Hormone-sensitive Lipase Activity Research Articles

Growth Associated Hormones Response and Fat Metabolism Change in Finishing Pigs Fed with n-Methyl-d, L-Aspartate

A trial was conducted to investigate the effect of dietary NMA on several growth associated hormones and fat metabolism in finishing pigs. A total of 84 crossbred finishing pigs (average initial BW of 56±0.37 kg) were divided into 6 pens, 14 pigs per pen (7 gilts and 7 barrows per pen). 3 pens of pigs were fed with control diet (corn-soybean meal) and the others were fed control diet addition with 50 mg/kg NMA. During the trial, all pigs were given free access to feed and water. After 44 days trial, 8 pigs from each treatment (4 gilts and 4 barrows, weight similar to average group weight, 86.94±0.71 kg for control group, and 90.55±1.51 kg for NMA treated group) were sacrificed to collect the sample of the liver, longissimus muscle, subcutaneous fat (10th rib). The addition of NMA in diet increased the IGF-I, Insulin, T3, T4 levels in serum by 50.68% (p<0.05), 38.36% (p<0.05), 123.33% (p<0.01), 60.58% (p<0.03), respectively. Meanwhile, IGF-I level in the liver and the muscle were increased with 17.83% (p<0.03) and 26.00% (p<0.03) with addition of NMA. The data from subcutaneous fat (10th rib) analysis showed that supplement of 50 mg/kg NMA decreased the total activities of malic dehydrogenase (MDH) by 20.54% (p<0.05), glucose-6- phosphate dehydrogenase (G-6-DPH) by 16.97% (p<0.05), and decreased the specific activities of MDH and G-6-DPH by 37.46% (p<0.01) and 35.06% (p<0.01), respectively. The hormone sensitive lipase (HSL) total activity was increased by 25.00% (p<0.05) in NMA treated pigs. These results indicated that addition of 50 mg/kg NMA to diet can induce the endocrine great change in finishing pigs, furthermore, inhibit the fat synthesis through suppressing lipogenic enzymes and promote the fat degradation by elevating HSL activity in finishing pigs. (Asian-Aust. J. Anim. Sci. 2002. Vol 15, No. 7 : 1026-1030)

Characterization of the Functional Interaction of Adipocyte Lipid-binding Protein with Hormone-sensitive Lipase

Hormone-sensitive lipase (HSL) is an intracellular lipase that plays an important role in the hydrolysis of triacylglycerol in adipose tissue. HSL has been shown to interact with adipocyte lipid-binding protein (ALBP), a member of the family of intracellular lipid-binding proteins that bind fatty acids and other hydrophobic ligands. The current studies have addressed the functional significance of the association and mapped the site of interaction between HSL and ALBP. Incubation of homogeneous ALBP with purified, recombinant HSL in vitro resulted in a 2-fold increase in substrate hydrolysis. Moreover, the ability of oleate to inhibit HSL hydrolytic activity was attenuated by co-incubation with ALBP. Co-transfection of Chinese hamster ovary cells with HSL and ALBP resulted in greater hydrolytic activity than transfection of cells with HSL and vector alone. Deletional mutations of HSL localized the region of HSL that interacts with ALBP to amino acids 192-200, and site-directed mutagenesis of individual amino acids in this region identified His-194 and Glu-199 as critical for mediating the interaction of HSL with ALBP. Interestingly, HSL mutants H194L and E199A, each of which retained normal basal hydrolytic activity, failed to display an increase in hydrolytic activity when co-transfected with wild type ALBP. Therefore, ALBP increases the hydrolytic activity of HSL through its ability to bind and sequester fatty acids and via specific protein-protein interaction. Thus, HSL and ALBP constitute a functionally important lipolytic complex.

The effect of exercise training on hormone-sensitive lipase in rat intra-abdominal adipose tissue and muscle.

