Pair-fed Research Articles

Prenatal Alcohol Exposure Disrupts Performance in a Differential Reinforcement of Low Rates Task Specifically in Adolescent Male Rats.

Prenatal alcohol exposure (PAE) can lead to a wide range of adverse effects in humans, including impaired self-control and increased impulsive behavior. Deficits in self-control can interfere with academic performance and have lasting impacts. In the present study, a rodent model of PAE was used to assess impulsivity through operant conditioning. Pregnant rats were assigned to one of three groups: ad-lib control (CON), pair-fed (PF), and alcohol-exposed (ALC). ALC rats were given a liquid diet containing 6% alcohol, PF rats were yoked to an ALC rat and given a CON liquid diet, and CON rats received ad libitum food. Operant conditioning was used to evaluate extinction in adolescents (Experiment 1) and differential reinforcement of low rates (DRL) in adolescents and adults (Experiment 2). PAE resulted in an increase in responses and resets during DRL testing, indicative of impaired self-control, an effect that was only observed in adolescent males. Females, regardless of age, did not show increased impulsivity following PAE. This indicates that children with PAE may exhibit attentional deficits similar to those with attention deficit hyperactivity disorder, with males at a higher risk.

Unraveling the role of long non-coding RNAs in chronic heat stress-induced muscle injury in broilers

BackgroundChronic heat stress (CHS) is a detrimental environmental stressor with a negative impact on the meat quality of broilers. However, the underlying mechanisms are not fully understood. This study investigates the effects of CHS on long non-coding RNA (lncRNA) expression and muscle injury in broilers, with a focus on its implications for meat quality.ResultsThe results showed that CHS diminished breast muscle yield, elevated abdominal fat deposition, induced cellular apoptosis (P < 0.05), and caused myofibrosis. Transcriptomic analysis revealed 151 differentially expressed (DE) lncRNAs when comparing the normal control (NC) and HS groups, 214 DE lncRNAs when comparing the HS and PF groups, and 79 DE lncRNAs when comparing the NC and pair-fed (PF) groups. After eliminating the confounding effect of feed intake, 68 lncRNAs were identified, primarily associated with cellular growth and death, signal transduction, and metabolic regulation. Notably, the apoptosis-related pathway P53, lysosomes, and the fibrosis-related gene TGF-β2 were significantly upregulated by lncRNAs.ConclusionsThese findings indicate that chronic heat stress induces cellular apoptosis and muscle injury through lncRNA, leading to connective tissue accumulation, which likely contributes to reduced breast muscle yield and meat quality in broilers.

Effects of chronic heat stress on Ca2+ homeostasis, apoptosis, and protein carbonylation profiles in the breast muscle of broilers

Heat stress (HS) largely impairs the quality of broiler breast meat through protein oxidative modification. This study aimed to investigate the carbonylation pattern of Ca2+ channels and apoptotic proteins in the breast muscle of heat-stressed broilers. A total of 144 twenty-eight-day-old male Arbor Acres broilers were randomly divided into three treatment groups. The normal control (NC) group was kept at 22°C and provided with unlimited feed. The HS group was exposed to 32°C and provided with unlimited feed. The pair-fed (PF) group was kept at 22°C and given an amount of feed equivalent to that consumed by the HS group on the previous day. Results showed that broilers under HS conditions had a higher respiratory rate than those in NC and PF groups (P < 0.05). HS disrupted the morphology and structure of breast muscle fibers by decreasing the average diameters and average density of myofibers compared to the NC group (P < 0.05). HS increased the mean fluorescence intensity of the positive carbonyl signal in breast muscle compared with the NC group (P < 0.05). Besides, the pectoral Ca2+ concentration in the sarcoplasmic reticulum, cytoplasm, and mitochondria was elevated by HS when compared with the NC group (P < 0.05). In comparison to the NC and PF groups, HS increased the apoptosis rate and caspase-3 activity in the breast muscle (P < 0.05). Furthermore, HS elevated the relative protein expressions of plasma membrane Ca2+-ATPase, Na+/Ca2+ exchanger 1, and sarco/endoplasmic reticulum calcium transport ATPase 1 compared to the NC group (P < 0.05). Higher relative protein expression of μ-calpain and lower relative protein expression of cytosolic cytochrome complex were found in the HS group than the NC group (P < 0.05). HS decreased the carbonylation levels of transient receptor potential canonical 1 and inositol 1,4,5-trisphosphate receptor compared to the NC group (P < 0.05). Additionally, the carbonylation levels of cleaved caspase-3 and precursor caspase-9 were increased and decreased, respectively, by HS treatment compared to the NC group (P < 0.05). In conclusion, HS damages the myofiber based on Ca2+ dyshomeostasis and apoptosis, which are potentially associated with protein carbonylation. These results shed new light on the possible mechanism behind the development of poor meat quality in broilers due to HS.

