Lipolytic Products Research Articles

Oral \u03b2-hydroxybutyrate increases ketonemia, decreases visceral adipocyte volume and improves serum lipid profile in Wistar rats

BackgroundKetosis can be induced in humans and in animals by fasting or dietary interventions, such as ketogenic diets. However, the increasing interest on the ketogenic state has motivated the development of alternative approaches to rapidly increase ketonemia using less drastic interventions. Here, it was tested whether oral intake of a β-hydroxybutyrate (βHB) mineral salt mixture could increase ketonemia in Wistar rats without any other dietary changes, thereby being a useful model to study ketones effects alone on metabolism.MethodsβHB salts were orally administered to provoke elevation in the ketonemia. Effects of this intervention were tested acutely (by gavage) and chronically (4 weeks in drinking water). Acutely, a concomitant glucose overload was used to suppress endogenous ketogenesis and verify whether βHB salts were really absorbed or not. Long-term administration allowed to weekly evaluate the impact on ketonemia, blood glucose and, after 4 weeks, on body weight, visceral fat mass, lipid blood profile, serum lipolysis products and adiponectinemia.ResultsβHB salts increased ketonemia in acute and long-term administrations, improved blood lipid profile by raising HDL-cholesterol concentration and decreasing LDL/HDL ratio, while reduced visceral adipocyte volume. Mean ketonemia correlated positively with HDLc and negatively with adipocyte volume and serum lipolysis products.ConclusionsOral βHB can rapidly increase ketonemia and, therefore, be used as an acute and long-term animal model of ketosis. Long-term treatment points to important beneficial effects of ketone bodies in serum lipid concentrations and visceral fat mass. These results may help to explain the metabolic adaptations following ketogenic diets, such as a better body fat control and a serum lipid profile improvement.

Interfacial behaviour of biopolymer multilayers: Influence of in vitro digestive conditions

Although multilayered emulsions have been related to reduced lipolysis, the involved interfacial phenomena have never been studied directly. In this work, we systematically built multilayers of whey protein and pectin, which we further subjected to digestive conditions, using two different techniques: droplet volume tensiometry to investigate interfacial rheology, and reflectometry to determine the amount of adsorbed material. Interfacial tension and dilatational rheology were linked to adsorption/desorption kinetics measured under static in vitro conditions.The interfacial tension and rheology of the multilayers was rather similar to those found for single whey protein layers, as well as their resistance to duodenal conditions and lipolytic components, which is explained by the rapid destabilisation of multilayers at neutral pH. Sequential adsorption of bile extract or lipase to pre-adsorbed films rapidly lowered the interfacial tension via co-adsorption and displacement, forming a viscoelastic film with low mechanical strength, and highly dynamic adsorption/desorption. When both were present, bile salts dominated the initial adsorption, followed by lipase co-adsorption and formation of lipolysis products that further lowered the interfacial tension, forming a complex interface (including biopolymers, bile salts, lipase, and lipolysis products), independent of pre-adsorbed biopolymer layers.Our study shows that the combination of drop volume tensiometry and reflectometry can be used to study complex interfacial behaviours under digestive conditions, which can lead to smart design of interfacial structures for controlled lipolysis in food emulsions.

Docosahexaenoic acid increases accumulation of adipocyte triacylglycerol through up-regulation of lipogenic gene expression in pigs