1. Adrenaline-stimulated lipolysis in adipose tissue may increase with training. The rate-limiting step in adipose tissue lipolysis is catalysed by the enzyme hormone-sensitive lipase (HSL). We studied the effect of exercise training on the activity of the total and the activated form of HSL, referred to as HSL (DG) and HSL (TG), respectively, and on the concentration of HSL protein in retroperitoneal (RE) and mesenteric (ME) adipose tissue, and in the extensor digitorum longus (EDL) and soleus muscles in rats. 2. Rats (weighing 96 +/- 1 g, mean +/- S.E.M.) were either swim trained (T, 18 weeks, n = 12) or sedentary (S, n = 12). Then RE and ME adipose tissue and the EDL and soleus muscles were incubated for 20 min with 4.4 microM adrenaline. 3. HSL enzyme activities in adipose tissue were higher in T compared with S rats. Furthermore, in RE adipose tissue, training also doubled HSL protein concentration (P < 0.05). In ME adipose tissue, the HSL protein levels did not differ significantly between T and S rats. In muscle, HSL (TG) activity as well as HSL (TG)/HSL (DG) were lower in T rats, whereas HSL (DG) activity did not differ between groups. Furthermore, HSL protein concentration in muscle did not differ between T and S rats (P > 0.05). 4. In conclusion, training increased the amount of HSL and the sensitivity of HSL to stimulation by adrenaline in intra-abdominal adipose tissue, the extent of the change differing between anatomical locations. In contrast, in skeletal muscle the amount of HSL was unchanged and its sensitivity to stimulation by adrenaline reduced after training.

Stimulation of Lipolysis and Hormone-sensitive Lipase via the Extracellular Signal-regulated Kinase Pathway

Hormonally stimulated lipolysis occurs by activation of cyclic AMP-dependent protein kinase (PKA) which phosphorylates hormone-sensitive lipase (HSL) and increases adipocyte lipolysis. Evidence suggests that catecholamines not only can activate PKA, but also the mitogen-activated protein kinase pathway and extracellular signal-regulated kinase (ERK). We now demonstrate that two different inhibitors of MEK, the upstream activator of ERK, block catecholamine- and beta(3)-stimulated lipolysis by approximately 30%. Furthermore, treatment of adipocytes with dioctanoylglycerol, which activates ERK, increases lipolysis, although MEK inhibitors decrease dioctanoylglycerol-stimulated activation of lipolysis. Using a tamoxifen regulatable Raf system expressed in 3T3-L1 preadipocytes, exposure to tamoxifen causes a 14-fold activation of ERK within 15-30 min and results in approximately 2-fold increase in HSL activity. In addition, when differentiated 3T3-L1 cells expressing the regulatable Raf were exposed to tamoxifen, a 2-fold increase in lipolysis is observed. HSL is a substrate of activated ERK and site-directed mutagenesis of putative ERK consensus phosphorylation sites in HSL identified Ser(600) as the site phosphorylated by active ERK. When S600A HSL was expressed in 3T3-L1 cells expressing the regulatable Raf, tamoxifen treatment fails to increase its activity. Thus, activation of the ERK pathway appears to be able to regulate adipocyte lipolysis by phosphorylating HSL on Ser(600) and increasing the activity of HSL.

The expression of hormone-sensitive lipase in clonal beta-cells and rat islets is induced by long-term exposure to high glucose.

Hormone-sensitive lipase (HSL) is expressed and enzymatically active in beta-cells and has been proposed to be involved in the generation of the lipid-derived signal that seems to be necessary for glucose-stimulated insulin secretion. In this study, we investigated whether the expression of HSL in INS-1 cells and in rat islets is affected by exposure to high glucose concentrations. Incubation of INS-1 cells in 25 mmol/l glucose for 16 and 32 h induced HSL protein expression twofold, whereas no effect was observed after 4 and 8 h of incubation. The HSL activity, defined as the diglyceride lipase activity inhibited by anti-rat HSL antibodies, constituted approximately 25% of total diglyceride lipase activity and was induced to a similar extent as HSL protein levels. The glucose effect at 16 h on HSL protein expression level was confirmed in freshly isolated rat islets. Exposure of INS-1 cells to different glucose concentrations for 16 h showed that the inductive effect on HSL protein levels was maximum at 20 mmol/l glucose (2- to 2.5-fold). Northern blot analysis demonstrated a more than threefold elevation of HSL mRNA levels. The induction was blocked by actinomycin D, and the half-life of the transcript seemed to be unchanged by high glucose, suggesting a transcriptional nature of the glucose effect on HSL gene expression. The nonmetabolizable glucose analog 2-deoxyglucose, which has no mitogenic effect, induced HSL approximately 1.3-fold, whereas mannose was similar to glucose, stimulating HSL expression 1.7- to 2-fold. The results suggest that HSL is involved in the beta-cell responses to hyperglycemia and also in generating the lipid signal that is needed in stimulus-secretion coupling.

