Release Of Lipoprotein Lipase Activity Research Articles

Release of Lipoprotein Lipase Induced by L-Arginine in Mouse Mammary Tumor FM3A Cells

The nitric oxide (NO)/cyclic guanosine 5'-monophosphate (GMP) pathway remains undefined regarding the regulation of lipoprotein lipase (LPL) release. Here, we investigated whether L-arginine (Arg) stimulates the release of LPL from mouse mammary tumor FM3A cells in a time- and dose-dependent manner. L-Arg-stimulated release of LPL activity was inhibited by NG-monomethyl-L-Arg monoacetate, which is an endothelial NO synthase (NOS) inhibitor. Furthermore, release of enzyme activity was also suppressed by various inhibitors of guanylate cyclase and adenylate cyclase, as well as cyclic GMP- and cyclic adenosine 5'-monophosphate (AMP)-dependent protein kinases (PKG and PKA). L-Arg also increased intracellular cyclic GMP contents as well as intracellular cyclic AMP contents. In addition, the increase in the cyclic AMP contents was reduced by inhibitors of guanylate cyclase and PKG. These results suggest that the stimulatory release of LPL from tumor cells by L-Arg is partly due to activation of cyclic AMP production and PKA activity caused by elevated cyclic GMP production and PKG activity.

An in situ perfusion protocol of rat epididymal adipose tissue useful in metabolic studies

Experimental approaches involving the perfusion of tissues and organs offer the advantage of improved physiological relevance over the use of isolated tissues or cells while at the same time being much more controlled and tissue-specific than studies in vivo. Nevertheless, there have been few metabolic studies performed in perfused white adipose tissue, largely because of the difficulty of the surgical technique involved. Although some methods have been described, they are difficult to use as perfusion protocols and their reproducibility is poor. We have modified a rat perfusion method, based on a modification of the Ho and Meng technique, for use with epididymal white adipose tissue (eWAT), and we present it here as a protocol to be reproduced. We also offer surgical solutions for the most common variants of vessel distributions in rats. Using the protocol described here, the perfused adipose tissue is viable and metabolically active, as indicated by the maintenance of tissue ATP levels and adiponectin secretion and by endogenous lipolysis regulation. Moreover, there is a high level of lipoprotein lipase activity in the endothelium of the tissue, which is heparin-releasable. Thus, this method is a useful and reproducible tool that allows the perfusion of eWAT for use in metabolic studies.

Effect of Oral Kohki Tea on Bladder Dysfunction Induced by Severe Partial Outlet Obstruction

Extracts of the leaves of Engelhardtia chrysolepis, a subtropical plant that grows wild in southern China, have been used medicinally in east Asia for hundreds of years. A standard extract named Kohki tea (Maruzen Pharmaceuticals, Onomichi City, Japan) is sold over the counter in Japan as a sweet tea shown to confer many beneficial effects on general health and well-being. The tea contains strong antioxidants, including several dihydroflavonol glycosides. The results of previous studies show that natural products with antioxidant activities provide protective effects on the bladder of rabbits with partial outlet obstruction. We determined in vivo and in vitro whether oral pretreatment of rabbits with Kohki tea protects the bladder from dysfunction induced by partial outlet obstruction. A total of 28 New Zealand White rabbits were separated into 4 groups of 7 each. Rabbits in groups 1 and 2 were treated by gavage with 100 mg./kg. Kohki tea daily in distilled water, while those in groups 3 and 4 were given distilled water. After 4 weeks of daily oral administration each rabbit was sedated, the bladder was catheterized and cystometry was performed at a filling rate of 1 ml. per minute. At the completion of cystometry the rabbits were immediately anesthetized. Moderate outlet obstruction was created in groups 1 and 3, and sham surgery was performed in groups 2 and 4. Treatment was continued for an additional 4 weeks, when each rabbit was sedated and cystometry was repeated. After cystometry the bladder was exposed through a midline incision, excised, weighed and 4 strips of bladder body were cut for contractility studies. The balance of the bladder was separated between smooth muscle and mucosa by blunt dissection, frozen in liquid nitrogen and stored at -70C for biochemical analyses. Partial outlet obstruction stimulated similar increases in the bladder weight of all obstructed rabbits. Partial outlet obstruction resulted in a significant decrease in bladder compliance in all obstructed animals. However, the bladder of obstructed rabbits given Kohki tea were significantly more compliant than those given water. Voiding pressures in the control group and the obstructed group given distilled water were approximately equal, while obstructed rabbits given Kohki tea showed significantly higher maximal voiding pressure. The contractile responses to all forms of stimulation were reduced by obstruction to a significantly greater degree in the rabbits not given tea than in those given tea. Sarcoplasmic reticulum Ca2+-adenosine triphosphatase enzyme activity of the bladder was significantly reduced in obstructed rabbits given vehicle but activity was not reduced in obstructed rabbits given Kohki tea. Kohki tea had a significant protective effect on bladder function, contractile responses and bladder biochemistry in rabbits with moderate to severe partial outlet obstruction.

