P144The role of prolyl-hydroxylase inhibition in rat aorta during ischemia/reperfusion

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Abstract

Purpose

Although, vascular grafts are frequently applicated in the cardiac surgery, the storage protocols need further improvement against ischemia/reperfusion (IR) injury. Endothelial dysfunction may result in postoperative graft failure and promote late graft vasculopathy. Low oxygen tension elicits a variety of complex cellular responses by altering the activity of many signaling pathways, such as the oxygen dependent prolyl-hyroxylase-domain containing enzymes (PHD). Reduction of PHD-activity during hypoxia leads to stabilisation and accumulation of hypoxia inducible factor 1-α, a transcription factor which regulates the expression of target genes in response to hypoxia. We investigated the effects of PHD-inhibitor dimethyloxalylglycine (DMOG) on the vasomotor responses of isolated rat aorta and aortic vascular smooth muscle cells (VSMC) in a model of cold ischemia/warm reperfusion.

Methods

Isolated rat aortic rings underwent a 24h cold ischemic preservation in NaCl or DMOG (10-5M) supplemented saline solution. In organ bath experiments we investigated endothelium-dependent and -independent vasorelaxation by using cumulative concentrations of acetylcholine (ACh) and sodium nitroprusside (SNP) on phenylephrine-precontracted isolated aortic rings with an additional external oxidant (hypochlorite, 200μM, 30min). VSMC under normoxic conditions are plated and 24h later the medium is changed to NaCl or DMOG (1mM) solution. The cells are incubated (4°C, 24h-cold ischemia) and after the medium is changed for a supplied standard medium (37°C, 6h-warm reperfusion). Different gene expression analysis was performed by quantitative real-time PCR.

Results

Compared with the control the NaCl-group showed a prominent endothelial dysfunction induced by hypoxia/reoxygenation, which was significantly improved by DMOG supplementation (maximal relaxation to ACh, control:90 ± 1% vs. NaCl:30 ± 6% vs. DMOG:62 ± 7%, p < 0.05). Following hypoxia/reoxygenation, concentration-response curves to SNP were significantly shifted to the right and DMOG has no effect (pD2, control:8.3 ± 0.1 vs. NaCl:8.0 ± 0.1 vs. DMOG:8.0 ± 0.1). Decreased mRNA-expression of heme oxygenase-1 in the NaCl-group as compared to control group was increased after DMOG-pretreatment (control:1.62 ± 0.11 vs. NaCl:0.50 ± 0.03 vs. DMOG:1.52 ± 0.40). The changes in the expression of vascular endothelial growth factor expression showed similar tendency (control:1.62 ± 0.14 vs. NaCl:0.37 ± 0.04 vs. DMOG:0.98 ± 0.23).

Conclusion

Inhibition of prolyl hyroxylase could be a new therapeutic avenue for protecting endothelium and vascular muscle cells against IR injury.

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