Absence of cardiac lipid accumulation in transgenic mice with heart-specific HSL overexpression

Hormone-sensitive lipase (HSL) hydrolyzes triglyceride (TG) in adipose tissue. HSL is also expressed in heart. To explore the actions of cardiac HSL, heart-specific, tetracycline (Tc)-controlled HSL-overexpressing mice were generated. Tc-responsive element-HSL transgenic (Tg) mice were generated and crossed with myosin heavy chain (MHC)alpha-tTA Tg mice, which express the Tc-responsive transactivator (tTA) in the heart. The double-Tg mice (MHC-HSL) were maintained with doxycycline (Dox) to suppress Tg HSL. Upon removal of Dox, cardiac HSL activity and protein increased 12- and 8-fold, respectively, and the expression was heart specific. Although cardiac TG content increased twofold in control mice after an overnight fast, it did not increase in HSL-induced mice. Electron microscopy showed numerous lipid droplets in the myocardium of fasted control mice, whereas fasted HSL-induced mice showed virtually no droplets. Microarray analysis showed altered expression of cardiac genes for fatty acid oxidation, transcription factors, signaling molecules, cytoskeletal proteins, and histocompatibility antigens in HSL-induced mice. Thus cardiac HSL plays a role in controlling accumulation of triglyceride droplets and can affect the expression of a number of cardiac genes.

Effects of acute and chronic ethanol administration on the response of mouse adipose tissue hormone-sensitive lipase to alpha(2)-adrenoceptor activation bu UK 14304.

Untreated (control) obese CBA mice had lower hormone-sensitive lipase (HSL) activity and cAMP levels in brown adipose tissue than normal lean mice, but white adipose tissue HSL activity and cAMP were similar in obese and lean mice. In the obese mice, chronic ethanol treatment increased HSL activity and cAMP levels in both brown and white adipose tissue to above the levels in lean mice. In the lean mice, chronic ethanol only stimulated white adipose tissue. UK 14304 [5-bromo-6-(2-imidazolin-2-ylamino)-quinoxaline: 2 mg/kg] inhibited HSL activity in both brown and white adipose tissues in lean mice, but a higher dose (3 mg/kg) was required to produce the same inhibition in obese mice. After chronic ethanol adipose tissues were more sensitive to UK 14304; only half the dose being required to produce the same level of lipase inhibition. We propose that, although chronic ethanol consumption increases cAMP levels in adipose tissue, particularly in obese mice, it also sensitizes the tissues to alpha(2)-adrenoceptor stimulation. These effects may explain the increased sympathetic nervous system activity observed in alcohol withdrawal.

Insulin Mediated Inhibition of Hormone Sensitive Lipase Activity in Vivo in Relation to Endogenous Catecholamines in Healthy Subjects

Insulin Mediated Inhibition of Hormone Sensitive Lipase Activity in Vivo in Relation to Endogenous Catecholamines in Healthy Subjects

Expression, activity, and subcellular localization of testicular hormone-sensitive lipase during postnatal development in the guinea pig.

The present work reports on testicular hormone-sensitive lipase (HSL), the biological significance of which has been documented in male fertility. The HSL protein levels and enzymatic activity were measured, respectively, by densitometry of immunoreactive bands in Western blots, performed with antibodies against recombinant rat HSL, and by spectrophotometry in seminiferous tubules (STf) and interstitial tissue (ITf) enriched fractions generated from neonatal, pubertal, and adult guinea pig testes. In addition, HSL was studied in subcellular fractions obtained from STf isolated from adult testes and in epididymal spermatozoa (Spz). A 104-kDa HSL protein was detected in STf and ITf, the expression and activity of which increased with testicular development. Three immunoreactive bands of 104, 110, and 120 kDa were detected in the lysosomal subfraction, and two bands of 104 and 120 kDa were detected in Spz. The HSL activity was positively correlated with free (FC) and esterified (EC) cholesterol ratios in STf and ITf, but not with triglyceride (TG) levels, during testicular development. Immunolabeling localized HSL to elongated spermatids and Sertoli cells, where its distribution was stage-dependent, and within the cells lining the excurrent ducts of the testis. The findings of the 104- and 120-kDa HSL immunoreactive bands and of HSL activity in Spz as well, as the detection of the 104-, 110-, and 120-kDa immunoreactive bands in lysosomes, suggest that part of HSL may originate from germ cells and be imported in Sertoli cells. The HSL protein levels and enzymatic activity in ITf and STf were positively correlated with serum testosterone levels during development. To the best of our knowledge, this study is the first to contribute insights regarding the impact of HSL on FC:EC cholesterol ratios and TG levels in the interstitial tissue and tubules in relation to serum testosterone levels during postnatal development, and regarding the immunolocalization of the enzyme in regions of the male gamete consistent with spermatozoa-oocyte interaction.

