Effect of Echium oil compared with marine oils on lipid profile and inhibition of hepatic steatosis in LDLr knockout mice

Botelho et al. Lipids in Health and Disease 2013, 12:38 http://www.lipidworld.com/content/12/1/38 RESEARCH Open Access Effect of Echium oil compared with marine oils on lipid profile and inhibition of hepatic steatosis in LDLr knockout mice Patrícia Borges Botelho1, Karina da Rocha Mariano1, Marcelo Macedo Rogero2 and Inar Alves de Castro1* Abstract Background: In an effort to identify new alternatives for long-chain n-3 polyunsaturated fatty acids (LC n-3 PUFA) supplementation, the effect of three sources of omega 3 fatty acids (algae, fish and Echium oils) on lipid profile and inflammation biomarkers was evaluated in LDL receptor knockout mice. Methods: The animals received a high fat diet and were supplemented by gavage with an emulsion containing water (CON), docosahexaenoic acid (DHA, 42.89%) from algae oil (ALG), eicosapentaenoic acid (EPA, 19.97%) plus DHA (11.51%) from fish oil (FIS), and alpha-linolenic acid (ALA, 26.75%) plus stearidonic acid (SDA, 11.13%) from Echium oil (ECH) for 4 weeks. Results: Animals supplemented with Echium oil presented lower cholesterol total and triacylglycerol concentrations than control group (CON) and lower VLDL than all of the other groups, constituting the best lipoprotein profile observed in our study. Moreover, the Echium oil attenuated the hepatic steatosis caused by the high fat diet. However, in contrast to the marine oils, Echium oil did not affect the levels of transcription factors involved in lipid metabolism, such as Peroxisome Proliferator Activated Receptor α (PPAR α) and Liver X Receptor α (LXR α), suggesting that it exerts its beneficial effects by a mechanism other than those observed to EPA and DHA. Echium oil also reduced N-6/N-3 FA ratio in hepatic tissue, which can have been responsible for the attenuation of steatosis hepatic observed in ECH group. None of the supplemented oils reduced the inflammation biomarkers. Conclusion: Our results suggest that Echium oil represents an alternative as natural ingredient to be applied in functional foods to reduce cardiovascular disease risk factors. Keywords: Atherosclerosis, Inflammation, Echium, Stearidonic, Omega 3, Steatosis Background The increased intake of omega-6 fatty acids during the 20th century as a result of an elevation in vegetal oil consumption (of more than 1,000-fold) contributed to a decline in the tissue concentration of long-chain n-3 polyunsaturated fatty acids (LC n-3 PUFA) [1,2], which might be associated with the increased incidence of inflammatory disorders, such as atherosclerosis [3]. The development of atherosclerotic plaques is associated with several clinical cardiovascular events. Considering the health effects of LC n-3 PUFA toward the reduction * Correspondence: 1 LADAF. NAPAN. Department of Food and Experimental Nutrition, Faculty of Pharmaceutical Sciences, University of São Paulo, Av. Lineu Prestes, 580, São Paulo B14 - 05508-900, Brazil Full list of author information is available at the end of the article of cardiovascular disease (CVD) risk [4,5], many industries have added eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) from marine oils to food formulations or supplements, aiming to explore this health claim. In 2004, the Food and Drug Administration (FDA) qualified the health claim of products containing EPA and DHA [6]. A similar recommendation was also provided by the American Heart Association (AHA), who suggested consumption of 1 g/day of EPA + DHA for patients with CVD and 2–4 g/day for patients with hypertriglyceridaemia [7]. The cardioprotective effects of LC N-3 PUFA appear to be due to a synergism between multiple mechanisms including triacylglycerol (TG) lowering, improving membrane fluidity, anti-inflammatory, antiarrhythmic and © 2013 Botelho et al.; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Botelho et al. Lipids in Health and Disease 2013, 12:38 http://www.lipidworld.com/content/12/1/38 antithrombotic effects [5]. The scientific evidence concerning the beneficial effects of the LC N-3 PUFA on lipid profile and inflammation were obtained from several studies using animal and human models. However, these effects and the mechanisms by which they occur are restricted to the action of EPA and DHA [8]. Other non-marine sources of omega 3 fatty acids (N-3 FA), such as alpha- linolenic acid (ALA) or stearidonic acid (SDA), can be converted in vivo to EPA and DHA by the desaturase and elongase enzymes in a tissue-dependent manner, the liver being the major site of this conversion [9]. It has been reported that the conversion rate of ALA is low (5-10% for EPA and < 1% for DHA), which diminishes the efficacy of these alternative sources in the reduction of cardiovascular risk [10-12]. However, due to dietary preferences, safety, sustainability, cost and oxidative stability aspects, other non-marine oils alternatives must be evaluated [3,9-11,13,14]. It has been suggested that the low rate by which ALA is converted to EPA is a result of the limited activity of Δ6-desaturase when linoleic acid (LNA) is also present [15]. However SDA, a precursor of EPA that is found in plants, such as Echium (Echium plantagineum), black currant seed and other genetically modified seeds, does not need Δ6-desaturase activity to be converted into EPA [3,10]. In an effort to identify new alternatives for LC N-3 PUFA supplementation, the objective of this study was to compare the effects of three sources of N-3 FA (algae, fish and Echium oil) on lipid composition and some inflammatory biomarkers using LDL receptor deficient mice (LDLr knockout mice) as model. Methods Oils and reagents The N-3 FA used in this study were commercial products: the algae oil containing 40% DHA (DHASCO) was obtained from Martek BiosciencesW (Winchester, KY, USA), the fish oil (EPA1T1600 MEG-3™) containing EPA (20%) + DHA (12%) was obtained from Ocean NutritionW (Dartmouth, NS, Canada) and the Echium oil containing 11.5% SDA (AW39144ECH) was obtained from Oil Seed ExtractionW (Ashburton, New Zealand). All reagents were purchased from Sigma Chemical Co. (St. Louis, MO, USA), Merck (Darmstadt, Germany), Calbiochem Technology Inc. (Boston, MA, USA) and GE Healthcare (Little Chalfont, Bucks, UK). The aqueous solutions were prepared with ultra-pure Milli-Q water (Millipore Ind. Com. Ltd., SP, Brazil), and the organic solvents were of HPLC grade. Fatty acids profile of the oils used in this study was analyzed by gas chromatography and is shown in Table 1. Animals and diets Forty male homozygous LDL receptor-deficient mice (LDLr Knockout mice, C57BL/6) weighing 25–29 g Page 2 of 10 Table 1 Fatty acids composition of the edible oils applied in this study Fatty acids (g/100 g total FA) Fish oil Alg (...truncated)