The forgotten variable? Does the euthanasia method and sample storage condition influence an organisms transcriptome - a gene expression analysis on multiple tissues in pigs.

BACKGROUND: Transcriptomic studies often require collection of fresh tissues post euthanasia. The chosen euthanasia method might have the potential to induce variations in gene expressions that are unlinked with the experimental design. The present study compared the suitability of 'nitrogen gas in foam' (ANOXIA) in comparison to a non-barbiturate anaesthetic, T-61® (T61), for euthanizing piglets used in transcriptome research. Further, the effect of common tissue storage conditions, RNAlater™ (RL) and snap freezing in liquid nitrogen (LN2), on gene expression profiles were also analysed. RESULTS: On comparison of the 3'mRNA-Seq data generated from pituitary, hypothalamus, liver and lung tissues, no significant differential expression in the protein coding genes were detected between the euthanasia methods. This implies that the nitrogen anoxia method could be a suitable alternative for euthanasia of piglets used in transcriptomic research. However, small nuclear RNAs (snRNAs) that constitute the eukaryotic spliceosomal machinery were found to be significantly higher (log2fold change ≥ 2.0, and adjusted p value ≤ 0.1) in pituitary samples collected using ANOXIA. Non-protein coding genes like snRNAs that play an important role in pre-mRNA splicing can subsequently modify gene expression. Storage in RL was found to be superior in preserving RNA compared to LN2 storage, as evidenced by the significantly higher RIN values in representative samples. However, storage in RL as opposed to LN2, also influenced differential gene expression in multiple tissues, perhaps as a result of its inability to inhibit biological activity during storage. Hence such external sources of variations should be carefully considered before arriving at research conclusions. CONCLUSIONS: Source of biological variations like euthanasia method and storage condition can confound research findings. Even if we are unable to prevent the effect of these external factors, it will be useful to identify the impact of these variables on the parameter under observation and thereby prevent misinterpretation of our results.

Effect of temperature manipulation during incubation on body weight, plasma parameters, muscle histology, and expression of myogenic genes in breast muscle of embryos and broiler chickens from two commercial strains.

1. This study evaluated the effect of a higher incubation temperature on body weight, plasma profile, histology and expression of myogenin (MYOG), insulin-like growth factor-I (IGF-I) and vascular endothelial growth factor A (VEGFA) genes in breast muscle of embryos and broilers from two commercial strains.2. A total of 784 eggs from Ross 308 and Cobb 500 broiler breeder flocks were used. Half of the eggs per strain were incubated at control temperature (37.8°C), whereas the other half were exposed to heat treatment (HT) of 38.8°C between embryonic day (ED) 10 and 14, for 6 h/day. Embryos and chicks were sampled on ED 19 and at hatch. A total of 480, one-day-old chicks per strain and incubation temperature were reared up to 42 d post-hatch.3. The HT increased hatch weight of Ross chicks and 42-d body weight of broilers from both strains. Lower plasma triacylglycerol levels were measured for HT embryos and broilers on ED 19 and 42 d post-hatch, respectively. HT reduced plasma T3 levels in Ross embryos and broilers for the same periods. Hepatic TBARS concentrations were elevated by HT compared to the control incubation.4. The HT reduced breast muscle VEGFA gene expression of Cobb embryos on ED 19, whereas expression was stimulated in day-old chicks. At 42 d post-hatch, fibre area was increased by HT regardless of strain. Compared to the control incubation, HT increased the breast yield of Ross broilers and leg yield of Cobb. Ross-HT broilers had a higher pH at 24 h after slaughter and better water holding capacity than Cobb-HT broilers.5. These results suggested that HT increased body weight, fibre area, IGF-I gene expression and lowered plasma triacylglycerol levels of broiler chickens from both strains at 42 d. However, HT influenced the expression of VEGF-A and MYOG genes and meat quality differently between the broiler strains.

Direct and maternal reduced balanced protein diet influences the liver transcriptome in chickens.

