Robert O. Scow

Lipoprotein lipase activity of adipose and mammary tissue and plasma triglyceride in pregnant and lactating rats

Abstract 1. 1. The effect of pregnancy and lactation on lipoprotein lipase activity in adipose and mammary tissues and on plasma triglyceride concentration were studied in the rat. 2. 2. Plasma triglyceride concentration, 0.9 mM in control rats, increased 3-fold between the 12th and 20th days of pregnancy to a peak of 3.3 mM, and then decreased 50% during the next 2 days. It increased again at parturition, on the 22nd–23rd days of pregnancy, to 3.0 mM, and then fell sharply and remained below 1.0 mM throughout lactation. 3. 3. Lipoprotein lipase activity in parametrial adipose tissue, 12 units/g in control rats, increased 2-fold between the 9th and 12th days of pregnancy and then decreased slightly during the next 7 days, (1 unit of lipoprotein lipase activity = 1 μmole of triglyceride hydrolyzed per h.) It decreased on the 20th day to 5 units/g, reached 2 units/g at parturition, and remained below 2 units/g throughout lactation. 4. 4. Lipoprotein lipase activity in the inguinal-abdominal mammary glands increased very slowly during the first 20 days of pregnancy, from 0.9 units/g to 7 units/g. It then increased the next day to 21 units/g, decreased sharply at parturition to 6 units/g, and increased after parturition to more than 40 units/g. Lipoprotein lipase activity in mammary gland remained high as long as the mother suckled. 5. 5. Nonsuckling for 9 or more h decreased mammary gland lipoprotein lipase activity to near zero and increased plasma triglyceride concentration to 3 mM. Nonsuckling also increased lipoprotein lipase activity in adipose tissue. 6. 6. Ligation of the lactiferous ducts of the inguinal-abdominal glands on one side markedly reduced the lipoprotein lipase activity in these glands but not that in the contralateral, suckled glands. 7. 7. Plasma triglyceride concentration was inversely related to the lipoprotein lipase activity in mammary tissue during the last 2 days of pregnancy and throughout lactation. 8. 8. It is suggested that the changes in lipoprotein lipase activity of adipose and mammary tissues that occur during late pregnancy and lactation serve to divert dietary lipid from storage in adipose tissue to mammary glands for milk formation.

Lingual Lipase and Its Role in the Digestion of Dietary Lipid

The serous glands of rat tongue were found to contain a potent lipolytic enzyme which hydrolyzed triglyceride to mostly diglyceride and free fatty acids (FFA) at pH 4.5-5.4. Homogenates of lingual serous glands from adult rats hydrolyzed 40-70 mmol of triglyceride/g per h. The soft palate, anterior oral pharyngeal wall, and lateral oral pharyngeal glands also contained the activity, but at a much lower level. The lipolytic activity was also found in saliva collected through an esophageal cannula and in stomach contents of rats fed a fat-rich meal. The stomach contained very little activity, however, when saliva was excluded. Lipolytic activity was not found in the stomach wall or in the parotid, submandibular, and sublingual glands. The findings suggest that the lingual serous glands secrete a lipase which catalyzes in the stomach the conversion of triglyceride to partial glycerides and FFA. It is proposed that this reaction is the first step in the digestion of dietary lipid.

Uptake of blood triglyceride by various tissues.

Triglycerides are transported in the blood in chylomicrons and very low density lipoproteins. Electron microscopic studies indicate that these particles, which range in diameter from 0.03–0.6 μ, cannot cross the capillary endothelium in most tissues. There is now considerable evidence that the triglycerides are hydrolyzed to free fatty acids (FFA) during uptake and that this process is catalyzed by lipoprotein lipase. The enzyme is found in nearly all tissues that utilize circulating triglyceride, and the level of activity, in individual tissues, varies with nutritional and physiological states that affect triglyceride uptake, such as fasting, diabetes and pregnancy. Studies in perfused adipose tissue with doubly labeled chylomicrons showed that hydrolysis occurs outside of the blood stream. Two-thirds of the fatty acids are incorporated into tissue triglyceride and the rest are release as FFA, with glycerol, to the blood. Infusion of heparin causes immediate release of lipoprotein lipase activity to the blood and decreases the amount of chylomicron-triglyceride hydrolyzed by the tissue. Electron microscopic cytochemical studies showed that hydrolysis of blood glycerides by lipoprotein lipase in adipose tissue occurs within the capillary endothelial cells and in the subendothelial space near the pericytes, but not in the capillary lumen or near the fat cells. The results indicate that the fatty acids of chylomicrons cross the capillary endothelium as glycerides and FFA, within a membrane-bounded system, and cross the extravascular space to the fat cells as FFA.