Insulin and leptin acutely regulate cholesterol ester metabolism in macrophages by novel signaling pathways.

Leptin is produced in adipose tissue and acts in the hypothalamus to regulate food intake. However, recent evidence also indicates a potential for direct roles for leptin in peripheral tissues, including those of the immune system. In this study, we provide direct evidence that macrophages are a target tissue for leptin. We found that J774.2 macrophages express the functional long form of the leptin receptor (ObRb) and that this becomes tyrosine-phosphorylated after stimulation with low doses of leptin. Leptin also stimulates both phosphoinositide 3-kinase (PI 3-kinase) activity and tyrosine phosphorylation of JAK2 and STAT3 in these cells. We investigated the effects of leptin on hormone-sensitive lipase (HSL), which acts as a neutral cholesterol esterase in macrophages and is a rate-limiting step in cholesterol ester breakdown. Leptin significantly increased HSL activity in J774.2 macrophages, and these effects were additive with the effects of cAMP and were blocked by PI 3-kinase inhibitors. Conversely, insulin inhibited HSL in macrophages, but unlike adipocytes, this effect did not require PI 3-kinase. These results indicate that leptin and insulin regulate cholesterol-ester homeostasis in macrophages and, therefore, defects in this process caused by leptin and/or insulin resistance could contribute to the increased incidence of atherosclerosis found associated with obesity and type 2 diabetes.

Structure-function relationships of hormone-sensitive lipase.

Research into the structure-function relationships of lipases and esterases has increased significantly during the past decade. Of particular importance has been the deduction of several crystal structures, providing a new basis for understanding these enzymes. The generated insights have, together with cloning and expression, aided studies on structure-function relationships of hormone-sensitive lipase (HSL). Novel phosphorylation sites have been identified in HSL, which are probably important for activation of HSL and lipolysis. Functional and structural analyses have revealed features in HSL common to lipases and esterases. In particular, the catalytic core with a catalytic triad has been unveiled. Furthermore, the investigations have given clear suggestions with regard to the identity of functional and structural domains of HSL. In the present paper, these studies on HSL structure-function relationships and short-term regulation are reviewed, and the results presented in relation to other discoveries in regulated lipolysis.

Decreased lipolysis and enhanced glycerol and glucose utilization by adipose tissue prior to development of obesity in monosodium glutamate (MSG) treated-rats.

To determine the metabolic alterations that lead to the neonatal administration of monosodium glutamate (MSG), which results in arrested growth and obesity. ANIMALS AND DESIGN: Wistar rats were injected 5 times, every other day, with 4 g of MSG/kg b.w. or with hyperosmotic saline (controls), within the first 10 days of life, and were studied at the age of 30 days. Body weight was lower, whereas adipocyte lipid content, cell diameter, surface area and volume were higher in MSG rats than in controls. Plasma glucose, insulin, NEFA, glycerol and triglyceride levels, and in vitro production of NEFA by lumbar fat pad pieces incubated under basal conditions or in the presence of epinephrine and epinephrine plus glucose in the media were lower in MSG than in control rats. In the same fat pad pieces, the conversion of 1-14C-glycerol into fatty acids was always enhanced and its conversion into glyceride glycerol was enhanced when incubations were carried out in the presence of epinephrine or glucose. Both the hormone sensitive lipase activity and mRNA expression were lower in adipose tissue from MSG rats. Besides, the number of insulin receptors, lipid synthesis from U14C glucose, 3H-2-deoxy D-glucose uptake and cellular GLUT4 translocation index were higher in adipocytes from MSG rats than from the controls. It is proposed that an enhanced insulin sensitivity in 1 month old MSG rats is responsible for the decreased lipolytic activity and enhanced glucose uptake. In addition, the enhanced lipogenesis and glycerol reutilization seen in their adipose tissue, disturbs the normal balance between fat depots breakdown and accumulation in favor of the latter.

