Dietary proteins modulate high-density lipoprotein characteristics in a sex-specific way in Apoe-deficient mice

Objectives: The type and amount of dietary protein have become a topic of renewed interest, considering their involvement in several diseases. However, little attention has been devoted to the effect of avian proteins despite their wide human consumption. In a previous study, we saw that compared with soybean protein, the consumption of avian proteins, depending on sex, resulted in similar or lower atherosclerosis with a higher paraoxonase 1 activity, an antioxidant enzyme carried by high-density lipoproteins (HDL). This suggests that under these conditions, the HDL lipoproteins may undergo important changes. The aim of this research was to study the influence of soybean, chicken, and turkey proteins on the characteristics of HDL. Methods: Male and female Apoe-deficient mice were fed purified Western diets based on the AIN-93 diet, differing only in the protein source, for 12 wk. After this period, blood and liver samples were taken for analysis of HDL composition and hepatic expression of genes related to HDL metabolism (Abca1, Lcat, Pltp, Pon1, and Scarb1). Depending on sex, these genes define a different network of interactions. Females consuming the turkey protein–containing diet showed decreased atherosclerotic foci, which can be due to larger very-low-density lipoproteins (VLDLs) calculated by molar ratio triacylglycerols/VLDL cholesterol and higher expression of Lcat. In contrast, in males, a higher ratio of paraoxonase1 to apolipoprotein A1 decreased the oxidative status of the different lipoproteins, and augmented Abca1 expression was observed. Conclusions: The source of protein has an effect on the development of atherosclerosis depending on sex by modifying HDL characteristics and the expression of genes involved in their properties

Pulsed electric field increases the extraction yield of extra virgin olive oil without loss of its biological properties

Introduction: Pulsed electric field (PEF) has been used for improving extraction of extra virgin olive oil (EVOO). However, the biological changes induced by the consumption of pulsed electric field-obtained extra virgin olive oil (PEFEVOO) have not been studied yet. Materials and methods: EVOO oils from Empeltre variety were prepared by standard (STD) cold pressure method involving crushing of the olives, malaxation and decanting and by this procedure including an additional step of PEF treatment. Chemical analyses of EVOO oils were done. Male and female Apoe-deficient mice received diets differing in both EVOOs for 12 weeks, and their plasma, aortas and livers were analyzed. Results: PEF application resulted in a 17% increase in the oil yield and minimal changes in chemical composition regarding phytosterols, phenolic compounds and microRNA. Only in females mice consuming PEF EVOO, a decreased plasma total cholesterol was observed, without significant changes in atherosclerosis and liver steatosis. Conclusion: PEF technology applied to EVOO extraction maintains the EVOO quality and improves the oil yield. The equivalent biological effects in atherosclerosis and fatty liver disease of PEF-obtained EVOO further support its safe use as a food

Hepatic galectin-3 is associated with lipid droplet area in non-alcoholic steatohepatitis in a new swine model

Non-alcoholic fatty liver disease (NAFLD) is currently a growing epidemic disease that can lead to cirrhosis and hepatic cancer when it evolves into non-alcoholic steatohepatitis (NASH), a gap not well understood. To characterize this disease, pigs, considered to be one of the most similar to human experimental animal models, were used. To date, all swine-based settings have been carried out using rare predisposed breeds or long-term experiments. Herein, we fully describe a new experimental swine model for initial and reversible NASH using cross-bred animals fed on a high saturated fat, fructose, cholesterol, cholate, choline and methionine-deficient diet. To gain insight into the hepatic transcriptome that undergoes steatosis and steatohepatitis, we used RNA sequencing. This process significantly up-regulated 976 and down-regulated 209 genes mainly involved in cellular processes. Gene expression changes of 22 selected transcripts were verified by RT-qPCR. Lipid droplet area was positively associated with CD68, GPNMB, LGALS3, SLC51B and SPP1, and negatively with SQLE expressions. When these genes were tested in a second experiment of NASH reversion, LGALS3, SLC51B and SPP1 significantly decreased their expression. However, only LGALS3 was associated with lipid droplet areas. Our results suggest a role for LGALS3 in the transition of NAFLD to NASH

Aceite de oliva virgen, el alimento con propiedades farmacológicas preventivas de la aterosclerosis

