摘要:

Apolipoprotein E-deficient (ApoE-/-) mice develop atherosclerotic lesions throughout the arterial tree, including the carotid bifurcation. Although the expression of adhesion molecules such as ICAM-1, vascular cell adhesion molecule-1 (VCAM-1), and P-selectin on endothelium that overlie atherosclerotic plaques has been implicated in monocyte recruitment to developing lesions, monocyte adhesion in atherosclerotic vessels has not been observed directly. To investigate which adhesion molecules may be important in monocyte adhesion to atherosclerotic lesions, an isolated mouse carotid artery preparation was developed and perfused with mononuclear cells. We show rolling and attachment of the human monocytic cell line U937 and the mouse monocyte-macrophage cell line P388D1 in carotid arteries from 10- to 12-week-old ApoE-/-and C57BL/6 wild-type mice fed a Western-type diet (21% fat wt/wt) for 4 to 5 weeks. No rolling was observed in carotid arteries from C57BL/6 or BALB/c wild-type mice fed a chow diet and little was observed in BALB/c mice fed a Western-type diet. This model represents early lesion development as shown by minimal macrophage infiltration in the intima of carotid arteries from ApoE-/-mice fed a Western-type diet. Rolling was observed at shear stresses that were characteristic of the low-shear recirculation zone near the carotid bifurcation. Mononuclear cell attachment and rolling were significantly inhibited by monoclonal antibody blockade of P-selectin or its leukocyte ligand P-selectin glycoprotein ligand-1. Rolling velocities increased after monoclonal antibody blockade of mononuclear cell alpha4-integrinor VCAM-1, which indicates that alpha4-integrininteracting with VCAM-1 stabilizes rolling interactions and prolongs monocyte transit times. (Circ Res. 1999;84:1237-1244.)

ISSN:0009-7330    

出版商:OVID    

年代:1999

数据来源: OVID

摘要:

We have previously shown that CD18 and alpha4integrin were important in the adherence of emigrated neutrophils to cardiac myocytes. Whether either of these molecules is important in myocyte dysfunction is unclear. In this study, we measured contractility as an index of myocyte function. Control contractility was compared with shortening response in myocytes exposed to neutrophils in the presence and absence of anti-CD18 or anti-alpha4antibodies. Control unloaded cell shortening, expressed as a percentage of resting cell length, measured 10.06 +/- 1.16% (n=10) at 5 minutes. Circulating neutrophils caused a 35% reduction in cell shortening, an event prevented by anti-CD18, but not by anti-alpha4antibody. When emigrated neutrophils were added to the myocytes, a profound reduction (50%) in unloaded cell shortening was noted. A significant increase in CD18 and alpha4integrin was found on emigrated neutrophils. Addition of anti-CD18 antibody did not protect the myocyte from the emigrated neutrophils, whereas the addition of an anti-alpha4antibody significantly reduced neutrophil-induced cell shortening, despite some neutrophils still adhering to the myocytes. Furthermore, emigrated neutrophils were able to cause myocytes to go into contracture within 5 minutes in the presence of neutrophils with or without anti-CD18 antibody. In addition to the impairment in unloaded cell shortening, at later times (10 minutes), neutrophils also caused a 40% reduction in the rate of contraction and relaxation. The addition of either anti-CD18 or anti-alpha4antibody protected the myocytes from these changes. The data suggest that immunosuppression of CD18 on emigrated neutrophils was only partially effective in reducing myocyte dysfunction. In contrast, immunosuppression of the alpha4integrin alone was sufficient to dramatically reduce all parameters of cell dysfunction measured in this study. (Circ Res. 1999;84:1245-1251.)