Relationships between lipolysis induced by various lipolytic agents and hormone-sensitive lipase in rat fat cells

Norepinephrine induced lipolysis in rat fat cells, in vitro, in a time- and concentration-dependent manner, without concomitantly increasing hormone-sensitive lipase (HSL) activity. It also induced, time and concentration dependently, HSL translocation from the cytosol to the lipid droplets in fat cells. Isoproterenol, forskolin, dibutyryl cyclic AMP, and theophylline also induced lipolysis in fat cells, but did not stimulate HSL activity. These agents also induced HSL translocation from the cytosol to the lipid droplets in fat cells: about 80% to 90% of all HSL was located in lipid droplets after incubation for 1 h. These results suggest that the critical event in lipolytic activation of fat cells induced by lipolytic agents is not an increase in the catalytic activity of HSL but translocation of HSL to its substrate on the surfaces of lipid droplets in fat cells.—Morimoto, C., K. Kameda, T. Tsujita, and H. Okuda. Relationships between lipolysis induced by various lipolytic agents and hormone-sensitive lipase in rat fat cells. J. Lipid Res. 2001. 42: 120–127.

Reduced hormone-sensitive lipase activity is not a major metabolic defect in Finnish FCHL families

The pathogenetic mechanisms behind familial combined hyperlipidemia (FCHL) are unknown. However, exaggerated postprandial lipemia and excessive serum free fatty acid (FFA) concentrations have drawn attention to altered lipid storage and lipolysis in peripheral adipose tissue. Hormone-sensitive lipase (HSL) is the enzyme responsible for intracellular lipolysis in adipocytes and a decrease of adipocyte HSL activity has been demonstrated in Swedish FCHL subjects. The aim of the study was to investigate if adipose tissue HSL activity had any effect on lipid phenotype and if low HSL activity and FCHL were linked in Finnish FCHL families. A total of 48 family members from 13 well-characterized Finnish FCHL families and 12 unrelated spouses participated in the study. FCHL patients with different lipid phenotypes (IIA, IIB, IV) did not differ in adipose tissue HSL activity from each other or from the 12 normolipidemic spouses ( P=0.752). In parametric linkage analysis using an affecteds-only strategy the low adipose tissue HSL activity was not significantly linked with FCHL phenotype. However, we found a significant sibling–sibling correlation for the HSL trait (0.51, P<0.01 ). Thus, a modifying or interacting role of HSL in the pathogenesis of FCHL could not be excluded.

Absence of perilipin results in leanness and reverses obesity in Lepr(db/db) mice.

Obesity is a disorder of energy balance. Hormone-sensitive lipase (HSL) mediates the hydrolysis of triacylglycerol, the major form of stored energy in the body. Perilipin (encoded by the gene Plin), an adipocyte protein, has been postulated to modulate HSL activity. We show here that targeted disruption of Plin results in healthy mice that have constitutively activated fat-cell HSL. Plin -/- mice consume more food than control mice, but have normal body weight. They are much leaner and more muscular than controls, have 62% smaller white adipocytes, show elevated basal lipolysis that is resistant to beta-adrenergic agonist stimulation, and are cold-sensitive except when fed. They are also resistant to diet-induced obesity. Breeding the Plin -/- alleles into Leprdb/db mice reverses the obesity by ncreasing the metabolic rate of the mice. Our results demonstrate a role for perilipin in reining in basal HSL activity and regulating lipolysis and energy balance; thus, agents that inactivate perilipin may prove useful as anti-obesity medications.

Changes in adipose tissue hormone-sensitive lipase activity and cAMP during ethanol withdrawal

The time course of the effects of ethanol withdrawal on brown and white adipose tissue hormone-sensitive lipase, cAMP production, and phosphodiesterase have been investigated after chronic drinking or liquid diet schedules. Chronic drinking significantly reduced brown adipose tissue hormone-sensitive lipase activity and cAMP levels from control. During withdrawal, there was a rebound increase to 200% control, peaking 9 h into withdrawal. White adipose tissue hormone-sensitive lipase activity and cAMP accumulation were significantly raised by both treatment schedules. Ethanol liquid diet produced a significant fall in adipose tissue hormone-sensitive lipase activity and cAMP accumulation. In brown fat, there was a rebound increase in hormone-sensitive lipase activity and cAMP; in white fat, no rebound was observed. In brown fat, the reductions in hormone-sensitive lipase activity and cAMP accumulation after chronic drinking coincided with an increase in phosphodiesterase activity. In white fat, the rise in cAMP and hormone-sensitive lipase activation coincided with a decrease in phosphodiesterase activity. We conclude that the effects of chronic ethanol on hormone-sensitive lipase activity are cAMP-dependent and mediated via alterations in phosphodiesterase activity.

