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Foetal life protein provision of mink (Neovison vison) changes the relative mRNA abundance of some hepatic enzymes regulating fat metabolism a

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Connie Frank Matthiesen , Maria Arantzazu Aguinaga Casañas Anne-Helene Tauson

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Department of Veterinary Clinical and Animal Sciences, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, Denmark b

Department of Physiology and Biochemistry of Animal Nutrition, Animal Nutrition Institute, Estación Experimental del Zaidín (CSIC), Armilla, Granada, Spain Published online: 20 Mar 2014.

To cite this article: Connie Frank Matthiesen, Maria Arantzazu Aguinaga Casañas & Anne-Helene Tauson (2014) Foetal life protein provision of mink (Neovison vison) changes the relative mRNA abundance of some hepatic enzymes regulating fat metabolism, Archives of Animal Nutrition, 68:2, 159-169, DOI: 10.1080/1745039X.2014.889506 To link to this article: http://dx.doi.org/10.1080/1745039X.2014.889506

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Archives of Animal Nutrition, 2014 Vol. 68, No. 2, 159–169, http://dx.doi.org/10.1080/1745039X.2014.889506

RESEARCH NOTE Foetal life protein provision of mink (Neovison vison) changes the relative mRNA abundance of some hepatic enzymes regulating fat metabolism Connie Frank Matthiesena, Maria Arantzazu Aguinaga Casañasa,b and Anne-Helene Tausona*

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a

Department of Veterinary Clinical and Animal Sciences, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, Denmark; bDepartment of Physiology and Biochemistry of Animal Nutrition, Animal Nutrition Institute, Estación Experimental del Zaidín (CSIC), Armilla, Granada, Spain (Received 27 October 2013; accepted 20 January 2014) The nutrient provision to pregnant females has high impact on the growth and metabolism of their offspring. The objective was to investigate if the expression of hepatic enzymes regulating the fat metabolism was affected in foetuses and adult female mink born by dams fed either a low or an adequate level of protein during late gestation. The relative abundances of acetyl coenzyme A carboxylase (ACC), fatty acid synthase (FAS) and carnitine palmitoyl transferase 1 (CPT1) mRNA were determined by qualitative polymerase chain reaction in the livers of F0- and F1-generation dams and in F1-generation foetuses. Low protein provision during foetal life resulted in a lower expression of FAS in foetal liver but a tendency towards increased expression in the liver of adult dams. There was a tendency towards an effect of life stage of the animal on the expression of ACC resulting in a higher expression among F1 foetuses exposed to low protein during foetal life than F0 dams fed a low protein diet during late gestation. The expression of CPT1 was significantly lower among dams exposed to low protein provision during foetal life than controls, possibly indicating a lower rate of mitochondrial β-oxidation. Further investigations are needed to clarify the consequences of these changes for the fat metabolism. Keywords: fat metabolism; foetal growth; gene expression; maternal nutrition; protein malnutrition

1. Introduction Poor foetal life environment such as maternal malnutrition can lead to changes in the growth or metabolism of the foetus in order to maximise the outcome of the given available nutrients and to ensure survival. If a mismatch in nutrient supply between the prenatal and postnatal environment appears, the foetal adaptations may become detrimental to health (Gluckman and Hanson 2004). A number of animal models, mainly rats, mice and sheep, have been used extensively to study prenatal alterations, caused by maternal malnutrition, on postnatal growth and metabolism. This also includes the low protein model in rats, which has resulted in reduced birthweight and intrauterine growth restriction (IUGR), reduced postnatal growth, *Corresponding author. Email: [email protected] Present address for Maria Arantzazu Aguinaga Casañas: Institute of Nutritional Physiology “Oskar Kellner”, Leibniz Institute for Farm Animal Biology (FBN), Dummerstorf, Germany © 2014 Taylor & Francis

