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This study investigated the diet-additive interactions of a Lactobacilli-based probiotic (Pro) and postbiotic (Post) on immune parameters and cecal microbiota composition, with subsequent effects on the metabolome in broilers. A completely randomized block design was employed with 2 diets [standard (SD), and challenge (CD)] and 3 additive conditions (Control, Pro, Post) involving 1,368 one-day-old male Ross 308 broilers equally distributed among 36 pens in a 42 d study. Diets were formulated to contain identical nutrient levels, with CD higher than SD in non-starch polysaccharide content by including rye and barley. Total non-specific serum Ig A, M and G concentrations were determined weekly from d14 to 35. Following vaccination, titres of specific antibodies binding Newcastle disease virus (NDV) and infectious bursal disease virus (IBDV) were measured. Microbiota composition was analyzed by 16S rRNA gene sequencing at d14 and 35, and α- and β-diversity indexes (Observed, Chao1, Bray, Jaccard) were calculated. Cecal short-chain fatty acids and the semi-polar metabolome were determined in the Control SD and all CD groups at d35. At d35, a diet-additive interaction was observed on cecal microbiota composition. Within SD, Pro and Post did not affect operational taxonomic units (OTU) abundance (adjusted-P > 0.05) and diversity indexes (P > 0.05). Within CD, Pro and Post affected the relative abundances of 37 and 44 OTUs, respectively (adjusted-P < 0.05), with Post but not Pro affecting β-diversity indexes (P = 0.041 and 0.064 for Bray and Jaccard, respectively). Within CD, Post increased cecal acetate (21%; P = 0.007) and butyrate (41%; P = 0.002) concentration and affected the concentration of 2 metabolites (adjusted-P < 0.05), while Pro affected 240 metabolites (adjusted-P < 0.05). No diet-additive interactions were observed on serum Ig (P > 0.05), except for IgM at d14 (P = 0.004). Diet composition, but not the additives, affected immune status parameters. The Pro and Post affected cecal microbiota composition only under dietary challenging conditions as previously reported for growth., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Dietary ingredient and nutrient composition may affect the efficacy of additives in broilers. Specific feed ingredients can represent dietary challenging conditions for broilers, resulting in impaired performances and health, which might be alleviated by dietary probiotics and postbiotics. We assessed the effects of a Lactobacilli probiotic (Pro) and postbiotic (Post) when added to a standard (SD) and challenge (CD) diet. A completely randomized block study with 2 diets (SD, CD) and 3 additive conditions (Control, Pro and Post) involving 1,368 one-day-old Ross male broilers, equally distributed among 36 pens, from d1 to d42 was conducted. Both diets were formulated to contain identical levels of nutrients, with CD formulated to be richer than SD in nonstarch polysaccharides using rye and barley as ingredients. Readout parameters included growth performance parameters, footpad lesions score, blood minerals and biochemical parameters, and tibia health, strength, and composition. Compared to SD, CD decreased BW (1,936 vs. 2,033 g; p = 0.001), increased FCR (p < 0.01) and impaired tibia health and strength (p < 0.05) at d35, thereby confirming the challenging effect of CD. Pro and Post increased BW in CD (+4.7 and +3.2%, respectively, at d35; P < 0.05) but not in the SD group, without affecting FCR. Independently of the diet, Pro increased plasma calcium, phosphorus and uric acid at d21 (+6.2, +7.4, and +15.5%, respectively) and d35 (+6.6, +6.2 and +21.0%, respectively) (P < 0.05) while Post increased plasma magnesium only at d21 (+11.3%; P = 0.037). Blood bile acids were affected by additives in an age- and diet-dependent manner, with some opposite effects between dietary conditions. Diet composition modulated Pro and Post effects on broiler growth performance. Additionally, Pro and Post affected animal metabolism and leg health diet-dependently for some but not all investigated parameters. Our findings show that the effects of pro- and postbiotics on the growth performance and physiology of broilers can be dependent on diet composition and thus