FFECTS OF HOMOCYSTEINE ON eNOS AND DDAH GENE EXPRESSION LEVELS IN PRIMARY HUMAN UMBILICAL ENDOTHELIAL CELL CULTURE

摘要

Aim: levated plasma homocysteine levels is an independent risk factor for myocardial infarction, stroke and peripheral vascular disease. Aim of this study is to investigate effects of homocysteine on nitric oxide (NO) synthesis, bioavailability and degradation by analysis gene expression levels of endothelial nitric oxide synthase (eNOS) and dimethylarginine dimethylaminohydrolase (DDAH) and also NO levels in endothelial cell line.Material and Methods: Human umbilical vein endothelial cell culture (HUVEC) is done and incubated with various homocysteine concentrations (10μM, 50μM, 100μM, 500μM, 1000μM). After that, total RNA extracted from these cells and converted to cDNA. These cDNA samples are amplified with PCR for eNOS and DDAH genes. Then the products are run in agarose gel electrophoresis and analysed for densitometric measurement by gel imaging system. In addition, NOx is analysed in cell culture mediums with electrochemiluminesans method.Results: All these studies show that homocysteine has no statistically significant effect on eNOS and DDAH gene expressions in endothelial cell culture. On the other hand, homocysteine has a minor suppressive effect on NOx levels.Conclusion: These data suggest that the effect of homocysteine on NO levels is not through gene expression levels of eNOS and DDAH but it is at substrate level.