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adaptive changes in organ size (Desai et al. 1996), alterations in the endocrine pancreas (Snoeck et al. 1990) and changes in the glucose (Desai et al. 1995; Ozanne and Hales 1999) and lipid metabolism (Lucas et al. 1996). The changes in the lipid metabolism due to protein malnutrition in foetal life seem to be both age and gender specific as evidenced by previous studies on hepatic (Maloney et al. 2003; Erhuma, Bellinger, et al. 2007; Qasem et al. 2010) and plasma triglyceride content (Maloney et al. 2003) in male and female rats. Offspring from protein-malnourished rats have also shown an altered expression of key hepatic enzymes important for the fat metabolism (Maloney et al. 2003; Erhuma, Bellinger, et al. 2007; Erhuma, Salter, et al. 2007). Metabolic adaptive changes caused by protein malnutrition during foetal life remain almost unexplored in the mink (Neovison vison) which, by being a strict carnivore, can be expected to be very responsive to protein shortage. The mink is a photo-periodic seasonal breeder with one annual breeding season. The ovulation is induced by mating and followed by an embryonic diapause which delays the implantation. After the implantation is completed the true gestation lasts for 31 days (Murphy and Douglas 1992). The reproductive performance of the mink is known to be highly responsive to changes in the energy supply (Tauson and Forsberg 2002). It gives, like the rat, birth to altricial young, and the kits are totally dependent on the energy and nutrient supply in mothers’ milk for the first four weeks of life. The few studies where mink dams exposed to gestational protein malnutrition have been used as an alternative animal model have resulted in reduced birthweight (Matthiesen, Blache, Thomsen, Hansen, et al. 2010; Vesterdorf et al. 2012) and revealed changes in the relative abundance of key hepatic enzymes in glucose metabolism in foetuses (Matthiesen, Blache, Thomsen, Hansen, et al. 2010) and mink kits post-weaning (Matthiesen et al. 2012). The objectives were to investigate whether maternal low protein provision resulted in changes in hepatic relative abundance of enzymes involved the lipid metabolism, both in foetal and in adult offspring. The working hypothesis was that the expression of enzymes involved in fat synthesis (acetyl coenzyme A carboxylase, ACC; and fatty acid synthase, FAS) would be up-regulated and that the one involved in β-oxidation (carnitine palmitoyl transferase 1, CPT1) would be down-regulated as an effect of foetal life protein shortage. 2. Materials and methods This study was an extension of previous investigations by Matthiesen, Blache, Thomsen, Hansen, et al. (2010) and Matthiesen, Blache, Thomsen, Tauson (2010) into the metabolic consequences for the offspring of gestational low protein (LP) provision of the mink dam. The experimental procedures followed the guidelines of the European Convention for the Protection of Vertebrate Animals used for Experimental and Other Scientific Purposes as well as Danish national legislation (European Convention for Protection… 1986). The experiment was conducted at the small animal laboratory facility, Rørrendegaard Experimental Farm, University of Copenhagen, Taastrup, Denmark. 2.1. Animals and experimental design A total of 12 pregnant mink dams were used in the experiment. Out of these, six dams had been adequately nourished for their entire life (F0-generation) and six dams (F1-generation) had been either adequately nourished their entire life (foetal life adequate – FA; n = 3) or exposed to LP provision during the last 17.9 ± 3.6 days of their 31 days foetal life (foetal life low – FL; n = 3). The dams were provided a conventional farm diet

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adequate in protein content at parturition and throughout lactation, and the offspring, including the F1 dams used here, were given diets sustaining their protein requirement from weaning onwards. The F0-generation dams were fed either a LP and or an adequate protein (AP) control diet from when implantation was anticipated to be completed (15 April) until euthanasia 30–34 days after mating corresponding to 7 or 13 days exposure to the experimental feeding. The F1 dams were given a conventional farm diet adequate in protein content from mating until euthanasia 37–40 days after mating. The treatment of the foetuses (F1) from the F0 dams were either adequately treated during foetal life (FA) or exposed to LP provision during foetal life. The F2 foetuses had been exposed to maternal foetal life adequate (MA) or maternal foetal life LP provision (ML) combined with foetal life adequate protein (AP) provision resulting in MAFA and MLFA F2 foetuses (Figure 1). The F0 dams were randomly chosen from an experiment by Matthiesen, Blache, Thomsen, Hansen, et al. (2010) with a total of 66 dams and where the litter size (AP: 5.2 vs. LP 4.3) was not significantly affected by the protein provision during late gestation, but AP-fed dams had a significantly higher number of kits per mated female (AP: 4.9 kits vs. 3.4 kits, p = 0.04) than the LP-fed dams. The birthweight was significantly lower (p = 0.004) among FL (10.3 g) than FA (11.3 g) F1 offspring (Matthiesen, Blache, Thomsen, Hansen, et al. 2010). The litter size of the F1 dams (n = 40) did not differ significantly between F1 FA (6.7 kits) and FL (7.2 kits) dams neither did the number of kits per mated female (FA: 6.3 kits vs. FL: 5.6). The birthweight