The T-cell receptor β chain CDR3 insights of bovine liver immune repertoire under heat stress.

The liver plays a dual role in regulating temperature and immune responses. Examining the influence of heat stress (HS) on liver T cells contributes significantly to understanding the intricate interplay between the immune system and hepatic tissues under thermal stress. This study focused on investigating the characteristics of the T-cell receptor (TCR) β chain CDR3 repertoire in bovine liver samples under both HS and pairfed (PF) environmental conditions. Sequencing data from six samples sourced from the GEO database underwent annotation. Utilizing immunarch and VDJtool software, the study conducted comprehensive analyses encompassing basic evaluation, clonality assessment, immune repertoire comparison, diversity estimation, gene usage profiling, VJ gene segment pairing scrutiny, clonal tracking, and Kmers analysis. All four TCR chains, namely α, β, γ, and δ, were detected, with the α chains exhibiting the highest detection frequency, followed closely by the β chains. The prevalence of αβ TCRs in bovine liver samples underscored their crucial role in governing hepatic tissue's physiological functions. The TCR β CDR3 repertoire showcased substantial inter-individual variability, featuring diverse clonotypes exhibiting distinct amino acid lengths. Intriguingly, HS cattle displayed heightened diversity and clonality, suggesting potential peripheral T cell migration into the liver under environmental conditions. Notably, differential VJ gene pairings were observed in HS cattle compared to the PF, despite individual variations in V and J gene utilization. Additionally, while most high-frequency amino acid 5-mers remained consistent between the HS and PF, GELHF, and YDYHF were notably prevalent in the HS group. Across all samples, a prevalent trend of high-frequency 5mers skewed towards polar and hydrophobic amino acids was evident. This study elucidates the characteristics of liver TCR β chain CDR3 repertoire under HS conditions, enhancing our understanding of HS implications.

Benefits of calorie restriction in mice are mediated via energy imbalance, not absolute energy or protein intake

Caloric restriction (CR) results in reduced energy and protein intake, raising questions about protein restriction’s contribution to CR longevity benefits. We kept ad libitum (AL)–fed male C57BL/6J mice at 27°C (AL27) and pair-fed (PF) mice at 22°C (22(PF27)). The 22(PF27) group was fed to match AL27 while restricted for calories due to cold-induced metabolism. The 22(PF27) mice had significantly lower body weight, lean mass, fat mass, leptin, IGF-1, and TNF-α levels than AL27 mice (p<0.001 for all). Manipulations over ~11 weeks resulted in significant differences in body temperature, physical activity, and expression of key genes linked to hunger in the hypothalamus. Survival was significantly greater in 22(PF27) compared to AL27 overall (p<0.001). CR in the context of equivalent energy and protein intake resulted in hormonal, metabolic, and physiological benefits and extended longevity. Hence, energy imbalance, rather than low energy or protein intake per se, mediates the benefits of CR.

