Preparation of vascular endothelium for scanning electron microscopy: a comparison of the effects of perfusion and immersion fixation

Gertz, S. David; Rennels, Marshall L.; Forbes, Michael S.; Nelson, Elizabeth

doi:10.1111/j.1365-2818.1975.tb04065.x

Cited by 31 publications

(7 citation statements)

References 11 publications

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“…The morphology of the endothelium of the sphereplugged vessels supports the hypothesis that the intravascular pressures are lower on the downstream side of the microspheres. The wrinkled surfaces and irregular cell margins we observed in the smaller vascular diameter side of the spheres are similar to those seen in arterial endothelial cells fixed at low intravascular pressures or by immersion fixation (Gertz et al, 1975;Clark & Glagov, 1976;Swinehart et al, 1976;Reidy & Schwartz, 1981;Garbarsch eta]., 1982). In contrast, the smooth appearance of the endothelial surfaces on the larger vascular diameter side of the microspheres is similar to that observed with endothelium fixed with elevated intravascular pressures.…”

Section: Discussionsupporting

confidence: 72%

A novel application of microsphere perfusion and scanning electron microscopy to the identification of pulmonary arterioles in guinea‐pig and rabbit lungs

Walker

Hosford

Mackenzie

1994

Journal of Microscopy

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Summary In arterioles of the lung the intravascular blood pressures are lower than in comparable vessels in the systemic circulation and the arteriole walls are thinner. Therefore, it is very difficult to distinguish between arterioles and venules of the same size using scanning electron microscopy. This study describes a novel application of latex microsphere perfusion and scanning electron microscopy which distinguishes between pulmonary arterioles and venules on the basis of endothelial cell morphology. Microspheres, 90 and 45 μm in diameter, were perfused into the arterial side of the pulmonary circulation of guinea‐pig and rabbit lungs. Scanning electron microscopy of the arterioles on both sides of the lodged microspheres indicated that the endothelial cells are spindle shaped. In contrast, the endothelial cells of equal diameter venules are polygonal. Furthermore, the nuclei of the arteriolar endothelial cells were significantly (P = 0·019) narrower than those of endothelial cells in venules of equal diameter. Finally, it was observed that the differences between arteriole and venule endothelial cells persisted distally to the capillaries.

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Section: Discussionsupporting

confidence: 72%

A novel application of microsphere perfusion and scanning electron microscopy to the identification of pulmonary arterioles in guinea‐pig and rabbit lungs

Walker

Hosford

Mackenzie

1994

Journal of Microscopy

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“…All carotid arteries were fixed in situ by intracardiac perfusion of 1.6% glutaraldehyde in 0.08 M Sorensen's sodium phosphate buffer (pH 7.4, 310 mosmol/liter, 120 mmHg, room temperature) without prior exsanguination (14). Arterial segments were excised from the sites of, and distal to, the application of CaCl2 (RCCA) and from similar segments ofcontrol vessels (LCCA) to which NaCI was applied.…”

Section: Methodsmentioning

confidence: 99%

Aneurysm of the rabbit common carotid artery induced by periarterial application of calcium chloride in vivo.

Gertz¹,

Kurgan²,

Eisenberg³

1988

J. Clin. Invest.

134

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Experimental aneurysmatic dilatation of the rabbit common carotid artery was induced by a single, periarterial application of calcium chloride in vivo. Vessels were fixed in situ after 3 d, 1 wk, 3 wk, 6 wk, and 12 wk by intracardiac perfusion of glutaraldehyde and tissues prepared for light, scanning, and transmission electron microscopy. Progressive focal aneurysmal dilatation was seen limited to the site of calcium application with endothelial damage and thrombus formation in areas of irregular luminal contour. Disruption of the elastic network of the intima and media was seen with varying degrees of intimal fibromuscular hyperplasia and medial disorganization.The calcium-elastic tissue complex was the focus of the inflammatory, arteriosclerotic reaction and subsequent aneurysm formation. The inflammatory cell infiltration initially included primarily neutrophils followed by lymphocytes, plasma cells, monocytes, and multinucleated giant cells. These studies support the hypothesis that disruption of the elastic tissue network of the vascular wall represents an important pathogenetic factor in the initiation of aneurysmal dilatation. In addition, the results of these studies suggest that interaction of calcium with the elastica of the arterial wall may represent an important pathogenetic factor in the initiation of giant cell arteritis.