Low protein diet (LP) during late gestaon

Adequate protein diet (AP) during gestaon Dams F0-generaon

F1-generaon FA - Foetus

F1-generaon FL - Foetus

Convenonelfarm feed from birth onwards F1-generaon FA – Dams

Convenonelfarm feed from birth onwards F1-generaon FL – Dams

Convenonelfarm feed during gestaon

Convenonelfarm feed during gestaon

F2-generaon MAFA - Foetus

F2-generaon MLFA - Foetus

Figure 1. Schematic overview of the experimental design. Adequately treated dams (F0-generation) were fed either adequate (AP) or low protein diet (LP) during late gestation. The F0 dams AP dams had F1 offspring adequately treated during foetal life (FA) and F0 LP-fed dams had F1 offspring exposed to LP levels during foetal life (FL). The F2 offspring were exposed to LP or adequately treated during maternal (MA or ML) foetal life and adequately treated during own foetal life (FA) resulting in MAFA or MLFA F2 offspring.

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of the F2 offspring was however significantly higher (p = 0.003) among MLFA (12.2 g) than MAFA (11.1 g) F2 offspring (Matthiesen, Blache, Thomsen, Tauson, et al. 2010).

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2.2. Diets The dams were fed ad libitum once a day and had free access to drinking water. The F0-generation dams were fed a conventional wet diet until implantation was completed and during the lactation. The F1-generation dams (FA, FL) were fed a conventional wet diet until euthanasia (Table 1). The diets were produced by Sjællands Pelsdyrfoder A.m.b.a. (Staarup, Denmark). The experimental diets were fed starting from 15 April, when it was anticipated that the implantation was completed, until euthanasia. The diets were produced at the experimental farm, Roerrendegaard (Taastrup, Denmark). The chickens were chopped in a mincing machine (BIZERBA, Bizerba-Werke Wilhelm Kraut GmbH & Co., Hamburg, Germany), and all the ingredients were mixed in a food mixer (Bjørn Varimixer, Denmark). The diets were prepared on one occasion, apportioned by weight in plastic Table 1.

Dietary and chemical composition of the experimental diets. Diet Adequate protein

Dietary composition [g/kg] Fresh chicken* Potato mash powder Steam rolled oats Fishmeal Sugar beet pulp Corn starch, gelatinised Corn oil Vitamin and mineral mixture† Water Chemical composition Dry matter (DM) [g/kg] Ash [g/kg DM] Crude protein [g/kg DM] Fat [g/kg DM] Carbohydrate [g/kg DM] Gross energy [MJ/kg DM] Metabolisable energy (ME) [MJ/kg DM] Protein : fat : carbohydrate ratio [% of ME] Conventional farm diet DM [g/kg] Crude protein [g/kg DM] Fat [g/kg DM] Carbohydrate [g/kg DM] Protein : fat : carbohydrate ratio [% of ME]

Low protein

600 40 40 150 30 40 60 3 37

250 70 100 70 30 150 100 3 227

446 85 423 291 200 25.12 19.35 29:56:15

386 47 212 269 472 23.58 18.49 14:51:35

F0 347 492 193 216 40:46:14

F1 352 494 170 233 42:42:16

Notes: *Chicken prepared for human consumption, i.e. without heads, feet, feathers or entrails; †Per kg of supplement: retinol 2,800,500 IU, cholecalciferol 280,000 IU, a-tocopherol 21,840 IU, thiamine 10 g, riboflavin 4.8 g, nicotiamide 80 g, pyridoxine 3.2 g, D-pantothenic acid 3.2 g, biotin 80 mg, folic acid 240 mg, choline chloride 60 g, cyanocobalamin 16 mg, para-aminobenzoic acid 800 mg, betaine 33.6 g, Fe (FeSO4) 19,712 mg, Cu (CuSO4) 1025 mg, Zn (ZnO) 12,560 mg and Mn (MnO) 6238 mg (Trouw Nutrition, Denmark A/S, Vejen, Denmark).

