Differentiated, quiescent vascular smooth muscle cells (VSMC) assume a dedifferentiated, proliferative phenotype in response to injury----one of the hallmarks of arteriosclerosis. Members of the LIM family of zinc-finger proteins are important in regulating development and differentiation in various cell types. For example, deletion of the LIM protein RBTN2 by homologous recombination blocks erythrocyte development, and antisense striated muscle LIM protein (MLP) prevents differentiation of myoblasts. We have cloned a developmentally regulated LIM protein, SmLIM, that is expressed preferentially in VSMC. Like RBTN2 and MLP, SmLIM contains two LIM domains. SmLIM mRNA levels were downregulated quickly when arterial smooth muscle cells became dedifferentiated in vitro and in vivo. Thus, we hypothesize that SmLIM may have an important role in regulating VSMC development and differentiation. The goals of our proposed work are to investigate the molecular mechanisms regulating preferential expression of SmLIM in VSMC, to study the significance of SmLIM in VSMC development and differentiation (both in vitro and in vivo), and to elucidate the molecular mechanisms by which SmLIM exerts its function. We will first characterize the specific nucleotide sequences (cis-acting elements) required for VSMC- restricted expression of the SmLIM gene. The specificity of these VSMC-specific cis-acting elements will also be tested in transgenic animals in vivo. Using these cis-acting elements, we will characterize and clone the nuclear proteins (trans-acting factors) binding to them. Since SmLIM may play an important role in maintaining the differentiated VSMC phenotype, we will study the effect of sense and antisense SmLIM on VSMC phenotype in culture. In addition, we will determine the effect on VSMC development and differentiation in mice of deletion of the SmLIM gene by homologous recombination. Finally, we will elucidate the molecular mechanisms by which SmLIM exerts its biologic function by cloning its interaction partners. These experiments will provide fundamentally important information about development and differentiation in VSMC, as well as critical insights into the treatment of arteriosclerosis and tumor angiogenesis. Furthermore, identification of VSMC-specific cis- and trans-acting factors will be crucial to the future success of efforts to restrict the expression of foreign genes to the blood vessel wall, for gene therapy of vascular disease.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL057977-05
Application #
6311139
Study Section
Physiological Chemistry Study Section (PC)
Project Start
1997-04-01
Project End
2002-03-31
Budget Start
2000-07-01
Budget End
2002-03-31
Support Year
5
Fiscal Year
2000
Total Cost
$310,772
Indirect Cost
Name
Brigham and Women's Hospital
Department
Type
DUNS #
071723621
City
Boston
State
MA
Country
United States
Zip Code
02115
Lin, Da-Wei; Chang, Il-Chi; Tseng, Alan et al. (2008) Transforming growth factor beta up-regulates cysteine-rich protein 2 in vascular smooth muscle cells via activating transcription factor 2. J Biol Chem 283:15003-14
Liu, Xiaoli; Wei, Jiao; Peng, David H et al. (2005) Absence of heme oxygenase-1 exacerbates myocardial ischemia/reperfusion injury in diabetic mice. Diabetes 54:778-84
Wei, Jiao; Gorman, Terri E; Liu, Xiaoli et al. (2005) Increased neointima formation in cysteine-rich protein 2-deficient mice in response to vascular injury. Circ Res 97:1323-31
Chang, Yung-Fu; Wei, Jiao; Liu, Xiaoli et al. (2003) Identification of a CArG-independent region of the cysteine-rich protein 2 promoter that directs expression in the developing vasculature. Am J Physiol Heart Circ Physiol 285:H1675-83
Layne, Matthew D; Yet, Shaw-Fang; Maemura, Koji et al. (2002) Characterization of the mouse aortic carboxypeptidase-like protein promoter reveals activity in differentiated and dedifferentiated vascular smooth muscle cells. Circ Res 90:728-36
Yet, Shaw-Fang; Melo, Luis G; Layne, Matthew D et al. (2002) Heme oxygenase 1 in regulation of inflammation and oxidative damage. Methods Enzymol 353:163-76
Chen, Y H; Layne, M D; Watanabe, M et al. (2001) Upstream stimulatory factors regulate aortic preferentially expressed gene-1 expression in vascular smooth muscle cells. J Biol Chem 276:47658-63
Yet, S F; Tian, R; Layne, M D et al. (2001) Cardiac-specific expression of heme oxygenase-1 protects against ischemia and reperfusion injury in transgenic mice. Circ Res 89:168-73
Layne, M D; Yet, S F; Maemura, K et al. (2001) Impaired abdominal wall development and deficient wound healing in mice lacking aortic carboxypeptidase-like protein. Mol Cell Biol 21:5256-61
Fukumoto, S; Hsieh, C M; Maemura, K et al. (2001) Akt participation in the Wnt signaling pathway through Dishevelled. J Biol Chem 276:17479-83