Localization of lipoprotein lipase in the diabetic heart: regulation by acute changes in insulin.

Vascular endothelium-bound lipoprotein lipase (LPL) is rate limiting for free fatty acid (FFA) transport into tissues. In streptozotocin (STZ)-diabetic rats, we have previously demonstrated an increased heparin-releasable LPL activity from perfused hearts. Because heparin can traverse the endothelial barrier, conventional Langendorff retrograde perfusion of the heart with heparin could release LPL from both the capillary luminal and abluminal surfaces. To determine the precise location of the augmented LPL, a modified Langendorff retrograde perfusion was used to isolate the enzyme at the coronary lumen from that in the interstitial effluent. In response to heparin, a 4-fold increase in LPL activity and protein mass was observed in the coronary perfusate after 2 weeks of STZ diabetes. Release of LPL activity into the interstitial fluid of control hearts was slow but progressive, whereas in diabetic hearts, peak enzyme activity was observed within 1 to 2 minutes after heparin, followed by a gradual decline. Immunohistochemical studies of myocardial sections confirmed that the augmented LPL in diabetic hearts was mainly localized at the capillary endothelium. To study the acute effects of insulin on endothelial LPL activity, we examined rat hearts at various times after the onset of hyperglycemia. An increased heparin-releasable LPL activity in diabetic rats was demonstrated shortly (6 to 24 hours) after STZ injection or after withdrawal from exogenous insulin. Heparin-releasable coronary LPL activity was also increased after an overnight fast. These studies indicate that the intravascular heparin-releasable fraction of cardiac LPL activity is acutely regulated by short-term changes in insulin rather than glucose. Thus, during short periods (hours) of hypoinsulinemia, increased LPL activity at the capillary endothelium can increase the delivery of FFAs to the heart. The resultant metabolic changes could induce the subsequent cardiomyopathy that is observed in the chronic diabetic rat.

A vanadyl sulfate-bovine serum albumin complex stimulates the release of lipoprotein lipase activity from isolated rat fat pads through an increase in the cellular content of cAMP and myo-inositol 1,4,5-trisphosphate.

A vanadyl sulfate-bovine serum albumin complex (vanadyl-BSA) prolonged the stability of the V4+ oxidation state, although vanadyl alone can readily change the oxidation state from V4+ to V5+ under physiological conditions. Vanadyl-BSA stimulated the release of lipoprotein lipase (LPL) activity from isolated rat fat pads and increased the cellular LPL activity in a time-dependent manner. These effects were independent of protein synthesis. Propranolol, quin 2-AM, ruthenium red, and neomycin all inhibited LPL release more potently than the increase in activity. In contrast, potent inhibition of the increase effect was observed with genistein and wortmannin. Short-term incubation of the fat pads with vanadyl-BSA showed a transient increase in the cellular content of cAMP and myo-inositol 1,4,5-trisphosphate (IP3), which was inhibited by propranolol and neomycin, respectively. These results suggest that vanadyl-BSA stimulates the release of LPL activity through an increase in the cellular content of cAMP and IP3, leading to an increased intracellular Ca2+ concentration, and that it also increases cellular LPL activity via process(es) sensitive to genistein and wortmannin.