possibly other factors affecting diet characteristics., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Copper is routinely supplemented to weanling pig diets at concentrations above nutritional requirements to enhance growth performance. We hypothesised that this effect depends on the source of Cu and its dietary concentration. We tested this in weaned pigs (26 d of age) over a 35-d period using a 2 × 3 factorial arrangement with two Cu-sources (CuSO 4 and Cu 2 O, monovalent copper oxide, CoRouge®) and three supplementary dietary Cu-levels (15, 80 and 160 mg Cu/kg) as respective factors. Increasing Cu level linearly increased (P < 0.001) final BW and daily gain. These effects tended (P = 0.09) to be greater with Cu 2 O than CuSO 4 . Feed conversion ratio decreased linearly (P < 0.001) with increasing dietary Cu content, independent of Cu source. Plasma Cu, Zn and Fe levels were unaffected, whereas liver Cu content increased quadratically (P < 0.001) with increasing dietary Cu content, with a larger increase (P < 0.001) with CuSO 4 than Cu 2 O. Bile Cu content increased quadratically (P = 0.025) with increasing Cu content, irrespective of Cu source. RT-qPCR analysis revealed that increasing Cu content quadratically (P = 0.009) increased duodenal but not ileal metallothionein 1A (MT1A) mRNA, with greater effect (P = 0.010) of CuSO 4 . Regardless of the Cu source, increasing Cu dose linearly increased (P = 0.006) duodenal DMT1/SLC11A2 mRNA but decreased ZIP4/SLC39A4 mRNA in duodenum (P < 0.001) and ileum (P < 0.005). ZnT10/SLC30A10 mRNA was significantly (P = 0.021) and numerically (P = 0.061) greater with Cu 2 O compared to CuSO 4 , in duodenum and ileum, respectively. Copper content quadratically modulated duodenal but not ileal transferrin receptor (P = 0.029) and ferric reductase CYBRD1 mRNA (P = 0.022). In hypothalamus, high Cu dose (P = 0.024) and Cu 2 O as source (P = 0.028) reduced corticotropin-releasing hormone (CRH) mRNA. Low versus high CuSO 4 increased corticotropin-releasing hormone receptor (CRHR2) mRNA, while low Cu 2 O had the opposite effect (P = 0.009). In conclusion, incremental Cu intake enhanced growth performance, with a tendency for a greater effect of Cu 2 O. The lower increase in duodenal MT1A mRNA and liver Cu content indicates that less Cu from Cu 2 O was absorbed by gut and sequestered in liver. Thus, high Cu absorption is not essential for its growth-promoting effect and dietary Cu may affect intestinal Fe and Zn absorption via the active, transcellular route. The effects on hypothalamic CRH and CRHR2 expression indicate a role for the hypothalamus in mediating the effects of Cu on growth performance., (Copyright © 2024 Wageningen University and Research (Netherlands). Published by Elsevier B.V. All rights reserved.)

The hypothesis that dietary inclusion of microbial phytase improves apparent calcium (Ca) digestibility thereby allowing a lower dietary Ca inclusion without compromising growth performance was tested. One-day-old male Ross 308 broilers (25 birds/pen, 9 pens/treatment) were assigned to 8 experimental diets containing one of 4 dietary Ca to retainable P (rP) ratios (1.3, 1.8, 2.3, and 2.8) with (1,000 FTU/kg) or without microbial phytase. On d 21 to 23, digesta from different intestinal segments of 8 birds per pen were collected to determine apparent Ca and P digestibility. Mid duodenal mucosa was collected for expression of Ca (CaBP-D28k, PMCA1) and P (NaPi-IIb, PiT-1, PiT-2, and XPR1) transporters by RT-qPCR. Dietary phytase inclusion in low Ca/rP diets increased Ca digestibility in the distal ileum (P interaction  = 0.023) but not the proximal or distal jejunum. Broilers receiving the lowest Ca/rP displayed the lowest body weight gain, highest feed conversion ratio (P < 0.001), and lowest tibia strength, irrespective of dietary phytase inclusion. Incremental dietary Ca/rP linearly reduced P digestibility to a greater extent in the absence of phytase in the distal jejunum and ileum (P interaction  = 0.021 and 0.001, respectively). Incremental dietary Ca/rP linearly reduced serum P more in phytase-free diets (P interaction < 0.001), and lowered duodenal expression of P transporters NaPi-IIb, PiT-2, and XPR1 (P = 0.052, 0.071 and 0.028, respectively). Incremental dietary Ca/rP linearly