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N J Severs

Publications and source records attributed to N J Severs.

At least 55 records · Page 3Linked to original sources
PubMed1998

Connexin expression in Huntington's diseased human brain.

In Huntington's diseased human brain, it is in the caudate nucleus (CN) and globus pallidus (GP) of the basal ganglia where nerve cell death is seen most dramatically. The distribution of five gap junction proteins (connexins 26, 32, 40, 43 and 50) has been examined in these areas in normal and Huntington's diseased human brain using immunohistochemical techniques. There was no Cx50 expression observed and Cx40 was localized in the endothelial cells of blood vessels, with the Huntington's diseased brains having more numerous and smaller blood vessels than normal tissue. Cx26 and Cx32 revealed a similar distribution pattern to each other in both normal and diseased brains with little labelling in the CN but clear labelling in the GP. Cx43, expressed by astrocytes, was the most abundant connexin type of those studied. In both normal and diseased brains Cx43 in the GP was homogeneously distributed in the neuropil. In the CN, however, Cx43 density was both increased with Huntington's disease and became located in patches. Glial fibrillary acidic protein(GFAP) staining of astrocytes was also highly increased in the CN compared with normal brains. These labelling patterns indicate a reactive astrocytosis around degenerating neurons with an increased expression of astrocytic gap junctions. The enhanced coupling state between astrocytes, assuming the junctions are functional, could provide an increased spatial buffering capacity by the astrocytes in an attempt to maintain a proper environment for the neurons, helping promote neuronal survival in this neurodegenerative disorder.

Animals↗
PubMed1998

Identification of two further gap-junctional proteins, connexin40 and connexin45, in human myometrial smooth muscle cells at term.

The powerful synchronous contractions of the uterus in labor depend on electrical coupling of myometrial smooth muscle cells by gap junctions. In humans and other mammals, gap junctions are scarce in the myometrium of the non-pregnant uterus, but become abundant at term and/or with the onset of labor. Previous work has shown that the gap-junctional protein (connexin) expressed by human myometrial smooth muscle cells is connexin43, the same connexin type that predominates in cardiac muscle. Here we show that two further gap junctional proteins, connexin40 and connexin45, are expressed by the myometrial smooth muscle cells of the human uterus at term. Transcripts encoding the human isoforms of these connexins were demonstrated by Northern blot analysis, and immunoconfocal microscopy enabled precise localization of the corresponding proteins to punctate contact points (i.e., gap junctions) between interacting smooth muscle cells. Double labeling demonstrated that, while some fluorescent spots comprise only connexin43, both connexin40 and connexin45 predominantly colocalize to connexin43-positive fluorescent spots. Triple labeling revealed that where all three connexin types were expressed, they frequently localized to the same gap junction spot. As gap-junctional channels composed of different connexin types have been demonstrated in vitro to have different functional properties, multiple connexin expression may contribute to modulation of gap junction function in human myometrial smooth muscle cells in vivo.

Blotting, Northern↗
PubMed1998

Co-localization of dystrophin and beta-dystroglycan demonstrated in en face view by double immunogold labeling of freeze-fractured skeletal muscle.

An absence of dystrophin causes Duchenne muscular dystrophy, but the precise mechanism underlying necrosis of the muscle cells is still unclear. Dystrophin and beta-dystroglycan are components of a complex of at least nine proteins, the dystrophin-glycoprotein complex (DGC), that links the membrane cytoskeleton to extracellular elements in skeletal and cardiac muscle. Biochemical studies indicate that dystrophin is bound to other components of the DGC via beta-dystroglycan, which suggests that the distribution of these two proteins should be almost identical. In this study, therefore, we examined the spatial relationship between dystrophin and beta-dystroglycan with a range of different imaging techniques to investigate the extent of the predicted co-localization. We used (a) double immunogold fracture-label, a freeze-fracture cytochemical technique that allows high-resolution face-on views of labeled membrane components in thin sections and in platinum-carbon replicas, (b) double immunogold labeling of cryosections and (c) confocal microscopy. Both dystrophin and beta-dystroglycan were found over the entire fiber surface and, when labeled singly, the nearest neighbor spacing of labeling sites for the two proteins was indistinguishable. With double labeling, very close co-localization could be demonstrated. The results support the conclusion that dystrophin and beta-dystroglycan directly interact at the muscle plasma membrane. (J Histochem Cytochem 46:945-953, 1998)

Animals↗
PubMed1998

Structural, biochemical and functional effects of distending pressure in the human saphenous vein: implications for bypass grafting.

