Inhibition of Endothelial Growth by Homocysteine

Summary

Principal Investigator: Hong Wang
Affiliation: Temple University
Country: USA
Abstract: Hyperhomocysteinemia is an independent risk for cardiovascular disease. Most of the reported biological effects of homocysteine (Hcy) in vascular cells have been attributed to oxidative mechanisms, which were observed at Hcy concentrations higher than 1mM, and can be mimicked by cysteine, another non-pathogenic biothiol. Thus, a biochemical mechanism unique to Hcy remains to be identified. Our previous work has demonstrated that Hcy at 10-50 muM, but not cysteine, arrests endothelial cell (EC) p21/ras, suppress cyclin A transcription in cell type specific manner. The basic hypothesis of this proposed project is that Hcy, at growth through a hypomethylation related mechanism, which blocks cell cycle progression and endothelium regeneration. This project will study this hypothesis utilizing three linked specific aims. First, in Aim 1, experiments are designed to elucidate the role of Ras demethylation-independent Ras over-expression on EC growth, and on cyclin A expression and promoter activity. Cell growth will be determine by thymidine uptake, flow cytometry and cell counting. Cyclin A expression and promoter activity will be determined by Northern, Western blot analysis and reporter gene transfection. Alternatively, DNA microarray will be used to identify other potential targets in Hcy signaling. Second, in Aim 2, studies are proposed to determine the biochemical mechanisms by which Hcy suppress cyclin A transcription. Experiments will be performed to study the RB phosphorylation and E2F expression, and the role of E2F and other transcription factors on cyclin A transcription by Western blot, reporter gene transfection, gel mobility shift, adenovirus-transduced E2F expression. The role of cyclin A in maintaining RB phosphorylation will be assessed by RNA interference. The promoter methylation pattern of cyclin A genes will be examined by bisufite genomic sequence and methylation sensitive restriction enzyme digestion. Finally, in Aim 3, a mouse endothelial denudation and regeneration model will be used in cystathionine beta-synthase (CBS) knockout and diet-induced hyperhomocysteinemic mice to assess the effect of Hcy in endothelial regeneration. Mouse blood will e examined for the concentrations of Hcy and SAH/SAM. Immunohistochemistry staining for leukocytes, macrophages, EC and smooth muscle cells will be performed on vessel sections to analyze the cellular composition of the lesion. In situ hybridization of immunohistochemistry staining for cyclin A will be performed to assess its involvement. The broad, long-term objective of this proposal i to elucidate Hcy signaling in EC growth inhibition, and to evaluate its importance on the role of atherogenesis in hyperhomocysteinemia. If we identify the key events in Hcy-induced arteriosclerosis, genetic or biochemical approaches to block these steps could lead to therapeutic advantage.
Funding Period: 2002-04-05 - 2008-03-31
more information: NIH RePORT

Top Publications

  1. ncbi Single perivascular delivery of mitomycin C stimulates p21 expression and inhibits neointima formation in rat arteries
    Juan F Granada
    The Methodist Research Institute, Houston, Texas, USA
    Arterioscler Thromb Vasc Biol 25:2343-8. 2005
  2. ncbi Hyperhomocystinemia impairs endothelial function and eNOS activity via PKC activation
    Xiaohua Jiang
    Department of Medicine, Baylor College of Medicine, Houston, TX 77030, USA
    Arterioscler Thromb Vasc Biol 25:2515-21. 2005
  3. ncbi Hyperhomocysteinemia inhibits post-injury reendothelialization in mice
    Hongmei Tan
    Department of Medicine, Baylor College of Medicine, USA
    Cardiovasc Res 69:253-62. 2006
  4. pmc Local administration of carbon monoxide inhibits neointima formation in balloon injured rat carotid arteries
    D A Tulis
    Cardiovascular Disease Research Program, J L Chambers Biomedical Biotechnology Research Institute, North Carolina Central University, Durham, North Carolina 27707, USA
    Cell Mol Biol (Noisy-le-grand) 51:441-6. 2005
  5. ncbi Hyperhomocysteinemia decreases circulating high-density lipoprotein by inhibiting apolipoprotein A-I Protein synthesis and enhancing HDL cholesterol clearance
    Dan Liao
    Department of Medicine, Baylor College of Medicine, Houston, TX, USA
    Circ Res 99:598-606. 2006
  6. pmc Homocysteine inhibits endothelial cell growth via DNA hypomethylation of the cyclin A gene
    M D S Jamaluddin
    Department of Pharmacology, Temple University School of Medicine, Philadelphia, PA 19140, USA
    Blood 110:3648-55. 2007
  7. ncbi Differential regulation of homocysteine transport in vascular endothelial and smooth muscle cells
    Xiaohua Jiang
    Temple University School of Medicine, Department of Pharmacology, 3420 North Broad Street, Philadelphia, PA 19140, USA
    Arterioscler Thromb Vasc Biol 27:1976-83. 2007
  8. pmc State-dependent calcium mobilization by urotensin-II in cultured human endothelial cells
    Eugen Brailoiu
    Department of Pharmacology, 3420 N Broad Street, Temple University School of Medicine, Philadelphia, PA 19140, USA
    Peptides 29:721-6. 2008
  9. pmc Hyperhomocysteinemia promotes inflammatory monocyte generation and accelerates atherosclerosis in transgenic cystathionine beta-synthase-deficient mice
    Daqing Zhang
    Department of Pharmacology and Cardiovascular Research Center, Temple University School of Medicine, Philadelphia, PA 19140, USA
    Circulation 120:1893-902. 2009

