Mary E. Todd

The ultrastructure of peripheral arteries during the development of DOCA hypertension in the rat.

SummaryThe ultrastructure of the unconstricted superficial epigastric and femoral arteries is described in normal rats and in animals with hypertension induced by unilateral nephrectomy, by subcutaneous injections of desoxycorticosterone acetate and drinking of 1% NaCl. The femoral artery showed by far the greater response to the DOCA-saline treatment. In both vessels, the smooth-muscle cells changed from the normal spindle shape to a blunt ended outline with numerous pinocytotic vesicles and prolific collagen production. With long term hypertension, particularly in the femoral artery, the smooth-muscle cell profiles became very irregular. Hypertrophy of the organelles of the smooth-muscle cells was associated with an increase in the intercellular material which gradually changed from a mainly collagenous character to mainly vesicular. Lysosomal activity indicated cell disintegration. White blood cells adhere to the endothelium in hypertensive rats and there was an increase in subendothelial material. The number of intimal smooth-muscle cells increased noticeably in the femoral artery. In both arteries, the adventitial fibroblasts hypertrophied in hypertensive rats. In animals with an elevated blood pressure the morphological response was observed as early as 4 to 7 days after initiation of treatment.

Preventing Intimal Hyperplasia With Photodynamic Therapy Using an Intravascular Probe

The purpose of this study was to determine the efficacy of intravascular photodynamic therapy (PDT) to prevent the development of intimal hyperplasia. Anesthetized New Zealand white rabbits underwent placement of Fogarty balloon catheters introduced via femoral artery cutdowns. Catheters were passed retrograde 10 cm into the lower abdominal aorta, inflated six times, and withdrawn toward the inguinal ligament. Rabbits were then randomly assigned to one of the following groups: group 1, drug with no light; group 2, no drug with 240 joules of light; group 3, drug plus 120 joules of light; or group 4, drug plus 240 joules of light. Uninjured carotid arteries served as negative control vessels (N) and injured but non-PDT-treated iliac artery segments served as positive controls (P). Porfimer sodium (Photofrin) was administered in a dose of 5.0 mg/kg. Light was provided by a fiberoptic probe with a 1 cm cylindric diffuser attached to an argon pumped dye laser tuned to 630 nm to provide 1 W of laser light for 120 or 240 seconds. One month after PDT, rabbits were killed, perfusion fixed with glutaraldehyde, and vessels removed and examined microscopically. Intimal thickness (mean ± SD) was calculated and expressed as ratios of the intima/media at four equal positions. Results for N, P, and groups 1, 2, 3, and 4 were 0.02±0.00, 1.18±0.71, 0.76±0.33, 0.96±0.43, 0.14±0.22, and 0.36±0.16, respectively. Intimal thickness was significantly reduced in groups 3 and 4 when compared with P, group 1, and group 2 (p<0.001, ANOVA). These results showed that intravascular PDT was effective in reducing intimal hyperplasia following arterial injury. This may be a practical method of delivering light for PDT.

The rat-tail artery maintained in culture: An experimental model

SummaryThe rat-tail artery was maintained in vitro for 2 weeks to investigate its suitability as an experimental model. The criteria were that (a) it should retain the overall histological organization with normal ultrastructural appearance of the smooth-muscle cells; (b) stored neurotransmitter which could be activated by experimental treatment should be absent; and (c) smooth-muscle ion transport mechanisms should fall within normal range. Vessels were maintained in Falcon tissue-culture dishes in Dulbeccos modified Eagles medium. Either 2% or no serum supplement was found to be more suitable than 10% serum due to the high rate of cell proliferation induced by the latter. Light and electron microscopy of cross sections of the vessels indicated that the overall normal vessel architecture was retained, and the ultrastructural features predicted normal function. There were no discernible differences dependent on the length (up to 8- to 10-cm lengths) of the cultured vessel. Preliminary experiments with fluorescent microscopy showed that stored neurotransmitter in the nerves of the vessel wall was no longer present after 48 hr. Ultrastructural examination revealed that storage vesicles in vitro lost their dense cores, representing noradrenalin, between 41 and 48 hr in culture. Normal ion transport mechanisms were retained in the smooth-muscle cells of the arteries in vitro for up to 2 weeks when tested with ion-specific electrodes. Morphological and physiological evidence support the suitability of the rat-tail artery as a model for experimental testing of vascular tissues.

Determining light dose for photodynamic therapy of atherosclerotic lesions in the Yucatan miniswine.

