The lipase/lipoxygenase bienzyme system in AOT reversed micelles in octane

In this work it is shown that the bienzyme lipase/lipoxygenase system can function in reversed micelles of bis(2-ethyl)hexyl sulfosuccinate (AOT) in octane. As a lipase substrate, a fish fat preparation (fat of sea mammals) with a high content of polyunsaturated fatty acids was used. It was demonstrated that the bienzyme reaction proceeded in a stationary mode and had a rate-limiting step catalyzed by lipase. Under optimal conditions, the efficacy of functioning of the bienzyme system was by an order of magnitude higher than that in water. The lipase/lipoxygenase bienzyme system can be used as a new method of spectrophotometric determination of lipase activity. The English version of the paper.     

A Spectrophotometric Lipase Activity Assay by Coupled Lipase-Lipoxygenase in Reverse Micelles

A new, continuous spectrophotometric method is described for determining lipase activity using a reverse micelle system, in which lipase (EC 3.1.1.3) and lipoxygenase (EC 1.13.11.12) are dissolved. The reverse micelle system consists of 2-ethyl hexyl sodium sulfosuccinate (AOT)-isooctane and water. Trilinolein is used as the lipase substrate; linoleate hydroperoxide is the end product of the oxidation catalyzed by lipoxygenase, which acts as an auxiliary coupled-enzyme of lipase. The method appears useful both for detailed kinetic studies of lipase and for serial analyses using sunflower oil, a cheaper substrate. This assay offers the typical advantages of the continuous direct photometric methods in that it is rapid, reproducible and sufficiently sensitive for measuring lipase activity even in some crude commercial preparations.     

A Spectrophotometric Lipase Activity Assay by Coupled Lipase-Lipoxygenase in Reverse Micelles

A new, continuous spectrophotometric method is described for determining lipase activity using a reverse micelle system, in which lipase (EC 3.1.1.3) and lipoxygenase (EC 1.13.11.12) are dissolved. The reverse micelle system consists of 2-ethyl hexyl sodium sulfosuccinate (AOT)-isooctane and water. Trilinolein is used as the lipase substrate; linoleate hydroperoxide is the end product of the oxidation catalyzed by lipoxygenase, which acts as an auxiliary coupled-enzyme of lipase. The method appears useful both for detailed kinetic studies of lipase and for serial analyses using sunflower oil, a cheaper substrate. This assay offers the typical advantages of the continuous direct photometric methods in that it is rapid, reproducible and sufficiently sensitive for measuring lipase activity even in some crude commercial preparations.     

Biomodulator-mediated susceptibility of endogenous lipid droplets from rat adipocytes to hormone-sensitive lipase

The amount of fatty acid release by a fat cell homogenate without pretreatment with epinephrine was found to be slightly more than that released from fat cells by epinephrine, suggesting that fat cells contain high lipolytic activity even in the absence of lipolytic agents. Fat cells contain high hormone-sensitive lipase activity (1383 mumole free fatty acids/g/hr) in the absence of epinephrine, and addition of epinephrine to the cells did not increase the activity, significantly. Like epinephrine, DBcAMP and/or theophylline also elicited marked release of glycerol from fat cells without activating the hormone-sensitive lipase activity. However, although fat cells contain a large amount of hormone-sensitive lipase, lipolysis was negligible in the absence of these lipolytic agents. These results suggest that lipolytic agents such as epinephrine, DBcAMP, and theophylline induce lipolysis in fat cells through some mechanism other than activation of hormone-sensitive lipase and that in the absence of lipolytic agents, some system in fat cells inhibits lipolysis of endogenous lipid droplets by hormone-sensitive lipase. The lipid droplets in fat cells consist mainly of triglyceride with phospholipids, cholesterol, carbohydrate, and protein as minor constituents. The phospholipid fraction was found to consist of 75% phosphatidylcholine and 25% phosphatidylethanolamine. Of the minor constituents of endogenous lipid droplets, only phosphatidylcholine strongly inhibited hormone-sensitive lipase activity in a [3H]triolein emulsion. These results suggest that phosphatidylcholine in endogenous lipid droplets may be responsible for inhibition of hormone-sensitive lipase. Then, a cell-free system was established in which epinephrine, DBcAMP, and theophylline stimulated lipolysis of endogenous lipid droplets from fat cells by lipase solution. In this system, these lipolytic agents did not induce lipolysis in the absence of added lipase. Lipolysis in the mixture of the endogenous lipid droplets and lipase solution was accelerated by phospholipase C with concomitant loss of epinephrine-induced lipolysis. After pretreatment of the endogenous lipid droplets with phospholipase C, these lipolytic agents no longer induced lipolysis. Pretreatment of the endogenous lipid droplets with phospholipase C reduced their phospholipid content with the formation of phosphorylcholine, but did not affect their triglyceride and cholesterol contents. Treatment of the endogenous lipid droplets with phospholipase D did not affect lipolysis in the cell-free system. These results suggest that phosphatidylcholine in the endogenous lipid droplets may inhibit their lipolysis by hormone-sensitive lipase in fat cells and also be involved in the mechanisms of the stimulatory effects of epinephrine, DBcAMP, and theophylline on lipolysis.     