BackgroundChanging dietary fatty acid composition in modern diet influences the prevalence of obesity. Increasing evidences suggest favorable effects of n-3 PUFA for protecting against obesity and the metabolic syndrome. However, the regulation of n-3 PUFA in adipose is still unclear. Thus, this study addressed metabolism of different dietary fats in the adipose tissue of porcine model.MethodsEight-week-old cross-bred pigs were randomly assigned to three groups and fed a 2% fat diet for 30 days from either soybean oil (SBO), docosahexaenoic acid (DHA) or beef tallow. An in vitro experiment was conducted in which linoleic acid (LA), DHA or oleic acid (OA) were added to represent the major fatty acid in the SBO-, DHA- or BT- diets, respectively. Adipocytes size and lipid metabolism related genes were analyzed.ResultsPlasma triacylglycerol (TAG) was lower in DHA- than in BT-fed pigs, and the product of lipolysis, glycerol was highest in BT-fed pigs. In addition, expression of the lipolytic genes, adipose triglyceride lipase and hormone sensitive lipase was higher in BT-fed pigs and with OA treatment in vitro. DHA promoted protein kinase A activity in pigs without affecting lipolytic genes. Adipocyte cell sizes, TAG content and expression of lipogenic-related genes including, adipose differentiated related protein (ADRP) and diacylglycerol acyltransferase 1 (DGAT1) were elevated by DHA in vivo and in vitro, indicating DHA promoted adipogenesis to trap TAG in adipose tissue. Fatty acid β-oxidation genes were increased in the DHA-fed pigs.ConclusionThis effect was partly explained by the effect of DHA to promote adipogenesis to trap TAG in adipocytes and also increase expression of genes involved in adipocyte fatty acid oxidation. Therefore, our results suggest a direct effect of DHA on adipocyte metabolism, resulting in TAG turnover and fatty acid dissipation to facilitate plasma lipid uptake from the circulation.

Triglyceride-rich lipoprotein lipolysis products increase blood-brain barrier transfer coefficient and induce astrocyte lipid droplets and cell stress.

Elevation of blood triglycerides, primarily as triglyceride-rich lipoproteins (TGRL), has been linked to cerebrovascular inflammation, vascular dementia, and Alzheimer's disease (AD). Brain microvascular endothelial cells and astrocytes, two cell components of the neurovascular unit, participate in controlling blood-brain barrier (BBB) permeability and regulating neurovascular unit homeostasis. Our studies showed that infusion of high physiological concentrations of TGRL lipolysis products (TGRL + lipoprotein lipase) activate and injure brain endothelial cells and transiently increase the BBB transfer coefficient (Ki = permeability × surface area/volume) in vivo. However, little is known about how blood lipids affect astrocyte lipid accumulation and inflammation. To address this, we first demonstrated TGRL lipolysis products increased lipid droplet formation in cultured normal human astrocytes. We then evaluated the transcriptional pathways activated in astrocytes by TGRL lipolysis products and found upregulated stress and inflammatory-related genes including activating transcription factor 3 (ATF3), macrophage inflammatory protein-3α (MIP-3α), growth differentiation factor-15 (GDF15), and prostaglandin-endoperoxide synthase 2 (COX2). TGRL lipolysis products also activated the JNK/cJUN/ATF3 pathway, induced endoplasmic reticulum stress protein C/EBP homologous protein (CHOP), and the NF-κB pathway, while increasing secretion of MIP-3α, GDF15, and IL-8. Thus our results demonstrate TGRL lipolysis products increase the BBB transfer coefficient (Ki), induce astrocyte lipid droplet formation, activate cell stress pathways, and induce secretion of inflammatory cytokines. Our observations are consistent with evidence for lipid-induced neurovascular injury and inflammation, and we, therefore, speculate that lipid-induced astrocyte injury could play a role in cognitive decline.

In vivo evaluation of lipid-based formulations for oral delivery of apomorphine and its diester prodrugs

In the present study, the differences in oral absorption of apomorphine and its diester prodrugs and the effect of lipid-based formulations on the absorption of apomorphine or its prodrugs were investigated. Apomorphine, dilauroyl apomorphine (DLA) and dipalmitoyl apomorphine (DPA) were orally administered (0.24mmol/kg) to rats as: DLA-o/w emulsion, DPA-o/w emulsion, apomorphine-o/w emulsion, apomorphine aqueous suspension, DLA-Maisine, DLA-soybean oil, DLA-self-emulsifying drug delivery systems (SEDDS), and DLA-w/o emulsion. The o/w and w/o emulsion consisted of Maisine 35-1 emulsified with water in 1:3 and 4:1 ratio, respectively. Tmax of diesters was significantly increased (p≤0.05) compared to apomorphine in o/w emulsion, suggesting that esterification yielded prolonged drug absorption. Cmax, AUC and the relative bioavailability of apomorphine after DLA-SEDDS administration was higher (p≤0.05) than after DLA-w/o administration, indicating that triglycerides and surfactants improved the oral absorption of DLA. Similarly, Cmax and AUC after dosing apomorphine-o/w were significantly higher (p≤0.05) than that of aqueous suspension. This suggested that lipids and lipolysis products possibly aided apomorphine micellar solubilization in intestinal fluids. A combination of prodrug strategy and lipid-based formulations facilitated a higher and prolonged absorption of apomorphine from its diester prodrugs.