Heparin-releasable lipoprotein lipase activity is increased in cardiomyocytes after culture

The activity of lipoprotein lipase (LPL) in adult rat heart cardiomyocytes after overnight culture on laminin-coated plates for 18-22 h was compared with enzyme activity in freshly isolated cardiomyocytes. LPL activity in cellular homogenates from cultured cardiomyocytes and freshly isolated cells was 240 and 233 nmol oleate released h-1.mg-1 protein, respectively. LPL specific activity (mU/ng LPL protein) was 0.07 in cultured cells compared with 0.42 in freshly isolated cells, indicating an increased content of inactive LPL mass after overnight culture. The heparin-induced release of LPL activity into the medium of cultured cardiomyocytes (198 nmol.h-1.mg-1) was much greater than heparin-releasable LPL (HR-LPL) activity (59 nmol.h-1.mg-1) from freshly isolated cells. HR-LPL activity from cultured cardiomyocytes was dependent on serum (16.3-fold activation) and was inhibited by high ionic strength (1 M NaCl) and by a polyclonal antibody to LPL. Cultured cardiomyocytes also had more immunodetectable LPL on the cell surface compared with freshly isolated cardiomyocytes, consistent with increased HR-LPL activity. Therefore, overnight culture may permit cardiomyocytes time to recover from the stress of isolation by increasing the content of LPL on the cell surface.

Enhancement of the vanadate-stimulated release of lipoprotein lipase activity by astilbin from the leaves of Engelhardtia chrysolepis.

Astilbin, a dihydroflavonol rhamnoside isolated from the leaves of Engelhardtia chrysolepis, enhanced the vanadate-stimulated release of lipoprotein lipase (LPL) activity from rat isolated fat pads. N-[2-(Methyl-amino)ethyl]-5-isoquinolinesulfonamide (H-8), a potent inhibitor of cAMP-dependent protein kinase (PKA), markedly inhibited the enhancement by astilbin. Lipolysis in the fat pads was stimulated by astilbin alone in a dose-dependent manner and this stimulation was suppressed in the presence of vanadate, probably due to its antilipolytic action. A significant enhancement by astilbin was observed with increasing effects of vanadate on cAMP content in the fat pads and on cAMP phosphodiesterase (PDE) activity in the particulate fraction although astilbin alone showed only a slight increase in the cellular cAMP content and PDE activity. Astilbin may enhance the vanadate-stimulated release of LPL activity through a synergistic effect on an increase in the cellular cAMP content produced by vanadate accompanied by more potent activation of PKA.

Involvement of adenosine in vanadate-stimulated release of lipoprotein lipase activity.

A short-term incubation of isolated rat fat pads with vanadate showed the stimulated release of lipoprotein lipase (LPL) activity and suppression of the rise in extracellular adenosine level. The addition of adenosine to the medium showed inhibition of both the stimulated release of LPL activity and an increase in intracellular cAMP content by vanadate. A progressive increase in 5'-nucleotidase activity in the particulate fraction containing plasma membrane was suppressed by vanadate in a time- and dose-dependent manner, suggesting that vanadate inhibits, in part, the production of adenosine based on a dephosphorylation of AMP. In adipocytes, the inhibition of adenylate cyclase via A1 adenosine receptor is more predominant than the stimulation of adenylate cyclase via A2 adenosine receptor (Londos C. et al., Proc. Natl. Acad. Sci. U.S.A., 75, 5362-5366 (1978)). Therefore, vanadate may stimulate the release of LPL activity from the fat pads by suppressing the rise in extracellular adenosine level, accompanied by the activation of adenylate cyclase activity.

