A DNA enzyme targeting Egr-1 inhibits rat vascular smooth muscle cell proliferation by down-regulation of cyclin D1 and TGF-β1

We have demonstrated that a synthetic DNA enzyme targeting early growth response factor-1 (Egr-1) can inhibit neointimal hyperplasia following vascular injury. However, the detailed mechanism of this inhibition is not known. Thus, the objective of the present study was to further investigate potenti...

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Main Authors: Y. Wu, W. Han, G.-N. Liu
Format: Article
Language:English
Published: Associação Brasileira de Divulgação Científica 2010-01-01
Series:Brazilian Journal of Medical and Biological Research
Subjects:
Online Access:http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0100-879X2010000100004&lng=en&tlng=en
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author Y. Wu
W. Han
G.-N. Liu
author_facet Y. Wu
W. Han
G.-N. Liu
author_sort Y. Wu
collection DOAJ
description We have demonstrated that a synthetic DNA enzyme targeting early growth response factor-1 (Egr-1) can inhibit neointimal hyperplasia following vascular injury. However, the detailed mechanism of this inhibition is not known. Thus, the objective of the present study was to further investigate potential inhibitory mechanisms. Catalytic DNA (ED5) and scrambled control DNA enzyme (ED5SCR) were synthesized and transfected into primary cultures of rat vascular smooth muscle cells (VSMCs). VSMC proliferation and DNA synthesis were analyzed by the MTT method and BrdU staining, respectively. Egr-1, TGF-β1, p53, p21, Bax, and cyclin D1 expression was detected by RT-PCR and Western blot. Apoptosis and cell cycle assays were performed by FACS. Green fluorescence could be seen localized in the cytoplasm of 70.6 ± 1.52 and 72 ± 2.73% VSMCs 24 h after transfection of FITC-labeled ED5 and ED5SCR, respectively. We found that transfection with ED5 significantly inhibited cultured VSMC proliferation in vitro after 24, 48, and 72 h of serum stimulation, and also effectively decreased the uptake of BrdU by VSMC. ED5 specifically reduced serum-induced Egr-1 expression in VSMCs, further down-regulated the expression of cyclin D1 and TGF-β1, and arrested the cells at G0/G1, inhibiting entry into the S phase. FACS analysis indicated that there was no significant difference in the rate of apoptosis between ED5- and ED5SCR-transfected cells. Thus, ED5 can specifically inhibit Egr-1 expression, and probably inhibits VSMC proliferation by down-regulating the expressions of cyclin D1 and TGF-β1. However, ED5 has no effect on VSMC apoptosis.
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spelling doaj.art-a598087b313a451f8e1b49edc02fed0c2022-12-22T01:15:16ZengAssociação Brasileira de Divulgação CientíficaBrazilian Journal of Medical and Biological Research1414-431X2010-01-014311724S0100-879X2010000100004A DNA enzyme targeting Egr-1 inhibits rat vascular smooth muscle cell proliferation by down-regulation of cyclin D1 and TGF-β1Y. Wu0W. Han1G.-N. Liu2China Medical UniversityChina Medical UniversityChina Medical UniversityWe have demonstrated that a synthetic DNA enzyme targeting early growth response factor-1 (Egr-1) can inhibit neointimal hyperplasia following vascular injury. However, the detailed mechanism of this inhibition is not known. Thus, the objective of the present study was to further investigate potential inhibitory mechanisms. Catalytic DNA (ED5) and scrambled control DNA enzyme (ED5SCR) were synthesized and transfected into primary cultures of rat vascular smooth muscle cells (VSMCs). VSMC proliferation and DNA synthesis were analyzed by the MTT method and BrdU staining, respectively. Egr-1, TGF-β1, p53, p21, Bax, and cyclin D1 expression was detected by RT-PCR and Western blot. Apoptosis and cell cycle assays were performed by FACS. Green fluorescence could be seen localized in the cytoplasm of 70.6 ± 1.52 and 72 ± 2.73% VSMCs 24 h after transfection of FITC-labeled ED5 and ED5SCR, respectively. We found that transfection with ED5 significantly inhibited cultured VSMC proliferation in vitro after 24, 48, and 72 h of serum stimulation, and also effectively decreased the uptake of BrdU by VSMC. ED5 specifically reduced serum-induced Egr-1 expression in VSMCs, further down-regulated the expression of cyclin D1 and TGF-β1, and arrested the cells at G0/G1, inhibiting entry into the S phase. FACS analysis indicated that there was no significant difference in the rate of apoptosis between ED5- and ED5SCR-transfected cells. Thus, ED5 can specifically inhibit Egr-1 expression, and probably inhibits VSMC proliferation by down-regulating the expressions of cyclin D1 and TGF-β1. However, ED5 has no effect on VSMC apoptosis.http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0100-879X2010000100004&lng=en&tlng=enDNA enzymeEgr-1VSMCProliferationApoptosis
spellingShingle Y. Wu
W. Han
G.-N. Liu
A DNA enzyme targeting Egr-1 inhibits rat vascular smooth muscle cell proliferation by down-regulation of cyclin D1 and TGF-β1
Brazilian Journal of Medical and Biological Research
DNA enzyme
Egr-1
VSMC
Proliferation
Apoptosis
title A DNA enzyme targeting Egr-1 inhibits rat vascular smooth muscle cell proliferation by down-regulation of cyclin D1 and TGF-β1
title_full A DNA enzyme targeting Egr-1 inhibits rat vascular smooth muscle cell proliferation by down-regulation of cyclin D1 and TGF-β1
title_fullStr A DNA enzyme targeting Egr-1 inhibits rat vascular smooth muscle cell proliferation by down-regulation of cyclin D1 and TGF-β1
title_full_unstemmed A DNA enzyme targeting Egr-1 inhibits rat vascular smooth muscle cell proliferation by down-regulation of cyclin D1 and TGF-β1
title_short A DNA enzyme targeting Egr-1 inhibits rat vascular smooth muscle cell proliferation by down-regulation of cyclin D1 and TGF-β1
title_sort dna enzyme targeting egr 1 inhibits rat vascular smooth muscle cell proliferation by down regulation of cyclin d1 and tgf β1
topic DNA enzyme
Egr-1
VSMC
Proliferation
Apoptosis
url http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0100-879X2010000100004&lng=en&tlng=en
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