HDL inhibits saturated fatty acid mediated augmentation of innate immune responses in endothelial cells by a novel pathway

Background and aimsPeripheral insulin resistance is associated with several metabolic abnormalities, including elevated serum fatty acids that contribute to vascular injury and atherogenesis. Our goals were to examine whether saturated fatty acids can modify innate immune responses to subclinical concentrations of lipopolysaccharide (LPS) in endothelial cells, and to explore the underlying pathway and determine whether it is modified by high density lipoprotein (HDL) and other factors commonly altered in insulin resistance. MethodsPhysiologic concentrations of palmitic acid were added to human aortic endothelial cells with and without a variety of inhibitors or HDL and measures of cell inflammation and function assessed. ResultsPalmitic acid significantly amplified human aortic endothelial cell inflammatory responses to LPS. Similar results were obtained from lipolysis products of triglyceride rich lipoproteins. Metabolism of palmitic acid to ceramide and subsequent activation of PKC-ζ, MAPK and ATF3 appeared critical in amplifying LPS induced inflammation. The amplified response to palmitic acid/LPS was decreased by HDL, dose dependently, and this inhibition was dependent on activation of PI3K/AKT and reduction in ATF3. ConclusionsThese results indicate that endothelial cell innate immune responses are modified by metabolic abnormalities commonly present in insulin resistance and provide evidence for a novel mechanism by which HDL may reduce vascular inflammation.

Influence of lipid composition on the oral bioavailability of cinnarizine sub‐microemulsions

The oral bioavailability of cinnarizine is usually limited by its poor solubility in the gastrointestinal tract. In this study, two kinds of cinnarizine sub‐microemulsions (SMEs) were prepared using medium (MCT) and long (LCT) chain triglycerides, respectively, and the effect of the lipid composition on the oral bioavailability of SMEs was investigated in terms of stability, in vitro lipolysis, in situ single‐pass intestinal perfusion, and in vivo pharmacokinetics. The SMEs increased cinnarizine stability in simulated gastrointestinal fluid and in vitro lipolysis products of LCT‐SMEs showed better cinnarizine solubilization capacity than that of MCT‐SMEs. In addition, the absorption rate (Ka) and the effective permeability coefficient (Peff) of LCT‐SMEs were significantly improved compared with MCT‐SMEs. The AUC0‐t value of LCT‐SMEs was 4132.6 ± 673.4 ng/mL · h, which was 1.94‐fold higher than that of the aqueous suspension. Moreover, the AUC0‐t value of LCT‐SMEs was also significantly greater than that of MCT‐SMEs (2490.7 ± 685.3 ng/mL · h), owing to the better drug solubilization capacity of the complex colloidal mixtures formed from digestion of LCT. Hence, LCT‐SMEs was successful in improving the oral bioavailability of cinnarizine and it was a superior drug vehicle for oral administration.Practical applications: Owing to poor solubility of cinnarizine in the gastrointestinal tract, its commercially available dosage forms including tablets and capsules exhibit low oral bioavailability. Here we propose SMEs to improve oral bioavailability of cinnarizine. In addition, the influence of lipid composition on the oral absorption of SMEs was investigated, and LCT‐SMEs significantly increased the oral bioavailability of cinnarizine compared with MCT‐SMEs. So, in this study we show that LCT‐SMEs is a promising drug vehicle for oral administration of cinnarizine.The colloidal structures generated from digested long chain triglycerides (LCT) have a better cinnarizine solubilisation ability than that generated from medium chain triglycerides (MCT) digestion, resulting in increased oral absorption for the LCT formulation compared with MCT.