BACKGROUND: Distension of the saphenous vein before and after coronary artery bypass grafting results in damage to mechanisms that regulate vascular tone. We have investigated the relationship between the magnitude of distending pressure and the degree of structural, biochemical and functional damage to the vessel wall. METHODS: Vessel segments that had been distended to either 100 or 300 mmHg were set up in isolated organ baths and the function of the smooth muscle and endothelial cells examined. All segments examined were then fixed for assessment of structural damage by scanning electron microscopy and for immunocytochemical localisation of endothelial nitric oxide synthase. RESULTS: Segments of saphenous vein distended to 100 mmHg retained their responsiveness to KCl (90 mmol/l) and phenylephrine (10(-6) mol/l), but those pressurised to 300 mmHg had significantly reduced responses to both agents. There was also a significant reduction in response to the endothelium-dependent dilators, acetylcholine (10(-10)-10(-6) mol/l) and bradykinin (10(-10)-10(-6) mol/l) in those segments distended to 300 mmHg. Quantitative studies of structural endothelial damage showed a significant loss of endothelium at 300 mmHg distension pressure. Remaining endothelial cells retained strong positive staining for endothelial nitric oxide synthase. By electron microscopic examination, those vessels distended to 100 mmHg showed lifting and rounding of individual cells, whereas segments distended to 300 mmHg revealed major areas of denuded endothelium. CONCLUSIONS: Distension of saphenous veins to pressures equivalent to those in the systemic circulation result in structural and biochemical changes in the endothelium that are not paralleled by immediate functional vasomotor changes.

Aged↗
PubMed1997

Disturbed connexin43 gap junction distribution correlates with the location of reentrant circuits in the epicardial border zone of healing canine infarcts that cause ventricular tachycardia.

BACKGROUND: Slow, nonuniform conduction caused by abnormal gap-junctional coupling of infarct-related myocardium is thought to be a component of the arrhythmogenic substrate. The hypothesis that changes in gap-junctional distribution in the epicardial border zone (EBZ) of healing canine infarcts define the locations of reentrant ventricular tachycardia (VT) circuits was tested by correlating activation maps of the surviving subepicardial myocardial layer with immunolocalization of the principal gap-junctional protein, connexin43 (Cx43). METHODS AND RESULTS: The EBZ overlying 4-day-old anterior infarcts in three dogs with inducible VT and three noninducible dogs was mapped with a high-resolution electrode array and systematically examined by standard histology and confocal immunolocalization of Cx43. The thickness of the EBZ was significantly less in the hearts with (538 +/- 257 microns) than without (840 +/- 132 microns; P < .05) VT. At the interface with the underlying necrotic cells, the EBZ myocardium showed a marked disruption of gap-junctional distribution, with Cx43 labeling abnormally arrayed longitudinally along the lateral surfaces of the cells. In the EBZ of all hearts, the disrupted Cx43 labeling extended part of the way to the epicardial surface, with the most superficial epicardial myocytes having the normal transversely orientated pattern. Only in the hearts with inducible VT did the disorganization extend through the full thickness of the surviving layer at sites correlating with the location of the central common pathways of the figure-of-8 reentrant VT circuits. CONCLUSIONS: Altered gap-junctional distribution is part of the early remodeling of myocardium after infarction, and by defining the location of the common central pathway of the reentrant VT circuits, it may be a determinant of VT susceptibility.

Animals↗
PubMed1997

Connexin43 gap junction levels during development of the thoracic aorta are temporally correlated with elastic laminae deposition and increased blood pressure.

A characteristic property of the vascular smooth muscle cell is its ability to modulate between a contractile phenotype, responsible for control of vascular tone, through to a synthetic phenotype, capable of migration and synthesis of extracellular matrix molecules. Smooth muscle cells are coupled by gap junctions, the membrane structures which permit direct intercelluar passage of ions and small molecules, and which play a role both in electrical coupling and intercellular communication during patterning and development. We have previously found that connexin43 type gap junction expression is upregulated in the synthetic phenotype smooth muscle cell in vitro and during atherosclerotic plaque formation in human coronary arteries. On the basis of immunohistochemical labelling, confocal laser scanning microscopy and digital image analysis, we now report that relatively high levels of connexin43 are expressed during development of the rat thoracic aorta, temporally correlating with reported periods of smooth muscle cell proliferation and secretion of elastic laminae. A major peak in expression occurs at seven days post-natal, with a second less pronounced peak at 72 days post-natal. The principal peak in gap junction levels appears to coincide with increased post-natal blood pressure and aorta media thickening. The amount of gap junction labelling falls off to normal adult levels as the smooth muscle cells modulate towards the contractile phenotype and growth is completed. The results indicate an association between direct cell-to-cell communication and synthetic phenotype smooth muscle cell activity during aortic growth and patterning.