Effect of insulin and acute diabetes on plasma FFA and ketone bodies in the fasting rat

The metabolism of FFA and ketone bodies was studied in fasted rats by infusing at a constant rate tracer amounts of FFA-(3)H, beta-hydroxybutyrate-(14)C or acetoacetate-(14)C for periods up to 2 hr. Blood that was removed for analyses was replaced by continuous transfusion. The rates of turnover of FFA, beta-hydroxybutyrate, and acetoacetate in rats fasted for 2 days were, respectively, 3.2, 5.6, and 2.5 mumoles/100 g body weight per min. Infusion of mannoheptulose with anti-insulin serum increased plasma glucose, FFA, and ketone body concentrations and decreased the specific activity of plasma FFA. Injection of insulin (20 mU i.v.) decreased almost simultaneously plasma glucose, FFA, and ketone body concentrations and increased the specific activity of FFA, but it did not affect the plasma concentration of FFA-(3)H. The findings indicate that insulin deprivation increased and insulin injection decreased the release of FFA from body tissues in fasting rats. The plasma FFA concentration in fasting rats was increased by infusing chylomicrons and heparin, but this had very little effect on either plasma ketone body or glucose concentrations. Insulin injection (20 mU i.v.) lowered the plasma ketone body concentration in these animals. Studies using beta-hydroxybutyrate-(14)C showed that insulin (50 mU i.v.) decreased ketogenesis in the presence of a sustained high plasma FFA concentration and had no effect on uptake of circulating ketone bodies. The results indicate that plasma FFA concentration is not the sole determinant of plasma ketone body concentration and that insulin can suppress ketone body production through some means other than lowering plasma FFA concentration.

Effect of albumin on products formed from chylomicron triacylglycerol by lipoprotein lipase in vitro

Abstract The effect of albumin and Ca2+ on the action of purified bovine milk lipoprotein lipase on chylomicron triacylglycerol in vitro was studied with rat lymph chylomicrons containing triacylglycerol labeled with [14C]oleic acid and [14H]glycerol. Lipoprotein lipase hydrolyzed chylomicron triacylglycerol to mostly glycerol and fatty acids when incubated in medium containing sufficient albumin to bind all of the fatty acids formed. There was, however, transient accumulation of monoacylglycerol during the first 5 min of incubation. It is suggested that the latter represented monoacylglycerol undergoing isomerization within chylomicrons before being hydrolyzed to glycerol and fatty acid. Hydrolysis of monoacylglycerol was markedly inhibited (67%) when the incubation mixture contained a 9-fold excess of albumin. This effect on hydrolysis probably resulted from binding of monoacylglycerol to albumin in the medium, thus making the monoacylglycerol inaccessible to the enzyme. When the incubation mixture contained limited albumin, hydrolysis of all acylglycerols was reduced, and diacylglycerol and monoacylglycerol accumulated. Although some of the monoacylglycerol was released to the medium, the molar ratio of monoacylglycerol to albumin in the medium never exceeded 0.35. In contrast, the molar ratio of fatty acids to albumin in the medium was sometimes as high as 7. Ca2+ was a poor substitute for albumin as an acceptor for fatty acids produced by the action of lipoprotein lipase on chylomicrons. The rate of hydrolysis of chylomicron triacylglycerol by lipoprotein lipase in vitro was related directly to the amount of enzyme added up to 1.7 μg (34 pmol) of enzyme per μmol of triacylglycerol. It was calculated that a nearly maximal rate was obtained when 43 molecules of enzyme were added per chylomicron.

Effects of Mannoheptulose on Glucose Metabolism of Isolated Tissues

Summary The mechanism of the diabetogenic action of mannoheptulose has been investigated by testing its action on glucose metabolism by isolated tissues. Mannoheptulose did not alter the in vitro metabolism of glucose by liver, kidney, diaphragm or epididymal fat pad. The stimulating effects of insulin on glucose metabolism of diaphragm and adipose tissue were not decreased by mannoheptulose.

Lipoprotein lipase and uptake of triacylglycerol, cholesterol and phosphatidylcholine from chylomicrons by mammary and adipose tissue of lactating rats in vivo.

