Studies on Atherosclerosis : II. Clinical Investigation on Lipid Mobilizer

Abstract

The author reported in the preceding paper (Studies on Atherosclerosis : Part 1) that the combined administration of lanolin and lipid mobilizer induces marked lipemia as well as atherosclerosis in the rabbits. Studies on lipid mobilizer (LM) in man have been reported by clinical findings with the quantitative changes of LM. The author attempted to measure the amount of LM in the human blood serum and to clearify its part in the development of atherosclerosis. Material and Methods 1) In vitro measurement of LM. LM was obtained and purified after Seifter's method from 2ml of the blood serum obtained from the hungry patients in the early morning. The amount of serum LM was evaluated according to its character to inhibit the activity of lipoprotein lipase. 0.3ml of lipoprotein lipase solution was added with 0.3ml of 10% human albumin solution (pH 8.5), 0.3ml of lipid mobilizer solution and 4.0ml of diluted cow's milk (1 in 600, dilutant : 0.25M NH3-NH4Cl, pH 3.5 diluted three times with saline), and incubated at 37°C for two hours. After the procedure the turbidity was compared. 2) Purification of lipoprotein lipase. Periepididymal and perirenal adipose tissue of male rats were put in the cold acetone (O°C, 30 times as much as the tissue), and homogenized at O°C, and then centrifuged in 6000 rpm for 20 minutes. The supernate was discarded. The precipitate was repeatedly washed in cold acetone and ether, and then dried under the decreased pressure in low temperature. This stock powder was dissolved in 0.025M NH3NH4Cl buffer solution (pH 8.6) at O°C and left for 60 minutes, and centrifuged in 8000 rpm for 30 minutes, and then the supernate was diluted in a proper way every time for use. 3) Preparation of anti-β-lipoprotein. Human blood serum containing a large amount of β-lipoprotein was added with 20 times as much volume of 4mg/dl dextran sulfate in physiological saline solution containing sodium citrate. 15-60 minutes later, this mixture was centrifuged in 6000 rpm for 30 minutes. The supernate was discarded. The same procedure was repeated 5 or 6 times to get the pure β-lipoprotein. 15-20 mg dose of this preparation was injected in to the rabbit subcutaneously 15 times. After the procedure, anti-β-lipoprotein-seurm was obtained. This serum reacted specifically with β-lipoprotein to form a single precipitation zone in agar gel double diffusion method (after Ouchterlony). 4) Patients with coronary disease, hypertension and cerebrovascular disease were subjected to this experiment. Healthy adult persons were examined as the control.