ISSN:0009-7330    

出版商:OVID    

年代:2000

数据来源: OVID

ISSN:0009-7330    

出版商:OVID    

年代:2000

数据来源: OVID

ISSN:0009-7330    

出版商:OVID    

年代:2000

数据来源: OVID

ISSN:0009-7330    

出版商:OVID    

年代:2000

数据来源: OVID

摘要:

In cultured vascular smooth muscle cells (VSMCs), Janus kinases (JAKs) and signal transducers and activators of transcription (STATs) are expressed constitutively and play a role in angiotensin II (Ang II)–induced intracellular signaling and proliferation. However, little is known regarding the relevance of these proteins to the process of vascular remodeling. The role of JAK and STAT proteins in vascular remodeling and their functional coupling with Ang II were examined in balloon-injured rat carotid artery. Immunoreactive Jak2, Tyk2, Stat1, and Stat3 were not detected in the intact artery. Immunohistostaining showed transient expressions of these JAKs and STATs in medial and neointimal VSMCs at days 2 and 5, respectively, with a peak at day 7 in both layers. The expressions declined to insignificant levels by day 14. Ang II type 1 receptors (AT1s) were coexpressed in the medial and neointimal VSMCs expressing Jak2 and Stat3. The Jak2 and Stat3 inductions in the injured artery were accompanied by constitutive Jak2 and Stat3 phosphorylations, which were enhanced by ex vivo Ang II stimulation via AT1. Additionally, a Jak2 inhibitor, AG490, blocked the Ang II–induced Stat3 phosphorylation. Furthermore, local treatment with AG490 inhibited constitutive Stat3 phosphorylation and neointimal VSMC replication and subsequently reduced neointima formation in the injured artery. In conclusion, JAK and STAT proteins were inducible in medial and neointimal VSMCs after vascular injury and were functionally coupled to AT1. The inductions of JAKs and STATs would be involved in the mechanisms of neointima formation after vascular injury. (Circ Res. 2000;87:12-18.)

ISSN:0009-7330    

出版商:OVID    

年代:2000

数据来源: OVID

摘要:

The antiatherogenic effect of estrogen is mediated, in part, by inhibitory effects on endothelial vascular cell adhesion molecule-1 (VCAM-1) expression. To determine the mechanism by which estrogen regulates VCAM-1 expression, we compared the effect of 17&bgr;-estradiol (E2) and of the glucocorticoid dexamethasone (Dex) on lipopolysaccharide (LPS)–induced VCAM-1 expression in human endothelial cells. E2decreased LPS-induced VCAM-1 mRNA and protein expression to a greater extent than Dex. Dex, but not E2, stabilized VCAM-1 mRNA. This correlated with inhibition of monocytoid U937 cell adhesion to endothelial cells. Transfection of endothelial cells with a functional VCAM-1 promoter construct showed that E2inhibited LPS-induced VCAM-1 gene transcription more potently than did Dex. However, using a truncated construct containing only the nuclear factor-&kgr;B (NF-&kgr;B)–responsive elements but lacking the consensus sequences for activator protein-1 (AP-1) and GATA, E2and Dex had similar inhibitory effects. Consistently, gel-shift assays showed that E2and Dex comparably inhibit LPS-induced activation of NF-&kgr;B, whereas E2inhibited LPS-induced activation of AP-1 and GATA to a greater extent than Dex. E2inhibition of NF-&kgr;B after LPS treatment was associated with decreased inhibitor &kgr;B (I&kgr;B) kinase activity and with a stabilization of the NF-&kgr;B inhibitor I&kgr;B&agr;. These results indicate that E2decreases VCAM-1 gene expression through the inhibition of NF-&kgr;B, AP-1, and GATA and suggest novel mechanisms for the antiatherogenic effect of estrogen on the vascular wall. (Circ Res. 2000;87:19-25.)

ISSN:0009-7330    

出版商:OVID    

年代:2000

数据来源: OVID

摘要:

Reactive oxygen species (ROS) play an important role in regulating vascular tone and intracellular signaling; the enzymes producing ROS in the vascular wall are, however, poorly characterized. We investigated whether a functionally active NADPH oxidase similar to the leukocyte enzyme, ie, containing the subunits p22phox and gp91phox, is expressed in endothelial cells (ECs) and smooth muscle cells (SMCs). Phorbol 12-myristate 13-acetate (PMA), a stimulus for leukocyte NADPH oxidase, increased ROS generation in cultured ECs and endothelium-intact rat aortic segments, but not in SMCs or endothelium-denuded arteries. NADPH enhanced chemiluminescence in all preparations. p22phox mRNA and protein was detected in ECs and SMCs, whereas the expression of gp91phox was confined to ECs. Endothelial gp91phox was identical to the leukocyte form as determined by sequence analysis. In contrast, mitogenic oxidase-1 (mox1) was expressed in SMCs, but not in ECs. To determine the functional relevance of gp91phox expression, experiments were performed in aortic segments from wild-type, gp91phox−/−, and endothelial NO synthase (eNOS)−/−mice. PMA-induced ROS generation was comparable in aortae from wild-type and eNOS−/−mice, but was attenuated in segments from gp91phox−/−mice. Endothelium-dependent relaxation was greater in aortae from gp91phox−/−than from wild-type mice. The ROS scavenger tiron increased endothelium-dependent relaxation in segments from wild-type, but not from gp91phox−/−mice. These data demonstrate that ECs, in contrast to SMCs, express a gp91phox-containing leukocyte-type NADPH oxidase. This enzyme is a major source for arterial ROS generation and affects the bioavailability of endothelium-derived NO. (Circ Res. 2000;87:26-32.)