increased (P < 0.001) serum Ca irrespective of phytase inclusion, accompanied by a lower (P < 0.001) duodenal expression of Ca transporters CaSR, CaBP-D28k and PMCA1 and Ca-pore forming claudins CLDN-2 and CLDN-12. Dietary phytase increased (P = 0.026) NaPi-IIb but reduced (P = 0.029) CLDN-2 expression. Incremental Ca/rP reduced Ca and P digestibility, increased serum Ca, lowered serum P and inhibited mRNA levels of Ca and P-related transporters, indicating that these transporters and CLDN contribute to the observed effect of dietary Ca and phytase on Ca and P absorption. Despite the improvement in Ca digestibility, dietary phytase did not restore the compromised growth performance and tibia strength of broilers fed a Ca-deficient diet, leading to rejection of the hypothesis., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

The impact of transgenerational effects on growth performance and immunity has not yet been studied extensively within the poultry husbandry sector. An important factor is the impact of the hens on the physical well-being and fitness to the environment of the offspring. This study is the first to investigate the effect of stimulating the maternal innate immune system with lipopolysaccharides (LPS) or β-glucan on growth performance and immune responses in the next generation. Transgenerational effects and consequences of these maternal treatments were further examined using a necrotic enteritis (NE) challenge model in the offspring. We show that offspring of LPS-treated broiler breeders have a higher feed efficiency from 14 to 21 days of age, that is, the period just after the NE challenge. Moreover, more broiler chickens with intestinal lesions after the NE challenge were found in the offspring of the LPS-treated broiler breeders. Both the LPS and β-glucan maternal treatments resulted in transgenerational effects on blood-derived monocytes by showing a tendency of decreased IL1β mRNA levels after ex vivo LPS stimulation. These data are a first indication that broiler breeder hens can affect immune responsiveness and feeding efficiency of their offspring in a transgenerational manner., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Fasting older broiler chickens (>7 d of age) enlarges the intestinal tight junction (TJ) pore size, resulting in high paracellular intestinal permeability. Broiler chickens often do not receive feed and water (nutrition) directly after hatch, which may result in fasting up to 72 h of age. Whether perinatal fasting affects intestinal permeability is minimally studied. We therefore investigated whether delayed access to nutrition after hatch increases intestinal permeability, compared with broilers receiving early access to nutrition. Therefore, 432 hatched broilers received nutrition 72 h after hatch (delayed nutrition [DN]) or directly after hatch (early nutrition [EN]) and were reared under similar conditions until 14 d of age. Two hours after application of an oral pulse dose (3.85 mg) of fluorescein isothiocyanate-dextran (4000 Da) at 4, 10, and 14 d of age, blood plasma concentrations of the marker were measured in 24 to 36 broilers per treatment and time point. Marker concentration in plasma did not differ between DN and EN broilers at any age. The villus width measured in at least 8 broilers per treatment was smaller in DN than in EN broilers at 4 d for both the ileum (92 ± 3 μm vs. 121 ± 4; P < 0.001) and colon (100 ± 3 vs. 120 ± 4; P < 0.01). Real-time quantitative PCR revealed that the expression of TJ protein claudin 3 in the ceca was elevated in DN, compared with EN broilers at 4 d of age, whereas that of zonula occludens 1 in the ileum was reduced. Expression of host defense-related genes was reduced in DN, compared with EN broilers, in the ileum (cyclo-oxygenase 2, mucin 2) and ceca (interleukin 1β, cyclo-oxygenase 2). We conclude that 72-hour DN reduced the BW up to 14 d of age, coinciding with transient effects on the villus width in the ileum and colon, and divergent expression of genes involved in TJ formation and host defense. These effects likely reflect the delayed onset of intestinal and immune development in DN, compared with EN broilers, while DN does not fundamentally alter intestinal permeability., (Copyright © 2020. Published by Elsevier Inc.)