The objective of this study was to evaluate, by means of RNA sequencing, the direct and transgenerational effect of a reduced balanced protein (RP) diet on broiler breeder metabolism. Chickens of the F0 generation were fed a control (C) or RP diet, and their F1 progeny was fed a C or RP diet as well, resulting in four groups of chickens: C/C, C/RP, RP/C and RP/RP. While both direct and maternal effects were seen on body weight, breast muscle weight and abdominal fat weight in the F1 generation, the direct effect was the most dominant one. The liver transcriptome in the F1 generation showed that amino acid metabolism was up-regulated in chickens that received the control feed when compared with their respective contemporaries that received the reduced protein diet. Interestingly, chickens hatched from control-fed hens but reared on the reduced protein diet (C/RP group) activated a fatty acid metabolism, expressing more fatty acid desaturase 1 gene, fatty acid desaturase 2 gene and elongation of very long-chain fatty acids protein 2 gene, when compared with control-fed chickens hatched from control-fed hens (C/C group), while chickens hatched from reduced protein-fed hens that received themselves the same reduced protein diet (RP/RP group) triggered their glucose metabolism more, showing elevated levels of phosphofructokinase gene, 6-phosphofructo-2-kinase/fructose-2,6-biphospatase 4 and fructose-biphosphate aldolase C mRNA compared with the chickens hatched from reduced protein-fed hens but reared on a control diet (RP/C group). This suggests that the maternal protein diet has an impact on the metabolism of broilers when they are reared on a RP diet.

The influence of thyroid state on hypothalamic AMP-activated protein kinase pathways in broilers.

To investigate whether there are important interactions in play in broilers between thyroid hormones and the central regulation of energy homeostasis through AMP-activated protein kinase (AMPK), we induced a functional hyperthyroid and hypothyroid state in broiler chicks, and quantified systemic and hypothalamic AMPK related gene expression and related protein. Thyroid state was manipulated through dietary supplementation of triiodothyronine (T3) or methimazole (MMI) for 7 days. A hypothalamic AMPK suppressor, 0.1% α-lipoic acid (α-LA) was used to assess the effects of the T3 and MMI feed formulations on the AMPK pathways. Feed intake and body weight were reduced in both hypothyroid and hyperthyroid conditions. In hyperthyroid conditions (T3 supplementation) expression of the AMPKα1 subunit increased, while in hypothyroid conditions (MMI supplementation) active phosphorylated AMPK levels in the hypothalamus dropped, but gene expression of the AMPKα1 and α2 subunit increased. For FAS and ACC (involved in fatty acid metabolism), and CRH, TRH and CNR1 (anorexigenic neuropeptides stimulating energy expenditure) there were indications that their regulation in response to thyroid state might be modulated through AMPK pathways. Our results indicate that the expression of hypothalamic AMPK as well as that of several other genes from AMPK pathways are involved in thyroid-hormone-induced changes in appetite, albeit differently according to thyroid state.

Lysolecithin, but not lecithin, improves nutrient digestibility and growth rates in young broilers.

1. The potential of lecithin and lysolecithin to improve lipid digestion and growth performance was investigated in three experiments: 1. an in vitro model that mimics the intestinal conditions of the chick, 2. a digestibility trial with chicks (5-7 days of age), and 3. a performance trial until 21 days of age. 2. In experiment 1, palm oil (PO), palm oil with lecithin (PO+L), and palm oil with lysolecithin (PO+LY) were subjected to in vitro hydrolysis and applied to Caco-2 monolayers to assess lipid absorption. 3. The in vitro hydrolysis rate of triglycerides was higher in PO+LY (k = 11.76 × 103/min) than in either PO (k = 9.73 × 103/min) or PO+L (k = 8.41 × 103/min), and the absorption of monoglycerides and free fatty acids was highest (P < 0.01) for PO+LY. In experiment 2, 90 broilers were assigned to three dietary treatments: a basal diet with 4% palm oil, and the basal diet supplemented with either 250 ppm lecithin or lysolecithin. 4. ATTD of crude fat was higher in broilers supplemented with lysolecithin, but was lower in broilers supplemented with lecithin. DM digestibility and AMEn in birds supplemented with lysolecithin were significantly higher (3.03% and 0.47 MJ/kg, respectively). 5. In experiment 3, 480 broilers were randomly allocated to four dietary treatments: basal diet with soybean oil (2%), basal diet with lecithin (2%), soybean oil diet with 250 ppm lysolecithin, or lecithin oil diet with 250 ppm lysolecithin. 6. Lecithin diets significantly reduced weight at day 10 and 21 compared with soybean oil. However, the addition of lysolecithin to lecithin-containing diets significantly improved bird performance. 7. The results of these studies showed that, in contrast to lecithin, lysolecithin was able to significantly improve the digestibility and energy values of feed in young broilers.

