Publications (19)110.02 Total impact
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Dataset: Network for activation of human endothelial cells by oxidized phospholipids supplement
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Article: Prooxidative Effects of Ambient Pollutant Chemicals Are Inhibited by HDL.
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ABSTRACT: Exposure to air pollution leads to adverse pulmonary and systemic vascular effects. High levels of plasma high-density lipoproteins (HDL) cholesterol reduce cardiovascular risk. We explored whether HDL could protect against the prooxidative effects of an organic extract of diesel exhaust particles (DEP) in vascular cells. We used a cell-free fluorescent assay to evaluate DEP oxidation by air, estimated by the degree of dichlorofluorescein (DCF) fluorescence and tested the ability of HDL to inhibit it. We also evaluated DEP prooxidative effects in bovine aortic endothelial cells and RAW264.7 macrophages by DCF fluorescence. DEP oxidation and prooxidative cellular effects occurred in concentration- and time-dependent manners. Normal HDL inhibited DEP oxidation and prooxidative cellular effects, whereas dysfunctional HDL failed to inhibit DEP oxidation and instead, it promoted further oxidation. In conclusion, DEP prooxidative effects in endothelial cells and macrophages are inhibited by normal HDL. Therefore, HDL may protect against air pollution mediated adverse vascular effects. © 2013 Wiley Periodicals, Inc. J BiochemMol Toxicol 27:172-183, 2013; View this article online at wileyonlinelibrary.com. DOI 10.1002/jbt.21475.Journal of Biochemical and Molecular Toxicology 02/2013; 27(2):172-83. · 1.38 Impact Factor -
Article: Heme oxygenase-1, oxidation, inflammation, and atherosclerosis.
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ABSTRACT: Atherosclerosis is an inflammatory process of the vascular wall characterized by the infiltration of lipids and inflammatory cells. Oxidative modifications of infiltrating low-density lipoproteins and induction of oxidative stress play a major role in lipid retention in the vascular wall, uptake by macrophages and generation of foam cells, a hallmark of this disorder. The vasculature has a plethora of protective resources against oxidation and inflammation, many of them regulated by the Nrf2 transcription factor. Heme oxygenase-1 (HO-1) is a Nrf2-regulated gene that plays a critical role in the prevention of vascular inflammation. It is the inducible isoform of HO, responsible for the oxidative cleavage of heme groups leading to the generation of biliverdin, carbon monoxide, and release of ferrous iron. HO-1 has important antioxidant, antiinflammatory, antiapoptotic, antiproliferative, and immunomodulatory effects in vascular cells, most of which play a significant role in the protection against atherogenesis. HO-1 may also be an important feature in macrophage differentiation and polarization to certain subtypes. The biological effects of HO-1 are largely attributable to its enzymatic activity, which can be conceived as a system with three arms of action, corresponding to its three enzymatic byproducts. HO-1 mediated vascular protection may be due to a combination of systemic and vascular local effects. It is usually expressed at low levels but can be highly upregulated in the presence of several proatherogenic stimuli. The HO-1 system is amenable for use in the development of new therapies, some of them currently under experimental and clinical trials. Interestingly, in contrast to the HO-1 antiatherogenic actions, the expression of its transcriptional regulator Nrf2 leads to proatherogenic effects instead. This suggests that a potential intervention on HO-1 or its byproducts may need to take into account any potential alteration in the status of Nrf2 activation. This article reviews the available evidence that supports the antiatherogenic role of HO-1 as well as the potential pathways and mechanisms mediating vascular protection.Frontiers in pharmacology. 01/2012; 3:119. -
Article: Network for activation of human endothelial cells by oxidized phospholipids: a critical role of heme oxygenase 1.