Green tea extract (AR25) inhibits lipolysis of triglycerides in gastric and duodenal medium in vitro

In this study, we aimed to evaluate in vitro the inhibitory activity of a green tea extract (AR25 standardized at 25% catechins) on gastric and pancreatic lipase activities. We first used tributyrin as a substrate to evaluate the capability of AR25 to induce digestive lipase inhibition. Gastric lipase was totally inhibited by 40 mg AR25/g tributyrin whereas pancreatic lipase inhibition was maximum (78.8 +/- 0.7%) with 80 mg AR25/g tributyrin. We then used triolein, a long-chain triglyceride, to check whether AR25 could alter lipase activities on a physiologic substrate. AR25 60 mg/g triolein induced a dramatic inhibition of gastric lipase (96.8 +/- 0.4%) whereas pancreatic lipase activity was partially reduced (66.50 +/- 0.92%). Finally, the concerted action of gastric and pancreatic lipases was studied with an excess of enzymes to mimic the physiologic conditions observed in vivo. Incubation of AR25 with an excess of digestive lipases resulted in a drastic decrease in gastric lipolysis but the inhibitory effect on pancreatic lipase was less marked. On the whole, as compared to the control, lipolysis of triolein under the successive action of the two digestive lipases was reduced by 37 +/- 0.6% in the presence of AR25. Because a lipid/water interface is necessary for lipolysis to occur, lipid emulsification and emulsion droplet size were measured in gastric and duodenal media in the presence of AR25. In gastric and duodenal conditions, AR25 inhibited the lipid emulsification process. From these data we conclude that (1) in vitro, fat digestion is significantly inhibited by 60 mg AR25/g triolein, and (2) gastric as well as pancreatic lipase inhibition could be related to altered lipid emulsification in gastric or duodenal media. The green tea extract AR25 exhibiting marked inhibition of digestive lipases in vitro is likely to reduce fat digestion in humans.     

Lipoxygenase inhibitors inhibit heparin-releasable lipoprotein lipase activity in 3T3-L1 adipocytes and enhance body fat reduction in mice by conjugated linoleic acid

The t10c12 isomer of conjugated linoleic acid (CLA) reduces lipid accumulation in adipocytes in part by inhibiting heparin-releasable lipoprotein lipase (LPL) activity. We now show that inhibitors of lipoxygenase (LOX) activity (2-[12-hydroxydodeca-5,10-diynyl]-3,5,6-trimethyl-p-benzoquinone; 5,8,11,14-eicosatetraynoic acid; salicylhydroxamic acid; indomethacin; nordihydroguaiaretic acid (NDGA)) produce a similar inhibitory effect on LPL activity in cultured 3T3-L1 mouse adipocytes. Additionally the LOX inhibitors had no effect on, or inhibited, lipolysis in this cell system (measured as glycerol release). Growing mice fed diet containing 0.1% NDGA for 4 weeks displayed 21% reduction in body fat, which was similar to 23% reduction in body fat produced by feeding diet containing a suboptimal amount of CLA (0.1%) for 4 weeks. Feeding diet containing both 0.1% NDGA and 0.1% CLA resulted in 51% reduction in body fat which was accompanied by significant increases in whole body water and protein. Aspirin, an inhibitor of cyclooxygenase 1 and 2, had no effect on LPL activity in 3T3-L1 adipocytes, did not affect body composition when fed to growing mice, and failed to influence the effects of CLA on LPL activity in 3T3-L1 cells or body composition in mice. These findings appear to provide new perspectives and insights into the relationships between CLA, eicosanoids, the control of lipid accumulation in adipocytes, and effects of CLA on the immune system.     