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bags, and frozen immediately. The dietary and chemical composition of the two experimental diets can be found in Table 1.

2.3. Tissue collection The F0-generation dams were euthanised 30–34 days after mating when they had been exposed to the two different protein contents (LP or AP) for 7 or 13 days. The F1-generation dams (FA, FL) were euthanised 37–40 days after mating. The animals were anaesthetised with 0.2 ml Ketaminol (50 mg/ml, InterVet International B.V. Boxmeer, The Netherlands) and 0.1 ml Narcoxyl (20 mg/ml, InterVet) by intramuscular injection. As soon as palpebral and toe pinch reflexes were absent the animals were killed. Foetal and maternal livers were quickly excised and frozen rapidly in liquid N2 and were stored at –80°C until RNA isolation. Body and liver weights of all dams were recorded. Body weight and body length of all the F1- and F2-generation foetuses and liver weights of the F2-generation foetuses were recorded. Liver tissue was collected from the F0- and F1-generation dams and from one F1-and one F2-generation foetus per litter. However, the foetal F2-generation hepatic tissue had to be discarded owing to technical problems.

2.4. Assessment of gene expression The RNA extraction method of the liver tissue, homogenisation, transcription into cDNA and estimation of the relative abundance of enzyme mRNA by means of real-time quantitative polymerase chain reaction (PCR) can be found elsewhere (Matthiesen, Blache, Thomsen, Hansen, et al. 2010). The used primers, all designed from canine mRNA sequences, are presented in Table 2. The chosen reference gene was β-actin. The PCR products of all the primer pairs used were sequenced to confirm product identity. Furthermore, the PCR efficiency was calculated for both the target and the reference genes by determining the fitting coefficients of the relative standard curve. The final relative quantification was efficiency corrected.

2.5. Statistics The statistical analyses of data were carried out using the mixed procedure in SAS 9.2 (Littell et al. 2006) according to the following complete model which was used for the gene expression analyses: Yijk ¼ μ þ αi þβj þ ðαβÞij þ"ijk where Yijk is the Yijkth observation, µ the general mean, αi the fixed effect of life stage (adult, foetus), βj the fixed effect of protein provision (AP, LP, FA, FL) during late gestation and foetal life, (αβ)ij the interactions between the fixed effects and εijk the residual error ~ N(0,σ 2i ). For the body weight and tissue data the non-significant interaction effects were removed from the model. Pairwise comparisons of LSmeans were made using the PDIFF option of SAS, and differences were denoted as significant if p < 0.05 and as a tendency if p < 0.10.

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PCR primers.

Gene

Primer

ACC

5´AGCACGCCAGGTTCTTATTG´3 5´GTGGTTGAGGTTGGAGGAGA´3 5´ACTACCTGGTGGACCACTGC´3 5´CAGGTTGCCATTCTCAGACA´3 5´GCTCACCAAGCCGTGGCCTT´3 5´CACGCCAGTGATGATGCCATTC´3 5´GATCCACACGGAGTACTTGC´3 5´ACCATGTACCCCGGCATC´3

FAS CPT1 β-Actin

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GenBank accession no. XM548250 XM540497 XM533208 AY460114