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ABSTRACT: Oxidized palmitoyl arachidonyl phosphatidylcholine (Ox-PAPC) accumulates in atherosclerotic lesions, is proatherogenic, and influences the expression of more than 1000 genes in endothelial cells. To elucidate the major pathways involved in Ox-PAPC action, we conducted a systems analysis of endothelial cell gene expression after exposure to Ox-PAPC. We used the variable responses of primary endothelial cells from 149 individuals exposed to Ox-PAPC to construct a network that consisted of 11 groups of genes, or modules. Modules were enriched for a broad range of Gene Ontology pathways, some of which have not been identified previously as major Ox-PAPC targets. Further validating our method of network construction, modules were consistent with relationships established by cell biology studies of Ox-PAPC effects on endothelial cells. This network provides novel hypotheses about molecular interactions, as well as candidate molecular regulators of inflammation and atherosclerosis. We validated several hypotheses based on network connections and genomic association. Our network analysis predicted that the hub gene CHAC1 (cation transport regulator homolog 1) was regulated by the ATF4 (activating transcription factor 4) arm of the unfolded protein response pathway, and here we showed that ATF4 directly activates an element in the CHAC1 promoter. We showed that variation in basal levels of heme oxygenase 1 (HMOX1) contribute to the response to Ox-PAPC, consistent with its position as a hub in our network. We also identified G-protein-coupled receptor 39 (GPR39) as a regulator of HMOX1 levels and showed that it modulates the promoter activity of HMOX1. We further showed that OKL38/OSGN1 (oxidative stress-induced growth inhibitor), the hub gene in the blue module, is a key regulator of both inflammatory and antiinflammatory molecules. Our systems genetics approach has provided a broad view of the pathways involved in the response of endothelial cells to Ox-PAPC and also identified novel regulatory mechanisms.Circulation Research 08/2011; 109(5):e27-41. · 9.49 Impact Factor -
Article: NF-E2-related factor 2 promotes atherosclerosis by effects on plasma lipoproteins and cholesterol transport that overshadow antioxidant protection.
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ABSTRACT: To test the hypothesis that NF-E2-related factor 2 (Nrf2) expression plays an antiatherogenic role by its vascular antioxidant and anti-inflammatory properties. Nrf2 is an important transcription factor that regulates the expression of phase 2 detoxifying enzymes and antioxidant genes. Its expression in vascular cells appears to be an important factor in the protection against vascular oxidative stress and inflammation. We developed Nrf2 heterozygous (HET) and homozygous knockout (KO) mice on an apolipoprotein (apo) E-null background by sequential breeding, resulting in Nrf2(-/-), apoE(-/-) (KO), Nrf2(-/+), apoE(-/-) (HET) and Nrf2(+/+), and apoE(-/-) wild-type littermates. KO mice exhibited decreased levels of antioxidant genes with evidence of increased reactive oxygen species generation compared with wild-type controls. Surprisingly, KO males exhibited 47% and 53% reductions in the degree of aortic atherosclerosis compared with HET or wild-type littermates, respectively. Decreased atherosclerosis in KO mice correlated with lower plasma total cholesterol in a sex-dependent manner. KO mice also had a decreased hepatic cholesterol content and a lower expression of lipogenic genes, suggesting that hepatic lipogenesis could be reduced. In addition, KO mice exhibited atherosclerotic plaques characterized by a lesser macrophage component and decreased foam cell formation in an in vitro lipid-loading assay. This was associated with a lower rate of cholesterol influx, mediated in part by decreased expression of the scavenger receptor CD36. Nrf2 expression unexpectedly promotes atherosclerotic lesion formation in a sex-dependent manner, most likely by a combination of systemic metabolic and local vascular effects.Arteriosclerosis Thrombosis and Vascular Biology 01/2011; 31(1):58-66. · 6.37 Impact Factor -
Article: Lipase Maturation Factor LMF1, Membrane Topology and Interaction with Lipase Proteins in the Endoplasmic Reticulum
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ABSTRACT: Lipase maturation factor 1 (LMF1) is predicted to be a polytopic protein localized to the endoplasmic reticulum (ER) membrane. It functions in the post-translational attainment of enzyme activity for both lipoprotein lipase and hepatic lipase. By using transmembrane prediction methods in mouse and human orthologs, models of LMF1 topology were constructed and tested experimentally. Employing a tagging strategy that used insertion of ectopic glycan attachment sites and terminal fusions of green fluorescent protein, we established a five-transmembrane model, thus dividing LMF1 into six domains. Three domains were found to face the cytoplasm (the amino-terminal domain and loops B and D), and the other half was oriented to the ER lumen (loops A and C and the carboxyl-terminal domain). This representative model shows the arrangement of an evolutionarily conserved domain within LMF1 (DUF1222) that is essential to lipase maturation. DUF1222 comprises four of the six domains, with the two largest ones facing the ER lumen. We showed for the first time, using several naturally occurring variants featuring DUF1222 truncations, that Lmf1 interacts physically with lipoprotein lipase and hepatic lipase and localizes the lipase interaction site to loop C within DUF1222. We discuss the implication of our results with regard to lipase maturation and DUF1222 domain structure.Journal of Biological Chemistry 11/2009; 284(48):33623-33633. · 4.77 Impact Factor -
Article: Novel LMF1 nonsense mutation in a patient with severe hypertriglyceridemia.