Changes in egg quality traits during the last phase of production: is there potential for an extended laying cycle?

1. A field study was conducted to investigate the deterioration of eggshell quality at the end of lay and examine the potential to extend the production cycle. A total of 1200 eggs were assessed originating from commercial farms in Belgium: eggs were random sampled at the collection belt at 57-65 weeks of age and at the end of the production cycle (74-92 weeks). 2. Based on a linear mixed model, egg quality was estimated for an average flock at 60 weeks of age and changes in quality traits were predicted until the end of the laying cycle. 3. Egg weight increased by 0.07 g every week from 60 weeks of age onwards and shape index decreased by 0.04 unit weekly (P < 0.001). Haugh unit values decreased weekly by 0.38 units whereas relative yolk weight was unaffected after 60 weeks of age. Relative albumen weight showed a weekly increase of 0.02% and relative shell decreased by 0.02% weekly until the end of lay. 4. Shell quality traits were also influenced by age: shell index decreased by 0.013 g weekly, shell thickness was calculated to be 0.23 µm thinner every week, deformation increased by 0.06 µm weekly from 60 weeks to the end of the laying cycle. Variability of the deformation of eggs indicated more heterogeneous shell quality at the end of the production cycle. Dynamic stiffness increased from 60 weeks of age on. 5. Laying hens were depopulated on average at 80 weeks of age (varying from 74-92 weeks). Although ageing had a significant effect on most of the egg quality traits, egg quality was still acceptable at the end of lay, indicating the potential to extend the laying cycle.

Can breeder reproductive status, performance and egg quality be enhanced by supplementation and transition of n-3 fatty acids?

The aim of this experiment was to investigate the effect of n-3 fatty acid (FA) supplemented diets on breeder performance, productivity and egg quality. Breeders (n = 480) were fed the supplemented diet from 18 weeks onwards; the inclusion level of n-3 FA was increased from 1.5% to 3.0% from 34 weeks of age onwards until 48 weeks of age. Ross-308 broiler breeders (n = 480) were fed one of four different diets: a basal diet rich in n-6 FA (control diet) or one of three diets rich in n-3 FA. For the n-3 FA diets, eicosapentaenoic acid (EPA, 20:5 n-3) and docosahexaenoic acid (DHA, 22:6 n-3) were fed to the broiler breeders at different ratios formulated to obtain EPA/DHA ratios of 1/1, 1/2 or 2/1. Differences in performance, reproduction and egg quality parameters due to n-3 supplementation were noted more for the 1.5% followed by the 3.0% fed broilers than their 1.5% supplemented counterparts. Egg weight (p < 0.001) and egg mass (p = 0.003) were significantly lower and feed conversion (p = 0.008) significantly higher for the n-3 FA (at 3.0% inclusion level) fed broilers compared to the control group. For the EPA- and DHA-fed breeders, a higher proportional abdominal fat percentage (p = 0.025) and proportional albumen weight (%) (p = 0.041) were found respectively. Dietary treatments did not affect reproduction. It can be concluded that the results of the present experiment indicate no significant differences between treatments at 1.5% inclusion levels. However, increasing this level to 3.0% is not recommended due to the rather negative effects on the measured parameters. It should be further investigated whether these adverse effects were obtained due to (i) the higher supplementation level, (ii) combining a supplementation level of 1.5% with 3% or (iii) the duration of supplementation.

In ovo L-arginine supplementation stimulates myoblast differentiation but negatively affects muscle development of broiler chicken after hatching.

