백서(白鼠)에 있어서 간부분절제 및 갑상선 기능저하가 지질분포에 미치는 영향

Abstract

[한글]

[영문]

It has long since been recognized that the development of atherosclerosis is

considered to be closely related to the degree and duration of

hypercholesterolemia, and lipid metabolism is important in the pathogenesis of

atherosclerosis.

Among the factors related to disturbance of lipid metabolism, there is abundant

evidence in the literature that one of the main factors which influences the

metabolism of lipids and cholesterol is the secretion of the endocrine glands

(Duell, 1955; Pincus, 1959). The role of the thyroid in controlling the level of

plasma lipids and cholesterol, is best established, and other endocrine glands,

such as pituitary, adrenals, gonads, and pancreas also effect the utilization of

lipids and cholesterol (Rosenman et al., 1951 & 1952b; Friedman et at.,1952a & b;

Stamler, 1953). The mechanism by which the thyroid hormone affects cholesterol

metabolism is not yet known, but there is evidence to suggest that cholesterol

destruction is elevated in hyperthyroid rats (Byers et al., 1952c). Hence, it is

assumed that the thyroid hormone acts on cholesterol metabolism cia the liver

(Rosenman et al., 1952b).

The liver is the organ where cholesterol synthesis predominantly occurs

(Cornforth, 1959), and is the main source for the plasma cholesterol (Friedman et.

al., 1951a & b). Although biliary concentration of cholesterol is a reliable index

of the rate of hepatic synthesis of this substance (Byers & Friedman, 1952a & b),

cholesterol synthesis may also occur elsewhere (friedman et at., 1955; Beher et al.

1956). This is strongly supported by the fact that the concentration of desmosterol

in several tissues may be higher than that observed in the liver and serum.

Therefore, the levels of circulating cholesterol seems to 1)e regulated by a

dynamic interchange of cholesterol between blood and tissue stores (reversible

pool), which would depend upon the amount of cholesterol available to the different

areas of the body.

Induced hypercholesterolemia, when sufficiently Prolonged, results in

atherosclerosis in many animals. Dogs and especially rats are resistant; resistance

in dogs, however, can be overcome by feeding large amounts of cholesterol during

induced hypothyroidism(Steiner St Kendall, 1946). The rat is generally considered

to be very resistant to development of atherosclerosis. Although numerous attempts

to produce atherosclerosis in this species have been recorded, most have been

completely unsuccessful (Horlick & Havel, 1948; Kendall, 1949; Marx et at., 1949;

Altschul, 1950; Katz & Stamper, 1953), and the few reported instances of deposition

of lipid in rat arteries have apparently had little resemblance to human

atherosclerosis (Page & Brown, 1952; Bragden & Boyle, 1952; Wissler, et at., 1954;

Fillios, et al., 1956).

The nature of this resistance to the development of experimental atherosclerosis

is unknown, and its elucidation on mechanisms is vital to further knowledge of the

pathogenesis of the disease. One of the factors involved in the process of

resistance may be hormonal(Steiner and Kendall, 1946).

Therefore, in rats a few causes for natural resistance may be considered: 1)

endogenous destruction of the additional cholesterol; 2) excretion of the

additional amount ingested via the bile and the gastrointestinal tract; 3)

diminished endogenous synthesis of cholesterol in balance the additional amount

ingested; 4) storage of the additional cholesterol outside the blood stream.

The majority of the studies, particularly in rats, have dealt mainly with the

fluctuation of plasma lipid levels and the production of atherosclerosis, butt very

little attent-ion huts been paid to lipid distribution within various organs. Also

it is recognized that the level of plasma lipid alone cannot adequately explain the

varied features of the diseases resulting from disturbances in lipid metabolism.

Therefore, study of the Pattern of tissue lipid distribution in hypothyroid and

partial hypatectomized, alone or combined, rats seems appropriate and important.

Moreover in order to consider the resistance of the production of atheromatous

arterial lesions in rats, it is important to evaluate the factors which regulate

lipid metabolism.

The purpose of this study is to characterize the patterns of tissue lipid

distribution and define the nature of the morphological changes in various organs

in rats, after prolonged administration of high concentrations of cholesterol. At

the same time, the effect of Tapazole (antithyroid) and of partial hepatectomy was

evaluated as to the mechanism(5) involved in their action on tissue lipid

alterations.

