摘要:

Regulation of [Ca2+]i and modulation of the [Ca2+]i sensitivity of myosin phosphorylation in smooth muscles may involve phosphorylation of one or more proteins on tyrosine residues. We tested this hypothesis by measuring tyrosine phosphorylation of proteins extracted from swine carotid artery and separated by SDS gel electrophoresis. Tyrosine phosphorylation was estimated by the binding of antiphosphotyrosine antibodies to proteins in SDS gels. We found four bands with approximate molecular weights of 120, 110, 85 and 75 kD in which tyrosine phosphorylation increased 1 min after histamine stimulation. After washout of histamine, dephosphorylation of the proteins in these four bands occurred at a slower rate than relaxation. Tyrosine phosphorylation of these four protein bands did not correlate with the [Ca2+]i sensitivity of myosin phosphorylation following agonist or high [K+]o stimulation. Phorbol dibutyrate stimulation also induced tyrosine phosphorylation of these four protein bands. These data suggest that tyrosine phosphorylation of the proteins in these four bands may be involved in the initial phase of swine carotid artery contraction. However, there was, at most, only a minor involvement of tyrosine phosphorylation of these four protein bands in the sustained phase of contraction or in the regulation of [Ca2+]i sensitivity of myosin phosphorylation in the swine carotid artery. These data do not rule out a role for other, less abundant tyrosine phosphoproteins in the regulation of sustained contraction or the [Ca2+]i sensitivity of myosin phosphorylation.

ISSN:1018-1172    

DOI:10.1159/000159196

出版商:S. Karger AG    

年代:1997

数据来源: Karger

ISSN:1018-1172    

DOI:10.1159/000159263

出版商:S. Karger AG    

年代:1997

数据来源: Karger

摘要:

Pituitary adenylate-cyclase-activating peptides (PACAPs) are potent dilators of arteries, including human coronary arteries. We tested the importance of specific K+ channel regulatory mechanisms in human arterial smooth muscle relaxation induced by PACAPs, using contraction and patch clamp measurements on human coronary artery vascular smooth muscle cells. PACAP27 and PACAP38 produced dose-dependent relaxations of 5 µM PGF2α-preconstricted rings, with half-maximal relaxations at 1.0 nM and 2.0 nM, respectively. Both peptides induced complete relaxation at 100 nM. Pretreatment of the vessels with the ATP-dependent K+ (Katp) channel blocker glibenclamide (1 µM) or with the Ca2+-activated K+ (KCa) channel blocker iberiotoxin (100 nM) inhibited PACAP27-induced relaxation in an additive manner. Moreover, in the patch clamp experiments on freshly isolated cells from human coronary arteries, PACAP27 (100 nM) induced a large, nonrectifying, outward (Ik(atp)) K+ current in a proportion of cells and a voltage-dependent outward (IK(Ca)) K+ current in other cells. The PACAP27-induced IK(ATP) was blocked by glibenclamide (3 µM), while the PACAP27-stimulated IK(Ca) was blocked by iberiotoxin (100 nM). These findings provide the first evidence that relaxation of arterial smooth muscle cells by PACAPs is mediated by opening of KATP and KCa channels. The data indicate that both KATP and KCa channels in vascular smooth muscle cells may serve as final common pathway to induce vasorelaxation by endogenous vasoactive signals in

ISSN:1018-1172    

DOI:10.1159/000159197

出版商:S. Karger AG    

年代:1997

数据来源: Karger

摘要:

The specialized functions of endothelium require intercellular communication between endothelial cells within the monolayer, and between endothelium and other cells present in the vessel wall. This is accomplished by a combination of paracrine soluble mediators and direct gap-junctional intercellular communication (GJIC) mediated by a family of connexin proteins. A prominent connexin expressed by vascular cells in vivo and in vitro is connexin 43 (Cx43). We have investigated the in vivo gene regulation of Cx43 in the context of vascular pathology, as a result of mechanical injury, hypercholesterolemia or both. The aortoiliac bifurcation in the rabbit was examined following three types of insult: (1) diet-induced hypercholesterolemia resulting in macrophage-rich fatty streak lesions, (2) mechanical, stretch-denudation injury resulting in intimal smooth muscle cell (SMC) proliferation and (3) mechanical injury superimposed on hypercholesterolemia resulting in a complex vascular lesion having characteristics of both interventions. The normal rabbit iliac artery expressed approximately equal levels of Cx43 mRNA in the medial SMC layers and in the endothelium. In hypercholesterolemia-induced atherosclerosis, Cx43 expression was most prominent in macrophage foam cells even though normocholesterolemic precursor monocytes did not express Cx43 mRNA. Antibodies directed specifically to Cx43 protein confirmed the expression of macrophage gap junction protein in these cells. Medial SMC in hypercholesterolemia exhibited less Cx43 than their normal counterparts in control animals. Mechanical injury in the absence of hypercholesterolemia resulted in intimal thickening in which Cx43 expression in the intimal SMC was equivalent to that in the subjacent medial SMC, both being approximately equivalent to normal uninjured rabbit medial SMC expression. Cell-specific expression of Cx43 in combined mechanical injury/hypercholesterolemia was similar to that observed in hypercholesterolemia alone: Cx43 upregulation in macrophages, while medial SMC were downregulated. Normo- and hypercholesterolemic alveolar macrophages of the lung and Kupffer cells of the liver did not exhibit induction of Cx43 mRNA, nor did macrophages isolated from peritoneal or bronchial lavage fluid of the same animals. This work extends our previous finding of Cx43 upregulation in human atherectomy tissue and demonstrates that atherosclerotic lesions in situ, in a controlled animal model of atherosclerosis, exhibit cell-specific changes in Cx43 gene expression. Changes in medial SMC migration, proliferation and phenotype, as well as enhanced interactions between adherent/infiltrating monocytes and endothelium may be related to modified GJIC pathways in the vessel wall.

ISSN:1018-1172    

DOI:10.1159/000159198

出版商:S. Karger AG    

年代:1997

数据来源: Karger

摘要:

Percutaneous transluminal angioplasty relieves discrete arterial stenosis but causes extensive vascular injury. There is denudation of the endothelium and variable medial disruption, but the effect on adventitial structures has not been studied in detail. We have investigated the innervation of the left and right carotid arteries after unilateral balloon-catheter-induced injury. Immunohistochemical examination of the arteries 1 day after Fogarty-catheter-induced injury of the left common carotid artery revealed a reduction in the density of protein gene product 9.5 (PGP)-, substance P (SP)- and calcitonin-gene-related peptide (CGRP)-containing nerves close to the medial smooth muscle of the injured vessel. At the same time, on the side contralateral to the injury, there was a substantial increase in the density of PGP-, SP- and CGRP-containing nerves innervating the carotid artery and vasa vasorum compared to controls. Immunoassay data from these vessels showed a selective increase in SP and CGRP contents of the contralateral carotid artery (SP, controls 0.02 ± 0.01, operated 0.59 ± 0.32 pmol/cm; CGRP, controls 0.03 ± 0.01, operated 0.14 ± 0.03 pmol/cm, n = 6, p& < 0.05). Neuropeptide Y levels were unchanged. Twenty-eight days after surgery, at which time a neointima was present, peptide levels were no different from controls, and the innervation of both the left and right carotid arteries and vasa vasorum was indistinguishable from the controls. In conclusion, balloon-catheter-induced injury includes damage to the perivascular nerves and induces a transient increase in the density of sensory neuropeptide-containing nerves innervating the contralateral, uninjured side. This novel observation may reflect neurocompensatory responses to vascular inj

ISSN:1018-1172    

DOI:10.1159/000159199

出版商:S. Karger AG    

年代:1997

数据来源: Karger

ISSN:1018-1172    

DOI:10.1159/000317489

出版商:S. Karger AG    

年代:1997

数据来源: Karger

摘要:

The potent growth factors and chemoattractants α-thrombin and transforming growth factor-β1 (TGF-β1) have both been identified at sites of arterial injury, however the interaction between these two factors has not been defined. By Northern hybridization analyses, accumulation of both a 1.9- and a 2.4-kb transcript of TGF-β1 were detected and occurred in a time- and dose-dependent fashion following α-thrombin stimulation of cultured vascular smooth muscle cells (VSMC). This induction of TGF-β1 mRNA required the proteolytic activity of thrombin and was mimicked by a thrombin-receptor-(TR)-activating peptide or TRAP (SFFLRNP). Increases in α-thrombin-induced TGF-β1 message expression were insensitive to cycloheximide, but sensitive to actinomycin D. Furthermore, the induction of TGF-β1. mRNA expression correlated with the production of latent TGF-β1 protein in α-thrombin-conditioned media. In summary, α-thrombin stimulation of VSMC induces transcriptional activation of the TGF-β1 gene through proteolytic activation of the cloned seven-transmembrane TR resulting in the formation of latent TGF-β1 protein. These results demonstrate a potential mechanism whereby α-thrombin may modulate the vascular response to injury through TGF-β1-depen