In this study, we tested the hypothesis that in ovo feeding (IOF) of L-arginine (L-Arg) enhances nitric oxide (NO) production, stimulates the process of myogenesis, and regulates post-hatching muscle growth. Different doses of L-Arg were injected into the amnion of chicken embryos at embryonic day (ED) 16. After hatching, the body weight of individual male chickens was recorded weekly for 3 weeks. During in vitro experiments, myoblasts of the pectoralis major (PM) were extracted at ED16 and were incubated in medium containing 0.01 mm L-Arg, 0.05 mm L-Arg, and (or) 0.05 mm L-nitro-arginine-methyl-ester (L-NAME), an inhibitor of nitric oxide synthase (NOS). When 25 mg/kg L-Arg/initial egg weight was injected, no difference was observed in body weight at hatch, but a significant decrease was found during the following 3 weeks compared to that of the non-injected and saline-injected control, and this also affected the growth of muscle mass. L-NAME inhibited gene expression of myogenic differentiation antigen (MyoD), myogenin, NOS, and follistatin, decreased the cell viability, and increased myostatin (MSTN) gene expression. 0.05 mm L-Arg stimulated myogenin gene expression but also depressed muscle cell viability. L-NAME blocked the effect of 0.05 mm L-Arg on myogenin mRNA levels when co-incubated with 0.05 mm L-Arg. L-Arg treatments had no significant influence on NOS mRNA gene expression, but had inhibiting effect on follistatin gene expression, while L-NAME treatments had effects on both. These results suggested that L-Arg stimulated myoblast differentiation, but the limited number of myoblasts would form less myotubes and then less myofibers, while the latter limited the growth of muscle mass.

Interaction between fat type and lysolecithin supplementation in broiler feeds.

Lysolecithins are added to poultry diets to promote the intestinal absorption of nutrients, in particular of dietary fats. Lysolecithins contain a mixture of phospho- and lysophospholipids and differ in composition depending on the conditions and source of the lecithin used for its production. The importance of the lysolecithin composition and its interaction with the fat type was investigated in vitro in a fat digestion model and in vivo in a digestibility trial with broilers (24 to 28 d age). The in vitro digestion of soybean oil and pig lard was investigated without and with the inclusion of soybean or rapeseed lysolecithin. Correspondingly, for the digestibility trial, 108 Ross 308 male broilers were assigned to 6 dietary treatments: a basal diet with either soybean oil (5.3%) or pig lard (5.8%), each basal diet supplemented with 250 ppm soybean lysolecithin, and each basal diet supplemented with 250 ppm rapeseed lysolecithin. In vitro pig lard digestibility was significantly lower compared to soybean oil digestibility. Although in vivo no significant difference was observed for crude fat digestibility, broilers fed the basal diet with pig lard had a lower (P < 0.05) DM digestibility, nitrogen retention, and AMEn compared to those fed the basal diet with soybean oil. Lysolecithin supplementation showed a significant interaction with the fat type, both in vitro and in vivo. The in vitro hydrolysis of pig lard, but not of soybean oil, increased (P < 0.001) with supplementation of soybean and rapeseed lysolecithin. Moreover, soybean and rapeseed lysolecithin supplementation improved (P < 0.05) DM digestibility of the basal diet with pig lard by 5.1 and 5.7%, respectively; nitrogen retention by 2.8 and 3.1 g/kg, respectively; and AMEn by 182 and 199 kcal/kg, respectively. Despite the major difference in molecular composition, there was, however, no impact of the lysolecithin composition on in vitro and in vivo fat digestibility. This study demonstrates that the improvements that can be made with lysolecithin supplementation are highly dependent on the fat incorporated in broiler feeds.

The interaction between maternal and post-hatch n-3 fatty acid supplementation in broiler diets.

This study investigated whether offspring from n-3-supplemented breeders have an enhanced performance and immune organ weight when fed a post-hatch n-3-enriched diet in comparison with their control-fed counterparts and the importance of timing of omega-3 supplementation. Therefore, 480 Ross-308 broiler breeder hens were fed one of four different diets (120/treatment). The control diet (CON) was a basal diet, rich in n-6 fatty acids (FA). The three other diets were enriched in n-3 FA, formulated to obtain a different EPA/DHA ratio of 1/1 (EPA = DHA), 1/2 (DHA) or 2/1 (EPA). At 33 weeks of age, eggs were incubated to obtain 1440 offspring. They were set up according to their maternal diet and sex in 48 pens of 30 chicks each (12 pens per maternal treatment: six male and six female). Half of the offspring were given a post-hatch control diet, whereas to other half received an n-3-supplemented diet. Zootechnical performance was followed for starter, grower and finisher phase, and at the end of each phase two, chicks per pen were sacrificed to determine the weight of the immune organs. No interaction was found between maternal and post-hatch n-3 treatment for zootechnical performance. An interaction arose between the maternal and post-hatch n-3 supplementation for proportional bursa weight at day 1 and day 14 and proportional liver weight at day 14, but effects on immune organ weight were rather limited. Offspring post-hatch n-3 supplementation did not enhance maternal n-3 supplementation.