Animals↗
PubMed1997

Angiotensin II receptor type 1 mRNA is upregulated in atria of patients with end-stage heart failure.

There is increasing evidence that pathological changes in the myocardium during chronic heart failure (CHF) are partly regulated through the activation of the renin-angiotensin system (RAS), an effect mediated by the angiotensin II type 1 receptor (AT1R). We examined the expression of cardiac AT1R mRNA in normal (atria, n=7; ventricle, n=3) and end-stage CHF human hearts (atria, n=8; ventricle, n=14). Tissue was snap-frozen immediately after explantation during orthotopic cardiac transplantation; control specimens were obtained from healthy donor hearts rejected for technical reasons. Northern blots of purified total mRNA from each tissue were hybridized with a random primed radiolabeled probe for the coding sequence of AT1R. Stringent conditions were used for both hybridization (5X SSC, 65 degrees C) and washing (0.5X SSC, 0.1% SDS, 65 degrees C) of the membrane. Left and right atrial tissue showed low levels of AT1R mRNA expression in the controls, with statistically significant upregulation of expression in tissue from pathological hearts; CHF atria 1.28+/-0.86 optical density (OD) units, control atria 0.56+/-0.31 OD units, P=0.05 (mean+/-s.d.). There were undetectable levels in ventricles from either control (2/2) or dilated hearts (7/7). The results were independent of the etiology of the heart failure and suggest that increased levels of atrial AT1R mRNA may occur in response to elevated atrial pressures in heart failure.

Adult↗
PubMed1997

Relative susceptibility of endothelium and myocardial cells to ischaemia-reperfusion injury.

We have investigated the timescale, extent and sequence of endothelial, myocardial and smooth muscle cell dysfunction following ischaemia-reperfusion in the isolated rat heart. Myocardial function in the form of aortic flow was evaluated in the working heart preparation. Vasodilatation induced by endothelium-dependent agent 5-hydroxytryptamine (5-HT) and endothelium-independent substance glyceryl trinitrate (GTN) was used to assess endothelial and smooth muscle function, respectively. The percentage recovery of 5-HT response and aortic flow (endothelial vs. myocardial function) plus 5-HT response and GTN effect (endothelial function vs. smooth muscle response) after a period of ischaemia was statistically analysed by Wilcoxon's signed rank test. A possible correlation between structural and functional changes in coronary vascular endothelium after ischaemia was also explored by comparing endothelial dysfunction with morphological assessment of endothelial damage seen after ischaemia. The experiments were performed at two clinically relevant temperatures of 20 degrees C and 4 degrees C. There was no significant difference in the percentage recovery of aortic flow and 5-HT response or in 5-HT response and GTN effect after 30 min of unprotected ischaemia at 4 degrees C. The same duration of ischaemia at 20 degrees C led to significantly better recovery of GTN effect as compared with 5-HT response, but there was no difference in the recovery of aortic flow and 5-HT response. Sixty minutes of unprotected ischaemia at both temperatures caused significantly better recovery of aortic flow and GTN effect as compared with 5-HT response; 60 min of ischaemia protected by an infusion of a cardioplegic solution (protected ischaemia) at 4 degrees C had no significant effect on the recovery of the three parameters. Periods of 90, 120, 180 and 240 min of protected ischaemia at 4 degrees C, and 60, 90 and 120 min of protected ischaemia at 20 degrees C resulted in significantly better recovery of aortic flow compared with 5-HT response and of GTN effect as compared to 5-HT response. This demonstrates varying susceptibility of different cell types to ischaemic injury and highlights the strong vulnerability of endothelium to ischaemic damage as compared with myocytes and smooth muscle cells. Furthermore, there was lack of correlation between post-ischaemic endothelial dysfunction and microscopically assessed structural damage.

Animals↗
PubMed1997

Quantitative immunoconfocal analysis of human myometrial gap junction connexin43 in relation to steroid hormone concentrations at term labour.

