P. Helin

Arteriosclerosis in Rabbit Aorta Induced By Systemic Hypoxia: Biochemical and Morphologic Studies

From the Connective Tissue Research Laboratories, University of Copenhagen, Department of Dermatology, Rigshospital, Copenhagen, and from Medical Department C, Gentofte Hospital, Hellerup, Copenhagen, Denmark. Supported by Grants from the Danish State Research Foundation and the Danish Heart Foundation. The statistical analyses were performed in Northern Europe University Computing Center, Technical University of Denmark, Lyngby, Denmark. ARTERIOSCLEROSIS IN RABBIT AORTA INDUCED BY

Repair in Arterial Tissue

Mechanical lesion of the descending thoracic aorta of male albino rabbits was performed with an embolectomy catheter and observed as the basic lesion in a correlative study by scanning electron microscopy (SEM) and light microscopy of injury and repair of arterial wall. Fixation and silver-staining were carried out with the aortas in situ. Specimens collected 3, 6, 14, 30, 60, and 180 days after injury were studied en face by SEM and light microscopy, and sectioned tissue was submitted to histochemical investigations, reaffirming previous observations on embolectomy catheter lesion. Shamoperated animals were used as controls. The observations by SEM and by surface light microscopy were the same: initial, severe trauma of the intimal surface followed by re-endothelialization from preserved endothelium on the aortic surface and around the intercostal arteries. In silver-stained specimens three main types of cellular patterns were recognized: hexagonal cells, foam-like patterns of cells, and polarized cells. The hexagonal cells were “true” endothelial cells, whereas the origin of the foam-like patterns of cells was less clear. From a comparison of the observations made in the present study and those recorded in morphological studies by other authors it is concluded that 1) re-endothelialization of rabbit thoracic aorta following extensive endothelial injury is slow, 2) re-endothelialization with “true” endothelium takes place from pre-existing endothelial areas, 3) the exposed arterial surface seems to be covered with sheets of cells from two sources, viz. pre-existing endothelium and cells originating from the blood and/or from the underlying tissue.

Repair in Arterial Tissue: MORPHOLOGICAL AND BIOCHEMICAL CHANGES IN RABBIT AORTA AFTER A SINGLE DILATATION INJURY

Male albino rabbits were subjected to a single mechanical dilatation of the descending thoracic aorta with a balloon catheter. The animals were killed 3, 6, 14, 30, and 60 days later. Between the sixth and the fourteenth days after injury, all the aortas which had been dilated developed severe, gross arteriosclerosis. Microscopic examination showed destruction and degeneration in the form of necrosis and calcification as well as regeneration and repair including new formation of cells, intercellular substance, and fibers. Biochemical and histochemical analyses revealed an early increase in hyaluronic acid and water followed by an increase in chondroitin-4, 6-sulfate and a later increase in heparan sulfate, dermatan sulfate, and collagen. These alterations were related to the focal lesions in the aortic wall. The permeability of the aorta to 125I-albumin increased to a maximum 3 days after the dilatation, whereupon it decreased rapidly. The alterations were interpreted as nonspecific processes of repair in the vascular connective tissue. The pronounced dependence of the alterations on the time elapsed after injury must be considered in the study of vascular diseases where injury and repair may be involved.

Arteriosclerosis and hypoxia

Summary Male albino rabbits were exposed to short daily periods of systemic hypoxia for 2 weeks. Thirteen out of 16 rabbits showed gross arteriosclerotic changes, while no gross changes were found in 14 controls. The microscopic changes were primarily located to the media and consisted of calcified necrotic foci surrounded by amorphous, metachromatic intercellular substance. Metachromatic intercellular substance was also found in the form of “lakes” between the elastic membranes. A similar increase in the metachromatical intercellular substance was seen in the intima. The meta-chromasia indicated accumulation of acid mucopolysaccharides, which in the present study appeared to be primarily chondroitin-4-sulphate and/or chondroitin-6-sulphate and some dermatan sulphate and/or heparitin sulphate. A distinct calcification of the endothelial lining was observed. The changes probably reflect a damage of the aortic wall following systemic hypoxia and secondary non-specific processes of repair in the vascular connective tissue. The alterations have features in common with human arteriosclerosis.

Arteriosclerosis in rabbit aorta induced by noradrenaline: The importance of the duration of the noradrenaline action☆

Abstract Gross and microscopic arteriosclerosis was induced in the aorta of male albino rabbits by daily intravenous infusions of noradrenaline for 2 weeks. In one experiment the rabbits had daily infusions in two periods of 15 min with an interval of 5 min. In another experiment the animals had daily infusions of noradrenaline in six periods of 5 min with an interval of 5 min between the individual periods. The daily injected amount of noradrenaline (1 mg) and the entire daily infusion period (30 min) were the same in both experiments. Similarly the integrated arterial hypertension curves of the two infusion types were identical. Nevertheless the gross and microscopic arteriosclerosis were considerably more extensive and frequent in the aortas of the 2 × 15-min experiment than in the aortas of the 6 × 5-min experiment. Thus a continuous action of high doses of noradrenaline seems to be more harmful to the aortic wall than an intermittent action. The observations support the theory of a hypoxic damage, but does not exclude a dilatation injury produced by the arterial hypertension.

