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Metabolic differences in hepatocytes of obese and lean pigs
- L. González-Valero, J. M. Rodríguez-López, M. Lachica, I. Fernández-Fígares
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There are important differences in terms of metabolic activity, energy utilization and capacity of protein and fat deposition when Iberian and modern pigs are compared. Primary culture of hepatocytes was used to evaluate hepatic function and sensitivity to hormones between breeds without the interference of circulating blood factors. Hepatocytes were isolated from pure Iberian (n=10) and Landrace (n=8) pigs of similar BW (24.5±12.1 and 32.9±6.1 kg BW, respectively), by collagenase perfusion. Monolayers were established in medium containing fetal bovine serum for 1 day and switched to serum-free medium for the remainder of the culture period. Hepatocytes were maintained in William’s E supplemented with β-mercaptoethanol (0.1 mM), glutamine (2 mM), antibiotics (gentamicin, penicillin, streptomycin and amphotericin B), dimethyl sulfoxide (1 µg/ml), dexamethasone (10−8 M), insulin (0.173 and 17.3 nM) and glucagon (0.287, 2.87 and 28.7 nM) for 24 to 48 h. Gluconeogenesis (GNG), glycogen degradation, triglycerides (TG) content and esterification, β-hydroxybutyrate (BHB) synthesis, IGF-1 synthesis, albumin and urea synthesis were determined. Iberian pigs had greater capacity of GNG than Landrace (24%, P<0.05), although no difference in glycogen degradation was found (P>0.10). TG content and esterification tended to be lower in hepatocytes from Iberian compared with Landrace pigs (12% and 31%, respectively; 0.10<P<0.05). Furthermore, addition of free fatty acids (CLA or linoleic acid, 0.2 mM) increased TG content (64%, P<0.001) although no difference between fatty acids was found. When free fatty acids were compared, a trend toward increased esterification (41%, P=0.078) was found for CLA. Although glucagon stimulated and insulin inhibited BHB synthesis, no difference between breeds was found (P>0.10). IGF-1 synthesis was diminished in hepatocytes from Iberian compared with Landrace pigs (16%, P<0.05). On the contrary, rate of albumin synthesis was greater in Iberian compared with Landrace pigs (58%, P<0.05). Finally, the capacity of urea synthesis was lower in hepatocytes of Iberian compared with Landrace pigs (37%, P<0.05). When ammonia was added to the media, urea concentration increased (648%, 1108% and 2791% when 0 mM was compared with 2.5, 5 and 10 mM, respectively). Urea synthesis increased on increasing ammonia content (55% and 325% when 0 mM was compared with 5 and 10 mM, respectively; P<0.0001). In conclusion, the genetic background accounts for important differences in protein and energy metabolism pathways found in primary culture of hepatocytes from lean and obese pigs.
Approaches for quantifying gastrointestinal nutrient absorption and metabolism in a native and a modern pig breed
- J. M. RODRÍGUEZ-LÓPEZ, M. LACHICA, L. GONZÁLEZ-VALERO, I. FERNÁNDEZ-FÍGARES
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- Journal:
- The Journal of Agricultural Science / Volume 151 / Issue 3 / June 2013
- Published online by Cambridge University Press:
- 15 August 2012, pp. 434-443
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When working with multi-catheterized animals, success and failure are separated by surgical procedures and minor details in catheter design and care. The current paper is a detailed description of novel approaches to multi-catheterization of pigs for investigations into nutrient absorption and metabolism of portal-drained viscera (PDV) in a native obese (Iberian) and a modern (Landrace) breed. Three Iberian and three Landrace gilts (25 kg average body weight; BW) were fitted with catheters in the carotid artery (CA), the portal vein (PV) and the ileal vein (IV). Tygon rings were attached to the catheter to mark the extent of introduction into the vessel and facilitate its fixing by means of a non-absorbable suture. The PV was catheterized through the visceral side of the left-lateral lobe of the liver and IV through a branch of the vein. The CA was secured directly in place with a purse-string suture where the artery was not occluded. Patency of the catheters was checked weekly and catheters filled with sterile heparinized saline and closed by two knots. Portal blood flow was determined to test the procedures. A 15 ml pulse dose of para-aminohippuric acid (PAH; 2% w/v) was infused into IV 45 min prior to blood sampling, followed by continuous infusion of 0·8 ml/min. Blood samples (4·5 ml) were taken simultaneously from CA and PV, using heparinized tubes, 5 min before feeding 0·25 of the total daily ration (barley–soybean meal diet; 160 g crude protein (CP)/kg; 14–14·5 MJ metabolizable energy (ME)/kg dry matter (DM); 2·4×ME for maintenance), and every 30 min for 4 h and then hourly until 6 h after feeding. Blood was centrifuged and plasma harvested and stored at −20 °C until PAH analysis. Whole-blood flow was based on the Fick principle. Post-prandial PDV blood flow was lower for the Iberian pigs than Landrace (866 and 1464 ml/min, respectively). The concurrence of access to the PV through the liver with a minimal wound, the non-occluded blood flow in CA, and the catheter design and care were all critical for the fast recovery of pigs and catheter patency. The procedures followed are recommended for studies of absorption of nutrients from the gastrointestinal tract and the impact of PDV on the metabolism of conscious, unrestrained, growing pigs.
Impact of dietary betaine and conjugated linoleic acid on insulin sensitivity, protein and fat metabolism of obese pigs
- I. Fernández-Fígares, M. Lachica, A. Martín, R. Nieto, L. González-Valero, J. M. Rodríguez-López, J. F. Aguilera
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To determine possible mechanisms of action that might explain the nutrient partitioning effect of betaine and conjugated linoleic acid (CLA) in Iberian pigs and to address potential adverse effects, twenty gilts were restrictively fed from 20 to 50 kg BW Control, 0.5% betaine, 1% CLA or 0.5% betaine + 1% CLA diets. Serum hormones and metabolites profile were determined at 30 kg BW and an oral glucose test was performed before slaughter. Pigs were slaughtered at 50 kg BW and livers were obtained for chemical and histological analysis. Decreased serum urea in pigs fed betaine and betaine + CLA diets (11%; P = 0.0001) indicated a more efficient N utilization. The increase in serum triacylglycerol (58% and 28%, respectively; P = 0.0098) indicated that CLA and betaine + CLA could have reduced adipose tissue triacylglycerol synthesis from preformed fatty acids. Serum glucose, low-density lipoprotein (LDL) cholesterol and non-esterified fatty acids were unaffected. CLA and betaine + CLA altered serum lipids profile, although liver of pigs fed CLA diet presented no histopathological changes and triglyceride content was not different from Control pigs. Compared with controls, serum growth hormone decreased (20% to 23%; P = 0.0209) for all treatments. Although serum insulin increased in CLA, and especially in betaine + CLA pigs (28% and 83%; P = 0.0001), indices of insulin resistance were unaffected. In conclusion, CLA, and especially betaine + CLA, induced changes in biochemical parameters and hormones that may partially explain a nutrient partitioning effect in young pigs. Nevertheless, they exhibited weak, although detrimental, effects on blood lipids. Moreover, although livers were chemically and histologically normal, pigs fed CLA diet challenged with a glucose load had higher serum glucose than controls.