Regression of poloxamer 407-induced atherosclerotic lesions in C57BL/6 mice using atorvastatin.

HMG-CoA reductase inhibitor drugs or 'statins' have been shown to effectively reduce plasma total cholesterol (CHOL), CHOL associated with low-density-lipoprotein (LDL), and triglycerides (TG). In addition, slight elevations in HDL-CHOL are also typically observed. Poloxamer 407 (P-407), a nonionic surfactant, effectively elevates both plasma CHOL and especially TG in a dose-controlled fashion and results in formation of atherosclerotic lesions in the aortas of C57BL/6 mice without the requirement of dietary cholic acid [1,2]. The purpose of the present study was to assess whether a typical statin, namely atorvastatin (Lipitor(R)) would significantly reduce P-407-induced hypercholesterolemia and hypertriglyceridemia as well as cause regression of atherosclerotic lesions resulting from administration of P-407 to C57BL/6 mice. C57BL/6 mice in the present study were treated with either normal saline (C, controls), 0.5 g/kg of P-407 (P), or a high-fat, high-cholesterol, cholate-containing diet (HF) for 120 days. Mice in all groups were then equally and randomly divided and treated with either atorvastatin or saline for an additional 120 days. Beginning at Day 121 and using mice in groups P and HF as an example, one-fourth of the mice in each group received 20 mg/kg per day of atorvastatin with either concomitant HF feeding or P-407 administration ('progression' treatment groups), one-fourth received 20 mg/kg per day of atorvastatin following cessation of HF feeding or P-407 administration, one-fourth received saline (placebo) with either simultaneous HF feeding or P-407 administration ('progression' placebo groups), and one-fourth received saline (placebo) following cessation of HF feeding or P-407 administration. Total plasma CHOL was significantly (P<0.01) lower for mice in groups P and HF when administered atorvastatin relative to saline, but remained significantly (P<0.05) elevated compared to total plasma CHOL of C mice. With discontinuation of either P-407 administration or HF feeding, total plasma CHOL declined rapidly in both P and HF mice with atorvastatin-treated mice generally demonstrating lower plasma CHOL concentrations relative to saline-treated mice. Total plasma TG was significantly (P<0.01) lower for mice in group P administered atorvastatin relative to saline, but remained significantly (P<0.05) elevated compared to plasma TG of C mice. With discontinuation of P-407 administration, total plasma TG declined rapidly in P mice with atorvastatin-treated mice typically demonstrating lower plasma TG concentrations relative to saline-treated P mice. Aortas of mice treated with 20 mg/kg per day of atorvastatin in both groups P and HF, whether maintained on the HF-diet or treated with P-407 from Day 120 to 240 or whether each treatment was terminated at Day 120, revealed no presence of atherosclerotic lesions relative to saline-treated mice and were indistinguishable from aortas retrieved from C mice. Atorvastatin at a dose of 20 mg/kg per day not only significantly reduced the plasma CHOL and TG concentrations, but also resulted in regression of atherosclerotic lesions induced in C57BL/6 mice by administration of P-407 or ingestion of a HF-diet containing cholic acid.

Potential downregulation of HMG-CoA reductase after prolonged administration of P-407 in C57BL/6 mice.