Destabilization of multilayered interfaces in digestive conditions limits their ability to prevent lipolysis in emulsions

Abstract Delivery of lipid fractions in the lower small intestine can induce feelings of satiety, but is only possible when lipids escape the highly efficient lipolysis and adsorption in the upper gastrointestinal (GI) tract. Our objective was to gain insight in the stability of multilayered interfaces in simulated GI conditions, and their suitability for intestinal delivery of undigested lipids. Oil-in-water emulsions ( d 32 ∼ 5–30 μm; one- to five-layered interfaces) were produced by sequentially adsorbing biopolymers with opposite charges at pH 3.0: whey protein isolate (WPI) (cationic), pectin (anionic), chitosan (cationic). Corresponding multilayered structures were characterized using reflectometry. Influence of layer composition and thickness on its protectiveness against lipolysis of emulsions was studied in simulated GI conditions. Multilayered WPI/pectin emulsions had an improved physical stability compared to WPI-stabilized emulsions, during both storage and in vitro gastric incubation, whereas chitosan-containing emulsions were physically unstable. Reflectometry and CLSM results showed that a greater number of layers increased the adsorbed amount, forming a mesoscopically heterogeneous structure. Under simulated intestinal conditions, however, outer layers instantaneously destabilized. Accordingly, similar initial lipolysis rates were recorded for all emulsions. Yet, compared to only WPI the final extent of lipolysis was lowered by addition of a second and a third layer under mild in vitro conditions. This moderate protective effect disappeared when harsher digestive conditions were applied. From this work, it became clear why multilayered interfaces (initially built under acidic pH) can improve gastric stability of emulsions, but are prone to disintegration under intestinal conditions. This knowledge is important for designing food systems that control release of lipolytic products in targeted locations of the GI tract; the emulsions reported here are expected to be suitable for duodenal release.

Lipotoxic brain microvascular injury is mediated by activating transcription factor 3-dependent inflammatory and oxidative stress pathways

Dysfunction of the cerebrovasculature plays an important role in vascular cognitive impairment (VCI). Lipotoxic injury of the systemic endothelium in response to hydrolyzed triglyceride-rich lipoproteins (TGRLs; TGRL lipolysis products) or a high-fat Western diet (WD) suggests similar mechanisms may be present in brain microvascular endothelium. We investigated the hypothesis that TGRL lipolysis products cause lipotoxic injury to brain microvascular endothelium by generating increased mitochondrial superoxide radical generation, upregulation of activating transcription factor 3 (ATF3)-dependent inflammatory pathways, and activation of cellular oxidative stress and apoptotic pathways. Human brain microvascular endothelial cells were treated with human TGRL lipolysis products that induced intracellular lipid droplet formation, mitochondrial superoxide generation, ATF3-dependent transcription of proinflammatory, stress response, and oxidative stress genes, as well as activation of proapoptotic cascades. Male apoE knockout mice were fed a high-fat/high-cholesterol WD for 2 months, and brain microvessels were isolated by laser capture microdissection. ATF3 gene transcription was elevated 8-fold in the hippocampus and cerebellar brain region of the WD-fed animals compared with chow-fed control animals. The microvascular injury phenotypes observed in vitro and in vivo were similar. ATF3 plays an important role in mediating brain microvascular responses to acute and chronic lipotoxic injury and may be an important preventative and therapeutic target for endothelial dysfunction in VCI.

