It is wellknown that postheparin plasma lipase appears rapidly in the blood. when heparin is injected intravenously. On the other hand, it has been recognized when epididymal adipose tissue or heart tissue are incubated in vitro with heparin, lipoprotein lipase (LPL) is rapidly released in the medium. After experimental conditions were determined referring to the method of Cherks et al., in vitro studies were made in order to investigate the effect of addition of heparin, Oxidized Starch Sulfate (OS-S) which was demonstrated to have lipemia clearing action in our previous paper and some other natural and synthetic polysaccharide sulfates on the rate of increase of LPL activity which appears in the medium when rat heart tissue is incubated with them.
Experimental methods: Male albino rats weighing about 200g were killed by blow on the head and the heart excised and trimmed, rinsed quickly with cold Krebs Ringer phosphate buffer (pH 7.4) and then sliced or homogenized. Slices or homogenate, each weighing approximately 100mg, were placed in Warburg flasks which contained 1.0
ml of Krebs Ringer phosphate buffer. To each flasks was added 200μg of heparin (Dai-ichi Chemicals, Japan), OS-S or other polysaccharide sulfates and preincubated for 60 minutes at 37°C under shaking and under air.
After preincubation, medium was centrifuged at 15, 000g for 10 minutes and 0.5
ml of the supernatant solution was added to centrifuge tube which contained 0.5
ml of artificial lipoprotein substrate which was made by mixing one part of 10% bovine serum albumin (pH 8.7), two parts of 5% coconut oil emulsion and equal parts of fresh rat serum with final adjustment of pH to 8.5 with diluted ammonia and incubating the mixture at 37°C for 30 minutes.
Incubation was carried out at 37°C and after 30 minutes reaction was stopped and deprotenized by adding 5% trichloroacetic acid. Mixture was centrifuged and glycerol in the supernatant was measured by means of Lambert-Neish method. Lipoprotein lipase activity was expressed as μg glycerol per 100mg of fresh tissue liberated in one hour.
Results: From the data obtained, specific activities of the rate of increase of LPL activity of the other polysaccharide sulfates were calculated and expressed in terms of heparin having an arbitrary value of 100. Chondroitin Sulfate=-19, Carrageenin=44, Fucoidin Sulfate=-39, Dextran Sulfate B.P.=41, Dextran Sulfate=70, and OS-S I-V which have same viscosities about 0.026 and S% 3.7, 6.5, 8.0, 10.1, and 12.7 respectively showed the rates of increase of LPL activity 17, 28, 50, 61, and 70, and these results are parallel with their sulfur content.
Further investigations on the increasing action of LPL activity in the medium by heparin or the other polysaccaride sulfates revealed that the action contains the action to activate LPL as well as that to release the enzyme from tissue.
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