This study investigated the potential alteration in the amount of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase messenger RNA (mRNA) and lipoprotein lipase (LPL) mRNA in the livers of C57BL/6 mice after long-term (200 days) treatment with the nonionic surfactant called poloxamer 407 (P-407). Previously, P-407 has been used to produce a dose-controlled hyperlipidemic state in C57BL/6 mice with subsequent formation of atherosclerotic lesions. Five groups of mice were studied; controls (C); mice fed a standard chow diet enriched with only cholic acid (CH); mice fed the high-cholesterol, high-fat Paigen diet (HF); mice treated with 0.5 g/kg P-407 every third day (P); and mice administered 0.5 g/kg P-407 every third day while consuming a diet identical to that of mice in group CH (PC). Neither a significant (p < 0.05) weight loss nor alteration in liver enzymes (AST and ALT) were observed for any group throughout the study when compared with the control mice. Total plasma cholesterol (CHOL) was significantly elevated compared with controls for mice in groups HF, P, and PC, whereas total plasma triglycerides (TG) were significantly increased for mice in only groups P and PC. Long-term ingestion of a high-fat diet or a diet enriched in cholic acid resulted in a significant (p < 0.05) reduction in HDL-CHOL when compared with controls. Plasma samples assayed at 200 days for mice in groups HF and P showed a shift in the lipoprotein fraction distribution primarily to VLDL-CHOL as compared with mice in group C in which, as expected, most of the CHOL was contained in the HDL fraction. The biologic activity of HMG-CoA reductase assayed in hepatic microsomal homogenates was significantly reduced for mice in groups CH (p < 0.01), HF (p < 0.01), and PC (p < 0.05), but not for mice in group P, when compared with control. A statistical analysis of the data demonstrated significant (p < 0.05) reductions in the HMG-CoA reductase mRNA levels in hepatic tissue for all treatment groups relative to mRNA levels determined for mice in group C. In contrast, no treatment group demonstrated a significant difference in hepatic LPL mRNA levels when compared with mRNA levels determined for control animals. These data demonstrate that P-407 administration to C57BL/6 mice significantly decreased the amount of HMG-CoA reductase mRNA detected in liver.

Poloxamer 407-induced atherogenesis in the C57BL/6 mouse.

Poloxamer 407 (P-407) induces hyperlipidemia in the rat. It was the purpose of this investigation to determine if chronic P-407 administration would produce atherogenic arterial lesions in the C57BL/6 mouse, a strain reported to be susceptible to hyperlipidemia-induced atherosclerotic plaque formation. One injection (i.p.) of P-407 (0.5g/kg) produced hypercholesterolemia in the mouse that peaked at 24 h and returned to control levels by 96 h following treatment. Four groups of mice were maintained: (1) saline injected (C); (2) P-407-injected (0.5g/kg every 3rd day) (P); (3) P-407 injected plus cholic acid in the diet (PC); and (4) mice fed a high cholesterol (CHOL) diet containing cholic acid (HF). Mice from each group were sacrificed following 90, 145, 200, or 300 days of treatment. Plasma lipid concentrations, hepatic CHOL concentrations (145 and 300 day), and aortic atherogenic lesion areas were measured. Plasma CHOL and triglyceride remained at control levels throughout the 300 days in the C group. CHOL of the HF animals plateaued at approximately 225 mg/dl. P-407 produced CHOL concentrations of 600 mg/dl in P mice and 1000-1500 mg/dl in PC animals. There was no lesion formation in C mice. However, by 90 days lesions were present in the three other groups. Size of the lesions progressed through day 300 with the largest lesions (184.33 + 27.99 mu2 x 10(-3)) being present in the PC mice. HF and P animals had lesions of 70.50 + 11.35 and 43.33 + 7.88 mu2 x 10(-3), respectively. This study provides an animal model where atherogenesis has been produced with hyperlipidemia induced using a chemical agent.

The poloxamer 407-induced hyperlipidemic atherogenic animal model.

We are attempting to develop a chemically-induced murine model for the study of atherosclerosis. Injection of poloxamer-407 (P-407) into rats and mice causes significant dose-dependent hypercholesterolemia and hypertriglyglyceridemia. The elevated triglycerides (TG) seem to result primarily from the compound's inhibition of lipoprotein lipase. P-407 also indirectly stimulates the activity of the rate limiting enzyme in cholesterol (CHOL) biosynthesis, HMG CoA reductase. In addition, P-407 promotes changes in the concentration of hepatic CHOL content. These date indicate that the hyper CHOL could be the result of increased CHOL synthesis, as well as a clearing of CHOL from the liver. Chronic injection into mice of P-407 for 145 d produced atherogenic lesions in the aortas of C57BL/6 mice. The response was equivalent to that seen in animals eating a high CHOL diet for 145 d. Cholic acid potentiated the P-407-induced atherogenesis. These data suggest that P-407 could be used as an agent for the study of hyperlipidemia-induced atherogenesis.

Effect of poloxamer 407 on the activity of microsomal 3-hydroxy-3-methylglutaryl CoA reductase in rats.