Arteriosclerosis in rabbit aorta induced by mechanical dilatation: Biochemical and morphological studies☆

Abstract The aortas of male albino rabbits were dilated with a Fogarty arterial embolectomy catheter. Two weeks after a single short-lasting dilatation, the animals developed severe gross arteriosclerosis of a type similar to that induced by catecholamines and exposure to systemic hypoxia. The microscopic alterations of the media were also identical to those induced by catecholamines and systemic hypoxia, whereas intimal thickening was more severe and frequent in the arteriosclerotic lesions induced by the mechanical dilatation. The aortic content of hexosamine, chondroitin-4,6-sulphates and hydroxyproline was increased. The uptake of [ 35 S] sulphate into the sulphated glycosaminoglycans was also increased, reflecting a stimulated synthesis of these substances. Finally there was an increased permability to 125 I labelled human serum albumin in the dilated aortas. The biochemical alterations were interpreted as 2-week-old repair processes of the vascular connective tissue. The similarities in the location and gross appearances of the arteriosclerotic lesions may indicate that dilatation of the aorta also plays a role in the development of arteriosclerosis induced by catecholamines and systemic hypoxia.

Arteriosclerosis and hypoxia: Part 2. Biochemical changes in mucopolysaccharides and collagen of rabbit aorta induced by systemic hypoxia

Summary Experimental arteriosclerosis was induced in the aorta of male albino rabbits by daily exposure to short periods of systemic hypoxia through 2 weeks. An increase of the inner surface area of aorta and gross arteriosclerotic changes in 13 out of 16 rabbits reflected an injury to the aortic wall produced by systemic hypoxia. The aortic content of acid mucopolysaccharides, the synthesis of sulpho-mucopolysaccharides, and the mucopolysaccharide to collagen ratio were increased. Regression analysis showed an increase of the mucopolysaccharides and collagen parallel to the increase of the inner surface area. This correlation supports the interpretation of the biochemical alterations as non-specific processes of repair elicited by a damage to the aortic wall.

Spontaneous aortic arteriosclerosis in rabbits of the Danish country strain

Abstract The occurrence of spontaneous arteriosclerosis of the aorta was studied in male albino rabbits of the Danish Country strain. The frequency of gross arteriosclerosis was compared with the frequency of spontaneous microscopic alterations in aorta. The distribution and frequency of gross aortic arteriosclerosis in 366 rabbits were as follows: the arch of aorta 10%, the descending thoracic aorta 2% and the abdominal aorta no gross changes. These frequencies are rather low in comparison to the frequency of spontaneous gross aortic arteriosclerosis in most other rabbit strains. A detailed investigation of the extent and type of the microscopic changes was done in 30 rabbits. In contrast to the gross changes the microscopic alterations showed the following frequencies: the arch of aorta 90%, the descending thoracic aorta 13% and the abdominal aorta 17%. Thus there is a great discrepancy between the frequencies of spontaneous gross and microscopic changes within the individual parts of aorta. The spontaneous aortic changes were similar to those described in a series of different types of experimental arteriosclerosis. The frequent occurrence of these microscopic alterations in the aortic arch makes this part of aorta unsuited for studies on experimental arteriosclerosis. The studies should be restricted to the descending thoracic aorta and the abdominal aorta which exhibit a low frequency of spontaneous gross and microscopic alterations.

Injury and Repair in Arterial Tissue in the Rabbit ANALYSIS OF DNA, RNA, HYDROXYPROLINE, AND LACTATE DEHYDROGENASE IN EXPERIMENTAL ARTERIOSCLEROSIS

The aortic wall in rabbits was injured by pulling an inflated balloon catheter through the aorta. This was followed by an increase of the RNA-DNA ratio in the aortic wall 3 days after the injury. At the same time, the cathodic lactate dehydrogenase (LDH) isoenzymes composed of M subunits increased. The changes in LDH may reflect tissue hypoxia at the early phase after the injury. At 6 days protocollagen proline hydroxylase activity was increased, suggesting increased capacity for collagen formation in the injured aorta. Two weeks after injury the total content of hydroxyproline and DNA were increased, indicating connective tissue proliferation and hyperplasia in the injured aorta. The biochemical changes were interpreted as a repair response of the arterial connective tissue to injury. Gross arteriosclerosis of the aorta developed simultaneously with the biochemical alterations.

The aortic glycosaminoglycans in arteriosclerosis induced by systemic hypoxia

Abstract Arteriosclerosis was produced by exposing male albino rabbits to daily periods of systemic hypoxia during 2 weeks. Biochemical analysis of aortae revealed an increase in the concentration of hyaluronic acid and chondroitin-4,6-sulphate. The total amount of glycosaminoglycans, i.e. hyaluronic acid, heparan sulphate, chondroitin-4,6-sulphate, and dermatan sulphate was also elevated. The increase in the aortic glycosaminoglycans probably reflects processes of repair following the arterial injury induced by systemic hypoxia.