The hypothesis was tested that an increased digestion of coarse compared with fine limestone can alleviate the negative effects of a low dietary Ca/P ratio on the growth performance and characteristics of tibia strength (CTS) in broilers. A total of 1,152 Ross 308 broiler chickens received a standard commercial starter feed from day 0 to 13. From day 14 onward, birds received 1 of 12 diets containing 1 of 6 Ca/P ratios (0.50, 0.75, 1.00, 1.25, 1.50, and 1.75) and 1 of 2 limestone particle sizes (<500 [fine] and 500 to 2,000 [coarse] μm) in a study with a 6 × 2 factorial arrangement of treatments. Total P content was fixed at 5.5 g/kg for all treatment diets. Each treatment was replicated 6 times with 16 birds per replicate pen. On day 20 and 21, twelve birds per pen were randomly selected from 4 of the 6 replicate pens for tibia analysis and digesta collection from different gut segments. The apparent Ca digestibility was higher for fine than coarse limestone in the jejunum (P = 0.043). However, this difference in Ca digestibility disappeared for the low, whereas it remained for the high Ca/P ratios in the proximal (P interaction = 0.067) and distal (P interaction = 0.052) ileum. In addition, coarse limestone improved apparent P digestibility in the proximal and distal ileum (P < 0.001) but not in the jejunum (P = 0.305). Regardless of limestone particle size, reducing dietary Ca/P ratio linearly improved apparent Ca and P digestibility in the proximal and distal ileum (P < 0.001). Moreover, decreasing dietary Ca/P ratio linearly (P < 0.001) and quadratically (P < 0.046) reduced the CTS. Reducing dietary Ca/P ratio linearly (P < 0.003) and quadratically (P ≤ 0.006) decreased body weight gain and increased feed conversion ratio. For both fine and coarse limestone, the optimal Ca/P ratio was 1.00 to 1.25 to optimize apparent Ca and P digestibility while maintaining growth performance and CTS. Reducing Ca/P ratio from 1.75 to 1.00 improved distal ileal Ca and P apparent digestibility from 36.6 to 53.7% and 48.0 to 58.3%, respectively. In conclusion, coarse limestone is equally digestible with fine limestone at a low Ca/P ratio but is less digestible at a high Ca/P ratio, and the optimal Ca/P ratio in the diet is 1.00 to 1.25 for both fine and coarse limestone., (Copyright © 2020. Published by Elsevier Inc.)

Objective: The new 2019 guideline of the European Society for Vascular Surgery (ESVS) recommends consideration for elective iliac artery aneurysm (eIAA) repair when the iliac diameter exceeds 3.5 cm, as opposed to 3.0 cm previously. The current study assessed diameters at time of eIAA repair and ruptured IAA (rIAA) repair and compared clinical outcomes after open surgical repair (OSR) and endovascular aneurysm repair (EVAR)., Methods: This retrospective observational study used the nationwide Dutch Surgical Aneurysm Audit (DSAA) registry that includes all patients who undergo aorto-iliac aneurysm repair in the Netherlands. All patients who underwent primary IAA repair between 1 January 2014 and 1 January 2018 were included. Diameters at time of eIAA and rIAA repair were compared in a descriptive fashion. The anatomical location of the IAA was not registered in the registry. Patient characteristics and outcomes of OSR and EVAR were compared with appropriate statistical tests., Results: The DSAA registry comprised 974 patients who underwent IAA repair. A total of 851 patients were included after exclusion of patients undergoing revision surgery and patients with missing essential variables. eIAA repair was carried out in 713 patients, rIAA repair in 102, and symptomatic IAA repair in 36. OSR was performed in 205, EVAR in 618, and hybrid repairs and conversions in 28. The median maximum IAA diameter at the time of eIAA and rIAA repair was 43 (IQR 38-50) mm and 68 (IQR 58-85) mm, respectively. Mortality was 1.3% (95% CI 0.7-2.4) after eIAA repair and 25.5% (95% CI 18.0-34.7) after rIAA repair. Mortality was not significantly different between the OSR and EVAR subgroups. Elective OSR was associated with significantly more complications than EVAR (intra-operative: 9.8% vs. 3.6%, post-operative: 34.0% vs. 13.8%, respectively)., Conclusion: In the Netherlands, most eIAA repairs are performed at diameters larger than recommended by the ESVS guideline. These findings appear to support the recent increase in the threshold diameter for eIAA repair., (Copyright © 2020 European Society for Vascular Surgery. Published by Elsevier B.V. All rights reserved.)