A single 300-mg i.p. injection of poloxamer 407 (P-407, also called Pluronic F-127) in rats produces a marked hypercholesterolemia for a minimum of 96 h. The purpose of this investigation was to determine mechanisms by which P-407 causes hypercholesterolemia. The enzyme targeted for investigation is the rate-limiting enzyme in cholesterolgenesis, 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase. Injection of P-407 in fasted rats resulted in a significant (p < 0.05) elevation in plasma cholesterol (61.2 +/- 4.2 mg/dl) as soon as 1 h after injection compared with sham-injected controls (50.1 +/- 3.7 mg/dl). Plasma cholesterol (CHO) 24 h after injection of P-407 was 449 +/- 57 mg/dl, with the fastest rate of accumulation occurring from 1 to 12 h (approximately 16.6 mg/dl/h). Over the concentration range of 0-5 mM, P-407 did not appear significantly to affect the activity of HMG-CoA reductase in vitro. However, the enzymatic activity assayed in microsomal fractions isolated from the livers of P-407-injected rats reached a maximum of 262 +/- 42.6 pmol mevalonate/min/mg approximately 15 h after injection, with a subsequent decline to control activity (94.1 +/- 8.7 pmol/min/mg) at approximately 40 h after injection. At 48 h after injection of P-407, the activity of HMG-CoA reductase significantly (p < 0.05) decreased below control values with a mean activity of 9.4 +/- 1.2 pmol/min/mg. The CHO concentrations in hepatic tissue were significantly (p < 0.01) increased at 2 h (3.26 +/- 0.19 mg/g) and 4 h (3.75 +/- 0.38 mg/g) and significantly (p < .01) reduced at 15 h (1.56 +/- 0.19 mg/g) after injection of P-407 compared with tissue CHO concentrations determined in control animals (2.65 +/- 0.18 mg/g). However, the hepatic CHO content appeared to return to control values by 24 h (mean +/- SEM, 2.61 +/- 0.08 mg/g) after injection. These data suggest that the activity of HMG-CoA reductase is regulated by some indirect mechanism(s) after injection of P-407 in rats.

The effect of pravastatin on hepatic 3-hydroxy-3-methylglutaryl CoA reductase obtained from poloxamer 407-induced hyperlipidemic rats.

A single 300-mg intraperitoneal injection of poloxamer 407 (P-407) to rats produces a marked hypercholesterolemia for a minimum of 96 hours and increases the activity of hepatic 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase compared with the enzyme activity found in microsomal homogenates of control animals. We attempted to determine whether inhibition of microsomal HMG-CoA reductase by pravastatin sodium would yield similar values for the maximum reaction in velocity (Vmax) and the HMG-CoA reductase-pravastatin dissociation constant (Ki) when the enzyme was in the activated state compared with the control state. Knowledge of the respective values for Vmax and Ki would allow us to determine whether P-407-induced hypercholesterolemia in the rat was refractory to pravastatin treatment. Over a pravastatin concentration range of 0.5-50 nM, enzyme activity in vitro decreased as the drug's concentration increased. A standard Dixon plot of mean values of reciprocal reaction velocity versus pravastatin concentration yielded Ki of 3.7 and 4.1 nM for the control and activated states, respectively. The Vmax for conversion of HMG-CoA to mevalonate in vitro in the presence of pravastatin was 3.5-fold greater when assayed in microsomal homogenates obtained from P-407-injected rats compared with control animals. Dixon plot analysis of the data resulted in Vmax of 58.1 and 202 pmol.min-1.mg-1 for the control and activated states, respectively. These data suggest that whereas the Vmax is affected, injection of P-407 to rats does not alter the binding affinity of pravastatin for receptor(s) contained in HMG-CoA reductase as reflected by similar Ki values. This experimental animal model may be an additional screen with which to rank order the relative potency of HMG-CoA reductase inhibitors by determining the drug's effectiveness when HMG-CoA reductase is in an activated state.

Effects of nicotinic acid on poloxamer 407-induced hyperlipidemia.