Cardiovascular diseases (including atherosclerosis) are the main cause of death in Spain and developed countries. Their association, during decades, with Western diet (high fat and cholesterol) resulted in the recommendation of low-fat diets, which prejudiced basic ingredients of the traditional Mediterranean Diet such as olive oil and nuts. However, conclusive studies not only disprove this theory, but, on the contrary, clearly confirm that the adherence to this diet, in which the extra virgin olive oil is the main source of fat, reduces cardiovascular diseases risk factors and death. In our laboratory, we have carried out several experiments using apoE-deficient mouse, an animal model that reproduces the development of human atherosclerosis, and, as such, allowed to study the effects of the different washing and refining steps that olive oil goes through, and of the physical processes (pressure or centrifugation) that produce oils with different content in minors compounds. Also, some studies have been carried out on the isolated effect of some of these minor compounds, such as hydroxytyrosol or squalene, on the development of atherosclerotic lesions. The protective properties of virgin olive oil against atherosclerosis can be partly attributed to the presence of these minor components, whose biological effect fades or disappears in processed oils that have eliminated them.Las enfermedades cardiovasculares (entre las que se incluye la aterosclerosis) representan la principal causa de muerte en España y en los países desarrollados. Durante décadas se las ha asociado con dietas de tipo occidental (ricas en grasas y colesterol), por lo que se han recomendado dietas bajas en grasa en las que se han visto perjudicados alimentos como el aceite de oliva y los frutos secos, ingredientes constituyentes de la tradicional dieta mediterránea. Sin embargo, estudios concluyentes de investigación no solo rechazan esta teoría, sino que confirman categóricamente que la adhesión a este tipo de dieta, en la que el aceite de oliva virgen extra es la principal fuen-te de grasas, reduce los factores de riesgo y las muertes por este tipo de enfer-medades. En nuestro laboratorio se han llevado a cabo una serie de experi-mentos empleando el ratón carente de apolipoproteína E, modelo animal que semeja el desarrollo de aterosclerosis humana, sobre el que se ha investigado el efecto de distintos procesos a que han sido sometidos los aceites de oliva (refinados, lavados) y de diferentes tratamientos físicos (prensado o centrifu-gado) con los que se han obtenido aceites con distinto contenido en compuestos minoritarios. También se ha investigado el efecto de algunos de estos com-puestos minoritarios aislados, como el hidroxitirosol o el escualeno, sobre el desarrollo de la lesión aterosclerótica. Las propiedades protectoras de la ate-rosclerosis del aceite de oliva virgen pueden atribuirse en parte a la presen-cia de esos componentes minoritarios y su efecto biológico se reduce o desa-parece en los aceites procesados que hayan eliminado dichos componente

Microarray analysis of hepatic genes differentially expressed in the presence of the unsaponifiable fraction of olive oil in apolipoprotein E-deficient mice

The hypothesis that the unsaponifiable fraction of olive oil dramatically influences hepatic gene expression was tested in mice. Two olive oils, obtained from the same olive cultivar but by different technological procedures, were characterized to show that they differed mainly in terms of the composition/quantity of this unsaponifiable fraction. Using DNA microarrays, hepatic gene expression was analysed in apoE-deficient mice fed one of two isoenergetic, isonitrogenous diets containing either 10% (w/w) olive oil or unsaponifiable fraction-enriched olive oil. To provide an initial screening of potential candidate genes involved in a differential response, only genes with remarkably modified expression (signal log2 ratio ≥3 or < -3) were further considered. The eleven genes fulfilling these prerequisites were confirmed by quantitative RT-PCR, and then analysed in apoE-deficient mice with a C57BL/6J genetic background. Orosomucoid and serum amyloid A2 were upregulated (to variable extents depending on the genetic background) in the absence of hepatic steatosis and inflammation. Fabp5 and Mt2 were also strongly upregulated. Several proteases were highly suppressed by the unsaponifiable-enriched olive diet, independent of the genetic background. The findings indicate that change in the expression of these genes is a good marker of the intake of the unsaponifiable fraction of olive oil. The results highlight the important biological effects of the unsaponifiable fraction of olive oil. The term 'monounsaturated fatty acid-enriched oil' no longer appears appropriate for describing all the oils to which it is currently applied since it does not adequately reflect that they have different biological effects. © The Authors 2007.This research was supported by grants FEGA-FEOGA (CAO99-014), Ministerio de Educación y Ciencia, CICYT (SAF2004-08 173-C03-02 and AGL2005-00 572), Junta de Andalucía (CAO01-002), FISS 01/0202, Redes FISS de investigación cooperativa C03-01 and G03-140 and by the Fundación Española del Corazón.Peer Reviewe

Apolipoprotein E determines the hepatic transcriptional profile of dietary maslinic acid in mice