Effects of maternal dietary EPA and DHA supplementation and breeder age on embryonic and post-hatch performance of broiler offspring: age and n-3 pufa affect embryonic and post-hatch performance.

Breeder age and nutrition are amongst the most important factors affecting progeny growth and development. The present experiment was carried out to evaluate the effects of n-3 fatty acid (FA), with special emphasis on the ratio of eicosapentaenoic (EPA, 20:5 n-3) and docosahexaenoic (DHA, 22:6 n-3) acid, provided to the diet of ageing broiler breeder hens at different ratios, on the incubation parameters and the performance of the offspring. Four hundred and eighty Ross-308 broiler breeder hens were fed one of four different diets (120/treatment), with an equal fat content. The control diet was a basal diet, rich in n-6 FAs (CON). Blends of fish oil were used to enrich the three other diets in n-3 FA and to obtain different EPA/DHA ratios of 1/1 (EPA=DHA), 1/2 (DHA) or 2/1 (EPA). Every 5 weeks, incubation parameters were assessed. Every 15 weeks, offspring was reared until slaughter age on a standard diet. Breeder age affected almost all incubation and post-hatch parameters, whereas n-3 FA treatment only lowered egg weight (p < 0.0001) and consequently hatched chick weight (p < 0.0001). Supplementation of EPA resulted in a higher proportional liver weight (p = 0.0219) at hatch, a lower body weight up to 28 days post-hatch (p = 0.0418), a lower daily weight gain (p = 0.0498) and a higher feed conversion ratio (p = 0.0395) during the starter period (p = 0.0498), resulting in a higher overall offspring feed conversion ratio (p = 0.0317) compared to the control diet. DHA supplementation, on the other hand, resulted in a lower residual yolk weight (p = 0.0220) and a higher overall offspring mortality (p = 0.0125). In conclusion, supplementation of n-3 FA could not counter the adverse effect of breeder flock age, but did not harm incubation or improve post-hatch performance, either. EPA and DHA affected offspring development differently during early post-hatch life.

Impact of a trace element supplementation programme on health and performance of cross-breed (Bos indicus x Bos taurus) dairy cattle under tropical farming conditions: a double-blinded randomized field trial.

Small-scale urban dairy farms (n = 16) in and around Jimma, Ethiopia with cross-bred (Bos indicus × Bos taurus) cows were enrolled in a double-blinded intervention study to investigate the effect of a trace element supplementation programme on trace element status and milk concentrations as well as performance [body condition score (BCS), milk yield, leptin], milk composition, antioxidant status (ferric-reducing ability of plasma (FRAP), thiobarbituric acid-reactive substances (TBARS)], blood biochemistry, serum proteins and immune response (antibody titre upon rabies vaccination). The farms were allocated to a (1) placebo or (2) Cu, Zn, Se, Co and I supplementation treatment for 150 d. On days 0 and 120, four lactating cows per farm were sampled for milk and plasma, and on day 150 for serum, following primo-vaccination. Cu deficiency was present in 17% and marginal Se deficiency in 30% of initially sampled cows, while no Zn shortage was detected. Over 120 days, trace element supplementation caused a bigger increase in plasma Se and Cu concentrations, but also a larger decrease of plasma Fe concentrations. A larger increase in milk Se concentrations was observed in the supplemented group, whereas none of the other elements were affected. BCS decreased more over time in the supplemented group. None of the other parameters of performance and antioxidant status nor milk composition or blood biochemistry was affected by treatment. Antibody response to rabies vaccination did not differ between groups, whereas α1-globulins tended to be lower and ß-globulins tended to be higher in the supplemented group. In conclusion, despite improved Cu and Se status and Se concentrations in milk, cows on tropical urban dairy farms did not seem to benefit from trace element supplementation, with respect to the parameters investigated.