MATERIALS AHD METHODS

Healthy male albino rats weighing around 200 Grams were used. A total of 59 rats

were subjected to the experiment. The animals were divided into 4 major groups:

Group Ⅰ consisted of 7 rats as normal controls; Group Ⅱ consisted of 6 rats

receiving cholesterol only; Group Ⅲ consisted of 8 rats receiving cholesterol and

Tapazole; Group Ⅳ consisted of 38 rats having a partial hepatectomy. Group Ⅳ was

subdivided into 5 minor groups: Group "a" consisted of 8 rats receiving partial

hepatectomy only; Group "b" consisted of 9 rats receiving cholesterol only after

partial hepatectomy; Group "c" consisted of 8 rats receiving Tapazole only after

partial hepatectomy; Group "d" consisted of 8 rats receiving cholesterol and

Tapazole after partial hepatectomy; Group "e" consisted of 5 rats receiving

cholesterol and thyroxine after partial hepatectomy.

Partial hepatectomy was performed by the method of Higgins and Anderson (1931),

removing the entire median lobes of the liver, around 40% of the entire liver

weight, under ether anesthesia and with aseptic precautions.

Cholesterol was given in 5 gm. doses daily per kg. until the end of the

experiment, and Tapazole in 0.17mg. doses daily per kg. parenterally, and

L-thyroxine sodium, in 0.2mg. doses per kg., was given intramuscularly every other

day until the end of the experiment.

During the 3 months' experimental period, body weight was measured weekly. The

surviving animals were sacrificed at the end of the experimental period. The total

serum cholesterol levels were determined by the method of Kingsley and

Schaffert(1949). The organs, such as heart, kidney, adrenal, and liver, were

weighed. For the study of tissue lipid distribution, Oil Red O stain after frozen

sections was performed on the adrenal, small intestine, liver, kidney, heart and

the aorta. For the histopathologic examination of the organs, paraffin sections of

all specimens, were prepared for hematoxylin-eosin stain and in addition, periodic

acid Schiff's reaction, aldehyde fuchsin stain of Gomori, and the colloidal-iron

method of Rinehart-Abul-Haj, were used for special tissue alterations.

RESULTS AND DISCUSSION

The level of total serum cholesterol was slightly elevated in the

cholesterol-only treated group compared with that of normal controls, and was more

elevated in the Tapazole treated groups such as Group Ⅲ, Group c and d, compared

with that of the cholesterol-only treated group. In contrast, the cholesterol level

appeared in the thyroxine treated group lower than that of the normal controls. In

all groups the fluctuation of the cholesterol levels was minimal, and partial

hepatectomy had no particular effect.

Histopathologic examination of the thyroid showed slight hyperplastic changes in

the cholesterol-only treated group with decreased colloid content and increased

P.A.S. positive granules in the lining epithelium. The thyroid gland in the

Tapazole treated group disclosed marked hyperplasia composed of irregular,

small-sized follicles with decreased colloid content and alto reduced P.A.S.

positive granules in the lining epithelium. However, the thyroxine treated group

showed intermingled minimal hyperplastic or hypoplastic changes with slight

increased P.A.S. positive granules in the lining epithelium, which appeared to be

an effect of cholesterol rather than thyroxine. Bernick et al.(1962) stated that

a1% cholesterol diet in several kinds of animals caused the thyroid glands of

hamster and rats to become hyperplastic, but the glands of rabbits and guinea pigs

to become hypo-plastic in appearance, and he concluded that the latter animals have

a relatively low thyroid activity and their liver is unable to efficiently

metabolize exogenous lipids or cholesterol. Hence, they show both hyperlipemia and

hypercholesterolemia, conditions which are factors in their susceptibility to

atherosclerosis.

The adrenal gland showed increased sudanophilic material particularly in the zona

fasciculata in the cholesterol-only treated group,―a finding which was, more

marked in the Tapazole treated groups and thyroxine treated group, but partial

hepatectomy had no particular effect. The adrenal gland of the thyroxine treated

group showed increased mitotic activity in the zona fasciculata, bona reticularis

and in the medulla.