Effects of Artemisia annua supplementation on the performance and gut health of laying hens challenged with mixed Eimeria species.

Coccidiosis outbreaks in susceptible laying hens can significantly decrease egg production and cause substantial economic loss to the egg industry. The supplementation of poultry diets with chemotherapeutic agents is limited due to antimicrobial resistance and residue in poultry meat or processed products. Therefore, alternative strategies to control coccidiosis are needed, and Artemisia annua (AA) might have the potential to be a phytogenic feed additive, an alternative to anticoccidial agents. This study aimed to investigate the effect of the dietary supplementation of powdered AA on the performance and gut health of laying hens infected with coccidiosis by Eimeria spp. A total of 225 Hy-Line W-36 laying hens at 23 weeks of age were allocated into 5 treatment groups: 1) control (NC), 2) pair-fed (PF) control, 3) challenged control (CC), 4) CC with dietary inclusion of 0.5% AA (0.5AA), and v) CC with dietary inclusion of 1% AA (1AA). The hens in the CC, 0.5AA, and 1AA groups were orally inoculated with sporulated oocysts of Eimeria maxima (12,500), Eimeria tenella (12,500), and Eimeria acervulina (62,500) at week 25. The PF hens received the same amount of feed consumed by the CC hens from 0-14days post-inoculation (dpi) of Eimeria spp. The performance of the laying hens, including body weight (BW), hen-day egg production (HDEP), feed intake (FI), and feed conversion ratio (FCR), was measured weekly. Additionally, markers of intestinal health, including gut permeability, lesion score, intestinal morphometry, and immune responses, were evaluated at 6, 14, and 21 dpi. At 6 and 14 dpi, laying hens challenged with Eimeria spp. had a lower BW than PF and NC hens (p < 0.0001). Supplementation of 1% AA improved the HDEP by 8.1% compared to CC hens; however, it was still 15.4% lower than that of PF hens (p < 0.0001). The inclusion of 1% AA did not have any beneficial effect on FI; however, the FCR was improved by 0.61 (2.46) than that of CC hens (3.07; p < 0.0001). The inclusion of 1% AA reduced the severity of the intestinal lesions and increased the recovery of intestinal villi (p < 0.05). Additionally, gut permeability was significantly different between the challenged and non-challenged hens; however, among the challenged hens, the inclusion of AA reduced the gut permeability by 29% compared to CC hens (p < 0.0001). Furthermore, the inclusion of 0.5% AA reduced the inflammatory responses in the infected hens. Dietary inclusion of AA partially restored the performance and gut health of the laying hens and modulated their inflammatory immune response following Eimeria infection; however, further studies are needed to better understand the mode of action and effective dosages to improve the gut health without negative impacts on the performance.

Paneth cell-specific TCF4 down-regulation potentiates alcohol-induced dysbiosis of gut microbiota and tissue injury at the gut-liver axis