Supersaturation of zafirlukast in fasted and fed state intestinal media with and without precipitation inhibitors

Poor water solubility is a bottle neck in the development of many new drug candidates, and understanding and circumventing this is essential for a more effective drug development. Zafirlukast (ZA) is a leukotriene antagonist marketed for the treatment of asthma (Accolate®). ZA is poorly water soluble, and is formulated in an amorphous form (aZA) to improve its solubility and oral bioavailability. It has been shown that upon dissolution of aZa, the concentration of ZA in solution is supersaturated with respect to its stable crystalline form (ZA monohydrate), and thus, in theory, the bioavailability increases upon amorphization of ZA. The polymers hydroxypropylmethylcellulose (HPMC) and polyvinylpyrrolidone (PVP), often used as stabilizers of the supersaturated state, are in the excipient list of Accolate®. It is not recommended to take Accolate® with food, as this reduces the bioavailability by 40%.The aim of this study was to investigate the effect of simulated fasted and fed state intestinal media as well as the effect of HPMC and PVP on the supersaturation and precipitation of ZA in vitro.Supersaturation of aZA was studied in vitro in a small scale setup using the μDiss Profiler™. Several media were used for this study: One medium simulating the fasted state intestinal fluids and three media simulating different fed state intestinal fluids. Solid state changes of the drug were investigated by small angle x-ray scattering. The duration wherein aZA was maintained at a supersaturated state was prolonged in the presence of HPMC and lasted more than 20h in the presence of PVP in a fasted state intestinal medium. The presence of PVP increased the concentration of drug dissolved in the supersaturated state. The duration of supersaturation was shorter in fed than in a fasted state simulated intestinal media, but the concentration during supersaturation was higher. It was thus not possible to predict any positive or negative food effects from the dissolution/precipitation curves from different media. Lipolysis products in the fed state simulated media seemed to cause both a negative effect on the duration of supersaturation, and an increased drug concentration during supersaturation. In contrast, when testing the effect of a fed state simulated medium compared to the fasted state medium, in the presence of PVP, a clear negative effect was seen on the dissolution/precipitation curved of the fed state medium. The drug concentration during supersaturation was marginally different in the two media, but a precipitation of ZA was seen in the fed state medium, which was not observed in the fasted state medium. Solid state transformation from aZA to ZA monohydrate (mhZA) upon precipitation of the supersaturated solutions was confirmed by small angle x-ray scattering. All of these results can explain the described in vivo behavior of ZA. For ZA simple dissolution experiments in vitro can be used to examine supersaturation, effectiveness of PI and potential food effects on these.

Lipoprotein Lipase Modulates Bone Marrow Myeloid Cell Proliferation by Affecting Colony‐Stimulating Factor Levels and Recruitment

Although inflammation is the body's basic response to ameliorate injury or infection, tissue macrophage‐mediated pro‐inflammatory responses promote metabolic diseases, such as atherosclerosis. Lipoprotein lipase (LpL) is the rate‐limiting enzyme that hydrolyzes circulating lipoprotein triglycerides (TG) to release free fatty acids. We have reported that loss of LpL expression reduced numbers of aortic macrophages and bone marrow (BM) myelopoiesis. Reconstitution of LpL‐expressing BM was able to replenish aortic macrophage density. Our current studies are directed toward understanding the molecular regulation of LpL in orchestrating lipoprotein metabolism and immune cell biology. Our data show that reduced aortic macrophage levels were associated with reduced blood white cells and leukocyte subsets, including macrophage precursor cells‐ monocytes. BM‐derived monocyte differentiation to macrophages was markedly reduced by 40% in the absence of LpL (p<0.05). We speculate that defective lipid metabolism induced by LpL deficiency can contribute to decreased BM myelopoiesis. In transgenic LpL knockout mice that are rescued from neonatal death by muscle‐specific LpL expression (MCKL0 mice), loss of LpL reduced BM monocyte/macrophage precursor (BMMP) cell TG uptake and accumulation by 22% and 50% (p<0.05), respectively. TG utilization was much lower (~40%) in MCKL0 BMMP cells compared to wild type (WT) BMMP cells. LpL deficiency significantly decreased BM expression of both macrophage‐colony‐stimulating factor (M‐CSF) and its receptor (M‐CSF‐R), required for myeloid cell proliferation and differentiation (p<0.05). LpL‐mediated lipolysis products (e.g., oleic acid) were able to stimulate expression of M‐CSF and M‐CSF‐R by > 35% (p<0.05) in WT BMMP cells. In addition, LpL and heparin, which releases LpL from the cell surface, modulated binding of M‐CSF to M‐CSF‐R in the immunobinding assay (p<0.05). We hypothesize that LpL also anchors M‐CSF to its receptor and triggers a series of downstream signaling pathways to stimulate monocyte differentiation to macrophages. We thus propose that LpL, in addition to being a lipolytic enzyme, also influences peripheral and BM immune cell responses. These studies provide valuable insight into the regulation of adverse inflammatory responses and have therapeutic applications for various blood cell diseases.