The small intestine revealed more increased sudanophilic material in the lining

epithelium and the stroma of the mucosa in the cholesterol-only treated group, the

Tapazole treated groups and the thyroxine treated group, as compared with normal

controls, without significant differences among the groups, and there was also no

particular effect following partial hepatectomy. Abell et at. (1956) studied

cholesterol metabolism in the dog, and obtained a result of 75 to 83% fecal

excretion of administered high cholesterol regardless of the thyroid state, they

found no demonstrable effect upon the intestinal absorption of cholesterol. They

concluded that the increase in bile acid output was equivalent to the increased

cholesterol absorption from the diet. Also it was noted that dogs receiving a high

cholesterol diet plus thiouracil have an impaired capacity to convert excess

dietary cholesterol into bile acids. In this study increased sudanophilic material

in the intestinal mucosa is considered microscopically to be evidence of increased

absorption of cholesterol on the high cholesterol diet. Moreover, it appeared that

there were no particular differences in the absorption of cholesterol among the

cholesterol-only treated group, the Tapazole treated groups, and the thyroxine

treated group.

In the cholesterol-only treated group, the liver, showed a minimal degree of

sudanophilic material in the Kupffer cells and immediately beneath the hepatic cell

membrane, particularly around the central veins. In the Tapazole treated groups,

sudanophilic material was markedly and diffusely increased throughout the entire

lobes, but in the thyroxine treated group, sudanophilic material had almost

completely disappeared. There were also no significant differences following

partial hepatectomy though it appeared to be slightly increased in the hepatectomy

groups compared with those of non-hepatectomy groups. Franz et al. (1954) studied

the relationship between the level of serum cholesterol and the concentration of

hepatic cholesterol. They found a relative constancy of the concentration of serum

cholesterol, despite large differences in the cholesterol content of the liver may

postulated the existance of a homeostatic mechanism, and stated that, the liver

appeared to act as a buffer for serum cholesterol. So, if large amounts of

cholesterol are absorbed from the intestinal tract, the liver quickly removed most

of the cholesterol from the blood. As the cholesterol content of the liver rises,

hepatic synthesis of cholesterol is depressed. Many investigators have noted the

depressed rate of hepatic cholesterol synthesis (endogenous) following the

administration of high cholestelol (exogenous) diet in dogs (Gould et at. 1953) and

in rats (Tomkins et al., 1953; Frantz et al., 1954). Thess is a negative feedback

mechanism which inhibition of endogenous cholesterol synthesis in the liver is

followed by administration of large amounts of exogenous cholesterol.

The kidney showed a minimal degree of sudanophilic material in the renal tubules

and glomeruli in the cholesterol-only treated group, and a moderate degree of

sudanophilic material in the Tapazole treated groups. In contrast, in the thyroxine

treated group, sudanephilic material was rarely seen. Partial hepatectomy was

followed by no significant change nor effect in and of these groups.

The coronary artery and aortas showed neither gross atheromatous changes nor

microscopic lipid deposits nor alteration of the acid mucopolysaccharides. However,

several instances, particularly of the Tapazole treated groups, showed minimal

interruption of the internal elastic membrane in the coronary artery. This appeared

to be similar to the coronary artery lesion described by Bernick et al. (1972) who

concluded it only minimal atherosclerotic lesions despite no atheromatous nor

lipomatous lesions of the coronary artery. However, in the face of these equivocal

findings further studies are needed to prove whether or not actual early

atheromatous changes are present.

CONCLUSION

1. Exogenous high cholesterol feeding did not elevate the total serum cholesterol

in normal and partial hepatectomized rats, but did cause significant alterations in

the tissue lipid distribution-particularly in the adrenal gland, small intestine,

liver, and kidney The thyroid gland became hyperplastic.

2. The combined administration of Tapazole and cholesterol caused only a slight

elevation of the serum cholesterol level as compared with that of controls, but

very significant alterations in the tissue lipid distribution in the adrenal gland

and liver.

3. The combined administration of thyroxine and cholesterol caused only a

tendency to minimal decrement of serum cholesterol level as compared with that of

controls, but produced a significant inhibition of tissue lipid accumulation in the

liver and kidney.

4. Partial hepatectomy caused neither the changes in serum cholesterol level, nor

effect of the tissue lipid distribution.

5. Lipid accumulation in the coronary artery and aorta could not be demonstrated,

although there was some alteration in the serum cholesterol level and in the tissue

lipid distribution.

It appears that, in the rats, there was no paricular alteration of the intestinal

absorption of cholesterol regardless of the thyroid status, and in the pathways of

cholesterol metabolism after absorption. Partial hepatectomy did not induce

particular effect, but changes in the thyroid function did show measurable effects.

However, the fact that there were no significant elevations in the serum

cholesterol and tissue lipid in the liver suggests that homeostatic mechanisms may

halve a greater role in the high resistance to the development of atherosclerosis

than does an actively functioning thyroid gland.