We attempted to determine the mechanism(s) of poloxamer (P)-407-induced hyperlipidemia in rats by administering a lipid-lowering drug with a known mechanism of action. Five weight-matched animals were assigned to each of four treatment groups. Two groups received P-407 300 mg/ml and two received saline 1 ml. One of the P-407 and one of the saline groups were administered nicotinic acid 100 mg/kg by intraperitoneal injection at 6-96 hours after blood sampling. Blood samples were collected at 7 points from time zero to 120 hours and analyzed for triglyceride and cholesterol concentrations. The detergent produces hypertriglyceridemia (HTG) increasing from 53.4 +/- 7.0 mg/dl (time zero) to 4026.9 +/- 42.1 mg/dl by 24 hours. The HTG response was significantly attenuated by nicotinic acid (at t = 24 hrs). This, however, was followed by an average triglyceride concentration increase of 2.8-fold from 72 to 120 hours. The detergent produces a dramatic hypercholesterolemia (HCHO), increasing cholesterol from 47.5 +/- 1.8 mg/dl to 468.5 +/- 27.9 mg/dl by 48 hours. The HCHO was significantly affected by nicotinic acid administration during the accumulation phase. Nicotinic acid reduced cholesterol concentration from 364.4 +/- 16.1 mg/dl to 276.8 +/- 16.4 mg/dl at 24 hours (p < 0.05). It is a potent antilipolytic agent, limiting the free fatty acids available for the synthesis of triglyceride and cholesterol. These data suggest that P-407 may act by stimulating the release of free fatty acids from the adipocyte for at least 24 hours after injection.

Effects of pravastatin on plasma lipid concentrations in poloxamer 407-induced hyperlipidemic rats.

A new animal model of hyperlipidemia is being developed using the nonionic surfactant poloxamer 407 (P-407). We investigated the impact of pravastatin on P-407-induced hyperlipidemia. Twenty rats received P-407 300 mg intraperitoneally to induce hyperlipidemia, and 20 control rats received saline injection. Pravastatin was administered orally to an equal number of rats in both groups using three different regimens. A fourth group did not receive pravastatin. At 24 hours after injection, total cholesterol levels in two of the pravastatin groups were 28% and 34% lower than those in animals that did not receive pravastatin (p < or = 0.01). At 48 hours, triglyceride levels were significantly lower in all pravastatin groups (21-44%) versus animals not receiving pravastatin. Pravastatin diminished the effects of P-407 on lipoproteins. This new animal model may be useful in screening for investigational antihyperlipidemic agents.

Expression of lipoprotein lipase during differentiation of cultured L6 muscle cells.

The presence of lipoprotein lipase (LPL) in L6 muscle cells is equivocal. Analysis of a 21-day time course indicates that these cells express both LPL activity and mRNA. Lipase activity peaked at 4 days after plating and decreased to a nadir at day 21 after plating. Characterization of lipase activity at 4 and 19 days after plating, corresponding to myoblasts and myotubes, respectively, indicated that most of the enzyme activity had the properties of LPL, including an alkaline pH optimum, a serum requirement, and inhibition by NaCl. LPL mRNA expression peaked at 7 days after plating and fell slightly (24%) at day 21. The primary LPL mRNA species in these cells is 3.7 kb in length. Lipase activity and LPL mRNA were highly correlated during the nine course (r = +0.82), suggesting transcriptional regulation of the enzyme. These data clearly demonstrate that L6 cells express LPL during differentiation.

Mechanism of poloxamer 407-induced hypertriglyceridemia in the rat.

