식이성 동맥경화증의 초기변화 발생 과정의 전자현미경적 연구

Abstract

[한글]

[영문]

Since the first experimental induction of dietary atherosclerosis in the rabbits by Ignatowsky (1908), numerous studies have been carried out on experimental cholesterol induced atherosclerosis. Most of these studies concentrated on the production of atheroma and the morphologic characteristics of advanced lesions.

However, the mechanism of atherogenesis can not be elucidated by studies on advanced atheroma owing to its degenerative changes and other secondary superimposed elements(Gets et al., 1969).

There has been considerable dispute as to wkich cellular element is primarily responsible for early atheroma formation as well as the origin of the responsible cells. Haust et al. (1965) and others considered these cells to be fibroblasts while Simpson (1969) and others thought they were macrophages derived from the

blood stream, and Daud (1968) and others regarded them as smooth muscle cells.

The present studies were undertaken in an attempt to identify the type of cells responsible for early lesions of cholesterol induced atheroma in the rabbits and to determine the origin of these cells.

Materials anti Methods

Albino rabbits weighing about 2.0kg. were used for the experiments, Cholesterol (Merck Co. product) in a dose of 1.5gm. per animal per day was given mixed with bean curd residue up to 60 days Complete ingestion of the cholesterol was confirmed The basic diet of the animals consisted of 300 gm. of bean curd residue per animal per day.

Five animals were sacrificed each time after the cholesterol feeding for 5,7,9,12,15,20,26,30,40,50 and 60 days Five control animals were killed at the 30th and 60th experimental day, respectively. Samples of blood were drawn immediately after the animals acre killed for determination of the serum total cholesterol level, which was carried out by Schoenheimer and Sperry method(1934).

Immediately upon killing the animals by air embolism, the aorta was perfused by injecting 10 ml. of normal saline followed by 5 ml, of 4% glutaraldehyde solution in phosphate buffer of pH 7.4. Then the aorta was opened for gross inspection and sampling for electron and light microscopy.

Samples were taken from the ascending aorta at early stage and from the atheromatous area at later stage. For the light microscopic examination, specimens were taken from the ascending, thoracic, and abdominal aorta. Samples for the electron microscopy were double fixed, first with glutar-aldehyde followed by osmium tetraoxide, and embedded in Epon 812 following dehydration with graded alcohol. Sections ware made by glass knife in 400 to 500 Å thickness and stained with uranyl acetate and lead hydroxide. Observation was made with Hitachi 11-E model electron microscope. Specimens for light microscopy were processed by routine H & E method. Alcian blue for acid mucopolysaccharides and aldehyde fuchsin staining for elastic fibers were applied Pieces of liver were taken for hematoxyll

and rosin and oil red-0 staining.

Result and Discussions

Serum cholesterol level rose rapidly following the cholesterol feeding, i.e. 347 mg% at the fifty day, 1,089 mg% at the 12th day, 2,000 mg% at the 20th dart and over 2,500 mg% thereafter.

A demonstrable lipid deposition appeared at the 7th day around the central vein of the liver and the lipid increased spreading to the periphery of the lobule as the cholesterol feeding continued

Light mlerosoople examinations of the aorta showed no demonstrable alteration until the 12th day, when splitting of Internal elastic membrane associated with increased acid mucopolysaccharides was observed. However, lipld deposition was

first noted after 20 days of cholesterol feeding. It was characteriged by a single or double layers of foam cells just beneath the endothelial cells as well was few of them attached to the luminal surface in between endothelial cells. As the cholesterol feeding eontinued numbers of foam cells increased to form a thick layer of atheroma formation extending into the inner media. These changes were most marked at the ascending portion, followed by thoracic and abdominal portions.

Electron microscopic examinations disclosed swelling of endothelial cells with increased pinocytotic acclivities an the 5th day, but no demonstrable lipid accumulation wart noted At the 7th day swelling of the endothelial cells increased with an occasional adhesion of monocyte to the luminal surface was noted. At this time the number of smooth muscle cells in subendothelial intima increased, containing a few lipid globules.

As the cholesterol feeding continued swelling and iregularity of the enddothelial cells increased with appearance of cytoplasmic vacuoles. The numbers of intimal smooth muscle cells increased with continuous accumulation of lipid. Lipid accumulation within smooth muscle cells of the media was also noted at the 12th day. At the 20th day a large amount of lipid accumlation was noted within the endothelial cells, and invasion of lipid laden monocytes through interrupted endothelial lining from the lumen was noted. At the 25th day a well formed atheroma

was noted, composed of foam cells derived from smooth muscle cells, monocytes, and endothelial cells. As the cholesterol fecding continued numbers of foam cells increased with more accumulation of lipids and finally degenerative changes of the

foam cells. The amount of lipid markedly increased in the smooth muscle cells of the media.

In summary, the earliest change of the aorta following cholesterol feeding was the swelling and increased pinocytotic activities of the endothelial cells associated with proliferation of smooth muscle cells in subendothelial intima. The first demonstrable lipid appeared within the smooth muscle of the intima, most likely derived from the lumen through pinocytosis of chylomicra by endothelial cells. This followed by accumulation of lipid in the endothelial cells, and immigration of lipid laden monocyte from the lumen. The atheroma formation was produced by these three cullular elements.