Adrenaline and glycogenolysis in skeletal muscle during exercise: a study in adrenalectomised humans.

The role of adrenaline in regulating muscle glycogenolysis and hormone-sensitive lipase (HSL) activity during exercise was examined in six adrenaline-deficient bilaterally adrenalectomised, adrenocortico-hormonal-substituted humans (Adr) and in six healthy control individuals (Con). Subjects cycled for 45 min at approximately 70% maximal pulmonary O2 uptake (VO2,max) followed by 15 min at approximately 86% VO2,max either without (-Adr and Con) or with (+Adr) adrenaline infusion that elevated plasma adrenaline levels (45 min, 4.49+/-0.69 nmol l(-1); 60 min, 12.41+/-1.80 nmol l(-1)). Muscle samples were obtained at 0, 45 and 60 min of exercise. In -Adr and Con, muscle glycogen was similar at rest (-Adr, 409+/-19 mmol (kg dry wt)(-1); Con, 453+/-24 mmol (kg dry wt)(-1)) and following exercise (-Adr, 237+/-52 mmol (kg dry wt)(-1); Con, 227+/-50 mmol (kg dry wt)(-1)). Muscle lactate, glucose-6-phosphate and glucose were similar in -Adr and Con, whereas glycogen phosphorylase (a/a + b x 100 %) and HSL (% phosphorylated) activities increased during exercise in Con only. Adrenaline infusion increased activities of phosphorylase and HSL as well as blood lactate concentrations compared with those in -Adr, but did not enhance glycogen breakdown (+Adr, glycogen following exercise: 274+/-55 mmol (kg dry wt)(-1)) in contracting muscle. The present findings demonstrate that during exercise muscle glycogenolysis can occur in the absence of adrenaline, and that adrenaline does not enhance muscle glycogenolysis in exercising adrenalectomised subjects. Although adrenaline increases the glycogen phosphorylase activity it is not essential for glycogen breakdown in contracting muscle. Finally, a novel finding is that the activity of HSL in human muscle is increased in exercising man and this is due, at least partly, to stimulation by adrenaline.

Stimulation of hormone-sensitive lipase activity by contractions in rat skeletal muscle

Because the enzymic regulation of muscle triglyceride breakdown is poorly understood we studied whether neutral lipase in skeletal muscle is activated by contractions. Incubated soleus muscles from 70 g rats were electrically stimulated for 60min. Neutral lipase activity against triacylglycerol increased after 1 and 5min of contractions [0.36±0.02 (basal) versus 0.49±0.05 (1min) and 0.54±0.05 (5min) m-unit·mg of protein-1, means±S.E.M., P &amp;lt; 0.05]. After 10min the neutral lipase activity (0.40±0.05m-unit·mg of protein-1) had decreased to basal values (P &amp;gt; 0.05). The contraction-mediated increase in lipase activity was increased by ≈ 110% when muscle was stimulated in the presence of okadaic acid. Conversely, treatment of muscle homogenate with alkaline phosphatase completely reversed the contraction-mediated lipase activation. Lipase activity did not change during contractions when analysed in the presence of anti-hormone-sensitive-lipase (HSL) antibody [0.17±0.02 (basal) versus 0.21±0.02 (5min) m-unit·mg of protein-1, P &amp;gt; 0.05]. Furthermore, immunoprecipitation with affinity-purified anti-HSL antibody reduced muscle-HSL protein concentration by 81±4% and caused similar reductions in lipase activity against triacylglycerol and in the contraction-induced increase in this activity. Neither prior sympathectomy [0.33±0.02 (basal) versus 0.53±0.06 (5min) m-unit·mg of protein-1, P &amp;lt; 0.05] nor propranolol impaired the lipase response to contractions. Glycogen phosphorylase activity in the absence of AMP increased after 1min [27.3±3.1 versus 8.9±1.8% (activity without AMP/total activity with AMP), P &amp;lt; 0.05] and returned to basal levels after 5min. In conclusion, skeletal-muscle-immunoreactive HSL is transiently stimulated by contractions and the mechanism probably involves phosphorylation. The time course of HSL activation is similar to that of glycogen phosphorylase. Apparently, the two enzymes are regulated in parallel by contraction-induced as well as hormonal mechanisms, allowing simultaneous recruitment of all major extra- and intra-muscular energy stores.