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“…Both common carotid arteries, the LAD, and the circumflex coronary arteries were then excised, immersed in 2.5% glutaraldehyde in 0.1 M phosphate buffer for at least 24 hours, and processed for SEM studies by the critical point drying technique. 4 The arteries were then opened longitudinally into three or four parts, mounted on aluminum studs, coated with gold palladium, and examined with a Cambridge S-180 scanning electron microscope.…”

Section: Methodsmentioning

confidence: 99%

Endothelial cell damage and thrombus formation after partial arterial constriction: relevance to the role of coronary artery spasm in the pathogenesis of myocardial infarction.

Gertz¹,

Uretsky²,

Wajnberg³

et al. 1981

Circulation

166

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SUMMARY The left anterior descending coronary artery of four dogs and the right common carotid artery of 15 rabbits were subjected to 40-60% reduction in transluminal diameter for 1 hour by partial ligation with suture thread. Scanning electron microscopic examination of the luminal surface of these vessels revealed endothelial craters and balloons, fragmentation and desquamation on the proximal slope of the constriction. Platelet attachment to exposed subendothelial tissues was clearly evident, and microthrombi were seen at the point of maximum constriction. Blood flow, as measured by electromagnetic flow probe, was virtually unchanged upon partial ligation. In control studies, where a second ligature was placed proximal to and before the first to reduce blood flow to the distal constriction site substantially or totally, endothelial desquamation was found in only one of 14 animals, and the number of craters and balloons was significantly reduced. We suggest that endothelial damage and thrombus formation may occur at the site of focal arterial constriction even when the reduction in transluminal diameter is insufficient to alter substantially the rate of flow.THE THREAT posed by coronary vasospasm for the initiation of myocardial ischemia or infarction is assumed to depend on the degree to which the reduction in transluminal diameter results in the interference with coronary blood flow. Further, it is thought that the thrombosis that may accompany vasospasm is a result of the grossly reduced blood flow secondary to a critical vascular constriction and that the interaction of platelets with a damaged vascular wall, with the ensuing release reaction, may further potentiate the coronary spasm." 2 In the present report we present evidence from scanning electron microscopic (SEM) studies that indicates that marked endothelial damage, extensive platelet deposition and thrombus formation may occur at the site of a focal arterial constriction even when the reduction in transluminal diameter is insufficient to alter flow substantially. MethodsFifteen New Zealand white rabbits (3.1 ± 0.4 kg, mean ± SD) were anesthetized with sodium pentobar- The RCCA of five rabbits and the LAD of two dogs were focally constricted 1 cm proximal to the probe by partial ligation with suture thread (0-cotton) to achieve a 50% reduction in transluminal diameter ( fig. 1). The desired degree of luminal constriction was achieved by the apposition of two points on the suture thread; the distance between these points was calculated from the vascular circumference, with the wall thickness (0.1 mm) considered negligible. The diameters were confirmed during SEM examination and varied from 40-60% luminal reduction. Group 2In a second group of five rabbits and two dogs, a second ligature was placed 1 cm proximal and immediately before the first ( fig. 1)

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Preparation of vascular endothelium for scanning electron microscopy: a comparison of the effects of perfusion and immersion fixation

Abstract: A comparison of currently used methods of tissue fixation for scanning electron microscopic study of vascular endothelium revealed that in situ fixation by intravascular perfusion is superior to immersion for the preservation of endothelial surface morphology.

Cited by 31 publications

References 11 publications

A novel application of microsphere perfusion and scanning electron microscopy to the identification of pulmonary arterioles in guinea‐pig and rabbit lungs

A novel application of microsphere perfusion and scanning electron microscopy to the identification of pulmonary arterioles in guinea‐pig and rabbit lungs

Aneurysm of the rabbit common carotid artery induced by periarterial application of calcium chloride in vivo.

Endothelial cell damage and thrombus formation after partial arterial constriction: relevance to the role of coronary artery spasm in the pathogenesis of myocardial infarction.

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