Transcellular transport of cobalamin in aortic endothelial cells.

Cobalamin [Cbl (or B)] deficiency causes megaloblastic anemia and a variety of neuropathies. However, homeostatic mechanisms of cyanocobalamin (CNCbl) and other Cbls by vascular endothelial cells are poorly understood. Herein, we describe our investigation into whether cultured bovine aortic endothelial cells (BAECs) perform transcytosis of B, namely, the complex formed between serum transcobalamin and B, designated as holo-transcobalamin (holo-TC).

HDL from apoA1 transgenic mice expressing the 4WF isoform is resistant to oxidative loss of function.

HDL functions are impaired by myeloperoxidase (MPO), which selectively targets and oxidizes human apoA1. We previously found that the 4WF isoform of human apoA1, in which the four tryptophan residues are substituted with phenylalanine, is resistant to MPO-mediated loss of function. The purpose of this study was to generate 4WF apoA1 transgenic mice and compare functional properties of the 4WF and wild-type human apoA1 isoforms in vivo.

Proteomics of vitamin B12 processing.

The causes of cobalamin (B12, Cbl) deficiency are multifactorial. Whether nutritional due to poor dietary intake, or functional due to impairments in absorption or intracellular processing and trafficking events, the major symptoms of Cbl deficiency include megaloblastic anemia, neurological deterioration and in extreme cases, failure to thrive and death. The common biomarkers of Cbl deficiency (hyperhomocysteinemia and methylmalonic acidemia) are extremely valuable diagnostic indicators of the condition, but little is known about the changes that occur at the protein level.

Both selenium deficiency and modest selenium supplementation lead to myocardial fibrosis in mice via effects on redox-methylation balance.

Scope: Selenium has complex effects in vivo on multiple homeostatic mechanisms such as redox balance, methylation balance, and epigenesis, via its interaction with the methionine-homocysteine cycle. In this study, we examined the hypothesis that selenium status would modulate both redox and methylation balance and thereby modulate myocardial structure and function.

Methods And Results: We examined the effects of selenium-deficient (<0.

Disulfide bond as a switch for copper-zinc superoxide dismutase activity in asthma.

Aim: Loss of superoxide dismutase (SOD) activity is a defining biochemical feature of asthma. However, mechanisms for the reduced activity are unknown. We hypothesized that loss of asthmatic SOD activity is due to greater susceptibility to oxidative inactivation.

The MMACHC proteome: hallmarks of functional cobalamin deficiency in humans.

Cobalamin (Cbl, B(12)) is an essential micronutrient required to fulfill the enzymatic reactions of cytosolic methylcobalamin-dependent methionine synthase and mitochondrial adenosylcobalamin-dependent methylmalonyl-CoA mutase. Mutations in the MMACHC gene (cblC complementation group) disrupt processing of the upper-axial ligand of newly internalized cobalamins, leading to functional deficiency of the vitamin. Patients with cblC disease present with both hyperhomocysteinemia and methylmalonic acidemia, cognitive dysfunction, and megaloblastic anemia.

The nutrigenetics of hyperhomocysteinemia: quantitative proteomics reveals differences in the methionine cycle enzymes of gene-induced versus diet-induced hyperhomocysteinemia.

Hyperhomocysteinemia has long been associated with atherosclerosis and thrombosis and is an independent risk factor for cardiovascular disease. Its causes include both genetic and environmental factors. Although homocysteine is produced in every cell as an intermediate of the methionine cycle, the liver contributes the major portion found in circulation, and fatty liver is a common finding in homocystinuric patients.

Exogenous thioredoxin prevents ethanol-induced oxidative damage and apoptosis in mouse liver.

Unlabelled: Ethanol-induced liver injury is characterized by increased formation of reactive oxygen species (ROS) and inflammatory cytokines, resulting in the development of hepatic steatosis, injury, and cell death by necrosis and apoptosis. Thioredoxin (Trx), a potent antioxidant and antiinflammatory molecule with antiapoptotic properties, protects animals from a number of inflammatory diseases. However, the effects of ethanol on Trx or its role in ethanol-induced liver injury are not known.

Hyperhomocysteinemia induced by methionine supplementation does not independently cause atherosclerosis in C57BL/6J mice.