One 300 mg i.p. injection of the nonionic surfactant poloxamer 407 (Pluronic F-127) produces a significant increase above control of both circulating cholesterol and triglyceride (TG) concentrations. The present study was conducted to determine the effect of poloxamer 407 (P-407) on the capacity to hydrolyze circulating TG by lipoprotein lipase (LPL) in an attempt to determine the mechanism of action of P-407. The concentration of TG in the rat following a single 300 mg i.p. injection of P-407 was marked, increasing from 84 +/- 10 to 3175 +/- 322 mg/dL at 24 hr. The maximal rate of TG accumulation (5.74 mg/dL/min) in the plasma of P-407-injected rats occurred between 2 and 4 hr post-injection. In vitro incubation of LPL with P-407 significantly inhibited enzyme activity with an inhibitory concentration at which 50% of the enzymatic activity was lost of approximately 24 microM. Concentrations of P-407 exceeding 350 microM in vitro completely inhibited LPL activity. The effects of P-407 on the enzymatic activity of LPL in post-heparin plasma obtained following a single 300 mg dose of P-407 to rats demonstrated greater than 95% suppression of LPL activity 3 hr post-injection compared with controls. Inhibition of LPL activity was greater than 90% as long as 24 hr following a single i.p. injection of P-407. However, while the heparin-releasable fraction of capillary-bound LPL was inhibited in the plasma, LPL activity significantly increased in cardiac and skeletal muscle in poloxamer-injected animals compared with sham-injected controls. Although there was no significant change in LPL activity in adipose tissue, testes, and lung resulting from P-407 treatment, LPL activity increased by 37% in myocardium, 69% in soleus, and 66% in gastrocnemius muscle in P-407-injected rats when compared with controls. Our studies would suggest that the predominant mechanism by which P-407 induced an increase in circulating TG was by a reduction in the rate at which TG was hydrolyzed due to inhibition of heparin-releasable LPL by the surfactant.

Regulation of lipoprotein lipase in muscle and adipose tissue during exercise.

Lipoprotein lipase (LPL) is regulated in a tissue-specific manner; exercise increases LPL activity in muscle at the same time it is reduced in adipose tissue. The purpose of this study was to determine the relationship between LPL activity and LPL mRNA in muscle and adipose tissue in rats exposed to one bout of exercise. Immediately after a 2-h swim, LPL activity [pmol free fatty acids (FFA).min-1.mg tissue-1] in the exercised animals was reduced 43% in adipose tissue (110 +/- 26 to 63 +/- 17) and increased almost twofold in the soleus muscle (203 +/- 26 to 383 +/- 59) compared with sedentary control animals. At the same time, LPL mRNA was reduced 42% in adipose tissue and increased 50 and 100% in the red vastus and white vastus muscles, respectively. Twenty-four hours after the swim, LPL activity had returned to control levels in adipose tissue and the soleus muscle. At hour 24 of recovery, LPL mRNA was still reduced 23% in the adipose tissue of exercised animals but was not significantly different between exercised and control animals in any of the muscle tissues analyzed. Changes in total RNA concentration could not account for the changes in relative LPL mRNA expression. The relationship between LPL enzyme activity and LPL mRNA in muscle and adipose tissue was +0.86 and +0.93 at 0 and 24 h postexercise, respectively. Thus the tissue-specific changes in enzyme activity induced by exercise could be mediated, in part, through pretranslational control.

Regulation of lipoprotein lipase in adipose and muscle tissues during fasting.

The purpose of this work was to determine the relationship between lipoprotein lipase (LPL) activity and LPL mRNA in muscle and adipose tissue in fed and fasted rats. In control animals, the correlation between enzyme activity and LPL mRNA for adipose tissue, heart, soleus, fast red vastus lateralis, and fast white vastus lateralis muscle was r = +0.97. Twenty-four hours of fasting increased LPL activity 38% in heart, reduced it 59% in adipose tissue, and had no effect on activity in the three skeletal muscles analyzed. At the same time, relative LPL mRNA concentrations were reduced 25% in adipose tissue and elevated in heart, soleus, red vastus, and white vastus muscles when compared with control concentrations. Prolonging the fast to 6 days was accompanied by a 64% reduction in adipose tissue LPL activity and an increase in the activities of slow-twitch soleus (83%) and fast-twitch red vastus lateralis muscles (193%), with no enzyme activity change in heart or white vastus lateralis muscle compared with values obtained from control fed animals. LPL mRNA concentration was reduced 66% in adipose tissue, increased more than twofold in heart, soleus, and white vastus muscle, and increased threefold in red vastus muscle. Changes in relative LPL mRNA concentration in adipose tissue induced by fasting could, in part, be accounted for by the increases seen in total RNA concentration. The relationships between enzyme activity and LPL mRNA in muscle and adipose tissue were r = 0.97 and 0.77 for 1-day and 6-day fasted animals, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of glucose and insulin on triacylglycerol metabolism in isolated normal and diabetic skeletal muscle.