On the role of bile salts in the digestion of emulsified lipids

The objective of this study was to understand quantitatively the role that bile salts play in the digestion of emulsified lipids. The behaviours of digestion by pancreatin (1.6 mg/mL) of sodium-caseinate-stabilized emulsions (0.5 wt% protein) and bile-extract-stabilized emulsions (0.2–5 mg/mL) as influenced by the addition of aqueous bile extract were studied under simulated intestinal conditions (37 °C; pH 7.5; 39 mM K2HPO4, 150 mM NaCl; with continuous agitation at ∼ 150 rev/min for 3 h). The droplet characteristics (size and ζ-potential) of the sodium caseinate- and bile extract-stabilized droplets were evaluated by light scattering techniques. The kinetics of the total fatty acids released by hydrolysis of the emulsified lipids were monitored by the pH-stat method with or without the presence of continuous phase bile extract. The results suggested that the presence of unadsorbed bile extract markedly enhanced the rate and the extent of lipid digestion. This could be attributed to considerable removal of lipolysis products (free fatty acids, mono- and/or di-acylglycerols) in mixed micelles, which are known to inhibit lipid digestion, by the unadsorbed bile salts. This study provides new insights for the lipid digestion of food formulations.

The role of plant cell wall encapsulation and porosity in regulating lipolysis during the digestion of almond seeds.

Previous studies have provided evidence that the physical encapsulation of intracellular nutrients by cell walls of plant foods (i.e. dietary fibre) plays a predominant role in influencing macronutrient bioaccessibility (release) from plant foods during human digestion. One unexplored aspect of this is the extent to which digestive enzymes can pass through the cell-wall barrier and hydrolyse the intracellular lipid in almond seeds. The purpose of the present study was to assess the role played by cell walls in influencing the bioaccessibility and digestibility of almond lipid using a range of techniques. Digestibility experiments were performed on raw and roasted almond cells as well as isolated almond oil bodies using in vitro gastric and duodenal digestion models. Residual triacylglycerols and lipolysis products were extracted after 1 h of incubation and analysed by thin layer chromatography. The lipolysis kinetics of almond cells and oil bodies were also investigated using the pH-stat technique. Finally, the potential penetration of pancreatic lipase through the cell wall matrix was investigated using confocal microscopy. Differences in the rates and extent of lipolysis were clearly seen between almond cells and oil bodies, and these differences were observed regardless of the lipase(s) used. These results also showed that almond cell walls that are completely intact limit lipid digestibility, due to an encapsulation mechanism that hinders the diffusion of lipase into the intracellular environment and lipolysis products out of the cells.

The In Vitro Lipolysis of Lipid-Based Drug Delivery Systems: A Newly Identified Relationship between Drug Release and Liquid Crystalline Phase.

