J. Physiol. Biochem., 59 (3),193-200,2003. Effects of conjugated linoleic acid on body fat accumulation and serum lipids in hamsters fed an atherogenic diet.
J. Physiol. Biochem., 59 (3),193-200,2003
Effects of conjugated linoleic acid on body fat accumulation and serum lipids in hamsters fed an atherogenic diet V. Navarro, A. Zabala, M. T. Macarulla, A. Fernandez-Quintela, V. M. Rodriguez, E. Simon and M. P. Portillo Department of Nutrition and Food Science, University of the Basque Country, Paseo de la Universidad 7, 01006 Vitoria (Spain) (Received on July 21, 2003)
V. NAVARRO, A. ZABALA, M. T. MACARULLA, A. FERNANDEZQUINTELA, V. M. RODRIGUEZ, E. SIMON and M. P. PORTILLO. Effects of conjugated linoleic acid on body fat accumulation and serum lipids in hamsters fed an atherogenic diet. J. Physiol. Biochem., 59 (3), 193-1200, 2003. Conjugated linoleic acid (CLA) refers to a mixture of naturally occurring positional and geometric isomers of linoleic acid that exist in dairy products and meat. The aim of the present work was to study the effects of c-9,t-ll and t-1 0,c-12 CLA isomers on body fat accumulation and serum lipids in hamsters fed an atherogenic diet. Hamsters were divided in four groups: one group was fed a chow diet (control) and the other three groups were given semi-purified atherogenic diets with 0.5% linoleic acid (LA), c-9,t-11 or t-10,c-12 CLA. Body weight and food intake were measured daily. After 6 weeks, adipose tissues from different anatomical locations and liver were dissected and weighed. Serum glucose, total cholesterol, HDL-c, LDL-c and triacylglycerollevels, as well as total and free cholesterol, triacylglycerol and phospholipid content in liver were determined by enzymatic methods. No differences in either energy intake or final body weight were found. The addition of t10,c-12 CLA reduced fat accumulation and led to lower serum cholesterol, as compared with LA group. Nevertheless the level remained higher than in the control animals. The reduction in serum cholesterol was limited to LDL-c. This isomer also reduced triacylglycerol content in liver but did not modify serum triacylglycerol level. In summary, the present study demonstrates that t-10,c-12 CLA is the biologically active agent when anti-obesity and hypocholesterolaemic properties of CLA are considered. In contrast, the isomer c-9,t-11 has no effect on lipid metabolism in hamsters. Key words: Conjugated linoleic acid,Body fat, Cholesterol,Triacylglycerols, Hamster.
Correspondence to M. P. Portillo (Tel.: +34945013 067; Fax: +34 945 013 014; e-mail: knppboamtsvc.ehu.es),
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Conjugated linoleic acid (CLA) refers to a mixture of naturally occurring positional (9 and 11 or 10 and 12) and geometric (cis or trans) isomers of linoleic acid (LA, 18:2 n-S) that exist in dairy products and meat (19). The principal isomers in commercial available mixtures are c-9,t-ll and t-l0,c-12. The former is the principal form of CLA found in the meat and milk of ruminant animals (5). A crude mixture of CLA isomers has been shown to have a variety of potential health benefits in both animals and cell culture models. These fatty acids are effective anticarcinogens, anti diabetics and potent modulators of immune function (20). Feeding diets containing CLA has been associated with marked changes in lipid metabolism. Rodents fed diets containing 0.5-1.2% CLA have enhanced feed efficiency, reduced body fat accumulation and increased lean body mass, in a dose response manner (2, 8, 9, 13, 25, 27). It should be emphasized that comparisons among studies on the effects of CLA on body fat in rodents, pigs or humans have sometimes led to contradictory conclusions. These discrepancies might originate from differences in gender, age, duration of supplementation or species responsiveness. Several authors have found that CLA also improves plasma lipoprotein metabolism and inhibits the progression of atherosclerosis (15, 16, 18). Nevertheless, studies on cholesterol metabolism are inconclusive. Some studies have shown that only specific CLA isomers are responsible for physiological effects (17). Most of the experimental results have been obtained by using different brands of CLA mixtures, which may greatly differ in their CLA isomer distribution and complexity (6). J. Physiol. Biochem., 59 (3), 2003