A causal relationship between diet-induced hyperhomocysteinemia (HHcy) and accelerated atherosclerosis has been established in apolipoprotein E-deficient (apoE(-/-)) mice. However, it is not known whether the proatherogenic effect of HHcy in apoE(-/-) mice is independent of hyperlipidemia and/or deficiency of apoE. In this study, a comprehensive dietary approach using C57BL/6J mice was used to investigate whether HHcy is an independent risk factor for accelerated atherosclerosis or dependent on additional dietary factors that increase plasma lipids and/or inflammation.

Elevated homocysteine, glutathione and cysteinylglycine concentrations in patients homozygous for the Chuvash polycythemia VHL mutation.

In Chuvash polycythemia, homozygous von Hippel-Lindau (VHL) 598C>T leads to increased hypoxia inducible factor-1alpha and 2alpha, thromboses and lower systemic blood pressures. Circulating homocysteine, glutathione, gamma-glutamyltransferase and cysteinylglycine concentrations were higher in 34 VHL598C>T homozygotes than in 37 normal controls and cysteine was lower. Multivariate analysis showed elevated homocysteine independently associated with higher mean systemic blood pressures and elevated glutathione was associated with lower pressures to a similar degree.

Targeting of metallothionein by L-homocysteine: a novel mechanism for disruption of zinc and redox homeostasis.

Objective: L-homocysteine and/or L-homocystine interact in vivo with albumin and other extracellular proteins by forming mixed-disulfide conjugates. Because of its extremely rich cysteine content, we hypothesized that metallothionein, a ubiquitous intracellular zinc-chaperone and superoxide anion radical scavenger, reacts with L-homocysteine and that homocysteinylated-metallothionein suffers loss of function.

Methods And Results: 35S-homocysteinylated-metallothionein was resolved in lysates of cultured human aortic endothelial cells in the absence and presence of reduced glutathione by SDS-PAGE and identified by Western blotting and phosphorimaging.

Homocysteine transport by human aortic endothelial cells: identification and properties of import systems.

Hyperhomocysteinemia is an independent risk factor for cardiovascular disease. Transport of L-homocysteine into and out of the human vascular endothelium is poorly understood. We hypothesized that cultured human aortic endothelial cells (HAEC) would import L-homocysteine on one or more of the L-cysteine transport systems.

Molecular targeting by homocysteine: a mechanism for vascular pathogenesis.

Hyperhomocysteinemia is an independent risk factor for cardiovascular disease. Although there is a growing body of evidence that homocysteine plays a causal role in atherogenesis, specific mechanisms to explain the underlying pathology have remained elusive. This review focuses on chemistry unique to the homocysteine molecule to explain its inherent cytotoxicity.

Modulation of cystathionine beta-synthase level regulates total serum homocysteine in mice.

Elevated total plasma homocysteine is an independent risk factor in the development of vascular disease in humans. Cystathionine beta-synthase (CBS) is an enzyme that condenses homocysteine with serine to form cystathionine. In this article, we describe the effects of modulating CBS activity using a transgenic mouse that contains the human CBS cDNA under control of the zinc-inducible metallothionein promoter (Tg-CBS).

The circulatory half-lives of alpha-profibrin and alpha-fibrin monomer, and comparisons with other fibrin(ogen) derivatives.

Previous studies showed that alpha-fibrin monomer (lacking both A-fibrinopeptides, FPA) is normally cleared from the circulation before it assembles into a clot. Recent studies indicate that substantial quantities of an intermediate, alpha-profibrin lacking only one of the two FPA are produced in the course of conversion of human fibrinogen to fibrin. Since clearance of the alpha-fibrin monomer is saturable and receptor mediated, the extent to which alpha-profibrin or other fibrin(ogen) derivatives might compete for monomer uptake was deemed important.

Allosteric effects potentiating the release of the second fibrinopeptide A from fibrinogen by thrombin.

Fibrin formation depends on the release of the two N-terminal fibrinopeptides A (FPA) from fibrinogen, and its formation is accompanied by an intermediate, alpha-profibrin, which lacks only one of the FPA. In this study, we confirm that the maximal levels of alpha-profibrin found over the course of thrombin reactions with human fibrinogen are only half of what would be expected if the first and second FPA were being released independently with equal rate constants. The rapidity of release of the fibrinopeptides by thrombin had been shown to depend on an allosteric transformation that is induced when Na(+) binds to a site defined by the 215-227 residues of thrombin, a transformation that results in the exposure of its fibrinogen-binding exosites transforming the thrombin from a slow to a fast acting form toward fibrinogen.