The purpose of this study was to offer a new insight into the microstructure changes during in vitro lipolysis of five lipid-based drug delivery formulations belonging to different lipid formulation types. Five lipid-based formulations of indomethacin were investigated using an in vitro lipolysis model. During lipolysis, microstructures of the intermediate phase formed by lipolytic products were observed. The results showed that the time of liquid crystal formation during in vitro digestion for these formulations was Type I > Type II > Type IIIB > Type IV > Type IIIA (p < 0.05). After lipolysis, the drug releases from these formulations were determined. The results showed that the amount of drug distributed in the aqueous phase, obtained by ultracentrifuge after lipolysis, was, astonishingly, in inverse rank order of the above mentioned, that is, Type IIIA > Type IV > Type IIIB > Type II > Type I (p < 0.05). These results showed that the liquid crystalline phase probably has a critical influence on the fate of the drug during in vitro lipolysis and suggested that the liquid crystalline phase facilitated drug precipitation. These findings may improve the understanding of lipolysis of lipid-based drug delivery systems for designing better delivery system.

Porous nanostructure controls kinetics, disposition and self-assembly structure of lipid digestion products

Combining1H NMR and sSAXS to discriminate the speciation and structure evolution of lipolysis products for submicron lipid droplets and lipid loaded in porous silica particles.

TGRL Lipolysis Products Induce Stress Protein ATF3 via the TGF-β Receptor Pathway in Human Aortic Endothelial Cells.

Studies have suggested a link between the transforming growth factor beta 1 (TGF-β1) signaling cascade and the stress-inducible activating transcription factor 3 (ATF3). We have demonstrated that triglyceride-rich lipoproteins (TGRL) lipolysis products activate MAP kinase stress associated JNK/c-Jun pathways resulting in up-regulation of ATF3, pro-inflammatory genes and induction of apoptosis in human aortic endothelial cells. Here we demonstrate increased release of active TGF-β at 15 min, phosphorylation of Smad2 and translocation of co-Smad4 from cytosol to nucleus after a 1.5 h treatment with lipolysis products. Activation and translocation of Smad2 and 4 was blocked by addition of SB431542 (10 μM), a specific inhibitor of TGF-β-activin receptor ALKs 4, 5, 7. Both ALK receptor inhibition and anti TGF-β1 antibody prevented lipolysis product induced up-regulation of ATF3 mRNA and protein. ALK inhibition prevented lipolysis product-induced nuclear accumulation of ATF3. ALKs 4, 5, 7 inhibition also prevented phosphorylation of c-Jun and TGRL lipolysis product-induced p53 and caspase-3 protein expression. These findings demonstrate that TGRL lipolysis products cause release of active TGF-β and lipolysis product-induced apoptosis is dependent on TGF-β signaling. Furthermore, signaling through the stress associated JNK/c-Jun pathway is dependent on TGF-β signaling suggesting that TGF-β signaling is necessary for nuclear accumulation of the ATF3/cJun transcription complex and induction of pro-inflammatory responses.

Characterizing blood-brain barrier perturbations after exposure to human triglyceride-rich lipoprotein lipolysis products using MRI in a rat model.

PurposePrevious studies indicated hyperlipidemia may be a risk factor for Alzheimer's disease, but the contributions of postprandial triglyceride‐rich lipoprotein (TGRL) are not known. In this study, changes in blood–brain barrier diffusional transport following exposure to human TGRL lipolysis products were studied using MRI in a rat model.MethodsMale Sprague‐Dawley rats (∼180–250 g) received an i.v. injection of lipoprotein lipase (LpL)‐hydrolyzed TGRL (n = 8, plasma concentration ≈ 150 mg human TGRL/dL). Controls received i.v. injection of either saline (n = 6) or LpL only (n = 6). The 1H longitudinal relaxation rate R1 = 1/T1 was measured over 18 min using a rapid‐acquired refocus‐echo (RARE) sequence after each of three injections of the contrast agent Gd‐DTPA. Patlak plots were generated for each pixel yielding blood‐to‐brain transfer coefficients, Ki, chosen for best fit to impermeable, uni‐directional influx or bi‐directional flux models using the F‐test.ResultsAnalysis from a 2‐mm slice, 2‐mm rostral to the bregma showed a 275% increase of mean Ki during the first 20 min after infusion of human TGRL lipolysis product that differed significantly compared with saline and LpL controls. This difference disappeared by 40 min mark.ConclusionThese results suggest human TGRL lipolysis products can lead to a transient increase in rat BBB permeability. Magn Reson Med 76:1246–1251, 2016. © 2015 The Authors. Magnetic Resonance in Medicine published by Wiley Periodicals, Inc. on behalf of International Society for Magnetic Resonance in Medicine. This is an open access article under the terms of the Creative Commons Attribution NonCommercial License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited and is not used for commercial purposes.