Lipoxygenase inhibitors inhibit heparin-releasable lipoprotein lipase activity in 3T3-L1 adipocytes and enhance body fat reduction in mice by conjugated linoleic acid.

The t10c12 isomer of conjugated linoleic acid (CLA) reduces lipid accumulation in adipocytes in part by inhibiting heparin-releasable lipoprotein lipase (LPL) activity. We now show that inhibitors of lipoxygenase (LOX) activity (2-[12-hydroxydodeca-5,10-diynyl]-3,5,6-trimethyl-p-benzoquinone; 5,8,11,14-eicosatetraynoic acid; salicylhydroxamic acid; indomethacin; nordihydroguaiaretic acid (NDGA)) produce a similar inhibitory effect on LPL activity in cultured 3T3-L1 mouse adipocytes. Additionally the LOX inhibitors had no effect on, or inhibited, lipolysis in this cell system (measured as glycerol release). Growing mice fed diet containing 0.1% NDGA for 4 weeks displayed 21% reduction in body fat, which was similar to 23% reduction in body fat produced by feeding diet containing a suboptimal amount of CLA (0.1%) for 4 weeks. Feeding diet containing both 0.1% NDGA and 0.1% CLA resulted in 51% reduction in body fat which was accompanied by significant increases in whole body water and protein. Aspirin, an inhibitor of cyclooxygenase 1 and 2, had no effect on LPL activity in 3T3-L1 adipocytes, did not affect body composition when fed to growing mice, and failed to influence the effects of CLA on LPL activity in 3T3-L1 cells or body composition in mice. These findings appear to provide new perspectives and insights into the relationships between CLA, eicosanoids, the control of lipid accumulation in adipocytes, and effects of CLA on the immune system.     

Evidence for the participation of arachidonic acid metabolites in trehalose efflux from the hormone activated fat body of the cockroach (Periplaneta americana)

The hypertrehalosemic hormones, HTH-I and HTH-II, activate trehalose synthesis and increase the rate of sugar efflux from Periplaneta americana fat body in vitro. These processes are unaffected by the diacylglycerol, 1-oleyl-2-acetyl-sn-glycerol, an activator of protein kinase C. Similarly, H-7 and spingosine, inhibitors of protein kinase C, are also inactive against trehalose efflux. The possibility that diacylglycerol lipase might generate an active fatty acid species was ruled out because of the failure of the inhibitor RHC-80267 to inhibit trehalose efflux. Activation of trehalose efflux from the intact fat body by HTH-I was strongly inhibited in a concentration dependent manner by the cyclooxygenase inhibitors indomethacin and diclofenac, but not by acetylsalicylic acid. Nordihydroguaiaretic acid, a lipoxygenase inhibitor, also blocked HTH-I activated trehalose efflux in a concentration dependent fashion. The phospholipase A(2) inhibitors mepacrine and 4'-bromophenacyl bromide were also effective in decreasing the efflux of trehalose from HTH-I challenged fat body. The data suggest possible roles for arachidonic acid metabolites in the regulation of trehalose synthesis and in the efflux of the sugar from the fat body.     