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ABSTRACT: Lipase maturation factor 1 (LMF1) gene is a novel candidate gene in severe hypertriglyceridemia. Lmf1 is involved in the maturation of lipoprotein lipase (LPL) and hepatic lipase in endoplasmic reticulum. To date only one patient with severe hypertriglyceridemia and related disorders was found to be homozygous for a nonsense mutation in LMF1 gene (Y439X). The objective of the study was to investigate LMF1 gene in hypertriglyceridemic patients in whom mutations in LPL, APOC2, and APOA5 genes had been excluded. The resequencing of LMF1 gene led to the discovery of a novel homozygous nonsense mutation in one patient with severe hypertriglyceridemia and recurrent episodes of pancreatitis. The mutation causes a G>A substitution in exon 9 (c.1395G>A), leading to a premature stop codon (W464X). LPL activity and mass were reduced by 76 and 50%, respectively, compared with normolipidemic controls. The proband over the years has shown a good response to treatment. The proband's son, heterozygous for the W464X, shows normal plasma triglyceride levels. We identified the second novel pathogenic mutation in LMF1 gene in a patient with severe hypertriglyceridemia. LPL deficiency in our patient was milder than in the carrier of the Y439X previously described.The Journal of clinical endocrinology and metabolism 10/2009; 94(11):4584-90. · 6.50 Impact Factor -
Article: Lipase maturation factor LMF1, membrane topology and interaction with lipase proteins in the endoplasmic reticulum.
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ABSTRACT: Lipase maturation factor 1 (LMF1) is predicted to be a polytopic protein localized to the endoplasmic reticulum (ER) membrane. It functions in the post-translational attainment of enzyme activity for both lipoprotein lipase and hepatic lipase. By using transmembrane prediction methods in mouse and human orthologs, models of LMF1 topology were constructed and tested experimentally. Employing a tagging strategy that used insertion of ectopic glycan attachment sites and terminal fusions of green fluorescent protein, we established a five-transmembrane model, thus dividing LMF1 into six domains. Three domains were found to face the cytoplasm (the amino-terminal domain and loops B and D), and the other half was oriented to the ER lumen (loops A and C and the carboxyl-terminal domain). This representative model shows the arrangement of an evolutionarily conserved domain within LMF1 (DUF1222) that is essential to lipase maturation. DUF1222 comprises four of the six domains, with the two largest ones facing the ER lumen. We showed for the first time, using several naturally occurring variants featuring DUF1222 truncations, that Lmf1 interacts physically with lipoprotein lipase and hepatic lipase and localizes the lipase interaction site to loop C within DUF1222. We discuss the implication of our results with regard to lipase maturation and DUF1222 domain structure.Journal of Biological Chemistry 09/2009; 284(48):33623-33. · 4.77 Impact Factor -
Article: A quantitative assay measuring the function of lipase maturation factor 1.