The aim of this study was to quantify gap junction expression in the human myometrium in relation to progesterone and oestradiol concentrations, and to establish whether oxytocin-resistant dystocia is due to an abnormality in gap junction expression. Three groups of patients were investigated: (i) before labour (at term), (ii) normal labour and (iii) oxytocin-resistant dystocia (eight patients per group). For each patient, the concentrations of oestradiol and progesterone in maternal blood and in myometrial tissue were measured, and the number and area of immunostained connexin43 gap junctions per unit volume of tissue determined by quantitative analysis of digital images obtained by confocal microscopy. No significant difference in connexin43 gap junction content was observed between the three patient groups. When all groups were pooled, there was a significant positive correlation (P < 0.05) between the quantity of immunolabelled gap junctions and the oestradiol:progesterone ratio, but there was no significant difference in this correlation between the groups. Gap junction immunolabelling was not correlated with the progesterone or oestradiol concentration in the maternal blood or the myometrium. These data suggest that in human myometrium: (i) dystocia is not due to a reduced level of immunodetectable connexin43 gap junctions, (ii) onset of labour is not associated with a sudden increase in immunodetectable gap junction protein and (iii) gap junctions can be expressed in the presence of high progesterone concentrations.

Adult↗
PubMed1997

Spatial distribution of dihydropyridine receptors in the plasma membrane of guinea pig cardiac myocytes investigated by correlative confocal microscopy and label-fracture electron microscopy.

Excitation-contraction coupling in cardiac muscle is thought to depend fundamentally on the spatial organization of sarcolemmal dihydropyridine receptors (L-type calcium channels) in relation to ryanodine receptors (calcium-release channels of the sarcoplasmic reticulum). In the present study, we have investigated the distribution of dihydropyridine receptors in the guinea pig myocyte plasma membrane by correlative immunoconfocal microscopy and label-fracture electron microscopy. Label-fracture, a method in freeze-fracture cytochemistry, permits immunogold localization of cell surface proteins in en face membrane views. Taken together, results from confocal microscopy and label-fracture replicas suggest that, in the peripheral plasma membrane, calcium channels are organized predominantly in the form of clusters. Confocal microscopy also suggests a similar organization in the transverse tubules. It is hypothesized that these clusters may lie adjacent to junctional sarcoplasmic reticulum, permitting the close coupling of influx of calcium through plasma membrane calcium channels to trigger release of calcium from the intracellular stores, as part of the mechanism of calcium-induced calcium release.

Animals↗
PubMed1997

Smooth muscle cells express granulocyte-macrophage colony-stimulating factor in the undiseased and atherosclerotic human coronary artery.

Granulocyte-macrophage colony-stimulating factor (GM-CSF), one of a family of cytokines that regulate proliferation in macrophages and other types of cells, has been implicated in the inflammatory-fibroproliferative response of atherosclerosis. However, previous studies have been restricted to cultured cells and animal models. In the present study, we investigated GM-CSF expression in undiseased and atherosclerotic human coronary arteries at both the mRNA and protein levels. Dual in situ hybridization/cell-marking experiments demonstrated that subpopulations of intimal smooth muscle cells (SMCs) and endothelial cells express the cytokine in the histologically normal human coronary artery and that augmented expression occurs at these sites, and in macrophage accumulations and medial SMCs, in the atherosclerotic vessel. Corresponding data were obtained by in situ hybridization and reverse transcription-polymerase chain reaction and Northern analyses of cultured cells. Cultured human coronary arterial SMCs showed constitutive expression of GM-CSF in cells that had adopted an activated synthetic phenotype. Electron microscope immunocytochemistry revealed that GM-CSF is a protein localized in the cytoplasmic matrix of SMCs of both the undiseased and atherosclerotic vessel wall; extracellular matrix was largely unlabeled, with only occasional small patches of amorphous immunopositive material. The expression of GM-CSF by subpopulations of intimal SMCs in the undiseased artery and the marked upregulation of GM-CSF apparent in atherosclerotic lesions suggest roles for the cytokine in the cellular events underlying initiation and progression of the human atherosclerotic lesion.

Adolescent↗
PubMed1997

Upregulation of connexin43 gap junctions between smooth muscle cells after balloon catheter injury in the rat carotid artery.