Effects of dietary cholesterol on adipose tissue lipoprotein lipase in the baboon

The effects of infant diet (breast milk or formula containing 2, 30 or 60 mg/dl cholesterol) and subsequent dietary cholesterol (0.02, 1.0 or 1.7 mg/kcal) and fat (saturated or unsaturated) on heparin-releasable lipolytic activity from omental adipose tissue was estimated from 99 baboons of 5-8 years of age. This lipase activity was characterized as lipoprotein lipase based on salt inhibition and apolipoprotein C-II activation. Lipoprotein lipase activity released from adipose tissue by heparin was significantly (P less than 0.002) lower in high cholesterol-fed baboons than in those fed low cholesterol. Most of this difference was due to impaired long-term heparin release of lipoprotein lipase. Adipose tissue lipoprotein lipase increased with increasing fat cell size regardless of diet, but there was no effect of diet on adipocyte size. There were no significant effects of infant cholesterol intake nor adult saturated or unsaturated fat on lipoprotein lipase activity. Adult baboons breast fed as infants had lower adipose tissue lipoprotein lipase activity (P less than 0.07) than adults fed formula as infants.     

Inhibition of human gastric lipase by intraduodenal fat involves glucagon-like peptide-1 and cholecystokinin

Seven healthy volunteers were intubated with two double lumen nasogastric tubes, one in the stomach, the other in the duodenum. This system allows simultaneous sampling of gastric juice and separate intraduodenal perfusion with a dietary fat (fish oil, 1269 kJ). Gastrin-17 was infused i.v. at a rate of 40 pmol/kg/h throughout the study. Gastric lipase was measured at 15-min intervals as activity (tributyrin) and as immunoreactivity (ELISA). Infusion of gastrin-17 resulted in a stable increase in the plasma concentration from a basal concentration of 8.3±0.8 pmol/l to 41.4±4.2 pmol/l. Perfusion with fat reduced gastric lipase activity from 24.2±5.3 to 7.2±2.5 kU/l (P<0.05), and immunoreactivity from 0.7±0.1 to 0.42±0.1 mg/l (P<0.05). After termination of fat perfusion, gastric lipase secretion increased again, though not reaching preinhibitory concentrations. During the intraduodenal perfusion with fat the plasma concentrations of glucagon-like peptide-1 (GLP-1) and cholecystokinin (CCK) increased from 6.9±0.5 to 15.1±1.5 pmol/l (P<0.05) and from 1.2±0.4 to 3.8±0.9 pmol/l (P<0.05). This study reveals a negative effect of fat in the duodenum on gastric lipase secretion. This effect may be mediated by GLP-1 and/or CCK.     

Activation of triacylglycerol lipase in the fat body of a beetle by adipokinetic hormone

The activation of triacylglycerol lipase and the stimulation of proline synthesis in the fat body of the fruit beetle Pachnoda sinuata by the endogenous octapeptide hormone Melme-CC (pQLNYSPDWa), which belongs to the family of insect adipokinetic hormones, were studied, and the correlation of both events investigated. At rest, the activity of triacylglycerol lipase in the fat body of the beetle was higher than in the fat body of the American cockroach, Periplaneta americana, but lower than in the migratory locust, Locusta migratoria. Triacylglycerol lipase of the beetle is activated by: (a) injection of synthetic Melme-CC and (b) the stimulus of flight. Activation of lipase by Melme-CC is time-dependent. Injection of cpt-cAMP activates triacylglycerol lipase in the fat body and causes an increase in the concentration of proline in the haemolymph at the expense of alanine. In contrast, injection of F-inositol-1,4,5-phosphate does not affect the activation state of lipase, nor the levels of amino acids in the haemolymph. High doses of octopamine do not activate lipase. Furthermore, activity of fat body lipase and proline concentration in the haemolymph both follow a circadian rhythm: both parameters are high in the morning, whereas they are low in the evening. When transfer of Melme-CC, released from the corpora cardiaca, to the thorax/abdomen is prevented by neck-ligation, the activity of lipase, as well as the circulating proline levels are low. Regression analysis revealed that activity of triacylglycerol lipase is positively correlated to proline concentration in the haemolymph, whereas there is a negative correlation of the enzyme activity and alanine level in the haemolymph. From these results we conclude that the activation of fat body triacylglycerol lipase by Melme-CC in P. sinuata stimulates proline synthesis. Proline is one of the major substrates to power flight activity in the beetle.     