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ABSTRACT: Newly synthesized lipoprotein lipase (LPL) and related members of the lipase gene family require an endoplasmic reticulum maturation factor for attainment of enzyme activity. This factor has been identified as lipase maturation factor 1 (Lmf1), and mutations affecting its function and/or expression result in combined lipase deficiency (cld) and hypertriglyceridemia. To assess the functional impact of Lmf1 sequence variations, both naturally occurring and induced, we report the development of a cell-based assay using LPL activity as a quantitative reporter of Lmf1 function. The assay uses a cell line homozygous for the cld mutation, which renders endogenous Lmf1 nonfunctional. LPL transfected into the mutant cld cell line fails to attain activity; however, cotransfection of LPL with wild-type Lmf1 restores its ability to support normal lipase maturation. In this report, we describe optimized conditions that ensure the detection of a complete range of Lmf1 function (full, partial, or complete loss of function) using LPL activity as the quantitative reporter. To illustrate the dynamic range of the assay, we tested several novel mutations in mouse Lmf1. Our results demonstrate the ability of the assay to detect and analyze Lmf1 mutations having a wide range of effects on Lmf1 function and protein expression.The Journal of Lipid Research 06/2009; 50(11):2265-9. · 5.56 Impact Factor -
Article: Age-accelerated atherosclerosis correlates with failure to upregulate antioxidant genes.
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ABSTRACT: Excess food intake leads to obesity and diabetes, both of which are well-known independent risk factors for atherosclerosis, and both of which are growing epidemics in an aging population. We hypothesized that aging enhances the metabolic and vascular effects of high fat diet (HFD) and therefore examined the effect of age on atherosclerosis and insulin resistance in lipoprotein receptor knockout (LDLR(-/-)) mice. We found that 12-month-old (middle-aged) LDLR(-/-) mice developed substantially worse metabolic syndrome, diabetes, and atherosclerosis than 3-month-old (young) LDLR(-/-) mice when both were fed HFD for 3 months, despite similar elevations in total cholesterol levels. Microarray analyses were performed to analyze the mechanism responsible for the marked acceleration of atherosclerosis in middle-aged mice. Chow-fed middle-aged mice had greater aortic expression of multiple antioxidant genes than chow-fed young mice, including glutathione peroxidase-1 and -4, catalase, superoxide dismutase-2, and uncoupling protein-2. Aortic expression of these enzymes markedly increased in young mice fed HFD but decreased or only modestly increased in middle-aged mice fed HFD, despite the fact that systemic oxidative stress and vascular reactive oxygen species generation, measured by plasma F2alpha isoprostane concentration (systemic) and dihydroethidium conversion and p47phox expression (vascular), were greater in middle-aged mice fed HFD. Thus, the mechanism for the accelerated vascular injury in older LDLR(-/-) mice was likely the profound inability to mount an antioxidant response. This effect was related to a decrease in vascular expression of 2 key transcriptional pathways regulating the antioxidant response, DJ-1 and forkhead box, subgroup O family (FOXOs). Treatment of middle-aged mice fed HFD with the antioxidant apocynin attenuated atherosclerosis, whereas treatment with the insulin sensitizer rosiglitazone attenuated both metabolic syndrome and atherosclerosis. Both treatments decreased oxidative stress. A novel effect of rosiglitazone was to increase expression of Nrf2 (nuclear factor [erythroid-derived 2]-like 2), a downstream target of DJ-1 contributing to enhanced expression of vascular antioxidant enzymes. This investigation underscores the role of oxidative stress when multiple atherosclerotic risk factors, particularly aging, converge on the vessel wall and emphasizes the need to develop effective strategies to inhibit oxidative stress to protect aging vasculature.Circulation Research 04/2009; 104(6):e42-54. · 9.49 Impact Factor -
Article: Differential roles of cardiomyocyte and macrophage peroxisome proliferator-activated receptor gamma in cardiac fibrosis.