Purpose: To determine the light dose required for photodynamic therapy of atherosclerotic lesions in the miniswine. Methods: Aortic atherosclerosis was created in seven Yucatan miniswine by a combination of balloon endothelial injury and 2% cholesterol and 15% lard for 7 weeks. Six animals received the photosensitizer PhotofrinR 2.5 mg/kg, while an additional swine received no drug. After 24 hours, the abdominal aorta was exposed and the aorta opened longitudinally in each animal. Three 1-cm spots were illuminated with energy densities of 60, 120, and 240 J/cm2 from an argon-pumped dye laser tuned to 630 nm with a laser output of 1 W. Four weeks later, the animals were killed, abdominal aortae removed, and intimal thickness determined by morphometry. Results: The percentage intimal thickness (mean ± SD) was 36.7 ± 27.1, 9.1 ± 5.0, and 6.4 ± 8.1 for the three energy densities, respectively. Although both 120 and 240 J/cm2 energy densities produced significant (p < 0.05) reduction in atheroma, considerable damage to the underlying media was also observed in the 240 J/cm2 group. Conclusions: A Photofrin dose of 2.5 mg/kg and 120 J/cm2 light are necessary for adequate ablation of atheroma while avoiding extensive medial damage.

An ultrastructural and fluorescence histochemical investigation of the innervation of retial arteries in Monodon monoceros

In this study, the innervation of cerebrally related retial arteries in the narwhal Monodon monoceros was examined. Vessels were processed for the demonstration of adrenergic nerve endings by fluorescence histochemistry, and the results were confirmed by electron microscopy. Innervation of cerebrally related retial arteries was compared to that of a system situated in the haemal canal and supplying the tail. The retial arteries were poorly innervated. Adrenergic nerve endings, as indicated by fluorescence, occurred only in caudal portions of the spinal rete. Ultrastructurally, nerves were found in most retial vessels examined. However, except for arteries from caudal portions of the spinal rete, nerve numbers were few and because they occurred in outer layers of the adventitia were probably not functionally significant. In contrast, vessels in the haemal canal were well innervated. Nerve endings possessing neurotransmitter vesicles were adjacent to the smooth muscle cells. The cetacean rete mirabile, a system which supplies blood to the entire central nervous system, is apparently not under extensive nervous control, even though most reports suggest there is a relationship, possibly based on the presence of adjacent nerve trunks. Any vasomotor activity that does occur, possibly does so in response to catecholamines or other vasoactive agents circulating in the blood.

Improved Ultrastructural Detail in Tissues Fixed with Potassium Permanganate

An improved method has been developed for fixation with potassium permanganate. Although this is one of the methods widely used to preserve the dense cores of adrenergic storage vesicles, fixation of other tissue components is usually poor. The main differences from previously reported methods using potassium permanganate are the use of a physiological saline as the vehicle for all solutions, and, following this, very rapid dehydration before infiltration with plastic. Cellular and intercellular details of tissue ultrastructure may, in general, be evaluated as satisfactorily as with conventional fixatives, with the exception of certain protein elements associated with ribosome, microtubule, and myofilament organization. Nerve endings with agranular or clear vesicles may be distinguished from adrenergic endings since the dense cores of the vesicles of the latter are preserved by this method.

Dosage and timing of Photofrin for photodynamic therapy of intimal hyperplasia

Photodynamic therapy has been recommended as a method of preventing intimal hyperplasia. The purpose of this study was to determine the dose and timing of Photofrin porfimer sodium needed to achieve a 3:1 or higher ratio between injured and control arteries after balloon endothelial injury. New Zealand White rabbits were anesthetized and their right femoral artery surgically exposed. A 4Fr Fogarty balloon catheter was passed retrograde into the lower abdominal aorta, inflated and pulled distally into the external iliac artery six times. All rabbits received heparin 100 IU/kg. Arteriotomies were closed and the animals recovered. Rabbits (n = 5 per group) were given intravenous Photofrin at a dose and time according to the following scheme: group I, 5.0 mg/kg immediately after balloon injury; group II, 2.5 mg/kg immediately after injury; group III, 5.0 mg/kg after 1 week; group IV, 5.0 mg/kg after 2 weeks; or group V, 2.5 mg/kg after 2 weeks. Animals were killed 24h after drug administration and the aortoiliac segments removed for spectrophotofluorometric determination of Photofrin levels from injured and control segments. Mean(s.d.) ratios of injured: control arteries for groups I to V were 4.8 (2.6), 2.8 (1.2), 3.0 (1.0), 1.4 (0.3) and 1.0 (0.0) respectively. This ratio was significantly higher for group I rabbits compared with groups IV and V (P < 0.01, ANOVA). Fluorescence and light microscopy showed that Photofrin was localized primarily in the tunica media, and that the drug must be administered before significant intimal hyperplasia occurs.(ABSTRACT TRUNCATED AT 250 WORDS)