Release of lipoprotein lipase from Ehrlich ascites tumor produced by an association with a rapid increase in cyclic AMP content

Although it is considered that the lipoprotein metabolism in tumor plays an important role in growth and multiplication, it is not clear as to the details. Lipoprotein lipase (LPL) is a key enzyme responsible for the hydrolysis of lipoprotein-triacylglyceride. In this study, we examined the regulatory step of LPL in lipoprotein metabolism of Ehrlich ascites tumor and especially the enzyme-release from the tumor cells. When LPL was stimulated to release from the tumor cells by the low molecular weight dextran sulfate (3.2 kDa), cyclic AMP content in the tumor cells was observed to increase rapidly in a time-dependent manner up to 30 s; its maximal effect was 1.5-fold higher than the basal level of cyclic AMP. The increase in cyclic AMP content was more enhanced in the presence of isobutylmethylxanthine and was never suppressed by propranolol. Moreover, cyclic AMP-dependent protein kinase (PKA) activity in the tumor cells was also recognized to elevate in a time- and dose-dependent manner. In addition, the release of LPL activity from the tumor cells was inhibited by 2′,5′-dideoxyadenosine. These results suggest that LPL in the tumor cells is released through a pathway involving an activation of PKA associated with the rapid increase in cyclic AMP content.

Involvement of the rapid increase in cAMP content in the vanadate-stimulated release of lipoprotein lipase activity from rat fat pads.

Mechanisms of the stimulatory release of lipoprotein lipase (LPL) activity from isolated rat fat pads by sodium orthovanadate (vanadate) were studied through a cAMP-dependent process. A potent inhibitor of insulin receptor tyrosine kinase, quercetin, inhibited the vanadate-increasing effect on the LPL activity in fat pads, but did not inhibit the vanadate-stimulated release of LPL activity from the fat pads. Propranolol and N-[2-(methylamino)ethyl]-5-isoquinolinesulfonamide (H-8) decreased the vanadate-stimulated release in a dose-dependent manner. Isoproterenol and dibutyryl cAMP (Bt2cAMP) stimulated the release of LPL activity from fat pads. Vanadate, as well as isoproterenol, rapidly increased the cAMP content in fat pads, and this increase was almost completely inhibited by propranolol. Vanadate increased the cAMP-dependent protein kinase (PKA) activity ratios calculated from the measurement in the presence or absence of cAMP or PKa inhibitor. These results suggest that the vanadate-stimulated release of LPL activity is associated with a process involving a rapid increase in the cAMP content accompanied by the activation of PKA.

Sodium orthovanadate increases phospholipase A2 activity in isolated rat fat pads: a role of phospholipase A2 in the vanadate-stimulated release of lipoprotein lipase activity.

Phospholipase (PL) A2 activity prepared from isolated rat fat pads incubated with sodium orthovanadate (vanadate) was increased in a time- and dose-dependent manner. The increasing effect of vanadate was reduced in the presence of tyrosine kinase inhibitors. Under the inhibition of protein synthesis by cycloheximide, vanadate still showed a full effect on the increase in PL A2 activity. Various PL A2 inhibitors, such as manoalide, quinacrine and p-bromophenacyl bromide, suppressed the stimulatory release of lipoprotein lipase (LPL) activity from the fat pads by vanadate. Moreover, the vanadate-stimulated release of LPL activity was decreased by the cyclooxygenase and thromboxane synthetase inhibitors, and a thromboxane A2 receptor antagonist, but was never suppressed by a lipooxygenase inhibitor. The stimulatory release of LPL activity by vanadate was also decreased in the presence of tyrosine kinase inhibitors. These results suggest that vanadate increases PL A2 activity, and the increase in PL A2 activity is partly involved in the vanadate-stimulated release of LPL activity with an association to the membrane tyrosine kinase.

Stimulatory release of lipoprotein lipase activity with activation of protein tyrosine kinase produced by low molecular weight dextran sulfate in Ehrlich ascites tumor cells.