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ABSTRACT: Cardiac fibrosis is an important component of diabetic cardiomyopathy. Peroxisome proliferator-activated receptor gamma (PPARgamma) ligands repress proinflammatory gene expression, including that of osteopontin, a known contributor to the development of myocardial fibrosis. We thus investigated the hypothesis that PPARgamma ligands could attenuate cardiac fibrosis. Wild-type cardiomyocyte- and macrophage-specific PPARgamma(-/-) mice were infused with angiotensin II (AngII) to promote cardiac fibrosis and treated with the PPARgamma ligand pioglitazone to determine the roles of cardiomyocyte and macrophage PPARgamma in cardiac fibrosis. Cardiomyocyte-specific PPARgamma(-/-) mice (cPPARgamma(-/-)) developed spontaneous cardiac hypertrophy with increased ventricular osteopontin expression and macrophage content, which were exacerbated by AngII infusion. Pioglitazone attenuated AngII-induced fibrosis, macrophage accumulation, and osteopontin expression in both wild-type and cPPARgamma(-/-) mice but induced hypertrophy in a PPARgamma-dependent manner. We pursued two mechanisms to explain the antifibrotic cardiomyocyte-PPARgamma-independent effects of pioglitazone: increased adiponectin expression and attenuation of proinflammatory macrophage activity. Adenovirus-expressed adiponectin had no effect on cardiac fibrosis and the PPARgamma ligand pioglitazone did not attenuate AngII-induced cardiac fibrosis, osteopontin expression, or macrophage accumulation in monocyte-specific PPARgamma(-/-) mice. We arrived at the following conclusions: 1) both cardiomyocyte-specific PPARgamma deficiency and activation promote cardiac hypertrophy, 2) both cardiomyocyte and monocyte PPARgamma regulate cardiac macrophage infiltration, 3) inflammation is a key mediator of AngII-induced cardiac fibrosis, 4) macrophage PPARgamma activation prevents myocardial macrophage accumulation, and 5) PPARgamma ligands attenuate AngII-induced cardiac fibrosis by inhibiting myocardial macrophage infiltration. These observations have important implications for potential interventions to prevent cardiac fibrosis.Diabetes 06/2008; 57(9):2470-9. · 8.29 Impact Factor -
Article: PPARdelta-mediated antiinflammatory mechanisms inhibit angiotensin II-accelerated atherosclerosis.
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ABSTRACT: Activation of the nuclear hormone receptor peroxisome proliferator-activated receptor delta (PPARdelta) has been shown to improve insulin resistance, adiposity, and plasma HDL levels. However, its antiatherogenic role remains controversial. Here we report atheroprotective effects of PPARdelta activation in a model of angiotensin II (AngII)-accelerated atherosclerosis, characterized by increased vascular inflammation related to repression of an antiinflammatory corepressor, B cell lymphoma-6 (Bcl-6), and the regulators of G protein-coupled signaling (RGS) proteins RGS4 and RGS5. In this model, administration of the PPARdelta agonist GW0742 (1 or 10 mg/kg) substantially attenuated AngII-accelerated atherosclerosis without altering blood pressure and increased vascular expression of Bcl-6, RGS4, and RGS5, which was associated with suppression of inflammatory and atherogenic gene expression in the artery. In vitro studies demonstrated similar changes in AngII-treated macrophages: PPARdelta activation increased both total and free Bcl-6 levels and inhibited AngII activation of MAP kinases, p38, and ERK1/2. These studies uncover crucial proinflammatory mechanisms of AngII and highlight actions of PPARdelta activation to inhibit AngII signaling, which is atheroprotective.Proceedings of the National Academy of Sciences 04/2008; 105(11):4277-82. · 9.68 Impact Factor -
Article: C-reactive protein induces apoptosis in human coronary vascular smooth muscle cells.
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ABSTRACT: Accumulating evidence suggests that C-reactive protein (CRP), in addition to being a predictor of coronary events, may have direct actions on the vessel wall in the evolution of atherosclerosis. Although accumulation of vascular smooth muscle cells (VSMCs) in the intima is a key event in the development of arterial lesions, apoptosis of VSMCs also plays an important role in progression of atherosclerotic lesions and contributes to increased plaque vulnerability. In the present study we demonstrate that CRP induces caspase-mediated apoptosis of human coronary VSMCs. DNA microarray analysis was used to identify CRP-regulated genes. The growth arrest- and DNA damage-inducible gene 153 (GADD153) mRNA expression was prominently upregulated by CRP. As confirmed by Northern blot analysis, CRP induced a time- and dose-dependent increase of GADD153 mRNA expression. GADD153, a gene involved in growth arrest and apoptosis in vascular and nonvascular cells, is regulated at both transcriptional and posttranscriptional levels. CRP regulation of GADD153 mRNA expression in VSMCs occurs primarily at the posttranscriptional level by mRNA stabilization. Small interfering RNA (siRNA) specifically targeted to GADD153 reduced CRP-induced apoptosis. GADD153 also specifically colocalized to apoptotic VSMCs in human coronary lesions, further supporting a functional role for GADD153 in CRP-induced cell death. These results demonstrate that GADD153 is a CRP-regulated gene in human VSMCs and plays a causal role in CRP-induced apoptosis. Pharmacological targeting of CRP expression or action may provide a novel therapy for atherosclerosis.Circulation 09/2004; 110(5):579-87. · 14.74 Impact Factor -
Article: Signaling pathways involved in induction of GADD45 gene expression and apoptosis by troglitazone in human MCF-7 breast carcinoma cells.