The effects of insulin and glucose on triacylglycerol (TG) metabolism in normal and diabetic isolated skeletal muscle were investigated in this study. Intracellular TG was continuously synthesized and hydrolyzed in both normal and diabetic skeletal muscle. In the absence of insulin and glucose, normal and diabetic skeletal muscle TG content and synthesis were decreased. In contrast, in the presence of insulin and glucose, the normal and diabetic TG contents were unchanged and triacylglycerol synthesis was increased as compared with the respective control values. However, insulin and glucose increased intramuscular TG content to a greater extent than could be accounted for by their stimulation of TG synthesis, indicating that insulin and glucose appear to inhibit TG hydrolysis in diabetic muscle, as well as in normal muscle. In addition, these data suggest that diabetes causes a defect in the ability of insulin and glucose to stimulate TG synthesis, as the increase in diabetic muscle TG synthesis in the presence of insulin and glucose was less than in normal muscle.

Characterization of serum-stimulated lipoprotein lipase from bovine heart.

1. A triglyceride (TG) lipase is present in whole homogenate and tissue extracts of beef myocardium with characteristics of lipoprotein lipase (LPL); i.e., activity is stimulated by serum, inhibited by NaCl and protamine sulfate, the protein binds to heparin-Sepharose, and the enzyme has an alkaline pH optimum. 2. This TG lipase, eluted from heparin-Sepharose at 0.9-1.0 M NaCl, has an apparent mol. wt of 64 K daltons. Its primary mRNA is 3.7 kb. 3. Expression of LPL mRNA and enzyme activities are in the ratio of approximately 20:8:1 for hearts of mouse, rat and beef, respectively and correlate with r = +0.99.

Lipase regulation of muscle triglyceride hydrolysis.

The cellular control of intramuscular triglyceride (TG) metabolism involves two major identified lipases: hormone-sensitive lipase (HSL) and lipoprotein lipase (LPL). Recently, the presence of HSL in muscle has been unequivocally demonstrated. However, although it is thought that HSL is responsible for intramuscular TG lipolysis, direct evidence for this is lacking. There is evidence to suggest that HSL and LPL are simultaneously activated under a variety of conditions. The two muscle lipases appear to be turned on by the same signal and function as a coordinated unit in meeting the energy demands of muscle. At a time when HSL is presumably hydrolyzing endogenous TG, LPL is sent to the capillary beds in search of substrate. TG uptake from circulation is highly related to muscle LPL activity. Exercise training increases LPL activity in plasma and in parenchymal cells in muscle. These results suggest that training may increase the capacity to clear TG from circulation and that LPL might have a role in replenishing muscle TG stores that have been decreased with exercise.

Electrical stimulation alters fatty acid metabolism in isolated skeletal muscle.

Little is known about the contribution of plasma free fatty acid (FFA) and intramuscular triacylglycerol (TG) as substrates for energy production during prolonged electrical stimulation of skeletal muscle. The purpose of this study was to investigate the effects of continuous and intermittent electrical stimulation protocols of different intensities on exogenous FFA oxidation, exogenous FFA incorporation into intracellular TG, and intracellular TG content in the isolated in vitro rat flexor digitorum brevis muscle preparation. Muscles were electrically stimulated for 0.5 h continuously at 0.2 Hz or intermittently (30 s on, 60 s off) at 0.2, 0.4, 0.8, and 5.0 Hz while incubated at 37 degrees C in 0.5 mM palmitate-3% bovine serum albumin medium (pH 7.4) in the presence of insulin (100 microU/ml) and glucose (11 mM). Control muscles were frozen immediately after excision or incubated for 0.5 h. At similar frequencies, less exogenous FFA esterification and more exogenous FFA oxidation occurred during continuous than during intermittent stimulation. As the frequency of intermittent stimulation increased, the amount of exogenous FFA esterified decreased and the amount of exogenous FFA oxidized increased. The data also indicate that at least a portion of TG was constantly being hydrolyzed during electrical stimulation. Under stimulation conditions in which exogenous FFA esterification was below the control (resting muscle) level, intramuscular TG content was significantly decreased compared with control TG content values. Thus both plasma FFA and intramuscular TG are substrates for energy production during electrical stimulation. However, the stimulation parameters employed affect the quantities utilized.