Influence of Surfactants on Lipase Fat Digestion in a Model Gastro-intestinal System

In the present study, we use a model gastro-intestinal system to study the influence of different food-grade surface-active molecules (Sn-2 monopalmitin, β-lactoglobulin, or lysophosphatodylcholine) on lipase activity. The interfacial activity of lipase and surfactants are assessed with the pendant drop technique, a commonly used tensiometry instrument. A mathematical model is adopted which enables quantitative determination of the composition of the water–oil interface as a function of bulk surfactant concentration in the water–oil mixtures. Our results show a decrease in gastric lipolysis when interfacially active molecules are incorporated into a food matrix. However, only the Sn-2 monopalmitin caused a systematic decrease in triglyceride hydrolysis throughout the gastro-intestinal tract. This effect is most likely due to exclusion of both lipase and triglyceride from the water–oil interface together with a probable saturation of the solubilization capacity of bile with monoglycerides. Addition of β-lactoglobulin or lysophopholipids increased the hydrolysis of fat after the gastric phase. These results can be attributed to an increasing interfacial area with lipase and substrate present at the interface. Otherwise, β-lactoglobulin, or lysophopholipids reduced fat hydrolysis in the stomach. From the mathematical modeling of the interface composition, we can conclude that Sn-2 monopalmitin can desorb lipase from the interface, which, together with exclusion of substrate from the interface, explains the gradually decreased triglyceride hydrolysis that occurs during the digestion. Our results provide a biophysics approach on lipolysis that can bring new insights into the problem of fat uptake.     

Testing of alkaline and enzymatic hydrolysis pretreatments for fat particles in slaughterhouse wastewater

Four pretreatments to hydrolyse and/or reduce the size of fat particles in slaughterhouse wastewater (SHW) were tested: sodium hydroxide and three lipases of plant, bacterial and animal (pancreatic) origin. Hydrolysing agents and SHW containing between 2.5 and 3 g/l of fat particles were mixed at room temperature for 4 h. Additions of 5-400 meq NaOH/l did not increase soluble COD (SCOD) in SHW, but the average particle size was reduced to 73% +/- 7% of the initial average particle size (D(in)) at NaOH concentrations ranging from 150 to 300 meq/l. Pretreatment with pancreatic lipase PL-250 reduced the average particle size to a maximum of 60% +/- 3% of D(in). As D(in) was decreased from 359 to 68 microns, the enzyme concentration required to obtain the maximum particle size reduction increased from 200 to 1000 mg/l. A 4-h pretreatment with PL-250 also increased the free long-chain fatty acid (LCFA) concentration to a maximum of 15.5 mg/l, indicating some solubilization of the pork fat particles in SHW. SCOD was not significantly increased by the pretreatment, but SCOD was not found to be a good indicator of enzymatic lipolysis because of enzyme adsorption on the fat particle surface. Pancreatic lipase appeared more efficient with beef fat than pork fat, possibly because beef fat contains less polyunsaturated fatty acids than pork fat. The bacterial lipase LG-1000 was also efficient in reducing average fat particle size, but high doses (> 1000 mg/l) were required to obtain a significant reduction after 4 h of pretreatment. SCOD was not increased by pretreatment with LG-1000. No particle size reduction or changes in SCOD were noted after 4 h of pretreatment with the plant lipase EcoSystem Plus. It was concluded that PL-250 was the best pretreatment to hydrolyse fat particles in SHW. However, its impact on the efficiency of a downstream anaerobic digestion process remains to be tested.     