Lipoprotein lipase (LPL), which is responsible for the hydrolysis of lipoprotein triacylglyceride, has been examined in Ehrlich ascites tumor cells. Low molecular weight dextran sulfate (DXS, M.W. 3.2 kDa) stimulates the release of the enzyme activity from the tumor cells into the incubation medium in a time-dependent manner. The stimulatory effect of DXS was markedly decreased by incubation with protein tyrosine kinase (TK) inhibitors, such as ST 638, biochanin A and amiloride. The activity of the partially purified TK preparation from the tumor cells was found to be increased following incubation with DXS in a manner which was both time- and dose-dependent. These results suggest that the stimulatory release of LPL activity by DXS is associated with the activation of TK in the tumor cells.

Low-Mr heparin is as potent as conventional heparin in releasing lipoprotein lipase, but is less effective in preventing hepatic clearance of the enzyme

This study compares a low-Mr heparin preparation with conventional heparin with respect to its interaction with lipoprotein lipase (LPL) in vitro and its effects on the enzyme in vivo. Both heparin preparations were polydisperse in binding to LPL, but on average the low-Mr preparation showed lower affinity. Thus both conventional and low-Mr heparin bound quantitatively to immobilized LPL, and were eluted as broad peaks when a salt gradient was applied, but the peak for low-Mr heparin was shifted towards lower salt concentrations. To displace LPL from immobilized heparin a higher concentration of low-Mr than of conventional heparin was needed. Injection of the low-Mr heparin into intact rats resulted in lower plasma LPL activity than did injection of an equal mass of conventional heparin, but when the liver was excluded from the circulation both heparin preparations resulted in similar plasma LPL activities. In perfused rat hearts, low-Mr heparin had at least the same effect on the release of LPL activity as did conventional heparin. In perfused livers, on the other hand, low-Mr heparin was less effective than conventional heparin in preventing the rapid uptake of exogenous labelled LPL. Hence the apparently lower average affinity of low-Mr heparin for LPL does not result in a demonstrably lower potency to release the enzyme from endothelial binding sites in peripheral tissues, but does result in a substantially decreased effect on the hepatic clearance of the enzyme.

Release of lipoprotein lipase and hepatic lipase activities. Effects of heparin and a low molecular weight heparin fragment

Unfractionated heparin and a low molecular weight heparin fragment (LMWH, mean molecular weight 5000) were compared with respect to the ability to release lipoprotein lipase (LPL) and hepatic lipase (HL) from tissue binding sites. The investigations were carried out in vivo and in vitro in man and in the rat. The in vivo release of LPL activity was greater with heparin in both species. The release of HL activity was equal with both heparins in man, but greater with unfractionated heparin in the rat. In the in vitro studies the LMWH fragment consistently released more LPL activity from the tissues investigated, i.e. fat, skeletal muscle and heart muscle. With isolated adipocytes, however, we could not demonstrate any difference between the two heparins with respect to their lipase-releasing effect. In liver tissue homogenate the two heparins showed the same ability to release HL activity. It is concluded that not only biochemical differences between the two heparins are of importance for the difference in the lipolytic effect. The possibility of different rheological behaviours of the two heparins in the capillary lumen, which might partly explain the greater in vivo lipolytic effects of heparin, is discussed. It is suggested that LMWH enters the tissue preparation more readily in vitro, thereby exerting a greater lipase-releasing effect than heparin.

Stimulatory release of lipoprotein lipase activity from rat fat pads by vanadate

Vanadate stimulated the release of lipoprotein lipase (LPL) activity from rat fat pads into the medium in a time- and dose-dependent manner. It exerted the synergetic effect with heparin. The stimulatory effects of vanadate and heparin were decreased by incubation in Na +- or Ca 2+-free media but were well preserved in K +-free medium. Amiloride inhibited the vanadate-stimulated release of LPL activity in a dose-dependent manner, but did not inhibit the heparin-stimulated release of LPL activity. Colchicine, antimycin A, and carbonyl cyanide m-chlorophenylhydrazone suppressed the stimulatory effect of vanadate, but cycloheximide did not. Preincubation of the fat pads with the tetrakis (acetoxymethyl) ester of quin 2 (quin 2-AM) inhibited the vanadate-stimulatory release of LPL activity without affecting basal activity. The concentration required for half-maximal inhibition of the action of vanadate by quin 2-AM was calculated to be 39 μ m, suggesting that the action of vanadate was dependent on intracellular Ca 2+ concentration. The heparin-stimulated release, on the other hand, was not inhibited even at higher concentrations of quin 2-AM (up to 200 μ m). These findings suggest that vanadate stimulates the release of LPL activity through mechanisms of action involving amiloride-sensitive and calcium-dependent pathways with a requirement of metabolic energy.