3. Results and discussion 3.1. Diets The ME intake in late gestation did not differ between F0 AP- and LP-fed dams (Matthiesen, Blache, Thomsen, Hansen, et al. 2010) or between FA or FL F1 dams (Matthiesen, Blache, Thomsen, Tauson, et al. 2010). The late gestation diets of the F0 dams differed in protein and carbohydrate content, whereas fat was almost similar between the diets (Table 1). Feeding a LP diet to rats resulted in a reduction in serum amino acids and an increase in plasma glucose as a consequence of higher carbohydrate content in the LP diet (Kwong et al. 2000). Furthermore, the plasma concentration of insulin was reduced in rats fed LP diets (Kwong et al. 2000) findings which are in accordance with pregnant LP mink dams (Matthiesen, Blache, Thomsen, Hansen, et al. 2010). The same study has shown that LP dams had numerically greater relative abundance of fructose-1,6-biphosphatase (Fru-1,6-P2ase) and pyruvate kinase (PKM2) mRNA than AP dams (1.2 vs. 0.5 and 0.6 vs. 0.3, respectively) and a 60% and 63% lower abundance of Fru-1,6-P2ase (p = 0.007) and of PKM2 (p = 0.002) mRNA in FL F1 than in FA F1 foetuses (Matthiesen, Blache, Thomsen, Hansen, et al. 2010). An increase in plasma glucose concentration in lactating mink dams fed different levels of protein (61% vs. 31% of ME) and carbohydrates (2% vs. 32% of ME) has been reported previously (p = 0.02) (Fink and Børsting 2002). If the LP dams had higher plasma glucose concentrations the transfer of glucose to the foetus would increase, which might lead to changes in the hepatic expression of enzymes important for the fat metabolism. Indeed, in weanling rats the expression of fat-synthesising enzymes has been reported to increase as a consequence of maternal LP feeding (Maloney et al. 2003).

3.2. Body and tissue collection data The LP provision during late gestation did not affect the body weight of the F0 dams during the differentiated feeding period or the body weight of F1 dams at euthanasia. The lowest liver weights and liver weights in per cent of body weight were found in the F0 LP dams, and these values were significantly lower than those of F1 FA dams (p < 0.05), whereas the liver weights in absolute terms and scaled to body weight among F1 dams were unaffected by foetal life protein provision. We have previously found that mink responds to LP diets by a reduced liver mass (dams: Larsson et al. 2012a; kits: Larsson et al. 2012b; Matthiesen et al. 2012) so the present LP dam data concur with previous findings and demonstrate that the response is rather rapid.

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Table 3. Body, liver and uterus weights and number of foetuses from dams (F0 generation) fed AP or LP diet during late gestation and of their F1-generation adult offspring exposed to either foetal life low or adequate protein provision during their first gestation (n = 12). Treatment F0 dams/F1 foetuses

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Adequate Low protein (AP) protein (LP) Body weight (BW) [g] Liver weight [g · kg0.75] Liver [% of BW] Uterus [g] Total litter size Number of live foetuses Live foetuses [% of total] Number of dead foetuses Dead foetuses [% of total] Weight of foetuses [g] Length of foetuses [g] Liver of foetuses [g]

1295

1274

F1 dams/F2 foetuses Foetal life adequate (FA) 1297

Foetal life low (FL) 1241

Square root of residuals 140

p-value treatment NS

31.3ab

26.9b

34.9a

32.5ab

4.1

0.05

2.9ab 117.8 10.0 9.0

2.5b 74.8 10.2 8.7

3.2a 101.7 7.7 7.5

3.1ab 103.9 8.2 8.0

0.4 31.9 3.3 3.5

0.05 NS NS NS

11.0

NS

90 1.0a 10

83 1.5a 17

94

97

0.2b

0.2b

6

3

0.8 11.0

NS NS

3.2ab

2.1b

6.7a

3.7ab

3.1

41.4ab

27.6b

54.6a

45.2ab

15.4

0.46

0.38

0.2





0.07

0.1 NS

Note: a,bMean values within a row with unlike lower-case superscript letters were significantly different (p < 0.05).

The weight of uterus with content was not significantly affected by diet or foetal life protein provision (Table 3), which differs from the findings of Vesterdorf et al. (2012), but the discrepancy can be explained by a large individual variation and that the stage of true gestation may have varied among groups in both investigations owing to the delayed implantation. The numbers of live foetuses were not affected by protein provision during gestation or foetal life, whereas the number of dead foetuses was significantly lower in F1 than in F0 dams irrespective of protein provision during gestation or foetal life, which corresponds to the numerically lower litter size. However, there were no significant differences between percentages of dead or live foetuses of total litter size, but they differed numerically (Table 3). The body weight and length of the foetuses were not significantly affected by protein provision during gestation or foetal life, but both the body weight and length of the foetuses were significantly lower of F0 LP than F1 FA dams. These differences were unlikely to be caused by the dietary treatments but rather an effect of the animals being in different stages of true gestation (Table 3).

