhibitors, while leaving cell viability unchanged, reduced proliferation and protein synthesis. This was paralleled by decreased Ki67 levels and increased p27 levels, suggesting that PHD inhibitors provoke growth arrest. Independently from this response, PHD inhibitors stabilized HIF‐1α and increased the production of VEGF. This increase of VEGF was observed in the presence of proinflammatory IL‐1 and pharmacological inhibitors of JNK and p38 signaling. Moreover, PHD inhibitors did not modulate expression of IL‐6 and the phosphorylation of JNK and p38. Conclusion: These results suggest that PHD inhibitors enhance the production of VEGF in periodontal fibroblasts, even in the presence of proinflammatory IL‐1. The data further suggest that PHD inhibitors do not provoke a significant proinflammatory or anti‐inflammatory response in this in vitro setting.