Prostaglandin production and lipolysis in isolated rat adipocytes as affected by dietary fat

The influence of dietary fat on prostaglandin production and lipolysis was tested in basal and norepinephrine stimulated adipocytes isolated from the epididymal fat pads of fasted rats. Seven diets varying in fat calories and polyunsaturation were utilized. No basal differences were noted for prostaglandin E₂ production or lipolysis. Norepinephrine stimulated prostaglandin E₂ and F_2α production was significantly (P < 0.01) increased with greater polyunsaturation of fat, but not by increased fat calories. Norepinephrine stimulated lipolysis was depressed by an increase in fat calories but was unaffected by the degree of polyunsaturation of fat. This is in vitro evidence against the concept that prostaglandins play a feedback regulator role in fat cell lipolysis since no correlation could be made between the two parameters.     

Membrane phospholipids and hormone-sensitive lipolysis in fat cells

Isolated fat cells from rats which have been made hypothyroid do not give a lipolytic response to catecholamines. A recent report has suggested that catecholamine-sensitive lipolysis may be correlated with an “unmasking” of receptors by linoleic acid rich phospholipids in the fat cell membrane. No apparent differences in phospholipid fatty acid composition could be found in membrane “ghosts” prepared from normal and hypothyroid rats.     

Effects of insulin on adrenoceptor binding and the rate of catecholamine-induced lipolysis in isolated human fat cells

The mechanisms by which insulin inhibits catecholamine-induced lipolysis in fat cells are unknown. In this study the possible role of an interaction between insulin and the adrenoceptors on human fat cells was investigated. Insulin inhibited, in a dose-dependent fashion, the specific binding of hydrophobic as well as hydrophilic nonselective beta-receptor radioligands but had no effect on the binding of alpha 2-selective radioligands. The results of saturation experiments and competition-inhibition experiments under both equilibrium conditions and nonequilibrium conditions revealed that insulin reduced the total number of beta-adrenergic binding sites (maximum effect 25%) without changing the beta-adrenoceptor affinity. This insulin effect was rapid and reversible; one-third of the effect occurred within 1 min of incubation and it was completely reversed within 30 min after withdrawal of insulin. It could be mimicked by a polyclonal rabbit insulin receptor antibody but not by insulin mimickers acting distal to the initial interaction between the hormone and its specific insulin-receptor binding site. The beta-adrenoceptor binding to a plasma membrane-enriched fraction decreased at the same time as it increased to a microsomal enriched fraction after insulin treatment, indicating a redistribution of beta-adrenoceptors in the cell. In lipolysis experiments performed under conditions like those in the binding experiments, insulin inhibited the rate of lipolysis with a lag period of 3 min. Furthermore, the hormone caused a dose-dependent maximum 10-fold shift to the right of the dose-response curve for isoprenaline-induced lipolysis without changing the amplitude of the curve. This effect of insulin was specific for the beta-adrenergic receptors system, since insulin markedly decreased the amplitude of the dose-response curve for parathyroid hormone-induced lipolysis. In addition, the effect of insulin on isoprenaline-induced lipolysis could be mimicked by long-lasting fractional inactivation of the beta-adrenoceptors. The dose-response relationships for the inhibitory effects of insulin on beta-adrenoceptor binding and the lipolytic sensitivity to isoprenaline were almost identical. Half-maximum and maximum effects occurred at about 5 and 100 microunits/ml of insulin, respectively. In conclusion, the exposure of human fat cells to physiological insulin doses is followed by a rapid and dose-dependent translocation of beta-adrenoceptors from the exterior to the interior of the cell and a subsequent dose-dependent decrease in the lipolytic sensitivity to beta-adrenergic agonists, without a change in maximum lipolysis.(ABSTRACT TRUNCATED AT 400 WORDS)     

Potentiation of epinephrine-induced lipolysis in fat cells from estrogen-treated rats