ISSN:0009-7330    

出版商:OVID    

年代:1999

数据来源: OVID

摘要:

We performed an initial screen of 11 rat strains by use of a standard balloon injury to the left iliac artery to observe whether genetically determined differences existed in the development of neointimal hyperplasia. Neointimal hyperplasia was assayed 8 weeks after the vascular injury on coded microscopic sections. Statistically significant differences in the percentages of the vascular wall cross-sectional areas composed of intima (percentage intima) secondary to neointimal hyperplasia were noted among the different rat strains (P<0.02), with the Brown-Norway (BN), Dark Agouti, and Milan normotensive strain rats having the highest and the spontaneously hypertensive rats (SHR) having the lowest percentages of intima. In a separate experiment, F1hybrids of SHRxBN strains and parental BN and SHR underwent the vascular injury, and the parental strains again showed a statistically significant difference from one another in the mean percentage of intima (P<0.0001). The F1hybrids showed an average percentage of intima intermediate between those of the parental strains. The average lumen size of the injured BN vessels were significantly smaller than that of the noninjured control vessels (P=0.044), but this significance disappeared when the circular areas of these vessels were calculated without taking neointimal growth into consideration (P=0.649). These results provide the groundwork for a genetic linkage analysis to identify the genes that influence the development of neointimal hyperplasia after vascular injury. (Circ Res. 1999;84:1252-1257.)

ISSN:0009-7330    

出版商:OVID    

年代:1999

数据来源: OVID

摘要:

Platelet-derived growth factor (PDGF) is a mitogen and chemoattractant for a wide variety of cell types. The genes encoding PDGF A chain (PDGF-A) and PDGF B chain (PDGF-B) reside on separate chromosomes and are independently regulated at the level of transcription. Regulatory events underlying inducible PDGF-A expression have been the focus of much investigation. However, mechanisms that inhibit transcription of this gene are not well understood. In this study, we report the capacity of a newly cloned DNA binding factor, GC factor 2 (GCF2), to repress expression driven by the human PDGF-A promoter. 5[prime] Deletion and transient cotransfection analysis in vascular endothelial cells revealed that GCF2 repression is mediated by a nucleotide region located in the proximal region of the PDGF-A promoter. Electrophoretic mobility shift assays demonstrate that GCF2 binds to this region in a specific and dose-dependent manner. Interestingly, the site bound by GCF2 overlaps those for specificity protein-1 (Sp1) and early growth response factor-1 (Egr-1), zinc finger transcription factors that direct basal and inducible expression of the PDGF-A gene. Gel shift experiments revealed that GCF2 competes with these factors for interaction with the PDGF-A promoter. Overexpression of GCF2 suppressed endogenous PDGF-A expression in vascular endothelial cells and smooth muscle cells. GCF2 was induced on mechanical injury of cells in culture as well as after balloon injury of the rat carotid artery wall. Time course studies revealed the sustained induction of GCF2 after injury while PDGF-A levels sharply returned to baseline. Smooth muscle cell proliferation was inhibited by GCF2, an effect reversed by the addition of exogenous PDGF-AA. These findings demonstrate negative regulation of PDGF-A expression by GCF2. This is the first report of the induction of an endogenous transcriptional repressor in the rat vessel wall. (Circ Res. 1999;84:1258-1267.)

ISSN:0009-7330    

出版商:OVID    

年代:1999

数据来源: OVID

摘要:

The vitronectin receptor (alphavbeta3) mediates several biological processes that are critical to the formation of a neointima after coronary interventions. Blockade of alphavbeta3could reduce neointima formation by inhibiting smooth muscle cell migration, decreasing transforming growth factor-beta (1) expression, enhancing apoptosis, or reducing neovasculature. The effects of short-term administration of Vitaxin, a humanized monoclonal antibody to alphavbeta3, on the responses to balloon injury were tested in hyperlipidemic rabbits. Balloon angioplasty was performed on the iliac arteries of male New Zealand White rabbits that were fed an atherogenic diet for 1 week before injury and until euthanization at 4 weeks. Rabbits were given either saline (control) or 1 of 2 dosing regimens of Vitaxin (high dose, 5.0 mg/kg, and low dose, 0.5 mg/kg), which were administered intra-arterially before injury and intramuscularly on days 2 and 3. High-dose and low-dose Vitaxin were equally effective in decreasing neointima formation even in the presence of hypercholesterolemia, a stimulus to alphavbeta3expression. Vitaxin reduced transforming growth factor-beta1and enhanced apoptosis in injured arteries. Despite these positive effects, Vitaxin administration was associated with a reduction in artery size, indicating a negative effect on remodeling. Vitaxin has a potential role in preventing intimal hyperplasia, especially if the negative effects on remodeling can be overcome, by dose adjustment or other strategies. (Circ Res. 1999;84:1268-1276.)