Effect of the ratio of dietary n-3 fatty acids eicosapentaenoic acid and docosahexaenoic acid on broiler breeder performance, egg quality, and yolk fatty acid composition at different breeder ages.

When added to the feed of broiler breeder hens, dietary polyunsaturated fatty acids (FA) can be incorporated into the yolk and therefore become available to the progeny during their early development. The mechanism involved in lipid metabolism and deposition in the egg may be influenced by breeder age. Before the effect of an elevated concentration of certain polyunsaturated FA on the embryo can be investigated, the effect at breeder level and egg quality must be further assessed. The aim of the present experiment was to evaluate the effects of dietary n-6/n-3 ratios and dietary eicosapentaenoic acid (EPA, 20:5 n-3) and docosahexaenoic acid (DHA, 22:6 n-3) ratios, provided to broiler breeder hens, in terms of their zoo technical performance, egg quality, and yolk FA composition. Starting at 6 wk of age, 640 Ross-308 broiler breeder hens were fed 1 of 4 different diets. The control diet was a basal diet, rich in n-6 FA. The 3 other diets were enriched in n-3 FA, formulated to obtain a different EPA/DHA ratio of 1/1 (EPA = DHA), 1/2 (DHA), or 2/1 (EPA). In fact, after analysis the EPA/DHA ratio was 0.8, 0.4, or 2.1, respectively. Dietary EPA and DHA addition did not affect the performance of the breeder hens, except for egg weight. Egg weight was lower (P < 0.001) for all n-3 treatments. Dietary EPA improved number of eggs laid in the first 2 wk of the production cycle (P = 0.029). The absolute and relative yolk weight of eggs laid by EPA = DHA fed hens was lowest (P = 0.004 and P = 0.025, respectively). The EPA and DHA concentrations in the yolk were highly dependent on dietary EPA and DHA concentrations with a regression coefficient equal to 0.89. It can be concluded that dietary EPA and DHA can be incorporated in the breeder egg yolk to become available for the developing embryo, without compromising the performance and egg quality of the flock.

Gene expression of heat shock protein 70 and antioxidant enzymes, oxidative status, and meat oxidative stability of cyclically heat-challenged finishing broilers fed Origanum compactum and Curcuma xanthorrhiza essential oils.

Heat stress in poultry is a serious problem in many countries and has been associated with oxidative stress. Hence, nutritional interventions with antioxidants might be beneficial. Therefore, the effects of dietary Curcuma xanthorrhiza (CX) and Origanum compactum (OC) essential oils on mRNA levels of heat shock protein 70 and antioxidant enzymes, oxidative status, and meat oxidative stability of heat-challenged broilers were studied. Starting on d 25 of age, a control diet and 4 diets containing 200 or 400 mg/kg feed of CX or OC (CX200, CX400, OC200, OC400 diets) were fed to 3 pen replicates of 20 Ross 308 chickens each. From d 28 of age on, the temperature was increased from 22 to 34°C with 50% RH for 5 h daily during 2 wk. Dietary CX or OC did not affect zootechnical performance. Feeding CX400 and both levels of OC increased the a* value in stored breast meat (P < 0.05), and OC diets tended to decrease the thiobarbituric acid reactive substances values in fresh breast meat (P = 0.061). Compared with control, at d 31, feeding CX400 and OC400 reduced mRNA levels of heat shock protein 70 and increased mRNA levels of catalase in kidney and liver (P < 0.05). The mRNA levels of superoxide dismutase were increased at d 31 on the OC400 diet in kidney and on the CX400 diet in heart (P < 0.05). In heart, at d 31, both dietary levels of CX and OC200 resulted in higher glutathione peroxidase activity (P < 0.05). Feeding CX400 increased superoxide dismutase activity in liver, kidney, and heart at d 31 (P < 0.05). Catalase activity was increased in the CX200 and OC400 groups at d 42 (P < 0.05). Feeding CX at both levels and OC200 decreased plasma malondialdehyde concentrations at d 42 (P < 0.05). In conclusion, dietary essential oils rich in simple phenolic compounds offer potential for improving the antioxidant defense against heat stress-induced changes.

