Homocysteine uptake by human umbilical vein endothelial cells in culture

Ewadh, Mufeed J.; Tudball, N.; Rose, F. A.

doi:10.1016/0167-4889(90)90097-w

Cited by 34 publications

(28 citation statements)

References 16 publications

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“…Because the latter requires 5-methyltetrahydrofolate as a methyl donor, the increase of NMDMC mRNA would contribute to activation of the remethylation pathway. This speculation is supported by the previous reports of existence of the homocysteine-uptake system (36) and homocysteine metabolic enzymes (37) in cultured EC. It would be a reasonable assumption that the same remethylation pathway would be activated in the cells of homocysteinemia patients.…”

Section: Identification Of Homocysteine-respondent Genes In Huvecsupporting

confidence: 76%

Homocysteine-respondent Genes in Vascular Endothelial Cells Identified by Differential Display Analysis

Kokame

Kato

Miyata

1996

Journal of Biological Chemistry

333

266

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An elevated blood level of homocysteine is associated with arteriosclerosis and thrombosis. The mechanisms by which homocysteine may promote vascular diseases have not been elucidated yet. In the present study, we have applied a modified nonradioactive differential display analysis to evaluate changes in gene expression induced by homocysteine treatment of cultured human umbilical vein endothelial cells (HUVEC). We identified six up-regulated and one down-regulated genes. One upregulated gene was GRP78/BiP, a stress protein, suggesting that misfolded proteins would accumulate in the endoplasmic reticulum because of redox potential changes caused by homocysteine. Another up-regulated gene encoded a bifunctional enzyme with activities of methylenetetrahydrofolate dehydrogenase and methenyltetrahydrofolate cyclohydrolase, which is involved in a homocysteine metabolism. A third up-regulated gene encoded activating transcription factor 4, and a fourth was a gene whose function is not identified yet. The remaining three were novel genes. We isolated a fulllength cDNA of one of the up-regulated genes from a HUVEC library. It encoded a novel protein with 394 amino acids, which was termed reducing agents and tunicamycin-responsive protein (RTP). Northern blot analysis revealed that RTP gene expression was induced in HUVEC after 4 h incubation with homocysteine. RTP mRNA was also observed in unstimulated cells and induced by not only homocysteine but also 2-mercaptoethanol and tunicamycin. The mRNA was ubiquitously expressed in human tissues. These observations indicate that homocysteine can alter the expressivity of multiple genes, including a stress protein and several novel genes. These responses may contribute to atherogenesis.

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Section: Identification Of Homocysteine-respondent Genes In Huvecsupporting

confidence: 76%

Homocysteine-respondent Genes in Vascular Endothelial Cells Identified by Differential Display Analysis

Kokame

Kato

Miyata

1996

Journal of Biological Chemistry

333

266

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“…Our data agree with this in that D-leucine has little influence on L-cystinemediated intracellular S-nitrosothiol formation, and D-cysteine has less effect than L-cysteine. L-homocysteine uptake has been shown to occur by those systems that transport L-cysteine, including system L and ASC in human umbilical vein endothelial cells (36). We found that addition of D͞L-homocysteine to HBSS enhances GSNO-stimulated intracellular S-nitrosothiol formation, suggesting that S-nitrosated homocysteine can also be transported into the cell through similar mechanisms as LCysNO.…”

Section: Discussionmentioning

confidence: 63%

The mechanism of transmembraneS-nitrosothiol transport

Zhang

Hogg

2004

Proc. Natl. Acad. Sci. U.S.A.