3.3. Gene expression in hepatic tissue The protein provision during the gestation of the F0 dams and the foetal life protein provision of the F1 offspring (foetuses and adults) led to some detectable changes in the

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relative mRNA abundance of investigated fat-metabolising enzymes. The relative abundance of ACC, which catalyses the limiting step in the de novo synthesis of malonyl-CoA, was not affected by the protein provision during gestation or foetal life, and hence the relative abundances did not differ among adult dams regardless of gestational or foetal dietary history (Figure 2a). However, a tendency (p = 0.06) for an effect of life stage of the animal (foetus vs. adult) appeared with the relative abundance of ACC tending to be higher in foetuses than adults. This resulted in a significantly higher relative abundance of ACC in F1 FL foetal liver than in their F0 LP dams (p = 0.03) and also higher than in adult F1 FA liver (p = 0.03). In F1 foetuses there was no significant difference in the expression of ACC between the FL and FA foetal life dietary treatments, but the expression of the mRNA level of ACC was numerically higher in FL than FA foetal tissue (Figure 2a). Our results were in contrast to those of Lane et al. (2001) where the mRNA level of ACC was lower in neonatal IUGR rat offspring than in controls. Other studies have found a twofold increase in ACC in weanling rats born by LP-fed dams (Maloney et al. 2003) which supports our numeric differences, whereas Erhuma, Salter, et al. (2007) found a lower expression of ACC in 9-month-old LP offspring, but at 18 months of age LP animals had a higher expression than controls. Therefore, the expression of ACC in rats was affected both by age of the animal and the interaction between maternal diet and age (Erhuma, Salter, et al. 2007). The relative abundance of FAS, which catalyses the synthesis of saturated fatty acids from malonyl-CoA, was influenced by life stage of the animals (foetus vs. adult) (p < 0.002)

Figure 2. The relative mRNA abundance normalised to β-actin of (a) acetyl coenzyme A carboxylase, (b) fatty acid synthase and (c) carnitine palmityol transferase 1 in the liver tissue of dams (F0 generation) fed either a low protein (LP) or an adequate protein (AP) diet in late gestation, their foetuses (F1 generation) and the adult F1-generation offspring. a,b,cWithin transcript, labelled LS means without a common letter differ at p < 0.05.

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and an interaction between protein provision and life stage (p = 0.03) (Figure 2b), such that F0 dams had the lowest and similar relative abundances, whereas their F1 foetuses had significantly higher relative abundances, which moreover were lower in F1 LP than in F1 AP (p = 0.04). In adult F1 dams, however, there was a tendency (p = 0.09) for the reverse, i.e. higher relative abundance of FAS in F1 FL than in F1 FA dams, which may indicate an increased formation of saturated fatty acids. Our findings are not completely comparable with other studies, but significantly lower expression of FAS among LP manipulated rat offspring at 9 months of age but significantly higher at 18 months of age than in controls (Erhuma, Salter, et al. 2007) indicates a similar effect of life stage. Contrary to these findings Maloney et al. (2003) found higher expression of FAS in weanling rat offspring exposed to a LP diet during foetal life and lactation. The relative abundance of CPT1 was affected by life stage of the animals (p = 0.001) with lower relative abundance in foetuses than in adults, but without effect of gestational or foetal life dietary treatment of the F0 dams and their F1 foetuses (Figure 2c). There was a tendency for an effect of protein provision (p = 0.09) and a tendency (p = 0.09) for interactions between them. This concurs partly with the findings by Guzmán et al. (1992) where the hepatic activity of CPT1 was similar to that of controls at birth but then increased until 21 days of age among offspring from the LP-diet-fed dams during the gestation and the lactation. Reversely, Maloney et al. (2003) reported that the expression of CPT1 was unaffected by maternal protein provision in weanling rats. Others have, however, found that newborn rat offspring exposed to IUGR by uteroplacental insufficiency had a lower level of hepatic CPT1 mRNA than controls (Lane et al. 2001). The significantly lower relative abundance of CPT1 in F1 FL than in F1 FA adults (p < 0.01) was in contrast to findings in 120-day-old adult female rats exposed to IUGR where the hepatic mRNA levels were similar to those of controls, but the fact that CPT1 was significantly lower in IUGR males at 120 days of age (Lane et al. 2001) may suggest that gender effects play a role. A lower relative abundance of CPT1 mRNA as in our F1 FL dams would suggest a lower mitochondrial β-oxidation, and thereby a possibility for a higher rate of body fat accretion. Such increased fat deposition as a consequence of maternal malnutrition has for instance previously been demonstrated in female (Zambrano et al. 2006) and male rats (Anguita et al. 1993), especially if catch up growth occurs rapidly during the suckling period (Bieswal et al. 2006; Bol et al. 2009).