Introduction: Alcohol misuse results in hepatitis, pancreatitis, and neurodegenerative disorders. Alcohol consumption alters gut microbiota and increases gut permeability, leading to endotoxemia and multiple organ damage. However, the cellular and molecular mechanisms of alcohol-induced microbiota dysbiosis and barrier dysfunction remain poorly defined. Antimicrobial peptides such as α-defensins, secreted by the Paneth cells, regulate gut microbiota homeostasis. We previously reported that ethanol (EtOH) feeding reduces α-defensin expression in mice. α-Defensin expression is regulated by Wnt signaling and T-cell factor 4 (TCF4). We investigated the role of Paneth cell TCF4 in alcohol-induced microbiota dysbiosis and tissue injury at the gut-liver axis. Materials and methods: TCF4fl/wt (WT) and TCF4fl/wt-Defa6Cre (KD; heterozygous Paneth cell-specific knockout of TCF4) mice were fed a Lieber-DiCarli liquid diet with or without EtOH (0% 2d, 1% 2d, 2% 2d, 4% 1wk, 5% 1wk, &amp; 6% 1wk) (EF). Control groups were pair-fed (PF) isocaloric EtOH-free diet. Intestinal permeability was measured by the vascular-to-luminal flux of FITC-inulin (6 kDa) in vivo, and tight junction (TJ) and adherens junction (AJ) integrity were assessed by confocal microscopy for occludin, ZO-1, E-cadherin, and β-catenin. Paneth cell function and mucosal inflammation were assessed by RT-PCR for defensins and cytokines. Gut microbiota was analyzed by 16S rRNA sequencing of colonic flushing. Plasma lipopolysaccharide (LPS) and cytokines were measured to evaluate endotoxemia and systemic inflammation. Liver damage was assessed by measuring plasma AST, liver triglyceride, and cytokine mRNA. Results and conclusion: EtOH reduced TCF4 expression in the ileum in both WT and KD mice; lowest TCF4 expression was observed in KD-EF mice. Similarly, the expression of DEFA5 and DEFA6 was reduced by EtOH in WT and KD mice, with the lowest expression in KD-EF mice. EtOH-induced mucosal permeability was associated with reduced staining for TJ and AJ proteins, indicating the mucosal barrier dysfunction. EtOH increased intestinal mRNA for IL-1β, IL-6, TNFα, and MCP-1, indicating the mucosal inflammatory response. EtOH-induced gut permeability, TJ/AJ disruption, and mucosal inflammation were significantly higher in KD-EF mice than in WT-EF mice. Shannon index analysis indicated that EtOH reduced α-diversity of microbiota in KD but not in WT mice. Firmicutes: Bacteriodetes ratio and the abundance of Verrucomicrobia were reduced by EtOH in WT but not in KD mice. The abundance of Bacillus was increased by EtOH both in WT and KD mice; however, this effect was more severe in KD mice. EtOH-induced plasma LPS and cytokine elevations were higher in KD mice than in WT mice. Elevation of plasma AST, liver triglyceride, and liver cytokine mRNA indicated EtOH-induced liver damage; the damage was more severe in KD mice. These data indicate that Paneth cell TCF4 downregulation exacerbates alcohol-induced microbiota dysbiosis, gut barrier dysfunction, endotoxemia, systemic inflammation, and liver damage; suggesting that Paneth cell Wnt signal downregulation plays a crucial role in alcohol-associated organ damage. NIH/NIAA - R01AA012307, AA029270; Veterans Administration - IO1BX003014. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Sex-Specific Effect of Ethanol on Colon Content Lipidome in a Mice Model Using Nontargeted LC/MS.

Consumption of alcohol has widespread effects on the human body. The organs that are most significantly impacted are the liver and digestive system. When alcohol is consumed, it is absorbed in the intestines and processed by the liver. However, excessive alcohol use may affect gut epithelial integrity, microbiome composition, and lipid metabolism. Despite past studies investigating the effect of ethanol on hepatic lipid metabolism, the focus on colonic lipid metabolism has not been well explored. In this study, we investigated the sex-specific effect of ethanol on the colonic content lipidome in a mouse model using nontargeted liquid chromatography-mass spectrometry. Comprehensive lipidome analysis of colonic flush samples was performed using ethanol-fed (EF) and pair-fed (PF) mice of each sex. Partial least-squares discriminant analysis revealed that ethanol altered colonic lipid composition largely in male mice compared with female mice. A significant increase in free fatty acids, ceramides, and hexosylceramides and decreased phosphatidylglycerols (PG) was observed in the EF group compared to the PF group in male mice. Phosphatidylethanolamine (PE) levels were increased significantly in the EF group of both sexes compared to the PF group. The volcanic plot shows that PG (O-15:1/15:0) and PE (O-18:2/15:0) are common markers that are increased in both sexes of the EF group. In addition, decreased fatty acid esters of hydroxy fatty acids (FAHFA) were observed specifically in the EF group of female mice. Overall, a significant variation in the mice colonic content lipidome between the EF and PF groups was observed. Target pathways, such as sphingolipid metabolism in males, FAHFA in females, and PE metabolism in both sexes, were suggested. This study provides new insight into the sex-dependent lipid change associated with alcohol-induced gut-microbiota dysfunction and its potential health impacts.