Scientific Experts

  • Eugen Brailoiu
  • Xiaohua Jiang
  • William Durante
  • Hong Wang
  • Xiaofeng Yang
  • Andrew I Schafer
  • Fan Yang
  • Hongmei Tan
  • Dan Liao
  • Daqing Zhang
  • M D S Jamaluddin
  • Zhaohui Li
  • D A Tulis
  • Juan F Granada
  • Pu Fang
  • Yan Yan
  • Warren D Kruger
  • Jian Song
  • Sapna Gupta
  • Xiaoming Liu
  • Irene Chen
  • Nae J Dun
  • Hieronim Jakubowski
  • Michael Jan
  • Lawrence Chan
  • Mark J Magera
  • Rutai Hui
  • Henry J Pownall
  • JoAnn Trial
  • John Gaubatz
  • K J Peyton
  • W Durante
  • Albert E Raizner
  • Xiao ming Liu
  • Robert M Bryan
  • Mohammed A Azam
  • Kelly J Peyton
  • H Wang
  • Jaspreet K Randhawa
  • Amit N Keswani
  • Grzegorz L Kaluza
  • Rolando E Rumbaut
  • A I Schafer
  • Diana Ensenat
  • A N Keswani

Detail Information

Publications9

  1. ncbi Single perivascular delivery of mitomycin C stimulates p21 expression and inhibits neointima formation in rat arteries
    Juan F Granada
    The Methodist Research Institute, Houston, Texas, USA
    Arterioscler Thromb Vasc Biol 25:2343-8. 2005
    ....
  2. ncbi Hyperhomocystinemia impairs endothelial function and eNOS activity via PKC activation
    Xiaohua Jiang
    Department of Medicine, Baylor College of Medicine, Houston, TX 77030, USA
    Arterioscler Thromb Vasc Biol 25:2515-21. 2005
    ..A risk factor for cardiovascular disease, hyperhomocystinemia (HHcy), is associated with endothelial dysfunction. In this study, we examined the mechanistic role of HHcy in endothelial dysfunction...
  3. ncbi Hyperhomocysteinemia inhibits post-injury reendothelialization in mice
    Hongmei Tan
    Department of Medicine, Baylor College of Medicine, USA
    Cardiovasc Res 69:253-62. 2006
    ..In this study, we established a mouse model of endothelial injury and reendothelialization and examined the role and mechanism of HHcy in endothelial repair...
  4. pmc Local administration of carbon monoxide inhibits neointima formation in balloon injured rat carotid arteries
    D A Tulis
    Cardiovascular Disease Research Program, J L Chambers Biomedical Biotechnology Research Institute, North Carolina Central University, Durham, North Carolina 27707, USA
    Cell Mol Biol (Noisy-le-grand) 51:441-6. 2005
    ....
  5. ncbi Hyperhomocysteinemia decreases circulating high-density lipoprotein by inhibiting apolipoprotein A-I Protein synthesis and enhancing HDL cholesterol clearance
    Dan Liao
    Department of Medicine, Baylor College of Medicine, Houston, TX, USA
    Circ Res 99:598-606. 2006
    ..These findings indicate that HHcy inhibits reverse cholesterol transport by reducing circulating HDL via inhibiting apoA-I protein synthesis and enhancing HDL-C clearance...
  6. pmc Homocysteine inhibits endothelial cell growth via DNA hypomethylation of the cyclin A gene
    M D S Jamaluddin
    Department of Pharmacology, Temple University School of Medicine, Philadelphia, PA 19140, USA
    Blood 110:3648-55. 2007
    ..In conclusion, Hcy inhibits cyclin A transcription and cell growth by inhibiting DNA methylation through suppression of DNMT1 in ECs...
  7. ncbi Differential regulation of homocysteine transport in vascular endothelial and smooth muscle cells
    Xiaohua Jiang
    Temple University School of Medicine, Department of Pharmacology, 3420 North Broad Street, Philadelphia, PA 19140, USA
    Arterioscler Thromb Vasc Biol 27:1976-83. 2007
    ..This study characterized and directly compared Hcy transport in cultured human aortic ECs (HAECs) and smooth muscle cells (HASMCs)...
  8. pmc State-dependent calcium mobilization by urotensin-II in cultured human endothelial cells
    Eugen Brailoiu
    Department of Pharmacology, 3420 N Broad Street, Temple University School of Medicine, Philadelphia, PA 19140, USA
    Peptides 29:721-6. 2008
    ..The result demonstrates a state-dependent effect of U-II in cultured HAEC, which may explain the variable responses to U-II under different experimental conditions...
  9. pmc Hyperhomocysteinemia promotes inflammatory monocyte generation and accelerates atherosclerosis in transgenic cystathionine beta-synthase-deficient mice
    Daqing Zhang
    Department of Pharmacology and Cardiovascular Research Center, Temple University School of Medicine, Philadelphia, PA 19140, USA
    Circulation 120:1893-902. 2009
    ..Monocytes display inflammatory and resident subsets and commit to specific functions in atherogenesis. In this study, we examined the hypothesis that HHcy modulates monocyte heterogeneity and leads to atherosclerosis...