We investigated concomitantly the effects of ethinyl-estradiol (EE₂) at low dose (1.2 μg/animal for 10 days) and high dose (120 μg/animal for 3 days) on body weight, weight of fat stores, triglyceridemia and fat cell lipoprotein lipase and hormone-sensitive lipase activities in female rats. At low dose, EE₂ increased triglyceridemia and lipoprotein lipase activity, whilst high dose decreased both parameters. At low and high doses, and regardless of whether triglyceridemia and lipoprotein lipase activity were increased and decreased, EE₂ caused depletion in fat stores. Fat cells isolated from depleted fat tissue elicited marked increases in the response of hormone-sensitivelipase to epinephrine. Taken together, the data suggest that the potentiation of epinephrine-induced lipolysis in fat cells is likely to represent a major cause to estrogen-induced fat depletion.     

Effects of adenosine deaminase on cyclic adenosine monophosphate accumulation, lipolysis, and glucose metabolism of fat cells.

In fat cells isolated from the parametrial adipose tissue of rats, the addition of purified adenosine deaminase increased lipolysis and cyclic adenosine 3':5'-monophosphate (cyclic AMP) accumulation. Adenosine deaminase markedly potentiated cyclic AMP accumulation due to norepinephrine. The increase in cyclic AMP due to adenosine deaminase was as rapid as that of theophylline with near maximal effects seen after only a 20-sec incubation. The increases in cyclic AMP due to crystalline adenosine deaminase from intestinal mucosa were seen at concentrations as low as 0.05 mug per ml. Further purification of the crystalline enzyme preparation by Sephadex G-100 chromatography increased both adenosine deaminase activity and cyclic AMP accumulation by fat cells. The effects of adenosine deaminase on fat cell metabolism were reversed by the addition of low concentrations of N6-(phenylisopropyl)adenosine, an analog of adenosine which is not deaminated. The effects of adenosine deaminase on cyclic AMP accumulation were blocked by coformycin which is a potent inhibitor of the enzyme. These findings suggest that deamination of adenosine is responsible for the observed effects of adenosine deaminase preparations. Protein kinase activity of fat cell homogenates was unaffected by adenosine or N6-(phenylisopropyl)adenosine. Norepinephrine-activated adenylate cyclase activity of fat cell ghosts was not inhibited by N6-(phenylisopropyl)adenosine. Adenosine deaminase did not alter basal or norepinephrine-activated adenylate cyclase activity. Cyclic AMP phosphodiesterase activity of fat cell ghosts was also unaffected by adenosine deaminase. Basal and insulin-stimulated glucose oxidation were little affected by adenosine deaminase. However, the addition of adenosine deaminase to fat cells incubated with 1.5 muM norepinephrine abolished the antilipolytic action of insulin and markedly reduced the increase in glucose oxidation due to insulin. These effects were reversed by N6-(phenylisopropyl)adenosine. Phenylisopropyl adenosine did not affect insulin action during a 1-hour incubation. If fat cells were incubated for 2 hours with phenylisopropyl adenosine prior to the addition of insulin for 1 hour there was a marked potentiation of insulin action. The potentiation of insulin action by prior incubation with phenylisopropyl adenosine was not unique as prostaglandin E1, and nicotinic acid had similar effects.     

Relationship between hormone-sensitive lipolysis and lipase activity in rat fat cells

An assay for total hormone-sensitive lipase (HSL) in rat fat cells was devised in which fat-associated HSL was solubilized with ether, and triolein or cholesteryloleate was used as substrate. Norepinephrine (NE) caused marked release of glycerol from fat cells but did not activate HSL as estimated using triolein or cholesteryloleate as substrate. Propranolol, a beta-blocker, inhibited NE-induced lipolysis in fat cells without a concomitant reduction in HSL activity. The antilipolytic action of insulin on NE-induced lipolysis could not be explained by a decrease in HSL activity. Neither ACTH-induced lipolysis in fat cells nor its inhibition by insulin was accompanied by matching fluctuations in HSL activity. These results indicate that neither NE and ACTH-induced lipolysis in fat cells, nor the antilipolytic actions of propranolol and insulin, involve fluctuations in HSL activity.     