ISSN:0009-7330    

出版商:OVID    

年代:1999

数据来源: OVID

摘要:

Three connexins, Cx43, Cx40, and Cx37, have been found by protein or mRNA analysis to be prominent in mammalian blood vessels, but electrophysiological characterization of gap junction channels in freshly isolated vascular smooth muscle cells (SMCs) has not previously been reported. We used a dual-perforated patch-clamp technique to study gap junction conductances in SMC pairs from rat basilar arteries. Macroscopic junctional conductance (Gj) measured in 98 cell pairs with either Cs+or K+ranged between 0.68 and 24.8 nS. In weakly coupled cells (Gj300 pS, were identified that have not been previously reported in vascular SMCs. Macroscopic recordings revealed currents that deactivated incompletely over a broad range of transjunctional potentials. In about half of the pairs, we identified macroscopic as well as single-channel currents that exhibited marked voltage asymmetry, consistent with nonhomotypic, ie, either heterotypic or heteromeric channels. Our data indicate that basilar artery SMCs are coupled in vivo in a richly complex manner, involving Cx43, Cx40, and other large-conductance channels, and that a significant number of couplings involve putative nonhomotypic channels. (Circ Res. 1999;84:1277-1284.)

ISSN:0009-7330    

出版商:OVID    

年代:1999

数据来源: OVID

摘要:

The atheroprotective effects of estrogen are well established and the presence of an estrogen receptor in vascular tissues has recently been reported. Therefore, we investigated the localization of the estrogen-producing enzyme aromatase in vascular tissues to assess the possible contribution of endocrine, paracrine, and autocrine modes of action. Aromatase was found in human vascular smooth muscle cells (SMCs) but not in endothelial cells on in situ hybridization. These observations were further supported by quantitative analysis of aromatase mRNA and the activity in 15 human vascular specimens. Only trace levels of expression were detected in the 3 infants examined, whereas 0.0088 to 0.0806 amol/[micro sign]g RNA of aromatase mRNA and 12.9 to 122.3 fmol [middle dot] h-1[middle dot] mg-1protein of the activity were detected in 12 of the adult individuals. The switching of tissue-specific exon 1 of the human aromatase gene was also observed in some cases. Aromatase was found to be expressed only in cultured SMCs and not in cultured endothelial cells of human aorta and pulmonary artery and to be regulated through dexamethasone and the signaling pathways of protein kinase A and C. Study results revealed the localized expression of aromatase in vascular SMCs, which indicated a possible direct action of locally produced estrogen in an autocrine or paracrine manner, with possible cross talk between smooth muscle and endothelial cells. (Circ Res. 1999;84:1285-1291.)

ISSN:0009-7330    

出版商:OVID    

年代:1999

数据来源: OVID

摘要:

Communication between the cytoplasm and nucleoplasm of cardiac cells occurs by molecular transport through nuclear pores. In lower eukaryotes, nuclear transport requires the maintenance of cellular energetics and ion homeostasis. Although heart muscle is particularly sensitive to metabolic stress, the regulation of nuclear transport through nuclear pores in cardiomyocytes has not yet been characterized. With the use of laser confocal and atomic force microscopy, we observed nuclear transport in cardiomyocytes and the structure of individual nuclear pores under different cellular conditions. In response to the depletion of Ca2+stores or ATP/GTP pools, the cardiac nuclear pore complex adopted 2 distinct conformations that led to different patterns of nuclear import regulation. Depletion of Ca2+indiscriminately prevented the nuclear import of macromolecules through closure of the nuclear pore opening. Depletion of ATP/GTP only blocked facilitated transport through a simultaneous closure of the pore and relaxation of the entire complex, which allowed other molecules to pass into the nucleus through peripheral routes. The current study of the structural plasticity of the cardiac nuclear pore complex, which was observed in response to changes in cellular conditions, identifies a gating mechanism for molecular translocation across the nuclear envelope of cardiac cells. The cardiac nuclear pore complex serves as a conduit that differentially regulates nuclear transport of macromolecules and provides a mechanism for the control of nucleocytoplasmic communication in cardiac cells, in particular under stress conditions associated with disturbances in cellular bioenergetics and Ca2+homeostasis. (Circ Res. 1999;84:1292-1301.)