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ABSTRACT: We previously reported that the PPARgamma agonist troglitazone (TRO) inhibits proliferation and induces apoptosis in human MCF-7 breast carcinoma cells. To understand the mechanisms of antiproliferative and pro-apoptotic effects of TRO, we screened a limited DNA array containing 23 genes involved in regulating either the cell cycle and/or apoptosis. Four of the 23 genes screened exhibited regulation by TRO, with growth arrest and DNA damage-inducible gene 45 (GADD45) being the most strongly upregulated. TRO induced GADD45 mRNA expression in a time- and dose-dependent manner. Depletion of GADD45 by siRNA abrogated TRO-induced apoptosis in MCF-7 cells demonstrating the physiological relevance of GADD45 upregulation. Signaling pathways mediating TRO-induced GADD45 were also investigated. Several mitogen-activated protein kinase (MAPK) pathways were involved in the induction of GADD45 by TRO. Inhibition of the c-jun N-terminal kinase MAPK pathway by SP600125 partially abolished TRO-induced GADD45 mRNA, and protein expression and apoptosis. In contrast, inhibition of the p38 MAPK pathway by SB203580, or through overexpression of a dominant-negative mutant of p38 MAPK, augmented GADD45 mRNA induction and GADD45 promoter activation as well as cell apoptosis by TRO. Blockade of the extracellular signal-regulated kinase MAPK pathway by PD98059 also enhanced TRO's effects on GADD45 and apoptosis. Two other PPARgamma agonists pioglitazone and rosiglitazone did not induce GADD45 expression. Our finding of GADD45 induction by TRO may provide a new insight concerning the mechanisms for TRO's antiproliferative and pro-apoptotic effects in breast cancer cells.Oncogene 07/2004; 23(26):4614-23. · 6.37 Impact Factor -
Article: Expression of minichromosome maintenance proteins in vascular smooth muscle cells is ERK/MAPK dependent.
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ABSTRACT: Proliferation of vascular smooth muscle cells (VSMC) represents a key event for the pathogenesis of postangioplasty restenosis. Minichromosome maintenance proteins (MCM) form essential components of the prereplicative complex at DNA replication origins and are regulated by E2F. The present studies were designed to investigate the signal transduction pathways controlling the expression of MCM6 and MCM7 in VSMC in response to mitogenic stimuli. MCM6 and MCM7 expression was substantially increased after stimulation with platelet-derived growth factor-BB and insulin. Pretreatment with PD98059, a specific inhibitor of the extracellular signal-regulated kinases (ERK)-mitogen-activated protein kinase (MAPK), competely inhibited the mitogen-induced MCM6 and MCM7 mRNA and protein expression, demonstrating a critical role for this pathway in transmitting transmembrane signals required for the initiation of DNA replication. The p38MAPK inhibitor SB203580, the phosphatidylinositol 3 kinase (PI3-kinase) pathway inhibitor wortmannin, and the protein kinase C pathway (PKC) inhibitor Gö 6976 did not significantly affect mitogen-induced MCM6 and MCM7 expression. Transient transfection experiments revealed that PD98059 inhibited mitogen-induced MCM6 and MCM7 transcriptional activation. In addition, blockade of ERK/MAPK signaling with PD98059 strongly inhibited phosphorylation of the retinoblastoma protein (Rb) and activity of a luciferase reporter plasmid driven by multiple E2F elements. Inhibition of mitogen-induced MCM6 and MCM7 expression by PD98059 was reversed by ectopic overexpression of E2F, indicating that ERK/MAPK signaling is required for events that occur upstream of E2F release from phosphorylated Rb. In combination, these data demonstrate that the ERK/MAPK signal transduction pathway plays a central role in regulating E2F-dependent MCM expression and DNA replication in VSMC.Experimental Cell Research 11/2003; 290(1):28-37. · 3.58 Impact Factor -
Article: Regulation of the growth arrest and DNA damage-inducible gene 45 (GADD45) by peroxisome proliferator-activated receptor gamma in vascular smooth muscle cells.