Coexistence of diabetes and hypertension results in unique structural alterations in the renal artery in rats beyond that found with diabetes alone

Numerous investigators have presented evidence of increased mortality in patients with diabetes mellitus due to cardiovascular disease. It is still unclear as to the reasons why there is a predisposition to vascular pathology that in the advanced state leads to atherosclerosis. Our hypotheses were: (1) The condition of diabetes mellitus in a streptozocin animal model may show vascular changes similar to early pathology in macrovessels and (2) since the model is normotensive, inducing hypertension will result in early atherogenic pathology. We carried out a quantitative analysis of the renal artery using light and electron microscopy to test the hypotheses. Male Wistar rats had diabetes mellitus induced using streptozocin and 1 week later half of the diabetic animals had hypertension induced with deoxycorticosterone acetate (DOCA). Samples were taken following 7 weeks of diabetes, or 7 weeks of diabetes with DOCA administration during the final 6 weeks. The renal artery from the diabetic group did not have any differences in wall or luminal dimensions from control, but did have proportionately more extracellular matrix than smooth muscle in the tunica media. This is evidence of structural change, in a large supply artery, as a manifestation of diabetes mellitus, similar to that seen in vascular disease. Vessels from the control/hypertensive had a significantly thickened tunica media as did the diabetic/hypertensive over control values. The latter also had proportionately even greater significant elevation of the extracellular matrix compared with either the diabetic or control/hypertensive. In addition, only the diabetic/hypertensive group showed marked subendothelial invasion of macrophage type cells and deposits of various shapes and densities. We have, therefore, demonstrated significant vascular alteration due to the diabetic condition in this animal models and also shown that with hypertension and diabetes combined, the early vascular pathology is exacerbated.

Smooth muscle cell characteristics: a computer-assisted analysis from serial sections

Accurate structural measurements of various smooth muscle cells, employing proven methodology, are relatively sparse, although some data have been provided, mainly for intestinal, uterine, and vascular cells. Morphological parameters by themselves are of only academic interest, unless their far-reaching functional significance is considered. For example, in smooth muscle cells of a given volume, if the relative proportion of surface area increases (an increase in the surface-area-to-volume ratio), then hypothetically, a greater calcium influx should occur, given that the proportion of calcium channels and all intracellular components remain constant. Conversely, if the surface-area-to-volume ratio decreases, then the reverse could be postulated. Another structural parameter, the orientation or angle of the cells within the vessel wall, will influence the plane in which the maximum force is developed.

Intraluminal Endothelium-Covered Bridges in Chronic Fat-Fed Balloon-Injured Yucatan Miniswine

The Yucatan miniswine has been recommended as an animal model of advanced atherosclerosis. Atherosclerotic plaques developed in this model demonstrate foam cells, widespread fibrosis, and calcification, features suggestive of human atherosclerosis. We have observed the occurrence of intraluminal projections that appear peculiar to this animal model. Forty-three miniswine, weighing between 20 and 30 kg, were rendered atherosclerotic with a combination of balloon endothelial injury of the aortoiliac segments and dietary supplementation with 2% cholesterol and 15% lard. Endothelial injury was created by retrograde balloon catheter injury of the aorta and both external iliac arteries via cutdowns on the femoral arteries. Serum cholesterol prior to starting the diet and at 1, 2, and 6 weeks following initiation of the diet was 2.0 +/- 0.4, 11.6 +/- 4.0, 15.9 +/- 5.0, and 16.4 +/- 4.2 mM, respectively (p < .0001, ANOVA). Angiographically significant lesions were apparent in 33 of 37 (89%) animals (occlusion 20/37, stenosis 17/37) at 6 weeks postinjury. In three of six (50%) animals followed up to 16 weeks postinjury, trabecular areas were seen in the external iliac arteries on angiography. Light and electron microscopy demonstrated that these areas were covered with normal endothelium and projected into the lumen or bridged with the adjacent arterial wall. Foam cells and calcification were not seen in these lesions. This finding is not typical of human atherosclerosis and appears peculiar to this type of animal model.