gw-501516 and sapropterin

Impaired endothelium-dependent relaxation (EDR) is a hallmark of endothelial dysfunction. A deficiency of tetrahydrobiopterin (BH. Gene expression was measured by using qRT-PCR and western blotting. Biopterins and ROS were determined by using HPLC. NO was measured with fluorescent dye and electron paramagnetic resonance spectroscopy. Vasorelaxation was measured by Multi Myograph System.. The PPARδ agonist GW501516 increased DHFR and BH. PPARδ prevented endothelial dysfunction by increasing DHFR and activating the BH

Topics: Animals; Aorta; Biopterins; Cells, Cultured; Endothelium, Vascular; Gene Expression; Human Umbilical Vein Endothelial Cells; Humans; Male; Mice, Inbred C57BL; Mice, Knockout; PPAR delta; Sulfones; Tetrahydrofolate Dehydrogenase; Thiazoles; Thiophenes; Thoracic Arteries

Existing evidence suggests that amyloid-β precursor protein (APP) causes endothelial dysfunction and contributes to pathogenesis of atherosclerosis. In the present study, experiments were designed to: (1) determine the mechanisms underlying endothelial dysfunction and (2) define the effects of peroxisome proliferator-activated receptor delta (PPARδ) ligand on endothelial function in transgenic Tg2576 mice overexpressing mutated human APP.. Confocal microscopy and western blot analyses of wild-type mice aortas provided evidence that APP protein is mainly present in endothelial cells. Overexpression of APP significantly impaired endothelium-dependent relaxations to acetylcholine and phosphorylation of endothelial nitric oxide synthase at Ser(1177) in aortas. HPLC analysis revealed that tetrahydrobiopterin (BH(4)) levels were reduced in Tg2576 mice aortas. This was caused by increased oxidation of BH(4) and reduced expression and activity of GTP-cyclohydrolase I. Furthermore, gp91phox protein expression and superoxide anion production were increased in aortas of Tg2576 mice. This augmented superoxide formation was completely prevented by the NADPH oxidase inhibitor VAS2870. Expression of copper-/zinc-superoxide dismutase (Cu/ZnSOD) and extracellular SOD was downregulated. Treatment with PPARδ ligand GW501516 (2 mg/kg/day) for 14 days significantly increased BH(4) bioavailability and improved endothelium-dependent relaxations in Tg2576 mice aortas. GW501516 also normalized protein expression of gp91(phox) and SODs, thereby reducing production of superoxide anion in the aortas.. Our results suggest that in APP transgenic mice loss of nitric oxide and increased oxidative stress are the major causes of endothelial dysfunction. The vascular protective effects of GW501516 in Tg2576 mice appear to be critically dependent on prevention of superoxide anion production.

Topics: Amyloid beta-Protein Precursor; Animals; Aorta; Biopterins; Blotting, Western; Chromatography, High Pressure Liquid; Dose-Response Relationship, Drug; Endothelium, Vascular; Enzyme Inhibitors; Female; GTP Cyclohydrolase; Humans; Membrane Glycoproteins; Mice; Mice, Transgenic; Microscopy, Confocal; Mutation; NADPH Oxidase 2; NADPH Oxidases; Nitric Oxide; Nitric Oxide Synthase Type III; Oxidative Stress; Phosphorylation; PPAR gamma; Prostaglandins; Serine; Superoxide Dismutase; Superoxides; Thiazoles; Vasodilation; Vasodilator Agents

Peroxisome proliferator-activated receptor delta (PPARδ) is ubiquitously expressed in the vasculature, including cerebral circulation. The role of PPARδ in metabolism of tetrahydrobiopterin (BH₄) has not been studied in the cerebral microvasculature. In the present study, the effects of PPARδ agonist GW501516 on uncoupling of endothelial nitric oxide synthase (eNOS) were determined in cerebral microvessels of BH₄-deficient hph-1 mice. Wild-type (B6CBA) and hph-1 mice were orally gavaged with a selective PPARδ activator, GW501516 (2 mg/kg/day) for 14 days, and thereafter, cerebral microvessels were isolated and studied. Treatment of hph-1 mice with GW501516 significantly reduced oxidation of BH₄ and increased the ratio of BH₄ to 7,8-BH₂ (P<0.05, n=6-9). Attenuation of L-NAME-inhibitable superoxide anion levels by GW501516 demonstrated that activation of PPARδ might prevent uncoupling of endothelial nitric oxide synthase (eNOS, P<0.05, n=6-9). Western blotting studies demonstrated that GW501516 selectively increased the endothelial expressions of CuZn superoxide dismutase (P<0.05, n=6-9) and catalase (P<0.05, n=6-8). PPARδ activation increased the total nitrite and nitrate (NO₂+NO₃) content in cerebral microvessels (P<0.05, n=6). Obtained results suggest that in vivo activation of PPARδ prevents eNOS uncoupling, restores bioavailability of NO and may help preserve endothelial function in the BH₄-deficient cerebral circulation.

Topics: Analysis of Variance; Animals; Biopterins; Blood Pressure; Cerebral Cortex; Enzyme Inhibitors; Gene Expression Regulation, Enzymologic; GTP Cyclohydrolase; In Vitro Techniques; Mice; Mice, Inbred BALB C; Mice, Transgenic; Microvessels; Mutation; NG-Nitroarginine Methyl Ester; Nitric Oxide Synthase Type III; Polycomb Repressive Complex 1; PPAR delta; Superoxides; Thiazoles