ISSN:0009-7330    

出版商:OVID    

年代:2000

数据来源: OVID

摘要:

Voltage-gated Na+channels are critical determinants of electrophysiological properties in the heart. Stimulation of &bgr;-adrenergic receptors, which activate cAMP-dependent protein kinase (protein kinase A [PKA]), can alter impulse conduction in normal tissue and promote development of cardiac arrhythmias in pathological states. Recent studies demonstrate that PKA activation increases cardiac Na+currents, although the mechanism of this effect is unknown. To explore the molecular basis of Na+channel modulation by &bgr;-adrenergic receptors, we have examined the effects of PKA activation on the recombinant human cardiac Na+channel, hH1. Both in the absence and the presence of h&bgr;1subunit coexpression, activation of PKA caused a slow increase in Na+current that did not saturate despite kinase stimulation for 1 hour. In addition, there was a small shift in the voltage dependence of channel activation and inactivation to more negative voltages. Chloroquine and monensin, compounds that disrupt plasma membrane recycling, reduced hH1 current, suggesting rapid turnover of channels at the cell surface. Preincubation with these agents also prevented the PKA-mediated rise in Na+current, indicating that this effect likely resulted from an increased number of Na+channels in the plasma membrane. Experiments using chimeric constructs of hH1 and the skeletal muscle Na+channel, hSKM1, identified the I-II interdomain loop of hH1 as the region responsible for the PKA effect. These results demonstrate that activation of PKA modulates both trafficking and function of the hH1 channel, with changes in Na+current that could either speed or slow conduction, depending on the physiological circumstances. (Circ Res. 2000;87:33-38.)

ISSN:0009-7330    

出版商:OVID    

年代:2000

数据来源: OVID

摘要:

Nitric oxide (NO), generated by platelets through stimulation of nitric oxide synthase (NOS), limits platelet adhesion and aggregation after a prothrombotic stimulus. Platelet &bgr;-adrenoceptors (&bgr;ARs) mediate inhibition of aggregation, but no direct link has been shown between these receptors and platelet adhesion or NO production. We examined NOS activity in human platelets from the conversion of l-[3H]-arginine to l-[3H]-citrulline, after &bgr;AR stimulation or cAMP elevation. Basal NOS activity was 0.11±0.03 pmol l-citrulline/108platelets. The &bgr;AR agonist isoproterenol 1 &mgr;mol/L and the adenylyl cyclase activator forskolin 1 &mgr;mol/L each increased NOS activity, to 0.26±0.04 and 0.23±0.03 pmol l-citrulline/108platelets, respectively (P<0.01 for each). Both responses were abolished by the adenylyl cyclase inhibitor SQ22536 50 &mgr;mol/L. NOS activation by isoproterenol or forskolin was not associated with a change in intracellular Ca2+. In functional studies, isoproterenol inhibited U46619-induced platelet aggregation in a concentration-dependent manner, but this effect was not significantly diminished by NOS inhibition. In contrast, thrombin-stimulated platelet adhesion to cultured human umbilical vein endothelial cell monolayers was inhibited by isoproterenol, and this effect was abolished by NOS inhibition (1.3±0.2% versus 2.6±0.2% respectively;P<0.001). Effects of isoproterenol on NOS activity, platelet aggregation, and adhesion were mediated exclusively through &bgr;2ARs, as determined by coincubation with &bgr;AR subtype-selective antagonists. We conclude that &bgr;2ARs activate platelet NOS by increasing cAMP, and that this activation is Ca2+-independent. &bgr;2ARs may contribute to modulation of platelet aggregation and adhesion to endothelium, and our findings suggest that activation of the l-arginine/NO system mediates the effects of &bgr;2ARs on adhesion but not aggregation. (Circ Res. 2000;87:39-44.)

ISSN:0009-7330    

出版商:OVID    

年代:2000

数据来源: OVID

摘要:

Editing of the nonprotein amino acid homocysteine by certain aminoacyl-tRNA synthetases results in the formation of the thioester homocysteine thiolactone. Here we show that in the presence of physiological concentrations of homocysteine, methionine, and folic acid, human umbilical vein endothelial cells efficiently convert homocysteine to thiolactone. The extent of this conversion is directly proportional to homocysteine concentration and inversely proportional to methionine concentration, suggesting involvement of methionyl-tRNA synthetase. Folic acid inhibits the synthesis of thiolactone by lowering homocysteine and increasing methionine concentrations in endothelial cells. We also show that the extent of post-translational protein homocysteinylation increases with increasing homocysteine levels but decreases with increasing folic acid and HDL levels in endothelial cell cultures. These data support a hypothesis that metabolic conversion of homocysteine to thiolactone and protein homocysteinylation by thiolactone may play a role in homocysteine-induced vascular damage. (Circ Res. 2000;87:45-51.)

ISSN:0009-7330    

出版商:OVID    

年代:2000

数据来源: OVID