ISSN:0009-7330    

出版商:OVID    

年代:1999

数据来源: OVID

摘要:

Heart failure is the leading cause of mortality in patients with transfusional iron (Fe) overload in which myocardial iron uptake ensues via a transferrin-independent process. We examined the ability of L-type Ca2+channel modifiers to alter Fe (2+) uptake by isolated rat hearts and ventricular myocytes. Perfusion of rat hearts with 100 nmol/L59Fe2+and 5 mmol/L ascorbate resulted in specific59Fe2+uptake of 20.4 +/- 1.9 ng of Fe per gram dry wt. Abolishing myocardial electrical excitability with 20 mmol/L KCl reduced specific59Fe2+uptake by 60 +/- 7% (P<0.01), which suggested that a component of myocardial Fe2+uptake depends on membrane voltage. Accordingly,59Fe2+uptake was inhibited by 10 [micro sign]mol/L nifedipine (45 +/- 12%, P<0.02) and 100 [micro sign]mol/L Cd (2+) (86 +/- 3%; P<0.001) while being augmented by 100 nmol/L Bay K 8644 (61 +/- 18%, P<0.01) or 100 nmol/L isoproterenol (40 +/- 12%, P<0.05). By contrast, uptake of 100 nmol/L ferric iron (59) Fe3+) was significantly lower (1.4 +/- 0.3 ng Fe per gram dry wt; P<0.001) compared with divalent iron. These data suggest that a component of Fe2+uptake into heart occurs via the L-type Ca2+channel in myocytes. To investigate this further, the effects of Fe2+on cardiac myocyte L-type Ca2+currents were measured. In the absence of Ca2+, noninactivating nitrendipine-sensitive Fe2+currents were recorded with 15 mmol/L [Fe2+] (o). Low concentrations of Fe2+enhanced Ca2+current amplitude and slowed inactivation rates, which was consistent with Fe2+entry into the cell, whereas higher Fe2+levels caused dose-dependent decreases in peak current. Fe3+had no effect on current amplitude or decay. Combined, our data suggest that myocardial Fe2+uptake occurs via L-type Ca2+channels and that blockade of these channels might be useful in the treatment of patients with excessive serum iron levels. (Circ Res. 1999;84:1302-1309.)

ISSN:0009-7330    

出版商:OVID    

年代:1999

数据来源: OVID

摘要:

Cardiac myosin heavy chain (MHC) isoforms are known to play a key role in defining the dynamic contractile behavior of the heart during development. It remains unclear, however, whether cardiac MHC isoforms influence other important features of cardiac contractility, including the Ca2+sensitivity of isometric tension development. To address this question, adult rats were treated chemically to induce the hypothyroid state and cause a transition in the ventricular cardiac MHC isoform expression pattern from predominantly the alpha-MHC isoform to exclusively the beta-MHC isoform. We found a significant desensitization in the Ca2+sensitivity of tension development in beta-MHC-expressing ventricular myocytes (pCa50=5.51 +/- 0.03, where pCa is -log[Ca2+], and pCa50is pCa at which tension is one-half maximal) compared with that in predominantly alpha-MHC-expressing myocytes (pCa50=5.68 +/- 0.05). No differences between the 2 groups were observed in the steepness of the tension-pCa relationship or in the maximum isometric force generated. Instantaneous stiffness measurements were made that provide a relative measure of changes in the numbers of myosin crossbridges attached to actin during Ca2+activation. Results showed that the relative stiffness-pCa relationship was shifted to the right in beta-MHC-expressing myocytes compared with the alpha-MHC-expressing cardiac myocytes (pCa50= 5.47 +/- 0.05 versus 5.76 +/- 0.05, respectively). We conclude that MHC isoform switching in adult cardiac myocytes alters the Ca2+sensitivity of stiffness and tension development. These results suggest that the activation properties of the thin filament are in part MHC isoform dependent in cardiac muscle, indicating an additional role for MHC isoforms in defining cardiac contractile function. (Circ Res. 1999;84:1310-1317.)

ISSN:0009-7330    

出版商:OVID    

年代:1999

数据来源: OVID