Natriuretic peptide-dependent lipolysis in fat cells is a primate specificity

We have recently demonstrated that natriuretic peptides (NPs), which are known for regulation of blood pressure via membrane guanylyl cyclase (GC) receptors, are lipolytic in human adipose tissue. In this study, we compared the NP control of lipolysis in adipocytes from humans, nonhuman primates (macaques), rodents (rats, mice, hamsters), and nonrodent mammals (rabbits, dogs). Isolated adipocytes from these species were exposed to increasing concentrations of atrial NP (ANP) or isoproterenol (beta-adrenergic agonist). Although isoproterenol was lipolytic in all of the species, ANP only enhanced lipolysis in human and macaque adipocytes. In primate fat cells, NP-induced lipolysis involved a cGMP-dependent pathway. Binding studies and real-time quantitative PCR assays revealed that rat adipocytes expressed a higher density of NP receptors compared with humans but with a different subtype pattern of expression; type-A GC receptors predominate in human fat cells. This was also confirmed by the weak GC-activity stimulation and the reduced cGMP formation under ANP exposure in rat adipocytes compared with human fat cells. In conclusion, NP-induced lipolysis is a primate specificity, and adipocytes from ANP-nonresponsive species present a predominance of "clearance" receptors and very low expression of "biologically active" receptors.     

, -Methylene-adenosine-5'-triphosphate--effects of a competitive inhibitor of adenylate cyclase on cyclic AMP accumulation and lipolysis in isolated fat cells

The rapid, transient rise in the intracellular concentration of cyclic AMP which follows addition of L-epinephrine to isolated fat cells is completely prevented by an ATP analog, α,β-methylene-adenosine-5′-triphosphate [Ap(CH₂)pp], a competitive inhibitor of adenylate cyclase activity in liver and fat cell membrane preparations. The concentration of cyclic AMP falls distinctly below that in the basal state after incubating fat cells for seven minutes in the presence of Ap(CH₂)pp. The results are consistent with the view that the ATP analog is also an effective $\text{in vivo}$ inhibitor of adenylate cyclase activity, and that intracellular cyclic AMP levels are normally delicately balanced by very rapid processes of synthesis and degradation. Epinephrine-induced lipolysis in fat cells is not inhibited but is instead enhanced by Ap(CH₂)pp. This is probably explained by the ability of the analog to act (like ATP) as a high-energy phosphate donor, an effect which is independent of its inhibition of adenylate cyclase activity. The predominant effect of this compound on glucose oxidation by fat cells also appears to be the result of this property since its effects are mimicked by ATP.     

Effect of aging and cellularity on lipolysis in isolated mouse fat cells

The effects of age and cellularity on lipolysis have been investigated in isolated epididymal fat cells from both Swiss albino mice and Sprague-Dawley rats. No significant lipolytic response to glucagon could be demonstrated with adipocytes from either young or old mice, while glycerol output was increased by this hormone with fat cells from young rats. Larger adipocytes from older mice showed significantly greater isoproterenol-stimulated lipolysis than those from younger animals if the glycerol output was expressed on a per cell basis. However, the lipolytic response per cell appeared to be equivalent in young and old rat adipocytes with either isoproterenol or ACTH-(1-24). In a complete aging study, relationships between body weight, epididymal fat pad weight and cellularity were examined covering the life span of the mouse. ACTH-(1-24)- and dibutyryl cyclic AMP-stimulated lipolysis increased with age and cell size but fell at senescence when adipocyte size diminished. Although an effect of aging per se cannot be ruled out with the experimental techniques used in the present study, a dominant influence of adipocyte size on the lipolytic process was demonstrated.     

Effects of thyroid hormone deficiency on lipolysis in chicken fat cells

Effect of lack of thyroid hormones on lipolysis in chicken fat cells was studied. Isolated fat cells from hypothyroid chickens in contrast to normal animals, have an impaired ability to give lipolytic response to glucagon. However activation of triglyceride lipolysis was induced to the same level by theophylline in hypothyroid and normal chickens.     