Russell's viper (Daboia russelii) is, together with Naja naja, Bungarus caeruleus and Echis carinatus, a member of the medically important 'Big Four' species responsible for causing a large number of morbidity and mortality cases across the Indian subcontinent. Despite the wide distribution of Russell's viper and the well-documented ubiquity of the phenomenon of geographic variability of intraspecific snake venom composition, Indian polyvalent antivenoms against the "Big Four" venoms are raised against venoms sourced mainly from Chennai in the southeastern Indian state of Tamil Nadu. Biochemical and venomics investigations have consistently revealed notable compositional, functional, and immunological differences among geographic variants of Russell's viper venoms across the Indian subcontinent. However, these studies, carried out by different laboratories using different protocols and involving venoms from a single geographical region, make the comparison of the different venoms difficult. To bridge this gap, we have conducted bioactivities and proteomic analyses of D. russelii venoms from the three corners of the Indian subcontinent, Pakistan, Bangladesh, and Tamil Nandu (India) and Sri Lanka, along with comparative in vivo neutralization and in vitro third-generation antivenomics of antivenoms used in India, Bangladesh and Sri Lanka. These analyses let us to propose two alternative routes of radiation for Russell's viper in the Indian subcontinent. Both radiations, towards the northeast of India and Bangladesh and towards south India and Sri Lanka, have a common origin in Pakistan, and provide a phylovenomics ground for rationalizing the geographic variability in venom composition and their distinct immunoreactivity against available antivenoms. BIOLOGICAL SIGNIFICANCE: Russell's viper (Daboia russelii), the Indian cobra (Naja naja), the common krait (Bungarus caeruleus), and the saw-scaled viper (Echis carinatus) constitute the 'Big Four' snake species responsible for most snakebite envenomings and deaths in the Indian subcontinent. Despite the medical relevance of Daboia russelii, and the well documented variations in the clinical manifestations of envenomings by this wide distributed species, which are doubtless functionally related to differences in venom composition of its geographic variants, antivenoms for the clinical treatment of envenomings by D. russelii across the Indian subcontinent are invariably raised using venom sourced mainly from the southeastern Indian state of Tamil Nadu. We have applied a phylovenomics approach to compare the venom proteomes of Russell's vipers from the three corners of the Indian subcontinent, Pakistan, Bangladesh, and South India/Sri Lanka, and have assessed the in vitro (third-generation antivenomics) and in vivo preclinical efficacy of a panel of homologous antivenoms. The identification of two dispersal routes of ancestral D. russelii into the Indian subcontinent provides the ground for rationalizing the variability in composition and immunoreactivity of the venoms of extant geographic variants of Russell's viper. Such knowledge is relevant for envisioning strategies to improve the clinical coverage of anti- D. russelii antivenoms., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Urea recycling, with urea originating from catabolism of amino acids and hepatic detoxification of ammonia, is particularly relevant for ruminant animals, in which microbial protein contributes substantially to the metabolizable protein supply. However, the quantitative contribution of urea recycling to protein anabolism in calves during the transition from preruminants (milk-fed calves) to ruminants [solid feed (SF)-fed calves] is unknown. The aim of this study was to quantify urea recycling in milk-fed calves when provided with low-protein SF. Forty-eight calves [164 ± 1.6 kg body weight (BW)] were assigned to 1 of 4 SF levels [0, 9, 18, and 27 g of dry matter (DM) SF · kg BW(-0.75) · d⁻¹] provided in addition to an identical amount of milk replacer. Urea recycling was quantified after a 24-h intravenous infusion of [¹⁵N₂]urea by analyzing urea isotopomers in 68-h fecal and urinary collections. Real-time qPCR was used to measure gene expression levels of bovine urea transporter B (bUTB) and aquaglyceroporin-3 and aquaglyceroporin-7 in rumen wall tissues. For every incremental gram of DM SF intake (g DM · kg(0.75)), nitrogen intake increased by 0.70 g, and nitrogen retention increased by 0.55 g (P < 0.01). Of this increase in nitrogen retention, 19% could be directly explained by urea recycling. Additionally, part of the observed increase in nitrogen retention could be explained by the extra protein provided by the SF and likely by a greater efficiency of postabsorptive use of nitrogen for gain. Ruminal bUTB abundance increased (P < 0.01) with SF provision. Aquaglyceroporin-3 expression increased (P < 0.01) with SF intake, but aquaglyceroporin-7 expression did not. We conclude that in addition to the increase in digested nitrogen, urea recycling contributes to the observed increase in nitrogen retention with increasing SF intake in milk-fed calves. Furthermore, ruminal bUTB and aquaglyceroporin-3 expression are upregulated with SF intake, which might be associated with urea recycling., (© 2014 American Society for Nutrition.)