The relationship between lipoprotein lipase activity and uptake of triacylglycerol, cholesterol and phosphatidylcholine from chylomicrons was studied in mammary gland and adipose tissue of rats lactating 6--7 days. 60% of triacylglycerol [14C]oleic acid, 13% of [3H]cholesterol and 8% of [32P]phosphatidylcholine in chylomicrons injected intravenously were taken up within 11 min by mammary gland whereas negligible amounts were taken up by adipose tissue. Non-suckling for 44 h decreased markedly uptake of all lipids by mammary gland and retarded clearance of chylomicrons from blood, while it increased significantly uptake of triacylglycerol fatty acids and cholesterol by adipose tissue. Non-suckling also decreased lipoprotein lipase activity in mammary gland from 7.7 to 0.4 units/g, while it increased activity in adipose tissue from 0.1 to 2.7 units/g. These findings indicate that lipoprotein lipase is involved in uptake of chylomicron triacyglycerol and cholesterol by mammary gland and adipose tissue, and also in uptake of chylomicron phosphatidylcholine by mammary gland. They also show that reciprocal changes in lipoprotein lipase activity in mammary gland and adipose tissue, as occur during lactation, result in diversion of chylomicron lipids from one tissue to the other.

Lipolysis and lamellar structures in white adipose tissue of young rats: lipid movement in membranes.

Lamellar structures with a periodicity of 40 A developed during incubation at 25°C in glutaraldehyde-fixed adipose tissue from young rats. The lamellae were found in transendothelial channels, at intercellular contacts, and in channels that extended from extracellular space to surfaces of lipid droplets in fat cells. They were also found in capillaries, associated with chylomicrons, in tissue from fed rats, and near lipid droplets of fat cells in tissue from fasted rats. The amount and distribution of lamellae varied with incubation time and metabolic state of the tissue donor. Development of lamellae under conditions causing lipolysis and accumulaton of fatty acids in fixed tissue indicates the lamellae were composed primarily of fatty acids. We conclude that fatty acids resulting from lipolysis accumulated in an interfacial continuum of external leaflets of cell membranes, and that when the continuum became crowded, fatty acids formed lamellar extensions of the continuum at different sites along its course through the tissue.

Regulation of Mammary and Adipose Tissue Lipoprotein Lipase and Blood Triacylglycerol in Rats during Late Pregnancy: EFFECT OF PROSTAGLANDINS

The effects of several prostaglandins on lipoprotein lipase activity of mammary gland and adipose tissue and serum triacylglycerol were studied during late pregnancy in rats. Prostaglandins were injected twice daily for 2 days before and once on the day of analysis. In rats pregnant 20 days, prostaglandin F(2alpha) (PGF(2alpha)) increased the activity of lipoprotein lipase in mammary gland fourfold, reduced the activity in adipose tissue about 60%, and decreased serum concentration of triacylglycerol 50%. PGF(2alpha) also reduced serum concentration of progesterone 90% and increased that of prolactin fivefold, but had no effect on serum concentrations of either immuno-reactive insulin or 17beta-estradiol. Injections of 13,14-dihydro-15-keto PGF(2alpha), a metabolite of PGF(2alpha), had similar effects in rats pregnant 20 days, whereas prostaglandins E(1) and E(2) did not. In rats pregnant 16 days, PGF(2alpha) did not affect lipoprotein lipase activity in the tissues or the concentration of triacylglycerol and prolactin in serum, although it decreased serum progesterone 80%.2-Br-alpha-ergocryptine prevented the increase in serum prolactin in response to PGF(2alpha), but did not alter the effect of PGF(2alpha) on lipoprotein lipase activity or serum triacylglycerol. Progesterone completely blocked the effects of PGF(2alpha) on lipoprotein lipase activity and serum triacylglycerol and prolactin concentrations. These findings indicate that the changes in lipoprotein lipase activity and serum triacylglycerol in PGF(2alpha)-treated rats are probably related to the inhibitory action of PGF(2alpha) on progesterone secretion. They also suggest that endogenous F prostaglandins may play a role in the regulation of lipoprotein lipase activity in mammary gland and adipose tissue near parturition.

Membrane continuities within cells and intercellular contacts in white adipose tissue of young rats

White adipose tissue of suckling rats was examined for evidence of continuity of membranes within cells, and areas of contact between adipocytes and capillary endothelial cells. Extracellular space, invaginations of cell surfaces, and lumens of channels and vesicles within both types of cells were marked with electron-opaque material in tissues which were treated with tannic acid after being fixed with glutaraldehyde. The electron-opaque material also marked channels that extended from extracellular space to the surface of intracellular lipid droplets in fat cells. Presence of this material in channels and vesicles, but not in cytoplasm, indicates they were open to extracellular space during treatment with tannic acid and, furthermore, that their membranes were continuous with plasma membrane. Cell processes of adipocytes and endothelium extended through basement membrane to make contact with each other. Apparent continuity of the external leaflet of plasma membrane of one cell with that of another cell was observed. These findings demonstrate membrane continuity between plasma membrane and membranes of intracellular channels in adipocytes and endothelial cells, and intercellular contacts in adipose tissue.