Intravenous lipopolysaccharide challenge in early- versus mid-lactation dairy cattle. II: The production and metabolic responses

Most immunometabolic research uses mid-lactation (ML) cows. Cows in early lactation (EL) are in a presumed state of immune suppression/dysregulation and less is known about how they respond to a pathogen. Study objectives were to compare the production and metabolic responses to i.v. LPS and to differentiate between the direct effects of immune activation and the indirect effects of illness-induced hypophagia in EL and ML cows. Cows in EL (n = 11; 20 ± 2 DIM) and ML (n = 12; 131 ± 31 DIM) were enrolled in a 2 × 2 factorial design containing 2 experimental periods (P). During P1 (3 d), cows were fed ad libitum and baseline data were collected. At the initiation of P2 (3 d), cows were randomly assigned to 1 of 2 treatments by lactation stage (LS): (1) EL (EL-LPS; n = 6) or ML (ML-LPS; n = 6) cows administered i.v. a single bolus of 0.09 µg LPS/kg of BW; Escherichia coli O55:B5 or (2) pair-fed (PF) EL (EL-PF; n = 5) or ML (ML-PF; n = 6) cows administered i.v. saline. Administering LPS decreased DMI and this was more severe in EL-LPS than ML-LPS cows (34% and 11% relative to baseline, respectively). By design, P2 DMI patterns were similar in the PF groups compared with their LPS counterparts. Milk yield decreased following LPS (42% on d 1 relative to P1) and despite an exacerbated decrease in EL-LPS cows on d 1 (25% relative to ML-LPS), remained similar between LS from d 2 to 3. The EL-LPS cows had increased milk fat content, but no difference in protein and lactose percentages compared with ML-LPS cows. Further, cumulative ECM yield was increased (21%) in EL-LPS compared with ML-LPS cows. During P2, EL-LPS cows had a more intense increase in MUN and BUN than ML-LPS and EL-PF cows. Administering LPS did not cause hypoglycemia in either EL-LPS or ML-LPS cows, but glucose was increased (33%) in EL-LPS compared with EL-PF. Hyperinsulinemia occurred after LPS, and insulin was further increased in ML-LPS than EL-LPS cows (2.2-fold at 12 h peak). During P2, circulating glucagon increased only in EL-LPS cows (64% relative to all other groups). Both EL groups had increased NEFA at 3 and 6 h after LPS from baseline (56%), but NEFA in EL-LPS cows gradually returned to baseline thereafter and were reduced relative to EL-PF until 36 h (50% from 12 to 24 h). Alterations in BHB did not differ between ML groups, but EL-LPS had reduced BHB compared with EL-PF from 24 to 72 h (51%). Results indicate that there are distinct LS differences in the anorexic and metabolic responses to immune activation. Collectively, EL cows are more sensitive to the catabolic effects of LPS than ML cows, but these exacerbated metabolic responses appear coordinated to fuel an augmented immune system while simultaneously supporting milk synthesis.