Lipoprotein lipase activity in neonatal-rat liver cell types.

The lipoprotein lipase activity in the liver of neonatal (1 day old) rats was about 3 times that in the liver of adult rats. Perfusion of the neonatal liver with collagenase decreased the tissue-associated activity by 77%. When neonatal-rat liver cells were dispersed, hepatocyte-enriched (fraction I) and haemopoietic-cell-enriched (fraction II) populations were obtained. The lipoprotein lipase activity in fraction I was 7 times that in fraction II. On the basis of those activities and the proportion of both cell types in either fraction, it was estimated that hepatocytes contained most, if not all, the lipoprotein lipase activity detected in collagenase-perfused neonatal-rat livers. From those calculations it was also concluded that haemopoietic cells did not contain lipoprotein lipase activity. When the hepatocyte-enriched cell population was incubated at 25 degrees C for up to 3 h, a slow but progressive release of enzyme activity to the incubation medium was found. However, the total activity (cells + medium) did not significantly change through the incubation period. Cycloheximide produced a time-dependent decrease in the cell-associated activity. Heparin increased the amount of lipoprotein lipase activity released to the medium. Because the cell-associated activity was unchanged, heparin also produced a time-dependent increase in the total activity. In those cells incubated with heparin, cycloheximide did not affect the initial release of lipoprotein lipase activity to the medium, but blocked further release. The cell-associated activity was also decreased by the presence of cycloheximide in those cells. It is concluded that neonatal-rat hepatocytes synthesize active lipoprotein lipase.

Studies with etofibrate in the rat. Part I: Effects on glycerol, free fatty acid and triacylglycerol metabolism

Etofibrate is the 1,2-ethandiol diester of clofibric acid and nicotinic acid that decreases circulating levels of triacylglycerols and cholesterol. To understand the mechanism by which the drug affects plasma triacylglycerols, normolipemic rats were treated daily with 300 mg of etofibrate /kg body weight or with the medium by a stomach tube. They were decapitated on the 10th day, and showed lower levels of plasma β-hydroxybutyrate, glycerol, free fatty acids (FFA), total triacylglycerols and cholesterol and VLDL triacylglycerols and cholesterol, whereas glucose and RIA-determined insulin levels were unmodified. Epididymal fat pad pieces from etofibrate-treated rats incubated in vitro released more glycerol but the same amount of FFA to the medium, and had greater uptake of [U- 14C]glycerol for [ 14C]acylglycerol formation. In the presence of heparin, they also showed an enhanced release of lipoprotein lipase activity to the medium. The disappearance from plasma of intravenously administered [1- 14C]palmitate was faster in the etofibrate-treated rats, and although they showed a decrease in 14C-esterified fatty acids of neutral lipids in both liver and plasma VLDL, there was an increase in liver 14C-labelled water-soluble components. After intravenous [U- 14C]glycerol administration, there was a decrease in plasma VLDL [ 14C]acylglycerol and [ 14C]glucose and in liver [ 14C]acylglycerol, but an increase in plasma [ 14C]lactate. In the liver, etofibrate treatment heightened the cytosolic glycerol-3-phosphate dehydrogenase activity and the total carnitine concentration, whereas it reduced triacylglycerol and cholesterol concentrations. It is proposed that etofibrate enhances the reesterification of fatty acids and glycerol in adipose tissue, which, together with its augmented lipoprotein lipase activity, may facilitate the clearance of circulating triacylglycerols. These effects may act concomitantly with the decreased synthesis of triacylglycerols, secondary to the increased utilization of their precursors, acyl-CoA and glycerol-3-phosphate, in other pathways, causing the reduction of plasma VLDL triacylglycerols produced by etofibrate treatment.