Phenotypic transformation of smooth muscle cells (SMCs) to the synthetic state in vitro and in human coronary atherosclerosis is reported to be associated with upregulation of connexin43 gap junctions. To determine whether cellular interactions mediated by gap junctions participate in the phenotypic transformation of SMCs in arterial injury and disease in general and to establish the spatial and temporal pattern of any such change in relation to neointimal development, we investigated SMC connexin43 gap junction expression during vascular healing in the rat carotid artery after balloon catheter injury. Quantitative immunoconfocal microscopy was applied to localize and to quantify connexin43 gap junctions 1, 3, 9, and 14 days after injury. Parallel studies were conducted by electron microscopy (direct morphological demonstration of SMC gap junctions) and immunoconfocal microscopy (localization of altered actin expression). Synthetic-state SMCs in the neointima (first apparent from 9 days postinjury) revealed abundant expression of gap junctions, with levels of immunodetectable connexin43 threefold greater than those of medial cells. However, the first detectable changes were found in the media, before neointimal formation; at 1 to 3 days postinjury, an increase in SMC gap junction expression was apparent in the innermost (subluminal) zone, the major site from which the cells subsequently found in the neointima are recruited. We conclude that upregulation of connexin43 gap junctions is intimately linked to SMC phenotypic transition and that interactions mediated by gap junctions may be a hitherto unrecognized contributor to the cellular mechanisms underlying the vascular response to injury.

Angioplasty, Balloon↗
PubMed1997

Dissociated spatial patterning of gap junctions and cell adhesion junctions during postnatal differentiation of ventricular myocardium.

Nonuniformity in the spatial patterning of gap junctions between heart muscle cells is now recognized as an important determinant of electromechanical function in working myocardium. Breakdown of the normal geometry of electrical intercellular connectivity in diseased myocardium correlates with reentry, arrhythmia, and conduction disturbance. The developmental mechanism(s) that determines this precise spatial order in gap junction organization in normal myocardium is at present unknown. To examine this question, we have used immunoelectron and immunoconfocal microscopy to analyze the spatial distributions of gap junctional (connexin43), desmosomal (desmoplakin), and adherens junctional (N-cadherin) components during maturation of rodent and canine left ventricular myocardium. In rats, a striking divergence in the distribution of gap junctions and cell adhesion junctions emerged within the first 20 days of postnatal life. It was found that although gap junctions initially demonstrated dispersed distributions across myocyte cell membranes, desmosomes and adherens junctions showed more rapid polarization toward cell termini (ie, nascent intercalated disks) after birth. Over subsequent postnatal development (20 to 90 postnatal days), gap junctions became progressively concentrated in these cell adhesion junction-rich zones of membrane. Quantitative analyses of this process in a series of rats aged 15 embryonic and 1, 5, 10, 20, 40, 70, and 90 postnatal days indicated that significantly higher levels (P < .01) of N-cadherin and desmoplakin than of connexin43 were immunolocalized to cell termini by as early as postnatal day 5. Although all three junctions types showed increasing polarization to myocyte termini with development, variation between junctions remained significant (P < .05) at all times points between 5 and 70 postnatal days. Only at 90 postnatal days, when the animals were nearly full grown, did the proportions of gap junction, desmosome, and adherens junction at intercalated disks become statistically similar (P > .05). Examination of myocardium from 1- and 3-month-old canines revealed that related differential changes to the spatiotemporal distribution of intercellular junctions occurred during postnatal maturation of the dog heart, suggesting that the process was not rodent specific. It is concluded that this progressive change in the organization and pattern of association between gap junctions and cell adhesion junctions is likely to be an important factor in maturation of electromechanical function within the mammalian heart.

Animals↗
PubMed1997

Dystrophin is not a specific component of the cardiac costamere.

Dystrophin is a key component of the subsarcolemmal skeleton of muscle cells, and lack of dystrophin is the direct cause of Duchenne muscular dystrophy. In skeletal muscle, dystrophin is reported to be localized specifically at costameres, transversely oriented riblike subsarcolemmal plaques that mechanically couple the contractile apparatus to the extracellular matrix. Costameres are characteristically rich in vinculin and are prominent in cardiac as well as skeletal muscle. To define the precise spatial relationship between dystrophin in relation to the costamere in cardiac muscle, we applied high-resolution single- and double-immunolabeling techniques, under a range of preparative conditions, with visualization of vinculin (as a costamere marker) and dystrophin by confocal microscopy and by the freeze-fracture cytochemical technique, fracture label. Immunoconfocal visualization revealed dystrophin as a continuous uniform layer at the cytoplasmic surface of the peripheral plasma membrane of the rat cardiac myocyte at both costameric and noncostameric regions. The pattern of labeling was reproducible with three different antibodies and was independent of time and antibody concentration. Platinum/carbon replicas and thin sections of fracture-label specimens permitted high-resolution visualization of the distribution of dystrophin in plane views of the freeze-fractured plasma membrane and in relation to the sarcomeric banding patterns of the underlying myofibrils. These results confirmed no preferential association of dystrophin with costameres or with any region of the sarcomeres of underlying myofibrils in rat cardiac tissue. We conclude that in contrast to skeletal muscle, dystrophin in cardiac muscle is not exclusively a component of the costamere.