The aim of the present work was to study the effects of the two main isomers of conjugated linoleic acid, c-9,t-ll and t10,c-12, on body fat accumulation and serum lipids in hamsters fed an atherogenic diet. Materials and Methods
Animals, diets and experimental design.- Thirty-two, 9-week-old, male Syrian Golden hamsters (105 ± 1 g) were Harlan Iberica purchased from (Barcelona, Spain). They were individually housed in polycarbonate metabolic cages (Techniplast Gazzada, Guguggiate, Italy) and placed in an air conditioned room (22 ± 2 "C) with a 12 hours daynight rhythm. After a 6-d adaptation period, hamsters were randomly divided into four groups of 8 animals each and fed the experimental diets for 6 weeks. One group was fed a commercial chow diet (Panlab, Barcelona, Spain). The other three groups were given semi-purified atherogenic diets consisting of 200 g/kg casein and 4 g/kg L-methionine (Sigma, St. Louis, MO, USA), 200 g/kg wheat starch (Vencasser, Bilbao, Spain), 404 g/kg sucrose (local market), 100 g/kg palm oil (Agra, Leioa, Spain), 30 g/kg cellulose (Vencasser, Bilbao, Spain) and 1 g/kg cholesterol (Sigma, St. Louis, MO, USA). Linoleic acid (from sunflower oil; LA group), c-9,t-ll conjugated linoleic acid or t-l0,c-12 conjugated linoleic acid (Natural Lipids Ltd., Hovdebygda, Norway) were added at a level of 0.5% to the atherogenic diets. Vitamin and mineral mixes were formulated according to AIN-93 guidelines (21) and supplied by ICN Pharmaceuticals (Costa Mesa, CA, USA). All animals had free access to food and water. Food intake was measured daily and body weight was recorded twice per week.
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Tissue removal and serum sampling.At the end of the experimental period hamsters were fasted 12 h and blood samples were collected under inhalation anaesthesia (diethyl ether) by cardiac puncture. Adipose tissue from different anatomical regions (interscapular, perirenal, epididymal and gluteal subcutaneous) and liver were dissected and weighed. Serum was obtained from blood samples after centrifugation (1,000 g for 10 min at 4 "C) and stored at -80°C until analysis. Liver lipids.- Hepatic lipids were extracted with chloroform-methanol (2:1) using the method of Folch et al. (11). The lipid extract was dissolved in isopropanol. Lipids were measured by using enzymatic analysis: a GPO-PAP kit for triglycerides (Spinreact, Sant Esteve de Bas, Spain), Choline-PAP kit for phospholipids (Spinreact, Sant Esteve de Bas, Spain) and BioSystems kit for free and total cholesterol. Total cholesterol was quantified by an enzymatic method (22), free cholesterol was analyzed by the same method, but lacking esterase enzyme, and esterified cholesterol was estimated as the difference between the total and the free cholesterol.
Serum analyse>- Serum glucose, total cholesterol, HDL-cholesterol and LDLcholesterol were measured using BioSystems kits (Barcelona, Spain) and triacylglycerols were measured using a GPOPAP kit (Spinreact, Sant Esteve de Bas, Spain). VLDL-c was calculated as the difference between total cholesterol and (HDL+LDL)-c. Statistical analysis.-Results are presented as means ± SEM. Statistical analysis was performed using SPSS 8.0 (SPSS Inc. Chicago, IL, USA). Data were analysed by one-way ANOVA followed by a N ewrnan- Keuls post-hoc test. Significance was assessed at the P