The role of neutral lipases in human adipose tissue lipolysis

PURPOSE OF REVIEW: The aim of this article is to describe the relative roles of hormone sensitive lipase and adipose triglyceride lipase in human fat cell lipolysis. RECENT FINDINGS: Until recently, only hormone sensitive lipase was considered important for the regulation of lipolysis within fat cells. Recent rodent studies have suggested that adipose triglyceride lipase may, however, be more important. The few human adipose triglyceride lipase studies that have been published point to species differences between humans and rodents. Selective inhibition of hormone sensitive lipase in human fat cells completely counteracts hormone-activated lipolysis, though there is a considerable (>50%) residual nonhormonal (basal) lipolysis. In rodents, adipose triglyceride lipase enzyme activity is stimulated by a cofactor termed CGI-58. In the absence of CGI-58, lipase activity in fat cells is much higher for hormone sensitive lipase than adipose triglyceride lipase. Hormone sensitive lipase expression is regulated by obesity and body weight reduction (decreased and increased, respectively), but this is not the case for adipose triglyceride lipase. A role of adipose triglyceride lipase in human lipolysis is suggested by studies of gene polymorphisms. SUMMARY: Two lipases the 'old' hormone sensitive lipase and the 'new' adipose triglyceride lipase are of importance for the regulation of lipolysis in rodent fat cells. In humans, adipose triglyceride lipase seems essential for maintaining basal lipolytic activity, while hormone sensitive lipase is the enzyme most responsive to stimulated lipolysis.     

Bienzyme HRP-GOx-modified gold nanoelectrodes for the sensitive amperometric detection of glucose at low overpotentials

Gold nanotubular electrode ensembles were prepared by using electroless deposition of the metal within the pores of polycarbonate track-etched membranes. Mono-enzyme (GOx) and monolayer/bilayer bienzyme (GOx/HRP) bioelectrodes were prepared by immobilizing the enzymes onto gold nanotubes surfaces modified with mercaptoethylamine. Batch amperometric responses to glucose for the different bioelectrodes were determined and compared. The response of the two geometries (monolayer and bilayer) of the bienzyme electrodes was shown to vary with regard to sensitivity at detection potentials above 0V. On the contrary, at detection potentials below 0V, no noticeable influence of the configuration of the bienzyme on the response intensity was observed. The mono-enzyme (650 microAmM-1 in benzoquinone (BQ) at -0.8 V versus Ag/AgCl) and the two bienzyme bioelectrodes (+/-400 microAmM-1 in hydroquinone (H2Q) at -0.2V versus Ag/AgCl) display remarkable sensitivities compared to a classical GOx-modified gold macroelectrode (13 microAmM-1 in BQ at -0.8 V versus Ag/AgCl). A remarkable feature of the bienzyme electrodes is the possibility to detect glucose at very low applied potentials where the noise level and interferences from other electro-oxidizable compounds are minimal. Another important characteristic of the monolayer bienzyme electrode is the possible existence of a direct electronic communication between HRP and the transducer surface.     

Tissue-Specific Lipoprotein Lipase: Relationships to Body Composition and Body Fat Distribution in Normal Weight Humans

Twenty-six normal weight subjects (22 female, 4 male) were studied to determine the relationships of fasting levels of lipoprotein lipase in gluteal adipose tissue (ATLPL) and skeletal muscle (SMLPL) to body composition and body fat distribution. No relationship was found between fasting gluteal ATLPL and percent (%) body fat There was, however, an inverse relationship between fasting SMLPL (from the vastus lateralis) and %body fat (p=0.005). A strong inverse correlation was also seen between fasting ATLPL and waist/hip ratio (p=0.0006), a measurement of body fat distribution. These relationships existed with or without the male subjects included. The tissue-specific relationships of lipoprotein lipase to body composition and body fat distribution could relate to the development of obesity or the maintenance of normal body weight by the effects of the lipase on the partitioning of lipoprotein triglyceride fatty acids.     