Dibutyryl cAMP-induced increases in triacylglycerol lipase activity in developing L8 myotube cultures.

Triacylglycerol (TG) lipase activity, with an alkaline pH optimum, has been identified in the cellular fraction of L8 myotube cultures. This TG lipase activity was stimulated by serum and inhibited by NaCl and protamine sulfate. These characteristics have been classically described for lipoprotein lipase. It was possible to increase the activity of this TG lipase three- to five-fold by incubating the cells with dibutyryl cAMP. Maximal enzyme activity was observed 16 h following the addition of 10-100 microM dibutyryl cAMP to the cultured cells. Enzyme activity returned to control levels 24 h after removal of the nucleotide from the culture medium. Serum-sensitive alkaline TG lipase activity was also identified in five other myotube preparations of cultured muscle cells. The highest levels of activity were found in rat skeletal muscle primary, H9, and L6 cell types. The finding that dibutyryl cAMP is an effective inducer of alkaline TG lipase activity provides us with a valuable model to investigate mechanisms regulating synthesis, compartmentalization, and transport of lipoprotein lipase in muscle.

Triacylglycerol metabolism in rat skeletal muscle after exercise.

The temporal relationships between triacylglycerol (TG) content and TG lipase activity in slow-twitch (STR) and fast-twitch red (FTR) muscles were determined in rats during recovery from a 2-h swim. Immediately after the exercise, plasma free fatty acid (FFA) was elevated and glycogen concentrations were decreased. TG content was decreased 40% in STR muscle and reduced 45% in FTR muscle. The TG concentration of STR muscle increased in a linear fashion throughout recovery so that control levels were reached within the first 24 h after exercise. TG lipase activity of STR muscle was elevated 36% above control immediately after the swim and continued to increase to 84% above control 24 h after the work. In STR muscle there was a net synthesis of TG, while lipase activity was elevated above that measured in muscle of control rats. TG content of FTR muscle remained 45% below control throughout the first 24 h of recovery, and TG lipase activity increased from 26% (P greater than 0.05) greater than control immediately after exercise to threefold above control 24 h after work. All parameters returned to control levels by 48 h of recovery. These data indicated that a net TG synthesis occurs in STR muscle when lipolytic activity is elevated. In FTR muscle, however, a gradual increase in TG lipase activity that occurs during the first 24 h of recovery accompanies a TG concentration well below the control level throughout this same time frame.(ABSTRACT TRUNCATED AT 250 WORDS)

Hepatic lipid metabolism in exercise and training.

The liver plays a central role in the metabolism of fat. The available data, though sometimes controversial, clearly indicate that muscular exercise affects almost every aspect of fat metabolism in this organ. Neither acute exercise nor training affects total lipid, phospholipid, or cholesterol concentrations in the liver of rats fed chow or low fat diets. However, exercise training reduces accumulation of total hepatic fat and cholesterol in rats fed a fat-rich diet. In addition, training seems to increase both the synthesis and catabolism of cholesterol in the liver in rats fed a chow diet. Production of ketones by the liver increases both during prolonged exercise and during recovery from exercise. Acute prolonged exercise reduces the activities of the enzymes involved in the synthesis of fatty acids and increases oxidation of fatty acids by the liver. This type of work also increases the esterification of fatty acids with the subsequent accumulation of triacylglycerols in this organ. Training does not affect triacylglycerol concentration in the liver of rats fed a chow diet but attenuates its accumulation after a fat-rich diet. Training reduces the postheparin plasma hepatic lipase activity. Finally, it reduces production of triacylglycerols and increases production of high density lipoprotein cholesterol by the liver. A large body of descriptive information has been published indicating that exercise has a dramatic effect upon hepatic lipid metabolism. The next step in this work is the identification of the molecular mechanisms responsible for these exercise-induced alterations.