Intravenous lipopolysaccharide challenge in early- versus mid-lactation dairy cattle. I: The immune and inflammatory responses

Cows in early lactation (EL) are purportedly immune suppressed, which renders them more susceptible to disease. Thus, the study objective was to compare key biomarkers of immune activation from i.v. LPS between EL and mid-lactation (ML) cows. Multiparous EL (20 ± 2 DIM; n = 11) and ML (131 ± 31 DIM; n = 12) cows were enrolled in a 2 × 2 factorial design and assigned to 1 of 2 treatments by lactation stage (LS): (1) EL (EL-LPS; n = 6) or ML (ML-LPS; n = 6) cows administered a single LPS bolus from Escherichia coli O55:B5 (0.09 µg/kg of BW), or (2) pair-fed (PF) EL (EL-PF; n = 5) or ML (ML-PF; n = 6) cows administered i.v. saline. After LPS administration, cows were intensely evaluated for 3 d to analyze their response and recovery to LPS. Rectal temperature increased in LPS relative to PF cows (1.1°C in the first 9 h), and the response was more severe in EL-LPS relative to ML-LPS cows (2.3 vs. 1.3°C increase at 4 h post-LPS; respectively). Respiration rate increased only in EL-LPS cows (47% relative to ML-LPS in the first hour post-LPS). Circulating tumor necrosis factor-α, IL-6, monocyte chemoattractant protein-1, macrophage inflammatory protein (MIP)-1α, MIP-1β, and IFN-γ-inducible protein-10 increased within the first 6 h after LPS and these changes were exacerbated in EL-LPS relative to ML-LPS cows (6.3-fold, 4.8-fold, 57%, 93%, 10%, and 61%, respectively). All cows administered LPS had decreased circulating iCa relative to PF cows (34% at the 6 h nadir), but the hypocalcemia was more severe in EL-LPS than ML-LPS cows (14% at 6 h nadir). In response to LPS, neutrophils decreased regardless of LS, then increased into neutrophilia by 24 h in all LPS relative to PF cows (2-fold); however, the neutrophilic phase was augmented in EL- compared with ML-LPS cows (63% from 24 to 72 h). Lymphocytes and monocytes rapidly decreased then gradually returned to baseline in LPS cows regardless of LS; however, monocytes were increased (57%) at 72 h in EL-LPS relative to ML-LPS cows. Platelets were reduced (46%) in LPS relative to PF cows throughout the 3-d following LPS, and from 24 to 48 h, platelets were further decreased (41%) in EL-LPS compared with ML-LPS. During the 3-d following LPS, serum amyloid A (SAA), LPS-binding protein (LBP), and haptoglobin (Hp) increased in LPS compared with PF groups (9-fold, 72%, and 153-fold, respectively), and the LBP and Hp responses were more exaggerated in EL-LPS than ML-LPS cows (85 and 79%, respectively) whereas the SAA response did not differ by LS. Thus, our data indicates that EL immune function does not appear "suppressed," and in fact many aspects of the immune response are seemingly functionally robust.

Chronic heat stress inhibits glycogen synthesis through gga-miR-212-5p/GYS1 axis in the breast muscle of broilers

Studies have demonstrated that chronic heat stress can accelerate glycolysis, decrease glycogen content in muscle, and affect muscle quality. However, the consequences of chronic heat stress on glycogen synthesis, miRNA expression in pectoralis major (PM) muscle, and its regulatory functions remain unknown. In this study, high-throughput sequencing and cell experiments were used to explore the effects of chronic heat stress on miRNA expression profiles and the regulatory mechanisms of miRNAs in glycogen synthesis under chronic heat stress. In total, 144 cocks were allocated into 3 groups: the normal control (NC) group, the heat stress (HS) group, and the pair-fed (PF) group. In total, 30 differently expressed (DE) miRNAs were screened after excluding the effect of feed intake, which were mainly related to metabolism, signal transduction, cell growth and death. Furthermore, the gga-miR-212-5p/GYS1 axis was predicted to participate in glycogen synthesis through the miRNA-mRNA analysis, and a dual-luciferase reporter test assay confirmed the target relationship. Mechanistically, chronic heat stress up-regulated gga-miR-212-5p, which could inhibit the expression of GYS1 in the PM muscle. Knocking down gga-miR-212-5p alleviates the reduction of glycogen content caused by chronic heat stress; overexpression of gga-miR-212-5p can reduce glycogen content. This study provided another important mechanism for the decreased glycogen contents within the PM muscle of broilers under heat stress, which might contribute to impaired meat quality.