4. Conclusion The results of this study suggest that the expression of some enzymes regulating the lipid metabolism might be altered by prenatal nutrient supply to mink, but also that life stagespecific changes occur. Tendencies for up-regulating of ACC in F1 LP foetuses and FL adult dams and the tendency for up-regulating of FAS in F1 FL adults, together with the finding of CPT1 being down-regulated in adult dams that were protein malnourished during foetal life, would support our hypothesis that fat-synthesising enzymes were likely to be upregulated, whereas those involved in β-oxidation were likely to be down-regulated as an effect of the experimental treatment. Such effects of foetal life nutrient supply may have consequences for the rate of fat accretion and oxidation in individuals that have experienced protein shortage during foetal life. These findings however need to be confirmed by plasma metabolites and phenotypic changes such as liver and body fat content.

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Funding This work was supported by the Danish Food Industry Agency of the Ministry of Food, Agriculture and Fisheries [grant number 5414114.95.304].

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References Anguita RM, Sigulem DM, Sawaya AL. 1993. Intrauterine food restriction is associated with obesity in young rats. J Nutr. 123:1421–1428. Bieswal F, Ahn M-T, Reusens B, Holvoet P, Raes M, Rees WD, Remacle C. 2006. The importance of catch-up growth after early malnutrition for the programming of obesity in male rat. Obesity. 14:1330–1343. Bol VV, Delattre A-I, Reusens B, Raes M, Remacle C. 2009. Forced catch-up growth after fetal protein restriction alters the adipose tissue gene expression program leading to obesity in adult mice. Am J Physiol Regul Comp Physiol. 297:R291–R299. Desai M, Crowther NJ, Lucas A, Hales CN. 1996. Organ-selective growth in the offspring of protein-restricted mothers. Br J Nutr. 76:591–603. Desai M, Crowther NJ, Ozanne SE, Lucas A, Hales CN. 1995. Adult glucose and lipid metabolism may be programmed during fetal life. Biochem Soc Trans. 23:331–335. Erhuma AM, Bellinger L, Langley-Evans SC, Bennett AJ. 2007. Prenatal exposure to undernutrition and programming of responses to high-fat feeding in the rat. Br J Nutr. 98:517–524. Erhuma AM, Salter AM, Sculley DV, Langley-Evans SC, Bennett AJ. 2007. Prenatal exposure to low-protein diet programs disordered regulation of lipid metabolism in aging rat. Am J Physiol. 292:E1702–1714. European convention for the protection of vertebrate animals used for experimental and other scientific purposes. 1986. European Treaty Series No. 123. Strasbourg. Fink R, Børsting CF. 2002. Quantitative glucose metabolism in lactating mink (Mustela vison) – effects of dietary levels of protein, fat and carbohydrates. Acta Agric Scand Sect A Animal Sci. 52:34–42. Gluckman PD, Hanson MA. 2004. Living with the past: evolution, development, and patterns of disease. Science. 305:1733–1736. Guzmán M, Azzolin IR, Moulin CC, Perry MLS. 1992. Pre- and postnatal protein undernutrition increases hepatic carnitine palmitoyltransferase I activity and decreases enzyme sensitivity to inhibitors in the suckling rat. Horm Metab Res. 24:471–473. Kwong WY, Wild AE, Roberts P, Willis AC, Fleming TP. 2000. Maternal undernutrition during the preimplantation period of rat development causes blastocyst abnormalities and programming of postnatal hypertension. Development. 127:4195–4202. Lane RH, Kelly DE, Gruetzmacher EM, Devaskar SU. 2001. Uteroplacental insufficiency alters hepatic fatty acid-metabolizing enzymes in juvenile and adult rats. Am J Physiol Regul Integr Comp Physiol. 280:R183–R190. Larsson C, Fink R, Matthiesen CF, Thomsen PD, Tauson A-H. 2012a. Metabolic and growth response of mink (Neovison vison) kits until 10 weeks of age when exposed to different dietary protein provision. Arch Anim Nutr. 66:237–255. Larsson C, Fink R, Matthiesen CF, Thomsen PD, Tauson AH. 2012b. Metabolic adaptation to different protein supply in mink (Neovison vison). Baltic J Comp Clin Syst Biol. 2:46–67. Littell RC, Milliken GA, Stroup WW, Wolfinger RD, Schabenberger O. 2006. SAS for mixed models. 2nd ed. Cary (NC): SAS Institute Incorporation. Lucas A, Baker BA, Desai M, Hales CN. 1996. Nutrition in pregnant or lactating rats programs lipid metabolism in the offspring. Br J Nutr. 76:605–612. Maloney CA, Gosby AK, Phuyal JL, Denyer GS, Bryson JM, Caterson ID. 2003. Site-specific changes in the expression of fat-partitioning genes in weanling rats exposed to a low-protein diet in utero. Obesity. 11:461–468. Matthiesen CF, Blache D, Thomsen PD, Hansen NE, Tauson A-H. 2010. Effect of late gestation low protein supply to mink (Mustela vison) dams on reproductive performance and metabolism of dam and offspring. Arch Anim Nutr. 64:56–76. Matthiesen CF, Blache D, Thomsen PD, Tauson A-H. 2010. Feeding mink (Neovison vison) a protein-restricted diet during pregnancy induces higher birth weight and altered hepatic gene expression in the F2 offspring. Br J Nutr. 104:544–553.