ISSN:1018-1172    

DOI:10.1159/000159200

出版商:S. Karger AG    

年代:1997

数据来源: Karger

ISSN:1018-1172    

DOI:10.1159/000159264

出版商:S. Karger AG    

年代:1997

数据来源: Karger

摘要:

The effects on rat aorta of EUK-8, a salen-manganese complex with high superoxide dismutase and catalase activities, were investigated. EUK-8 protected the acetylcholine-induced relaxation of rat aortic rings from inhibition by superoxide anions and reduced H2O2-induced relaxation. Moreover, EUK-8 dose-dependently relaxed rat aorta precontracted with phenyl-ephrine (10–6M) and decreased the vascular tone of noncontracted aortic rings. The relaxant effect of EUK-8 was significantly potentiated by endothelium abrasion and/or preincubation with N-nitro-L-arginine methyl ester (10–5M and 5 × 10–4M), an inhibitor of nitric oxide synthase. lndomefhacin (10–5M)had no effect on the action of EUK-8, showing that it was not dependent on prostacyclin synthesis. Methylene blue (10–5M), an inhibitor of soluble guanylate cyclase, partly abolished relaxation induced by EUK-8. Incubation of rat aorta with EUK-8 (10–4M)induced an increase in vascular cyclic AMP content. The lack of inhibition by dl-propranolol showed that adenylate cyclase activation by EUK-8 was not mediated through β-adrenergic receptors. The inhibition of the effects of EUK-8 by tetraethylammonium (10–2M)and glibenclamide (10–5 and 2 × 10–5M)showed the implication of potassium channels in the intracellular cascade triggered by EUK-8. The vasorelaxant activity of EUK-8 was neither affected by xanthine oxidase inhibition (incubation with oxypurinol 25 µM)nor by superoxide anion scavenging (incubation with oxypurinol 125 µM). Finally, the ligand for EUK-8 (EUK-8 without manganese), which has the same aromatic structure as EUK-8 without its antioxidant activities because of the absence of manganese, conversely potentiated phenylephrine-induced contraction of aortic rings. We conclude that the vasorelaxant effect of EUK-8 observed under our experimental conditions is essentially mediated through an activation of adenylate cyclase and soluble guanylate cyclase of smooth muscle cells and is different from a classical antioxidant effect of prot

ISSN:1018-1172    

DOI:10.1159/000159201

出版商:S. Karger AG    

年代:1997

数据来源: Karger

摘要:

The involvement of inflammatory mechanisms in the progression of atherosclerosis has recently been suggested. Monocyte chemotactic protein 1 (MCP-1) is a soluble protein which is implicated in acute and chronic inflammatory processes, including atherosclerosis. We evaluated the effect of human recombinant MCP-1 on the in vitro proliferation of rat vascular smooth muscle cells (VSMCs). Incubation of VSMCs with MCP-1 (50-200 ng/ml) in the presence of 0.5% FCS significantly increased cell proliferation, [3H]-thymidine incorporation and the proliferative S fraction, measured by flow cytometry, compared to control cells. The proliferative effect of MCP-1 was specific, as shown by inhibition with a rabbit polyclonal serum to MCP-1. Moreover, the mitogenic effect of MCP-1 was significantly inhibited by downregulation of protein kinase C (PKC) activity and by incubation with H-7, a protein kinase inhibitor, suggesting the involvement of the PKC system. Verapamil, a Ca2+ channel blocker, also reduced the stimulatory effect of MCP-1 on cell proliferation. This study demonstrates that MCP-1 does not merely have a chemotactic activity, but also a mitogenic effect on cultured rat VSMCs.

ISSN:1018-1172    

DOI:10.1159/000159202

出版商:S. Karger AG    

年代:1997

数据来源: Karger