The cat as a model for human obesity: insights into depot-specific inflammation associated with feline obesity.

According to human research, the location of fat accumulation seems to play an important role in the induction of obesity-related inflammatory complications. To evaluate whether an inflammatory response to obesity depends on adipose tissue location, adipokine gene expression, presence of immune cells and adipocyte cell size of subcutaneous adipose tissue (SAT) and visceral adipose tissue (VAT) were compared between lean and obese cats. Additionally, the present study proposes the cat as a model for human obesity and highlights the importance of animal models for human research. A total of ten chronically obese and ten lean control cats were included in the present study. Body weight, body condition score and body composition were determined. T-lymphocyte, B-lymphocyte, macrophage concentrations and adipocyte cell size were measured in adipose tissue at different locations. Serum leptin concentration and the mRNA expression of leptin and adiponectin, monocyte chemoattractant protein-1, chemoligand-5, IL-8, TNF-alpha, interferon-gamma, IL-6 and IL-10 were measured in blood and adipose tissues (abdominal and inguinal SAT, and omental, bladder and renal VAT). Feline obesity was characterised by increased adipocyte cell size and altered adipokine gene expression, in favour of pro-inflammatory cytokines and chemokines. Consequently, concentration of T-lymphocytes was increased in the adipose tissue of obese cats. Alteration of adipose tissue was location dependent in both lean and obese cats. Moreover, the observed changes were more prominent in SAT compared with VAT.

Partial albumen removal early during embryonic development of layer-type chickens has negative consequences on laying performance in adult life.

To examine the importance of albumen as a protein source during embryonic development on the posthatch performance of laying hens, 3 mL of the albumen was removed. At hatch, no difference in BW could be observed. Chicks from the albumen-deprived group had a lower residual yolk weight due to higher yolk utilization. During the rearing phase (hatch to 17 wk of age), the BW of the albumen-deprived pullets was lower compared with the control and sham pullets. The feed intake of the albumen-deprived pullets was also lower than the control pullets. However, during the laying phase (18 to 55 wk of age) these hens exceeded the control and sham hens in BW, although this was not accompanied by a higher feed intake. The albumen-deprived hens exhibited a lower egg production capacity as demonstrated by the reduced egg weight, laying rate, and egg mass and increased number of second grade eggs. In addition, the eggs laid by the albumen-deprived hens had a higher proportional yolk and lower proportional albumen weight. In conclusion, prenatal protein deprivation by albumen removal caused a long-lasting programming effect, possibly by differences in energy allocation, in favor of growth and maintenance and impairing reproductive performance.

Effects of interaction of incubator CO2 levels and mixing hatching eggs of different embryo growth trajectory on embryo physiological and hatching parameters.

1. Two experiments were designed to investigate the effects of mixing eggs of different genotype and incubating with different CO2 concentrations on embryonic parameters. Half of the eggs were incubated in a CO2 controlled incubator (VCO2) during the first 10 d of incubation and the other half was incubated at standard incubation ventilation rate (SV). 2. From 10 to 18 d of incubation, thick albumen and embryos were weighed. Blood samples were collected at d 18 of incubation, at internal pipping and at hatch for determination of triiodothyronine (T3), thyroxine (T4) and corticosterone concentration. During the last 2 d of incubation, hatching events of individual eggs were monitored every 2 h. 3. From 4 to 5 d of incubation in the SV group of Experiment 1, pH values of Ross strain chickens were lower than those of Isa Brown. From d 12 of incubation onwards, Ross embryos grew faster than those of Isa Brown. At 14 and 16 d of incubation, Ross eggs in the CO2 controlled incubator had lower albumen weights compared to all other treatments. 4. T3 concentrations in Ross embryos were higher than those of Isa Brown embryos. Chicken weight at hatch was in the following order: Isa Brown SV < Isa Brown VCO2 = Ross SV < Ross VCO2. 5. In Experiment 2, incubation time of VCO2 eggs until internal pipping, external pipping and hatch was significantly shorter than that of SV eggs. 6. It was concluded that mixing of hatching eggs of differential embryo developmental trajectory affects their hatching process.

Voluntary feed intake and leptin sensitivity in ad libitum fed obese ponies following a period of restricted feeding: a pilot study.