Downloaded by [UOV University of Oviedo] at 08:45 17 October 2014

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Matthiesen CF, Blache D, Thomsen PD, Tauson A-H. 2012. Foetal life protein restriction in male mink (Neovison vison) kits lowers post-weaning protein oxidation and the relative abundance of hepatic fructose-1,6-bisphosphatase mRNA. Animal. 6:50–60. Murphy BD, Douglas DA. 1992. Reproduction in female mink. In: Tauson A-H, Valtonen M, editors. Reproduction in carnivorous fur bearing animals. Nordic association of agricultural scientists. Report No. 75. Copenhagen: Jordbrugsforlaget. Ozanne SE, Hales CN. 1999. The long-term consequences of intrauterine protein malnutrition for glucose metabolism. Proc Nutr Soc. 58:615–619. Qasem RJ, Cherala G, D’Mello AP. 2010. Maternal protein restriction during pregnancy and lactation in rats imprints long-term reduction in hepatic lipid content selectively in the male offspring. Nutr Res. 30:410–417. Snoeck A, Remacle C, Reusens B, Hoet JJ. 1990. Effect of a low protein diet during pregnancy on the fetal rat endocrine pancreas. Biol Neonate. 57:107–118. Tauson AH, Forsberg M. 2002. Body-weight changes are clearly reflected in plasma concentrations of leptin in female mink (Mustela vison). Br J Nutr. 87:101–105. Vesterdorf K, Harrison A, Matthiesen CF, Tauson A-H. 2012. Effects of protein restriction in utero on the metabolism of mink dams (Neovison vison) and on mink kit survival as well as on postnatal growth. Open J Anim Sci. 02:19–31. Zambrano E, Bautista CJ, Deás M, Martínez-Samayoa PM, González-Zamorano M, Ledesma H, Morales J, Larrea F, Nathanielsz PW. 2006. A low maternal protein diet during pregnancy and lactation has sex- and window of exposure-specific effects on offspring growth and food intake, glucose metabolism and serum leptin in the rat. J Physiol. 571:221–230.

Foetal life protein provision of mink (Neovison vison) changes the relative mRNA abundance of some hepatic enzymes regulating fat metabolism.

The nutrient provision to pregnant females has high impact on the growth and metabolism of their offspring. The objective was to investigate if the ex...
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