Stimulation of hormone-sensitive lipase activity by contractions in rat skeletal muscle.

Because the enzymic regulation of muscle triglyceride breakdown is poorly understood we studied whether neutral lipase in skeletal muscle is activated by contractions. Incubated soleus muscles from 70 g rats were electrically stimulated for 60 min. Neutral lipase activity against triacylglycerol increased after 1 and 5 min of contractions [0.36 +/- 0.02 (basal) versus 0.49 +/- 0.05 (1 min) and 0.54 +/- 0.05 (5 min) m-unit.mg of protein(-1), means +/- S.E.M., P < 0.05]. After 10 min the neutral lipase activity (0.40 +/- 0.05 m-unit.mg of protein(-1)) had decreased to basal values (P > 0.05). The contraction-mediated increase in lipase activity was increased by approximately 110% when muscle was stimulated in the presence of okadaic acid. Conversely, treatment of muscle homogenate with alkaline phosphatase completely reversed the contraction-mediated lipase activation. Lipase activity did not change during contractions when analysed in the presence of anti-hormone-sensitive-lipase (HSL) antibody [0.17 +/- 0.02 (basal) versus 0.21 +/- 0.02 (5 min) m-unit.mg of protein(-1), P > 0.05]. Furthermore, immunoprecipitation with affinity-purified anti-HSL antibody reduced muscle-HSL protein concentration by 81+/-4% and caused similar reductions in lipase activity against triacylglycerol and in the contraction-induced increase in this activity. Neither prior sympathectomy [0.33+/- 0.02 (basal) versus 0.53 +/- 0.06 (5 min) m-unit.mg of protein(-1), P < 0.05] nor propranolol impaired the lipase response to contractions. Glycogen phosphorylase activity in the absence of AMP increased after 1 min [27.3 +/- 3.1 versus 8.9 +/- 1.8% (activity without AMP/total activity with AMP), P < 0.05] and returned to basal levels after 5 min. In conclusion, skeletal-muscle-immunoreactive HSL is transiently stimulated by contractions and the mechanism probably involves phosphorylation. The time course of HSL activation is similar to that of glycogen phosphorylase. Apparently, the two enzymes are regulated in parallel by contraction-induced as well as hormonal mechanisms, allowing simultaneous recruitment of all major extra- and intra-muscular energy stores.

Masoprocol decreases rat lipolytic activity by decreasing the phosphorylation of HSL.

Masoprocol (nordihydroguaiaretic acid), a lipoxygenase inhibitor isolated from the creosote bush, has been shown to decrease adipose tissue lipolytic activity both in vivo and in vitro. The present study was initiated to test the hypothesis that the decrease in lipolytic activity by masoprocol resulted from modulation of adipose tissue hormone-sensitive lipase (HSL) activity. The results indicate that oral administration of masoprocol to rats with fructose-induced hypertriglyceridemia significantly decreased their serum free fatty acid (FFA; P < 0.05), triglyceride (TG; P < 0.001), and insulin (P < 0.05) concentrations. In addition, isoproterenol-induced lipolytic rate and HSL activity were significantly lower (P < 0.001) in adipocytes isolated from masoprocol compared with vehicle-treated rats and was associated with a decrease in HSL protein. Incubation of masoprocol with adipocytes from chow-fed rats significantly inhibited isoproterenol-induced lipolytic activity and HSL activity, associated with a decrease in the ability of isoproterenol to phosphorylate HSL. Masoprocol had no apparent effect on adipose tissue phosphatidylinositol 3-kinase activity, but okadaic acid, a serine/threonine phosphatase inhibitor, blocked the antilipolytic effect of masoprocol. The results of these in vitro and in vivo experiments suggest that the antilipolytic activity of masoprocol is secondary to its ability to inhibit HSL phosphorylation, possibly by increasing phosphatase activity. As a consequence, masoprocol administration results in lower serum FFA and TG concentrations in hypertriglyceridemic rodents.