Electrically Contacted Bienzyme‐Functionalized Mesoporous Carbon Nanoparticle Electrodes: Applications for the Development of Dual Amperometric Biosensors and Multifuel‐Driven Biofuel Cells

The capping of electron relay units in mesoporous carbon nanoparticles (MPC NPs) by crosslinking of different enzymes on MPC NPs matrices leads to integrated electrically contacted bienzyme electrodes acting as dual biosensors or as functional bienzyme anodes and cathodes for biofuel cells. The capping of ferrocene methanol and methylene blue in MPC NPs by the crosslinking of glucose oxidase (GOx) and horseradish peroxidase (HRP) yields a functional sensing electrode for both glucose and H₂O₂, which also acts as a bienzyme cascaded system for the indirect detection of glucose. A MPC NP matrix, loaded with ferrocene methanol and capped by GOx/lactate oxidase (LOx), is implemented for the oxidation and detection of both glucose and lactate. Similarly, MPC NPs, loaded with 2,2′‐azino‐bis(3‐ethylbenzo­thiazoline‐6‐sulphonic acid), are capped with bilirubin oxidase (BOD) and catalase (Cat), to yield a bienzyme O₂ reduction cathode. A biofuel cell that uses the bienzyme GOx/LOx anode and the BOD/Cat cathode, glucose and/or lactate as fuels, and O₂ and/or H₂O₂ as oxidizers is assembled, revealing a power efficiency of ≈90 μW cm^?2 in the presence of the two fuels. The study demonstrates that multienzyme MPC NP electrodes may improve the performance of biofuel cells by oxidizing mixtures of fuels in biomass.     

Cetilistat (ATL-962), a novel pancreatic lipase inhibitor, ameliorates body weight gain and improves lipid profiles in rats.

Cetilistat is a novel inhibitor of pancreatic lipase. The aim of this report is to evaluate the anti-obesity action of cetilistat in diet-induced obesity (DIO) rats. Cetilistat inhibited rat and human pancreatic lipase activity with an IC (50) of 54.8 nmol/l, and 5.95 nmol/l, respectively, meaning that it is 9.2 times more potent for human pancreatic lipase than for that of rat. Cetilistat was orally administered simultaneously with fat emulsion to Sprague-Dawley rats. Plasma triglyceride (TG) concentrations were measured before and after oral fat loading. The elevation in plasma triglyceride concentration by oral fat loading was reduced by cetilistat in a dose-dependent manner at 3, 10, 30, and 100 mg/kg, indicating that cetilistat reduces intestinal fat absorption in rats. Cetilistat was administered to DIO F344 rats as food admixture in a high-fat diet at 4.9, 14.9, or 50.7 mg/kg/day for three weeks. Both triglyceride and nonesterified fatty acid content in the feces were dose-dependently and drastically increased, suggesting the intestinal breakdown of fat and excretion. Body weight (BW) gain and white adipose tissue (WAT) weight were reduced in a dose-dependent manner. In addition, leptin, TG, and total cholesterol (TC) in plasma were reduced and there were no reports of oily stools. These results suggest that cetilistat ameliorates obesity and hyperlipidemia in DIO rats, a plausible animal model of the most common type of human obesity.     

Fat absorption in cystic fibrosis mice is impeded by defective lipolysis and post-lipolytic events

Cystic fibrosis (CF) is frequently associated with progressive loss of exocrine pancreas function, leading to incomplete digestion and absorption of dietary fat. Supplementing patients with pancreatic lipase reduces fat excretion, but it does not completely correct fat malabsorption, indicating that additional pathological processes affect lipolysis and/or uptake of lipolytic products. To delineate the role of such (post) lipolytic processes in CF-related fat malabsorption, we assessed fat absorption, lipolysis, and fatty acid uptake in two murine CF models by measuring fecal fat excretion and uptake of oleate- and triolein-derived lipid. Pancreatic and biliary function was investigated by determining lipase secretion and biliary bile salt (BS) secretion, respectively. A marked increase in fecal fat excretion was observed in cftr null mice but not in homozygous DeltaF508 mice. Fecal BS loss was enhanced in both CF models, but biliary BS secretion rates were similar. Uptake of free fatty acid was delayed in both CF models, but only in null mice was a specific reduction in lipolytic activity apparent, characterized by strongly reduced triglyceride absorption. Impaired lipolysis was not due to reduced pancreatic lipase secretion. Suppression of gastric acid secretion partially restored lipolytic activity and lipid uptake, indicating that incomplete neutralization of gastric acid impedes fat absorption. We conclude that fat malabsorption in cftr null mice is caused by impairment of lipolysis, which may result from aberrant duodenal pH regulation.