Animals↗
PubMed1997

Gap junction localization and connexin expression in cytochemically identified endothelial cells of arterial tissue.

Vascular endothelial cells interact with one another via gap junctions, but information on the precise connexin make-up of endothelial gap junctions in intact arterial tissue is limited. One factor contributing to this lack of information is that standard immunocytochemical methodologies applied to arterial sections do not readily permit unequivocal localization of connexin immunolabeling to endothelium. Here we introduce a method for multiple labeling with specific endothelial cell markers and one or more connexin-specific antibodies which overcomes this limitation. Applying this method to localize connexins 43, 40, and 37 by confocal microscopy, we show that the three connexin types have quite distinctive labeling patterns in different vessels. Whereas endothelial cells of rat aorta and coronary artery characteristically show extensive, prominent connexin40, and heterogeneous scattered connexin37, the former, unlike the latter, also has abundant connexin43. The relative lack of connexin43 in coronary artery endothelium was confirmed in both rat and human using three alternative antibodies. In the aorta, connexins43 and 40 commonly co-localize to the same junctional plaque. Even within a given type of endothelium, zonal variation in connexin expression was apparent. In rat endocardium, a zone just below the mitral valve region is marked by expression of greater quantities of connexin43 than surrounding areas. These results are consistent with the idea that differential expression of connexins may contribute to modulation of endothelial gap junction function in different segments and subzones of the arterial system.

Animals↗
PubMed1996

Effect of neuromimetics upon the release of atrial natriuretic peptide granules: are multiple pathways involved in secretion?

The release of atrial natriuretic peptide (ANP) in response to the application of neurohumoral agonists (neuromimetics) is directly demonstrated and quantified at the cellular level, using an ultrastructural assay developed to quantify secretion. The assay uses an in situ tannic acid perfusion technique to arrest the exocytosis of atrial secretory granules in the anesthetized rat. The animal is perfused with the neuromimetic, and secretory granules, which retain the capacity to undergo exocytosis throughout the subsequent 30 min tannic acid perfusion, accumulate at the cell surface in a state of fusion with the plasma membrane. Quantification of arrested granules thus provides a measure of the rate of granule release and allows the responses to different agents to be assessed. The actions of three different agents were investigated: isoproterenol, phenylephrine, and acetylcholine. In previously published studies, investigations of the actions of these agents on ANP release has produced unclear and sometimes contradictory results. Using our ultrastructural assay, it was found that during the 30 min perfusion period neither isoprenaline nor phenylephrine caused a significant change in the rate of secretory granule release, whereas acetylcholine significantly decreased the rate of granule release. A new model of secretion is proposed to integrate these findings with previous results and help clarify the complex picture of atrial natriuretic peptide release.

Acetylcholine↗
PubMed1996

Effects of calcium on the migration and recruitment of macrophages and macrophage-derived foam cells.

Calcium is thought to play an important role in the genesis of atherosclerotic lesions, but the precise mechanisms involved are unclear. In the present investigation, we have used in vitro systems to investigate the effects of calcium on one key aspect of lesion development: the migration of macrophages and macrophage/foam cells. Using agarose plate migration assays, the migratory characteristics of macrophages exposed to 1) no lipoprotein, 2) low density lipoprotein (LDL), 3) acetylated low density lipoprotein (acLDL), and ) oxidized low density lipoprotein were examined. The most marked stimulatory effect on macrophage mobility was observed when freshly isolated cells were exposed to acLDL during the migration assay. High levels of exogenous calcium were found to suppress the stimulatory effect of acLDL on migration. As the responses of macrophages exposed to a uniform concentration of agents in the surrounding medium may differ from chemotactic responses to a concentration gradient, the migration of macrophages, with and without preexposure to acLDL or LDL, was studied using microchemotaxis Boyden chambers. Under these conditions, calcium acted as a highly potent chemoattractant, especially to cells that had been preincubated with acLDL. These results suggest how elevated external calcium concentration leads initially to macrophage recruitment, and subsequently to foam cell aggregation and lipid core formation, in association with calcification, in the developing atherosclerotic plaque.

Animals↗