Stearoyl-CoA desaturase (SCD) in the bovine mammary gland introduces a cis-double bond at the Δ9 position in a wide range of fatty acids (FA). Several long-chain polyunsaturated fatty acids (PUFA) inhibit expression of SCD, but information on the effect of short-chain fatty acids on mammary SCD expression is scarce. We used a bovine mammary cell line (MAC-T) to assess the effect of acetic acid (Ac) and β-hydroxybutyric acid (BHBA) in comparison with the effect of various long-chain fatty acids on the mRNA expression of the lipogenic enzymes SCD, acetyl-CoA carboxylase (ACACA), fatty acid synthase (FASN) and their associated gene regulatory proteins sterol regulatory element binding transcription factor 1 (SREBF1), insulin-induced gene 1 protein (INSIG1) and peroxisome proliferator-activated receptor alpha (PPARA)and peroxisome proliferator-activated receptor delta (PPARD) by quantitative real-time PCR. MAC-T cells were treated for 12 h without FA additions (CON) or with either 5 mM Ac, 5 mM BHBA, a combination of 5 mM Ac + 5 mM BHBA, 100 μM C16:0, 100 μM C18:0, 100 μM C18:1 cis-9, 100 μM C18:1 trans-11, 100 μM C18:2 cis-9,12 or 100 μM C18:3 cis-9,12,15. Compared with control, mRNA expression of SCD1 was increased by Ac (+61%) and reduced by C18:1 cis-9 (-61%), C18:2 cis-9,12 (-84%) and C18:3 cis-9,12,15 (-88%). In contrast to native bovine mammary gland tissue, MAC-T cells did not express SCD5. Expression of ACACA was increased by Ac (+44%) and reduced by C18:2 cis-9,12 (-48%) and C18:3 cis-9,12,15 (-49%). Compared with control, FASN expression was not significantly affected by the treatments. The mRNA level of SREBF1 was not affected by Ac or BHBA, but was reduced by C18:1 cis-9 (-44%), C18:1 trans-11 (-42%), C18:2 cis-9,12 (-62%) and C18:3 cis-9,12,15 (-68%) compared with control. Expression of INSIG1 was downregulated by C18:0 (-37%), C18:1 cis-9 (-63%), C18:1 trans-11 (-53%), C18:2 cis-9,12 (-81%) and C18:3 cis-9,12,15 (-91%). Both PPARA and PPARD expression were not significantly affected by the treatments. Our results show that Ac upregulated mRNA expression of SCD1 and ACACA in MAC-T cells. The opposite effect of the PUFA C18:2 cis-9,12 and C18:3 cis-9,12,15 on the these genes and the failure of Ac to mimic the PUFA-inhibited SREBF1 and INSIG1 mRNA expression, suggest that Ac can stimulate mammary lipogenesis via a transcriptional regulatory mechanism different from PUFA.

Epidermal growth factor receptor (EGFR) activation is negatively regulated by protein kinase C (PKC) signaling. Stimulation of A431 cells with EGF, bradykinin or UTP increased EGFR phosphorylation at Thr654 in a PKC-dependent manner. Inhibition of PKC signaling enhanced EGFR activation, as assessed by increased phosphorylation of Tyr845 and Tyr1068 residues of the EGFR. Diacylglycerol is a physiological activator of PKC that can be removed by diacylglycerol kinase (DGK) activity. We found, in A431 and HEK293 cells, that the DGKθ isozyme translocated from the cytosol to the plasma membrane, where it co-localized with the EGFR and subsequently moved into EGFR-containing intracellular vesicles. This translocation was dependent on both activation of EGFR and PKC signaling. Furthermore, DGKθ physically interacted with the EGFR and became tyrosine-phosphorylated upon EGFR stimulation. Overexpression of DGKθ attenuated the bradykinin-stimulated, PKC-mediated EGFR phosphorylation at Thr654, and enhanced the phosphorylation at Tyr845 and Tyr1068. SiRNA-induced DGKθ downregulation enhanced this PKC-mediated Thr654 phosphorylation. Our data indicate that DGKθ translocation and activity is regulated by the concerted activity of EGFR and PKC and that DGKθ attenuates PKC-mediated Thr654 phosphorylation that is linked to desensitisation of EGFR signaling., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

