ISSN:0009-7330    

出版商:OVID    

年代:2002

数据来源: OVID

ISSN:0009-7330    

出版商:OVID    

年代:2002

数据来源: OVID

ISSN:0009-7330    

出版商:OVID    

年代:2002

数据来源: OVID

ISSN:0009-7330    

出版商:OVID    

年代:2002

数据来源: OVID

ISSN:0009-7330    

出版商:OVID    

年代:2002

数据来源: OVID

摘要:

Growth hormone-releasing peptides (GHRPs) are known as potent growth hormone secretagogues whose actions are mediated by the ghrelin receptor, a G protein-coupled receptor cloned from pituitary libraries. Hexarelin, a hexapeptide of the GHRP family, has reported cardiovascular activity. To identify the molecular target mediating this activity, rat cardiac membranes were labeled with a radioactive photoactivatable derivative of hexarelin and purified using lectin affinity chromatography and preparative gel electrophoresis. A binding protein of Mr84 000 was identified. The N-terminal sequence determination of the deglycosylated protein was identical to rat CD36, a multifunctional glycoprotein, which was expressed in cardiomyocytes and microvascular endothelial cells. Activation of CD36 in perfused hearts by hexarelin was shown to elicit an increase in coronary perfusion pressure in a dose-dependent manner. This effect was lacking in hearts from CD36-null mice and hearts from spontaneous hypertensive rats genetically deficient in CD36. The coronary vasoconstrictive response correlated with expression of CD36 as assessed by immunoblotting and covalent binding with hexarelin. These data suggest that CD36 may mediate the coronary vasospasm seen in hypercholesterolemia and atherosclerosis.

ISSN:0009-7330    

出版商:OVID    

年代:2002

数据来源: OVID

摘要:

Previous attempts to delineate the consequences of G&agr;qactivation in cardiomyocytes relied largely on molecular strategies in cultures or transgenic mice. Modest levels of wild-type G&agr;qoverexpression induce stable cardiac hypertrophy, whereas intense G&agr;qstimulation induces cardiomyocyte apoptosis. The precise mechanism(s) whereby traditional targets of G&agr;qsubunits that induce hypertrophy also trigger cardiomyocyte apoptosis is not obvious and is explored with recombinantPasteurella multocidatoxin (rPMT, a G&agr;qagonist). Cells cultured with rPMT display cardiomyocyte enlargement, sarcomeric organization, and increased atrial natriuretic factor expression in association with activation of phospholipase C, novel protein kinase C (PKC) isoforms, extracellular signal-regulated protein kinase (ERK), and (to a lesser extent) JNK/p38-MAPK. rPMT stimulates the ERK cascade via epidermal growth factor (EGF) receptor transactivation in cardiac fibroblasts, but EGF receptor transactivation plays no role in ERK activation in cardiomyocytes. Surprisingly, rPMT (or novel PKC isoform activation by PMA) decreases basal Akt phosphorylation; rPMT prevents Akt phosphorylation by EGF or IGF-1 and functionally augments cardiomyocyte apoptosis in response to H2O2. These results identify a G&agr;q-PKC pathway that represses basal Akt phosphorylation and impairs Akt stimulation by survival factors. Because inhibition of Akt enhances cardiomyocyte susceptibility to apoptosis, this pathway is predicted to contribute to the transition from hypertrophy to cardiac decompensation and could be targeted for therapy in heart failure.

ISSN:0009-7330    

出版商:OVID    

年代:2002

数据来源: OVID

摘要:

Chromatin acetylation and deacetylation catalyzed by histone acetyltransferases (HATs) and histone deacetylases (HDACs) are closely related to eukaryotic gene transcription. Although the binding of serum response factor (SRF) to the CArG boxes in the promoter region is necessary for SM22 expression, it has never been examined whether the local chromatin modification is involved in SM22 gene regulation. In this study, we used the SM22 gene as a model to address whether transcriptional activation of the gene can be manipulated through adjusting histone acetylation of the chromatin template and whether SRF- and HAT-containing coactivators can be recruited onto the SM22 promoter region during gene activation. Here, we showed that the stimulation of the SM22 promoter by the coactivator CREB-binding protein (CBP) was dependent on HAT activity. Overexpression of HDACs decreased SM22 promoter activity, whereas trichostatin A, an HDAC inhibitor, stimulated SM22 promoter activity in a CArG box-dependent manner and induced endogenous SM22 gene expression. Chromatin immunoprecipitation assays showed that trichostatin A treatment in 10T1/2 cells induces chromatin hyperacetylation in the SM22 gene. Although histone hyperacetylation of the SM22 gene occurred during SM22 gene expression and SRF and CBP immunocomplexes possess HAT activities in smooth muscle cells, both SRF and CBP were recruited to the CArG box-containing region of the promoter. This study provides evidence that chromatin acetylation is involved in smooth muscle cell-specific gene regulation.

ISSN:0009-7330    

出版商:OVID    

年代:2002

数据来源: OVID

摘要:

Protein-protein interactions with the molecular chaperone hsp90 and phosphorylation on serine 1179 by the protein kinase Akt leads to activation of endothelial nitric oxide synthase. However, the interplay between these protein-protein interactions remains to be established. In the present study, we show that vascular endothelial growth factor stimulates the coordinated association of hsp90, Akt, and resultant phosphorylation of eNOS. Characterization of the domains of hsp90 required to bind eNOS, using yeast 2-hybrid, cell-based coprecipitation experiments, and GST-fusion proteins, revealed that the M region of hsp90 interacts with the amino terminus of eNOS and Akt. The addition of purified hsp90 to in vitro kinase assays facilitates Akt-driven phosphorylation of recombinant eNOS protein, but not a short peptide encoding the Akt phosphorylation site, suggesting that hsp90 may function as a scaffold for eNOS and Akt. In vivo, coexpression of adenoviral or the cDNA for hsp90 with eNOS promotes nitric oxide release; an effect eliminated using a catalytically functional phosphorylation mutant of eNOS. These results demonstrate that stimulation of endothelial cells with vascular endothelial growth factor recruits eNOS and Akt to an adjacent region on the same domain of hsp90, thereby facilitating eNOS phosphorylation and enzyme activation.

ISSN:0009-7330    

出版商:OVID    

年代:2002

数据来源: OVID

摘要:

The atrial natriuretic peptide (ANP) is a cardiovascular hormone possessing antiinflammatory potential due to its inhibitory action on the production of inflammatory mediators, such as tumor necrosis factor-&agr; (TNF-&agr;). The aim of this study was to determine whether ANP is able to attenuate inflammatory effects of TNF-&agr; on target cells. Human umbilical vein endothelial cells (HUVECs) were treated with TNF-&agr; in the presence or absence of ANP. Changes in permeability, cytoskeletal alterations, phosphorylation of p38 MAPK and HSP27, and expression of MKP-1 were determined by macromolecule permeability assay, fluorescence labeling, RT-PCR, and immunoblotting. Antisense studies were done by transfecting cells with MKP-1 antisense oligonucleotides. Activation of HUVECs with TNF-&agr; lead to a significant increase of macromolecule permeability and formation of stress fibers. Treatment of cells with ANP (10−8to 10−6mol/L) significantly reduced the formation of stress fibers and elevated permeability. Both TNF-&agr;–induced effects were shown to be mediated via the activation of p38 using SB203580, a specific inhibitor of p38. ANP significantly reduced the TNF-&agr;–induced activation of p38 and attenuated the phosphorylation of HSP27, a central target downstream of p38. ANP showed no effect on p38 upstream kinases MKK3/6. However, a significant induction of the MAPK phosphatase MKP-1 mRNA and protein could be observed in ANP-treated cells. Antisense experiments proved a causal role for MKP-1 induction in the ANP-mediated inhibition of p38. These data show the inhibitory action of ANP on TNF-&agr;–induced changes in endothelial cytoskeleton and macromolecule permeability involving an MKP-1–induced inactivation of p38 MAPK. These effects point to an antiinflammatory and antiatherogenic potential of this cardiovascular hormone.

ISSN:0009-7330    

出版商:OVID    

年代:2002

数据来源: OVID