Impact of colloidal structure of gastric digesta on in-vitro intestinal digestion of whey protein emulsion gels

Two whey protein emulsion gels containing 10 mM NaCl (soft gel) or 200 mM NaCl (hard gel) were formed and subjected to sequential digestion, including an in vitro gastric step followed by an in vitro intestinal step. The breakdown of gel particles, stability of oil droplets, microstructure and free fatty acid release during digestion were examined. The results showed that after 60 and 240 min gastric digestion the digesta of both gels had different physicochemical forms. The 60 min gastric digesta of both gels was mainly composed of gel particles ranging from ∼1 to ∼1000 μm. The 240 min gastric digesta of the soft gel mainly consisted of individual oil droplets and small gel particles (∼10 μm) while that of the hard gel mainly consisted of small gel particles (∼10 μm). During intestinal digestion, the breakup and coalescence of oil droplets occurred simultaneously for all gastric digesta. The remaining network structure of gel particles hindered their breakdown especially in the hard gel during intestinal digestion, which delayed lipid digestion significantly. Lipolytic products appeared to accumulate around oil droplet surfaces. Needle-shaped crystals that may be composed of free fatty acids were also found during intestinal digestion.

Elucidating the Molecular Interactions Occurring during Drug Precipitation of Weak Bases from Lipid-Based Formulations: A Case Study with Cinnarizine and a Long Chain Self-Nanoemulsifying Drug Delivery System.

The aim of this study was to investigate if molecular interactions between the weak base cinnarizine and lipolysis products were affecting the morphology of precipitated drug formed during in vitro lipolysis. In vitro lipolysis studies of a self-nanoemulsifying drug delivery system with or without cinnarizine were conducted. The digestion phases (aqueous phase and pellet phase) were separated by ultracentrifugation, and the pellet was isolated and lyophilized. The lyophilized pellets were examined by X-ray powder diffraction, (13)C solid-state nuclear magnetic resonance ((13)C NMR), (1)H liquid-state NMR ((1)H NMR) spectroscopy and differential scanning calorimetry (DSC). The (13)C NMR data indicated that the carbonyl groups and aliphatic part of the lipids undergo structural changes when the pellet contains cinnarizine. The (1)H NMR data suggests interactions occurring around the nitrogens on cinnarizine and the carboxylic group of fatty acids. DSC thermograms showed cinnarizine to be homogeneously incorporated into the lipids of the pellet, and no free amorphous cinnarizine was present. The three techniques (13)C NMR, (1)H NMR, and DSC complement each other and suggest interactions to occur between fatty acids and cinnarizine, which in turn favors amorphous precipitation.

Dynamic modeling of in vitro lipid digestion: Individual fatty acid release and bioaccessibility kinetics

The aim of this study was to gain knowledge about the role of triacylglycerol (TAG) composition in fatty acids (FA) of o/w emulsions on both the pancreatic lipolysis kinetics and the bioaccessibility of released products (i.e. contained within the bile salt micellar phase). A mathematical model was developed and its predictions were compared to a set of experimental data obtained during an in vitro digestion of a whey protein stabilized emulsion. Modeling results show that FA residues of TAG were hydrolyzed at specific rates, inducing different bioaccessibility kinetics. The estimated lipolysis rate constants of the studied FA (C8:0, C10:0≫C18:1 n-9≫C12:0>C14:0>C16:0≈C16:1 n-7>C22:6 n-3) were in close agreement with the available literature on the substrate specificity of pancreatic lipase. Results also suggest that lipolysis products are very rapidly solubilized in the bile salt mixed micelles with no fractionation according to the FA carbon chain.