The relation between plasma leptin and daily ad libitum roughage intake was evaluated during a 14-day period in eight obese Shetland ponies. When the feeding strategy was changed from maintenance feeding to ad libitum feeding, feed intake increased the first day, decreased the second day and increased again during the following days to reach a constant level after 8 days. Plasma leptin concentration increased during the first 2 days, but remained constant afterwards. Although the same pattern was found in all ponies, the magnitude of the increase in leptin on day 1 and the resulting decrease in feed intake on day 2 differed between ponies. A lower anorectic effect was seen in ponies with higher initial leptin concentration, suggesting the presence of different degrees in leptin sensitivity in obese ponies. High leptin production in a attempt to compensate for the decrease in leptin sensitivity might explain large variations in plasma leptin among obese ponies with similar body condition score. Further research is necessary to clarify whether the reduced leptin sensitivity precedes obesity in equines or vice versa.

Supplementation of guanidinoacetic acid to broiler diets: effects on performance, carcass characteristics, meat quality, and energy metabolism.

Creatine, (CREA) a central constituent in energy metabolism, is obtained from dietary animal protein or de novo synthesis from guanidinoacetic acid (GAA). Especially in all-vegetable diets, supplemental CREA or GAA may restore the CREA availability in tissues, and hence, improve performance. In this study, 768 one-d-old male Ross 308 broilers were assigned to 1 of 4 diets: negative control, all-vegetable corn-soybean-based; negative control supplemented with either 0.6 or 1.2 g of GAA per kilogram of feed; and positive control (60, 30, and 30 g/kg of fish meal in the starter, grower, and finisher diets, respectively). Each treatment was replicated in 6 pens of 32 birds each. At the end of the grower period (d 26), 2 birds per pen were euthanized for metabolic measurements. Four broilers per pen were selected at slaughter age (d 39) to determine carcass characteristics and meat quality. Compared with the negative control, GAA supplementation resulted in an improved gain:feed ratio (P < 0.05) and ADG (P < 0.05; + 2.7 and + 2.2% for GAA at 0.6 and 1.2 g/kg, respectively) throughout the entire period. Breast meat yield was higher for the GAA diets compared with that of the negative control birds (P < 0.05; 30.6 vs. 29.4%) and was comparable with that of the positive control birds (30.2%). With regard to meat quality, lower ultimate pH values, higher cooking and press fluid losses, and higher color L* values were observed for the GAA diets compared with those of the negative control diet (P < 0.05). These effects were small, however. The GAA and CREA levels in breast meat were lower and higher, respectively, in GAA-fed birds compared with those of the control birds (P < 0.01). The diets did not affect plasma metabolic traits, except that plasma insulin-like growth factor I concentrations were almost twice as high in animals fed 1.2 g/kg of GAA compared with those of all other treatments. The GAA included in all-vegetable diets improved animal performance for the whole rearing period and increased breast meat yield.

Propionate absorbed from the colon acts as gluconeogenic substrate in a strict carnivore, the domestic cat (Felis catus).

In six normal-weight and six obese cats, the metabolic effect of propionate absorbed from the colon was assessed. Two colonic infusions were tested in a crossover design with intervals of 4 weeks. The test solution contained 4 mmol sodium propionate per kg ideal body weight in a 0.2% NaCl solution. Normal saline was given as control solution. Solutions were infused into the hindgut over 30 min. Blood samples were obtained prior to and at various time points after starting the infusion. As body condition did not affect evaluated parameters, all data were pooled. Plasma glucose concentrations showed differences neither over time nor during or after infusion with propionate or control. Plasma amino acid concentrations rose over time (p < 0.001), but were similar for both infusions. Plasma propionylcarnitine rose markedly towards the end of the propionate infusion and decreased afterwards (p < 0.001), whereas 3-hydroxy-3-methylglutarylcarnitine was lower 30 (p = 0.005) and 60 min (p = 0.032) after ending propionate infusions and acetylcarnitine tended to fall at the same time points (p = 0.079; p = 0.080), suggesting inhibition of gluconeogenesis from pyruvate and amino acids, but initiation of propionate-induced gluconeogenesis. In conclusion, propionate absorbed from the colon is hypothesized to act as gluconeogenic substrate, regardless of the cat's body condition.