164

191

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S-nitrosothiols have been suggested to play an important role in nitric oxide (NO)-mediated biological events. However, the mechanisms by which an S-nitrosothiol (or the S-nitroso functional group) is transferred across cell membrane are still poorly understood. We have demonstrated previously that the degradation of S-nitrosoglutathione (GSNO) by cells absolutely required the presence of cystine in the extracellular medium and proposed a mechanism that involved the reduction of cystine to cysteine, followed by the reaction of cysteine with GSNO to form S-nitrosocysteine (CysNO), mixed disulfides, and nitrosyl anion. In the present study we have assessed the effect of cystine on the transfer of the S-nitroso functional group from the extracellular to the intracellular space. Using RAW 264.7 cells, we found that the presence of L-cystine enhanced GSNO-dependent S-nitrosothiol uptake, increasing the intracellular S-nitrosothiol level from Ϸ60 pmol͞mg of protein to Ϸ3 nmol͞mg of protein. A lthough S-nitrosothiols have been thought to play an important role in many nitric oxide (NO)-mediated biological events, the processes by which S-nitrosothiol or their bound S-nitroso groups are transferred between the extracellular and intracellular spaces are still poorly understood. Transport mechanisms involving cell-surface protein disulfide isomerase (1, 2), ␥-glutamyl transpeptidase (3, 4), or anion exchanger (5) have been proposed.Previous studies have shown that some cellular effects of either S-nitrosoglutathione (GSNO) or S-nitroso-N-acetylpenicillamine (SNAP) depend on the presence of L-cysteine (6-9). It was suggested that cysteine can increase the efficiency of NO release from GSNO by the formation of unstable Snitrosocysteine (CysNO) through a transnitrosation reaction (9). Increases in cytosolic GSNO and cysteine levels were observed during the incubation of CysNO with human red blood cells (RBCs), implying the transport of CysNO into RBCs and the transnitrosation between CysNO and cytosolic glutathione (GSH) (10). Mallis and Thomas (8) indicated that a stereospecific transporter is not involved in the direct transfer of CysNO across the cell membrane, because the combination of GSNO with L-cysteine or D-cysteine gave a similar CysNO-like effect. However, other studies indicate that amino acid transporter system L (L-AT) is involved in the uptake of CysNO by cells (7,(11)(12)(13). 2-Aminobicyclo[2.2.1]-heptane-2-carboxylate (BCH), a specific L-AT inhibitor, and L-leucine, a substrate for L-AT, were shown to inhibit CysNO-stimulated cellular effects. L-leucine uptake in PC12 cells was inhibited by CysNO, not by SNAP, sodium nitroprusside, 3-morpholinosydnonimine (SIN-1)⅐HCl, (ϩ or Ϫ)-N-{(E)-4-ethyl-2-[(Z)-hydroxyimino]-5-nitro-3-hexene-1-yl}-3-pyridinecarboximide (NOR-4), or sodium nitrite (12). Although these studies suggest the involvement of specific transport mechanisms for S-nitrosothiols, the change of intracellular S-nitrosothiol level was not examined. Consequently, the direct involvement of these pathways...

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“…In addition, the effect of HC on '251-t-PA binding was completely abrogated by coincubation of cells with excess L-cysteine, a competitive inhibitor of HC uptake by endothelial cells (Fig. 2 E) (28). Halfmaximal restoration of 1251-t-PA binding to HC-treated cells (5 mM) was observed upon coincubation with roughly equimolar doses of L-cysteine ( -4.8 mM).…”

Section: Introductionmentioning

confidence: 84%

Homocysteine-induced modulation of tissue plasminogen activator binding to its endothelial cell membrane receptor.

1993

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Endothelial cells impart thromboresistance to the blood vessel wall. As modulators of fibrinolytic activity, these cells synthesize and secrete tissue plasminogen activator (t-PA) as well as its physiologic inhibitor, plasminogen activator inhibitor-i. In addition, endothelial cells support membrane-associated assembly of plasminogen and tissue plasminogen activator. Recently, an M, -40,000 protein expressed on endothelial cells has been shown to interact noncompetitively through disparate mechanisms with both t-PA and plasminogen, suggesting trimolecular assembly of enzyme, substrate, and receptor (Hajjar, K. A. 1991. J. BioL. Chem. 266:21962-21970). In the present study, treatment of cultured endothelial cells with DL-homocysteine was specifically associated with a selective reduction in cellular binding sites for t-PA. This 65% decrease in binding was associated with a 60% decrease in cell-associated t-PA activity. No change in affinity for t-PA or plasminogen or in the maximal number of binding sites for plasminogen was observed. Matrix-associated t-PA binding sites were not affected. These data suggest a new mechanism whereby homocysteine may perturb endothelial cell function, thus promoting a prothrombotic state at the surface ofthe blood vessel wall. (J.

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Homocysteine uptake by human umbilical vein endothelial cells in culture

Cited by 34 publications

References 16 publications

Homocysteine-respondent Genes in Vascular Endothelial Cells Identified by Differential Display Analysis

Homocysteine-respondent Genes in Vascular Endothelial Cells Identified by Differential Display Analysis

The mechanism of transmembraneS-nitrosothiol transport

Homocysteine-induced modulation of tissue plasminogen activator binding to its endothelial cell membrane receptor.

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