Effects of supplemental citrulline on thermal and intestinal morphology parameters during heat stress and feed restriction in growing pigs.

Study objectives were to characterize the effects of citrulline (CIT) on physiological and intestinal morphology metrics during heat stress (HS) and feed restriction. Forty crossbred gilts (30 ± 2kg body weight [BW]) were assigned to one of five treatments: (1) thermoneutral (TN) fed ad libitum (AL) with control (CON) supplement (TNAL; n = 8), (2) TN pair-fed (PF) with CON (PF-CON; n = 8), (3) TN PF with CIT (PF-CIT; n = 8), (4) HS AL with CON (HS-CON; n = 8), and (5) HS AL with CIT (HS-CIT; n = 8). During the period (P) 1 (7 d), pigs were in TN conditions (23.6 °C) and fed AL their respective supplemental treatments. During P2 (2.5 d), HS-CON and HS-CIT pigs were fed AL and exposed to cyclical HS (33.6 to 38.3 °C), while TNAL, PF-CON, and PF-CIT remained in TN and were fed either AL or PF to their HS counterparts. Citrulline (0.13g/kg BW) was orally administered twice daily during P1 and P2. HS increased rectal temperature (Tr), skin temperature (Ts), and respiration rate (RR) relative to TN pigs (0.8 °C, 4.7 °C, and 47 breaths/min, respectively; P < 0.01). However, HS-CIT had decreased RR (7 breaths/min, P = 0.04) and a tendency for decreased Tr (0.1 °C, P = 0.07) relative to HS-CON pigs. During P2, HS pigs had decreased feed intake (22%; P < 0.01) and a tendency for decreased average daily gain (P = 0.08) relative to TNAL pigs, and by experimental design, PF pigs followed this same pattern. Circulating lipopolysaccharide-binding protein tended to be decreased (29%; P = 0.08) in PF relative to TNAL pigs and was increased (41%; P = 0.03) in HS compared to PF pigs. Jejunum villus height was decreased in PF relative to TNAL pigs (15%; P = 0.03); however, CIT supplementation improved this metric during feed restriction (16%; P = 0.10). Jejunum mucosal surface area decreased in PF (16%; P = 0.02) and tended to decrease in HS (11%; P = 0.10) compared to TNAL pigs. Ileum villus height and mucosal surface area decreased in HS compared to TNAL pigs (10 and 14%, respectively; P ≤ 0.04), but both parameters were rescued by CIT supplementation (P ≤ 0.08). Intestinal myeloperoxidase and goblet cell area remained similar among treatments and intestinal segments (P > 0.24). In summary, CIT supplementation slightly improved RR and Tr during HS. Feed restriction and HS differentially affected jejunum and ileum morphology and while CIT ameliorated some of these effects, the benefit appeared dependent on intestinal section and stressor type.

Zearalenone exposure differentially affects the ovarian proteome in pre-pubertal gilts during thermal neutral and heat stress conditions.