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ABSTRACT: Peroxisome proliferator-activated receptor (PPAR) gamma is activated by thiazolidinediones (TZDs), widely used as insulin-sensitizing agents for the treatment of type 2 diabetes. TZDs have been shown to induce apoptosis in a variety of mammalian cells. In vascular smooth muscle cells (VSMCs), proliferation and apoptosis may be competing processes during the formation of restenotic and atherosclerotic lesions. The precise molecular mechanisms by which TZDs induce apoptosis in VSMCs, however, remain unclear. In the present study, we demonstrate that the TZDs rosiglitazone (RSG), troglitazone (TRO), and a novel non-TZD partial PPARgamma agonist (nTZDpa) induce caspase-mediated apoptosis of human coronary VSMCs. Induction of VSMC apoptosis correlated closely with an upregulation of growth arrest and DNA damage-inducible gene 45 (GADD45) mRNA expression and transcription, a well-recognized modulator of cell cycle arrest and apoptosis. Using adenoviral-mediated overexpression of a constitutively active PPARgamma mutant and the irreversible PPARgamma antagonist GW9662, we provide evidence that PPARgamma ligands induce caspase-mediated apoptosis and GADD45 expression through a receptor-dependent pathway. Deletion analysis of the GADD45 promoter revealed that a 153-bp region between -234 and -81 bp proximal to the transcription start site, containing an Oct-1 element, was crucial for the PPARgamma ligand-mediated induction of the GADD45 promoter. PPARgamma activation induced Oct-1 protein expression and DNA binding and stimulated activity of a reporter plasmid driven by multiple Oct-1 elements. These findings suggest that activation of PPARgamma can lead to apoptosis and growth arrest in VSMCs, at least in part, by inducing Oct-1-mediated transcription of GADD45. The full text of this article is available online at http://www.circresaha.org.Circulation Research 09/2003; 93(4):e38-47. · 9.49 Impact Factor -
Article: Peroxisome proliferator-activated receptor gamma inhibits expression of minichromosome maintenance proteins in vascular smooth muscle cells.
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ABSTRACT: Using a cDNA array consisting only of cell cycle genes, we found that a novel nonthiazolidinedione partial peroxisome proliferator-activated receptor gamma (PPARgamma) agonist (nTZDpa) inhibited expression of minichromosome maintenance (MCM) proteins 6 and 7 in vascular smooth muscle cells. MCM proteins are required for the initiation and elongation stages of DNA replication and are regulated by the transcription factor E2F. Mitogen-induced MCM6 and MCM7 mRNA expression was potently inhibited by nTZDpa and to a lesser degree by the full PPARgamma agonist, rosiglitazone. Inhibition of MCM6 and MCM7 expression by nTZDpa and rosiglitazone paralleled their effect to inhibit phosphorylation of the retinoblastoma protein and cell proliferation. Transient transfection experiments revealed that the nTZDpa inhibited mitogen-induced MCM6 and MCM7 promoter activity, implicating a transcriptional mechanism. Adenoviral-mediated E2F overexpression reversed the suppressive effect of nTZDpa on MCM6 and MCM7 expression. Furthermore, activity of a luciferase reporter plasmid driven by multiple E2F elements was inhibited by nTZDpa, indicating that their down-regulation by nTZDpa involves an E2F-dependent mechanism. Overexpression of dominant-negative PPARgamma or addition of a PPARgamma antagonist, GW 9662, blocked nTZDpa inhibition of MCM7 transcription. Adenovirus-mediated overexpression of constitutively active PPARgamma inhibited MCM7 expression in a similar manner as the nTZDpa. These findings provide strong evidence that activation of PPARgamma attenuates MCM7 transcription and support the important role of this nuclear receptor in regulating vascular smooth muscle cell proliferation.Molecular Endocrinology 07/2003; 17(6):1005-18. · 4.54 Impact Factor -
Article: Rapamycin inhibits E2F-dependent expression of minichromosome maintenance proteins in vascular smooth muscle cells.