Influence of lactate on isoproterenol-induced lipolysis and beta-adrenoceptors distribution in human fat cells

The influence of lactate on human adipocytes lipolysis and the possible relationship between lactate-induced metabolic effects and beta-adrenoceptor binding sites were investigated. beta-sites were identified in membranes with (125I)-cyanopindolol and in intact cells with (125I)-cyanopindolol and (3H)-CGP 12177. Lactate reduced isoproterenol-induced lipolysis in a dose-response fashion and such inhibition became significant only at 16 mmol/l lactate. Exposure of human fat cells to 16 mmol/l lactate significantly reduced beta-adrenoceptors density on crude membranes. When the binding assay was performed on intact cells using (125I)-cyanopindolol at 37 degrees C, the radioligand identified the same number of receptors, regardless of the presence of lactate in the preincubation medium. When (3H)-CGP 12177 was used, it bound to about 35% less receptors in lactate pre-treated cells than in control. Seemingly, at 37 degrees C, because of its lipophilicity, (125I)-cyanopindolol can cross the plasma membrane and bind to intracellular sites whereas, (3H)-CGP 1277, due to its hydrophilicity, identifies surface receptors only. Thus, the present in vitro study provides evidence that high levels of lactate, similar to the concentrations usually achieved in overt lactic acidosis, are able per se to inhibit human lipolysis and to redistribute beta-adrenoceptors from cell surface to a domain not accessible to hydrophilic ligands.     

Resveratrol directly affects in vitro lipolysis and glucose transport in human fat cells

Resveratrol is a naturally occurring polyphenol found in many dietary sources and red wine. Recognized as a cancer chemoprevention agent, an anti-inflammatory factor and an antioxidant molecule, resveratrol has been proposed as a potential anti-obesity compound and to be beneficial in diabetes. Most of the studies demonstrating the anti-adipogenic action of resveratrol were performed as long-term treatments on cultured preadipocytes. The aim of this study was to analyse the acute effects of resveratrol on glucose uptake and lipolysis in human mature adipocytes. Samples of subcutaneous abdominal adipose tissue were obtained from overweight humans and immediately digested by liberase. Fat cells were incubated (from 45 min to 4 h) with resveratrol 1 μM–1 mM. Then, glycerol release or hexose uptake was determined. Regarding lipolysis, the significant effects of resveratrol were found at 100 μM, consisting in a facilitation of isoprenaline stimulation and an impairment of insulin antilipolytic action. At 1 and 10 μM, resveratrol only tended to limit glucose uptake. Resveratrol 100 μM did not change basal glucose uptake but impaired its activation by insulin or by benzylamine. This inhibition was not found with other antioxidants. Such impairment of glucose uptake activation in fat cells may led to a reduced availability of glycerol phosphate and then to a decreased triacylglycerol assembly. Therefore, resveratrol increased triacylglycerol breakdown triggered by β-adrenergic activation and impaired lipogenesis. Consequently, our data indicate that resveratrol can be considered as limiting fat accumulation in human fat cells and further support its use for the mitigation of obesity.     

Effect of PCO2 on epinephrine-induced lipolysis in isolated fat cells.

The effect of different pHs obtained by changing the PCO2 and the effect of PCO2 at constant pH on the lipolysis induced by epinephrine in isolated fat cells have been investigated. An inhibition of activated lipolysis was found in acidosis while in alkalosis no significant change was detected. When the experiments were performed at different PCO2s but at constant pH, the results showed an inhibition of lipolysis by high PCO2 whereas low PCO2 did not affect it. It is concluded that either acidosis or high PCO2 lead to an inhibition of the lipolysis induced by epinephrine in isolated fat cells. As regards alkalosis and low PCO2 it seems likely that the intracellular pH is not affected to the same extent as in alkalosis by high [HCO(-3)] or under the conditions of the present experiments the [H+] needed to alterate lipolysis was not reached.