repeated observation that it is altered by a variety of pathophysiological conditions. Such alterations of NO production and/or bioavailability have been shown to occur both in experimental animal models and in human subjects, in the setting of such diverse disorders as hypertension, hypercholesterolemia, aging, cigarette smoking, diabetes, and heart failure (1). The mechanisms underlying the alteration of this important function of the endothelium are varied and likely multifactorial. During the past several years an enormous amount of research has been devoted to understanding these abnormalities, which has led to new insights into regulation of vascular tone, redox state, inflammation, growth, and the prothrombotic/antithrombotic properties of the vessel wall. This Perspective will highlight some of these important new observations, as they relate to the pathology of the endothelial cell L -arginine/NO synthase (NOS) system. In addition, future directions of research that may be particularly informative will be indicated. Alterations of the substrate for the NOS enzyme The substrate for NOS is the basic amino acid L -arginine (with a K m of approximately 5 m M, reference 2). L -Arginine is syn

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Cellular and molecular mechanisms of endothelial cell dysfunction.

Tetrahydrobiopterin (BH % ) cofactor is essential for various processes, and is present in probably every cell or tissue of higher organisms. BH % is required for various enzyme activities, and for less defined functions at the cellular level. The pathway for the de noao biosynthesis of BH % from GTP involves GTP cyclohydrolase I, 6-pyruvoyl-tetrahydropterin synthase and sepiapterin reductase. Cofactor regeneration requires pterin-4a-carbinolamine dehydratase and dihydropteridine reductase. Based on gene cloning, recombinant expression, mutagenesis studies, structural analysis of crystals and NMR studies, reaction mechanisms for the biosynthetic and recycling enzymes were proposed. With regard to the regulation of cofactor biosynthesis, the major controlling point is GTP cyclohydrolase I, the expression of which may be under the control of cytokine induction. In the liver at least, activity is inhibited by BH % , but stimulated by phenylalanine through the GTP cyclohydrolase I feedback regulatory protein. The enzymes that depend on BH % are the phenylalanine, tyrosine and tryptophan hydroxylases, the latter

/pdf/tetrahydrobiopterin-biosynthesis-regeneration-and-functions-d9u3xq4b8n.pdf

Tetrahydrobiopterin biosynthesis, regeneration and functions.

Nitric oxide (NO) signalling has pleiotropic roles in biology and a crucial function in cardiovascular homeostasis. Tremendous knowledge has been accumulated on the mechanisms of the nitric oxide synthase (NOS)-NO pathway, but how this highly reactive, free radical gas signals to specific targets for precise regulation of cardiovascular function remains the focus of much intense research. In this Review, we summarize the updated paradigms on NOS regulation, NO interaction with reactive oxidant species in specific subcellular compartments, and downstream effects of NO in target cardiovascular tissues, while emphasizing the latest developments of molecular tools and biomarkers to modulate and monitor NO production and bioavailability.

Nitric oxide signalling in cardiovascular health and disease

BH4 (6R-L-erythro-5,6,7,8-tetrahydrobiopterin) is an essential cofactor of a set of enzymes that are of central metabolic importance, including four aromatic amino acid hydroxylases, alkylglycerol mono-oxygenase and three NOS (NO synthase) isoenzymes. Consequently, BH4 is present in probably every cell or tissue of higher organisms and plays a key role in a number of biological processes and pathological states associated with monoamine neurotransmitter formation, cardiovascular and endothelial dysfunction, the immune response and pain sensitivity. BH4 is formed de novo from GTP via a sequence of three enzymatic steps carried out by GTP cyclohydrolase I, 6-pyruvoyltetrahydropterin synthase and sepiapterin reductase. An alternative or salvage pathway involves dihydrofolate reductase and may play an essential role in peripheral tissues. Cofactor regeneration requires pterin-4a-carbinolamine dehydratase and dihydropteridine reductase, except for NOSs, in which the BH4 cofactor undergoes a one-electron redox cycle without the need for additional regeneration enzymes. With regard to the regulation of cofactor biosynthesis, the major controlling point is GTP cyclohydrolase I. BH4 biosynthesis is controlled in mammals by hormones and cytokines. BH4 deficiency due to autosomal recessive mutations in all enzymes, except for sepiapterin reductase, has been described as a cause of hyperphenylalaninaemia. A major contributor to vascular dysfunction associated with hypertension, ischaemic reperfusion injury, diabetes and others, appears to be an effect of oxidized BH4, which leads to an increased formation of oxygen-derived radicals instead of NO by decoupled NOS. Furthermore, several neurological diseases have been suggested to be a consequence of restricted cofactor availability, and oral cofactor replacement therapy to stabilize mutant phenylalanine hydroxylase in the BH4-responsive type of hyperphenylalaninaemia has an advantageous effect on pathological phenylalanine levels in patients.

Tetrahydrobiopterin: biochemistry and pathophysiology.

How does ascorbic acid prevent endothelial dysfunction

The in vitro antioxidative activity of 5,6,7,8-tetrahydrobiopterin (BPH4) was measured and the ability of BPH4 to prevent paraquat-induced cell damage was examined in cultured hepatocytes. The scavenging activity of BPH4 against superoxide anion radicals was assayed in two systems, i.e., xanthine/xanthine oxidase (X/XOD) and rat macrophage/phorbol myristate acetate (MξPMA) radical-generating systems. BPH4 showed an extremely strong superoxide anion radical-scavenging activity in both assay systems. Biopterin (BP) itself did not show any activity in the X/XOD system, but was effective in the MξPMA system. The antioxidative activities of BPH4 against both superoxide anion and hydroxyl radicals were confirmed by spin trapping-ESR spectrometry. BPH4 also protected rat brain homogenate against auto-oxidation. We further examined the effect of BPH4 on paraquat-induced cell toxicity in cultured rat hepatocytes. The paraquat-induced elevation of the release of lactate dehydrogenase (LDH), a marker enzyme for cyto...

S. Ona

Papers

Antioxidative activity of 5,6,7,8-tetrahydrobiopterin and its inhibitory effect on paraquat-induced cell toxicity in cultured rat hepatocytes.