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ABSTRACT: Rapamycin inhibits vascular smooth muscle cell (VSMC) proliferation and rapamycin-eluting stents represent a novel therapeutic strategy for preventing postangioplasty restenosis. The precise molecular mechanism, for rapamycin-mediated inhibition of VSMC cell cycle progression and DNA replication remain to be elucidated. Minichromosome maintenance proteins (MCM) are essential regulators of DNA replication and the objective of this study was to examine the effect of rapamycin on their expression in rat aortic VSMC. Rapamycin substantially inhibited mitogen-induced MCM6 and MCM7 mRNA and protein expression in a dose-dependent fashion. Transient transfection experiments revealed that rapamycin inhibited MCM6 and MCM7 promoter activity, implicating a transcriptional mechanism. MCM6 and MCM7 transcriptional activation is regulated by E2F and activity of a luciferase reporter plasmid driven by four E2F elements was also significantly inhibited by rapamycin. The inhibitory effect of rapamycin on MCM6 and MCM7 was reversed by overexpression of E2F, indicating that their downregulation by rapamycin involves an E2F-dependent mechanism. These observations suggest that rapamycin inhibits MCM6 and MCM7 expression by blocking their E2F-dependent transactivation which may contribute importantly to the inhibition of VSMC DNA synthesis by rapamycin.Biochemical and Biophysical Research Communications 04/2003; 303(1):251-8. · 2.48 Impact Factor -
Article: Atorvastatin inhibits expression of minichromosome maintenance proteins in vascular smooth muscle cells.
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ABSTRACT: 3-Hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors have been reported to inhibit vascular smooth muscle cell growth, a key event in the pathogenesis of proliferative vascular diseases. The mechanism by which HMG-CoA reductase inhibitors exert their antiproliferative activity is not fully understood, especially their effect on DNA replication. We therefore investigated the effects of atorvastatin on minichromosome maintenance (MCM) protein 6 and 7 expression in vascular smooth muscle cells, two proteins essential for initiation of DNA replication. Stimulation of quiescent rat aortic vascular smooth muscle cells with fetal bovine serum induced MCM6 and MCM7 protein and mRNA expression, which was potently attenuated by atorvastatin in a dose-dependent fashion. Mevalonate completely abrogated the inhibitory effect on serum-induced MCM6 and MCM7 expression, demonstrating that biosynthesis of isoprenoids was likely the specific pathway blocked by atorvastatin. Transient transfection experiments revealed that atorvastatin inhibited MCM6 and MCM7 promoter activity, implicating a transcriptional mechanism. The MCM6 and MCM7 promoters contain several E2F sites critical for their transcriptional activation. Activity of a luciferase reporter plasmid containing four E2F elements was also strongly inhibited by atorvastatin. The inhibitory effect of atorvastatin on MCM6 and MCM7 was reversed by adenoviral-mediated overexpression of E2F, indicating that their downregulation by atorvastatin involves an E2F-dependent mechanism. These findings demonstrate that MCM proteins play an essential role during the proliferation of vascular smooth muscle cells and may provide a novel therapeutic target for proliferative vascular diseases. Inhibition of MCM6 and MCM7 expression by blocking E2F function may contribute importantly to the inhibition of vascular smooth muscle cell DNA synthesis by atorvastatin.European Journal of Pharmacology 03/2003; 462(1-3):15-23. · 2.52 Impact Factor
Top co-authors
Top Journals
Institutions
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2003–2013
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University of California, Los Angeles
- • Division of Cardiology
- • Department of Medicine
Los Angeles, CA, USA -
CSU Mentor
Long Beach, CA, USA -
University of Southern California
Los Angeles, CA, USA
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