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Roles for endothelial cell and macrophage Gch1 and tetrahydrobiopterin in atherosclerosis progression.

ABSTRACT: Aims:GTP cyclohydrolase I catalyses the first and rate-limiting reaction in the synthesis of tetrahydrobiopterin (BH4), an essential cofactor for nitric oxide synthases (NOS). Both eNOS and iNOS have been implicated in the progression of atherosclerosis, with opposing effects in eNOS and iNOS knockout mice. However, the pathophysiologic requirement for BH4 in regulating both eNOS and iNOS function, and the effects of loss of BH4 on the progression of atherosclerosis remains unknown. Methods and results:Hyperlipidemic mice deficient in Gch1 in endothelial cells and leucocytes were generated by crossing Gch1fl/flTie2cre mice with ApoE-/- mice. Deficiency of Gch1 and BH4 in endothelial cells and myeloid cells was associated with mildly increased blood pressure. High fat feeding for 6?weeks in Gch1fl/flTie2CreApoE-/- mice resulted in significantly decreased circulating BH4 levels, increased atherosclerosis burden and increased plaque macrophage content. Gch1fl/flTie2CreApoE-/- mice showed hallmarks of endothelial cell dysfunction, with increased aortic VCAM-1 expression and decreased endothelial cell dependent vasodilation. Furthermore, loss of BH4 from pro-inflammatory macrophages resulted in increased foam cell formation and altered cellular redox signalling, with decreased expression of antioxidant genes and increased reactive oxygen species. Bone marrow chimeras revealed that loss of Gch1 in both endothelial cells and leucocytes is required to accelerate atherosclerosis. Conclusion:Both endothelial cell and macrophage BH4 play important roles in the regulation of NOS function and cellular redox signalling in atherosclerosis.

SUBMITTER: Douglas G

PROVIDER: S-EPMC6054219 | biostudies-literature | 2018 Aug

REPOSITORIES: biostudies-literature

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Roles for endothelial cell and macrophage Gch1 and tetrahydrobiopterin in atherosclerosis progression.

Douglas Gillian G Hale Ashley B AB Patel Jyoti J Chuaiphichai Surawee S Al Haj Zen Ayman A Rashbrook Victoria S VS Trelfa Lucy L Crabtree Mark J MJ McNeill Eileen E Channon Keith M KM

Cardiovascular research 20180801 10

<h4>Aims</h4>GTP cyclohydrolase I catalyses the first and rate-limiting reaction in the synthesis of tetrahydrobiopterin (BH4), an essential cofactor for nitric oxide synthases (NOS). Both eNOS and iNOS have been implicated in the progression of atherosclerosis, with opposing effects in eNOS and iNOS knockout mice. However, the pathophysiologic requirement for BH4 in regulating both eNOS and iNOS function, and the effects of loss of BH4 on the progression of atherosclerosis remains unknown.<h4>M ...[more]

PMID: 29596571

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Project description:Inducible nitric oxide synthase (iNOS) is a key enzyme in the macrophage inflammatory response, which is the source of nitric oxide (NO) that is potently induced in response to proinflammatory stimuli. However, the specific role of NO production, as distinct from iNOS induction, in macrophage inflammatory responses remains unproven. We have generated a novel mouse model with conditional deletion of Gch1, encoding GTP cyclohydrolase 1 (GTPCH), an essential enzyme in the biosynthesis of tetrahydrobiopterin (BH4) that is a required cofactor for iNOS NO production. Mice with a floxed Gch1 allele (Gch1(fl/fl)) were crossed with Tie2cre transgenic mice, causing Gch1 deletion in leukocytes (Gch1(fl/fl)Tie2cre). Macrophages from Gch1(fl/fl)Tie2cre mice lacked GTPCH protein and de novo biopterin biosynthesis. When activated with LPS and IFN?, macrophages from Gch1(fl/fl)Tie2cre mice induced iNOS protein in a manner indistinguishable from wild-type controls, but produced no detectable NO, as judged by L-citrulline production, EPR spin trapping of NO, and by nitrite accumulation. Incubation of Gch1(fl/fl)Tie2cre macrophages with dihydroethidium revealed significantly increased production of superoxide in the presence of iNOS expression, and an iNOS-independent, BH4-dependent increase in other ROS species. Normal BH4 levels, nitric oxide production, and cellular redox state were restored by sepiapterin, a precursor of BH4 production by the salvage pathway, demonstrating that the effects of BH4 deficiency were reversible. Gch1(fl/fl)Tie2cre macrophages showed only minor alterations in cytokine production and normal cell migration, and minimal changes in basal gene expression. However, gene expression analysis after iNOS induction identified 78 genes that were altered between wild-type and Gch1(fl/fl)Tie2cre macrophages. Pathway analysis identified decreased NRF2 activation, with reduced induction of archetypal NRF2 genes (gclm, prdx1, gsta3, nqo1, and catalase) in BH4-deficient Gch1(fl/fl)Tie2cre macrophages. These findings identify BH4-dependent iNOS regulation and NO generation as specific requirements for NRF2-dependent responses in macrophage inflammatory activation.

Dataset Information

Roles for endothelial cell and macrophage Gch1 and tetrahydrobiopterin in atherosclerosis progression.

Publications

Roles for endothelial cell and macrophage Gch1 and tetrahydrobiopterin in atherosclerosis progression.

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