The hypothesis that the maslinic acid (MA) of olive oil (OO) dramatically influences hepatic gene expression was tested in mice. Two OOs only differing in the presence of MA were prepared. Using DNA microarrays, we analyzed hepatic gene expression in apolipoprotein E (apoE)-deficient mice with a C57BL/6J genetic background that were fed with isocaloric, isonitrogenous diets containing either 10% (w/w) OO or 10% MA-enriched OO. As an initial screening of potential candidate genes involved in a differential response, this study further considered only genes with remarkably modified expression (signal log2 ratio higher than1.5 or lower than −1.5). The nine genes fulfilling these prerequisites were confirmed by quantitative reverse transcriptase polymerase chain reaction and analyzed in C57BL/6J wild-type mice. Only Cyp2b9, Cyp2b13 and Dbp expressions appeared significantly increased, and Marco was significantly decreased in apoE-deficient mice receiving the MA-enriched diet. Dbp was up-regulated to an extent depending on the genetic background of the mice and negatively associated with the expression of Marco, a gene strongly up-regulated by the absence of apoE. These expression changes could be used as markers of the intake of the MA-enriched OO and are influenced by genetic background generated by the absence or the presence of apoE. Overall, these results (a) indicate that MA in virgin OO is highly active in controlling hepatic gene expression and (b) highlight the important interaction between the response to MA and the presence of apoE. They also confirm that virgin OO cannot be simplistically classified as monounsaturated fatty-enriched oil without paying attention to its active minor components.This research was supported by grants from FEDER-CICYT (SAF2007-60173, AGL2005-00572 and AGL2008-02285/ALI), Redes DGA (B-69) and FISS de Investigación Cooperativa (C03-01 and G03-140) and by Fundación Española del Corazón and Instituto Aragonés de Ciencias de la Salud. S.A. and N.G. were recipients of DGA and Fundación Cuenca Villoro fellowships.Peer reviewe

Dietary oleanolic acid mediates circadian clock gene expression in liver independently of diet and animal model but requires apolipoprotein A1

Oleanolic acid is a triterpene widely distributed throughout the plant kingdom and present in virgin olive oil at a concentration of 57 mg/kg. To test the hypotheses that its long-term administration could modify hepatic gene expression in several animal models and that this could be influenced by the presence of APOA1-containing high-density lipoproteins (HDLs), diets including 0.01% oleanolic acid were provided to Apoe- and Apoa1-deficient mice and F344 rats. Hepatic transcriptome was analyzed in Apoe-deficient mice fed long-term semipurified Western diets differing in the oleanolic acid content. Gene expression changes, confirmed by reverse transcriptase quantitative polymerase chain reaction, were sought for their implication in hepatic steatosis. To establish the effect of oleanolic acid independently of diet and animal model, male rats were fed chow diet with or without oleanolic acid, and to test the influence of HDL, Apoa1-deficient mice consuming the latter diet were used. In Apoe-deficient mice, oleanolic acid intake increased hepatic area occupied by lipid droplets with no change in oxidative stress. Bmal1 and the other core component of the circadian clock, Clock, together with Elovl3, Tubb2a and Cldn1 expressions, were significantly increased, while Amy2a5, Usp2, Per3 and Thrsp were significantly decreased in mice receiving the compound. Bmal1 and Cldn1 expressions were positively associated with lipid droplets. Increased Clock and Bmal1 expressions were also observed in rats, but not in Apoa1-deficient mice. The core liver clock components Clock-Bmal1 are a target of oleanolic acid in two animal models independently of the diets provided, and this compound requires APOA1-HDL for its hepatic action

Effect of squalene on lipoproteins from the three experimental models.

<p>Plasma was obtained following 11 weeks of consuming control or squalene- enriched semipurified diets and after a four-hour fast. Two independent pools of all mice per experimental group were prepared, except for <i>Apoe</i>-deficient mice, where three plasma pools were utilized. Lipoproteins were separated by FPLC, and collected fractions analyzed for total cholesterol (A, G, L), esterified cholesterol (B, H, M), phosphatidylcholine (C, I, N), sphingomyelin (D, J, O), APOA1 (E, P) and APOA4 (F, K, Q). Representative profiles are shown from WT mice, left panels (control and squalene pools consisting of plasma from 6 and 7 mice, respectively), <i>Apoa1</i>-deficient mice, middle panels (n = 7 for control and n = 7 for squalene plasma pool) and Apoe-deficient mice, right panels (n = 13 for control, n = 13 for 0.25 g/kg and n = 14 for 1 g/kg squalene plasma pool). Fraction numbers 1–6 corresponded to VLDL/chylomicron remnants, 7–13 to low density lipoproteins, 14–21 to cholesterol-rich HDL and 22–27 to cholesterol-poor HDL (pHDL).</p

Effect of the experimental diets on somatic variables in male mice of the three experimental models.

<p>Results are shown as mean values with their standard deviations. Statistical analysis was carried out using Mann Whitney U test.</p><p>* p<0.05 <i>vs</i> control.</p