Plasma kinetics of lipoprotein lipase and hepatic lipase activities induced by heparin and a low molecular weight heparin fragment

Intravenous injections of conventional heparin and a low molecular weight heparin fragment (LMWH, mean molecular weight 4000-6000) were given to six male volunteers at doses of 10, 50 and 100 U (antiFXa)/kg body wt. The plasma kinetics of lipoprotein lipase (LPL) and hepatic lipase (HL) were analysed. The peak values, as well as the accumulated release of LPL activity, were dose dependent and were twice as high after heparin as after LMWH. The plasma half-life of LPL activity followed first order kinetics and was similar for both heparin preparations when given in comparable doses. The peak values and the plasma half-life of HL activity were the same for heparin and LMWH in the clinically relevant doses (50 and 100 U (antiFXa)/kg). Compared with LMWH, the total release of HL was twice as large after the heparin injections, possibly due to mobilization of an additional enzyme pool by the conventional heparin. It is concluded that the use of LMWH as an anticoagulant is associated with a lower plasma lipolytic activity than with standard heparin.

Heparin-independent release of lipoprotein lipase activity from perfused rat hearts

Heparin-independent release of lipoprotein lipase activity from isolated perfused rat hearts was measured and related to the rapid turnover of the enzyme. Hearts consistently released lipoprotein lipase activity (2.1 ± 0.2 U/g released per min) during 60 min of nonrecirculating perfusion without heparin. This rate of release did not significantly differ from that measured in heparin-perfused hearts after the first 10 min of perfusion (2.2 ± 0.2 U/g released per min). The fractional release rate of lipoprotein lipase activity during nonheparin perfusion was 1.3% per min, which was higher than that calculated for alkaline phosphatase (0.002%) and creatine kinase (0.03%) activities. The lipase activity released was activated 4-fold by serum and inhibited 94 and 88% by 0.5 M NaCl and 3 mg/ml protamine sulfate, respectively. Lipoprotein lipase activity in the 1-min heparin-releasable (extracellular) and residual (intracellular) compartment remained stable during the last 40 min of nonheparin perfusion. During this period total heart, intracellular and extracellular enzyme t 1 2 were calculated to be 52, 42 and 10 min, respectively. The results are consistent with the postulate that continuous release of lipoprotein lipase into the vascular compartment may be an important determinant of its rapid turnover in the heartand possibly other tissues.

Changes with starvation in the rat of the lipoprotein lipase activity and hydrolysis of triacylglycerols from triacylglycerol-rich lipoproteins in adipose tissue preparations

Lipoprotein lipase activity was higher in fat-pad pieces than in isolated adipocytes from the same fed rats, whereas hydrolysis of triacylglycerols from triacylglycerol-rich lipoproteins was similar in the two preparations when incubated either in basal conditions or in the presence of heparin. In both preparations there was a similar release of lipoprotein lipase activity into the medium during basal incubation, enhanced by the presence of heparin. In fat-pad pieces, but not in isolated adipocytes, incubation with heparin produced a decrease in the lipoprotein lipase activity measured in the tissue preparation. In fat-pad pieces from 24 h-starved rats, lipoprotein lipase activity was the same as in isolated adipocytes from the same animals and incubation with heparin did not affect the appearance of lipoprotein lipase in the medium or the utilization of triacylglycerols from triacylglycerol-rich lipoproteins. These results support the following conclusions. (1) The effectiveness of lipoprotein lipase in adipose tissue preparations in vitro depends more on its availability to the substrate than on its total activity. (2) Heparin acts on adipose tissue preparations from fed animals both by enhancing the release of pre-existing extracellular enzyme (which is absent in isolated adipocytes) and by enhancing the transfer outside the cells of the intracellular (and mainly undetectable) enzyme that is activated in the secretion process. (3) In adipose tissue from starved animals there is not only a decrease in the active extracellular form of lipoprotein lipase activity but also a reduction in the intracellular (and mainly undetectable) pool of the enzyme.