Studies suggested that in human adults, linoleic acid (LA) inhibits the biosynthesis of n-3 long-chain polyunsaturated fatty acids (LC-PUFA), but their effects in growing subjects are largely unknown. We used growing pigs as a model to investigate whether high LA intake affects the conversion of n-3 LC-PUFA by determining fatty acid composition and mRNA levels of Δ5- and Δ6 desaturase and elongase 2 and -5 in liver and brain. In a 2 × 2 factorial arrangement, 32 gilts from eight litters were assigned to one of the four dietary treatments, varying in LA and α-linolenic acid (ALA) intakes. Low ALA and LA intakes were 0.15 and 1.31, and high ALA and LA intakes were 1.48 and 2.65 g/kg BW0.75 per day, respectively. LA intake increased arachidonic acid (ARA) in liver. ALA intake increased eicosapentaenoic acid (EPA) concentrations, but decreased docosahexaenoic acid (DHA) (all P < 0.01) in liver. Competition between the n-3 and n-6 LC-PUFA biosynthetic pathways was evidenced by reductions of ARA (>40%) at high ALA intakes. Concentration of EPA (>35%) and DHA (>20%) was decreased by high LA intake (all P < 0.001). Liver mRNA levels of Δ5- and Δ6 desaturase were increased by LA, and that of elongase 2 by both ALA and LA intakes. In contrast, brain DHA was virtually unaffected by dietary LA and ALA. Generally, dietary LA inhibited the biosynthesis of n-3 LC-PUFA in liver. ALA strongly affects the conversion of both hepatic n-3 and n-6 LC-PUFA. DHA levels in brain were irresponsive to these diets. Apart from Δ6 desaturase, elongase 2 may be a rate-limiting enzyme in the formation of DHA.

The aim of this study was to determine the effects of supplementing unprotected dietary unsaturated fatty acids (UFAs) from different plant oils on gene expression in the mammary gland of grazing dairy cows. A total of 28 Holstein-Friesian dairy cows in mid-lactation were blocked according to parity, days in milk, milk yield and fat percentage. The cows were then randomly assigned to four UFA sources based on rapeseed, soybean, linseed or a mixture of the three oils for 23 days, after which, all 28 cows were switched to a control diet for an additional 28 days. On the last day of both periods, mammary gland biopsies were taken to study genome-wide differences in gene expression on Affymetrix GeneChip® Bovine Genome Arrays (no. 900493) by ServiceXS (Leiden, The Netherlands). Supplementation with UFAs resulted in increased milk yield but decreased milk fat and protein percentages. Furthermore, the proportion of de novo fatty acids (FAs) in the milk was reduced, whereas that of long-chain FAs increased. Applying a statistical cut-off of false discovery rate of q-values <0.05 together with an absolute fold change of 1.3, a total of 972 genes were found to be significantly affected through UFA supplementation, indicating that large transcriptional adaptations occurred in the mammary gland when grazing dairy cows were supplemented with unprotected dietary UFA. Gene sets related to cell development and remodeling, apoptosis, nutrient metabolic process, as well as immune system response were predominantly downregulated during UFA supplementation. Such molecular knowledge on the physiology of the mammary gland might provide the basis for further functional research on dairy cows.

A method for the identification and quantitation of nucleotide pools in lymphocytes and leukemic blasts is described. Separation of these metabolites was performed by anion-exchange high-performance liquid chromatography using a pH and concentration gradient consisting of several linear steps. The mono-, di- and triphosphates of adenosine, cytidine, guanosine, inosine, uridine and xanthosine could conveniently be separated together with NAD+, cyclic AMP, NADP+ and uridinediphosphoglucose (UDPG). In addition, data on the accuracy and precision of the method are given and its potentials for use in the analysis of nucleotide pools in leukemic lymphoblasts are illustrated.

A method is presented for the separation and quantitative determination of compounds normally related to purine and pyrimidine metabolism in biological material. The retention behaviour of nucleobases, ribonucleosides, deoxyribonucleosides and cyclic ribonucleotides has been systematically investigated by reversed-phase high-performance liquid chromatography using a non-linear gradient. Ultimately a separation of the purine and pyrimidine compounds was achieved in a 35-min run with an average detection limit of 5-10 pmol per injection. Recoveries of standards added to urine, plasma or serum were 96 +/- 5%.