Zearalenone (ZEN), a nonsteroidal estrogenic mycotoxin, causes endocrine disruption and porcine reproductive dysfunction. Heat stress (HS) occurs when exogenous and metabolic heat accumulation exceeds heat dissipation. Independently, HS and ZEN both compromise swine reproduction; thus, the hypothesis investigated was two-pronged: that ZEN exposure would alter the ovarian proteome and that these effects would differ in thermal neutral (TN) and HS pigs. Pre-pubertal gilts (n = 38) were fed ad libitum and assigned to either (TN: 21.0 ± 0.1 °C) or HS (12h cyclic temperatures of 35.0 ± 0.2 °C and 32.2 ± 0.1 °C). Within the TN group, a subset of pigs were pair-fed (PF) to the amount of feed that the HS gilts consumed to eliminate the confounding effects of dissimilar nutrient intake. All gilts orally received a vehicle control (CT) or ZEN (40 μg/kg/BW) resulting in six treatment groups: thermoneutral (TN) vehicle control (TC; n = 6); TN ZEN (TZ; n = 6); PF vehicle control (PC; n = 6); PF ZEN (PZ; n = 6); HS vehicle control (HC; n = 7); or HS ZEN (HZ; n = 7) for 7 d. When compared to the TC pigs, TZ pigs had 45 increased and 39 decreased proteins (P ≤ 0.05). In the HZ pigs, 47 proteins were increased and 61 were decreased (P ≤ 0.05). Exposure to ZEN during TN conditions altered sec61 translocon complex (40%), rough endoplasmic reticulum membrane (8.2%), and proteasome complex (5.4%), asparagine metabolic process (0.60%), aspartate family amino acid metabolic process (0.14%), and cellular amide metabolic process (0.02%) pathways. During HS, ZEN affected cellular pathways associated with proteasome core complex alpha subunit complex (0.23%), fibrillar collagen trimer (0.14%), proteasome complex (0.05%), and spliceosomal complex (0.03%). Thus, these data identify ovarian pathways altered by ZEN exposure and suggest that the molecular targets of ZEN differ in TN and HS pigs.

Effects of Dietary Zinc Deficiency and Supplementation on Prepubertal Rat Testes: Sulfhydryl and Antioxidant Status.

The study was designed to investigate the effects of dietary zinc deficiency and supplementation on antioxidant system viz. superoxide-dismutase, glutathione reductase, glutathione peroxidase, glutathione- S-transferase, catalase and sulfhydryls levels (GSH, TSH, NPSH and PBSH) in testes of Wistar rats. Pre-pubertal rats were divided into two groups with 6 sub-groups each viz. zinc control (ZC), pair fed (PF), zinc deficient (ZD), zinc control supplementation (ZCS), pair-fed supplementation (PFS) and zinc deficient supplementation (ZDS). Experiments were set for 2- and 4-weeks followed by 4weeks of zinc supplementation. The zinc deficient group animals exhibited significant decrease in gonado-somatic index (2- and 4- weeks), sulfhydryls levels, GSH, GPx, GR (2 and 4-weeks) and GST concentration (2-weeks). However, after zinc supplementation significant improvement in gonadosomatic index, SH, GSH, antioxidant enzyme levels (GR, GPx, and GST) in deficient groups has been observed. Zinc deficiency during pre-pubertal period affected growth and caused dysregulation of the glutathione antioxidant system. The significant alterations in the levels of antioxidant enzymes and non-enzymatic antioxidant system (GSH and SH) in zinc deficient groups could be due to alleviated generation of free radicals, causative factor for increased oxidative stress which may lead to infertility as oxidative stress is a common pathology seen during infertility. Altered antioxidant system and sulfhydryls levels in testes due to dietary zinc deficiency reflect the significance of optimum zinc for maintaining homeostatic balance in gonadal physiology. Supplementing zinc for 4weeks could reduce the redox imbalance which may help in alleviating oxidative stress induced alterations in testes.

102 Growth Performance in Pigs fed a High Soybean Meal Diet During Heat Stress

Abstract In the U.S. swine industry, feeding programs are focused on using feed ingredients that are easily digestible and cost effective. With the predicted increases in soybean processing, pig producers may have increased availability of a cost-effective protein source in soybean meal (SBM). This expanded availability of SBM may result in feeding protein above the ostensible requirements, which would likely intensify the thermic effect of digestion, potentially increasing the susceptibility of the pig to heat stress (HS). Heat stress has damaging effects on feed intake and growth performance, costing the swine industry &amp;gt;$900 million annually. Therefore, the study objectives were to evaluate the effects of dietary crude protein diets (in the form of SBM) during HS on growth performance in growing pigs. Growing pigs (n = 40; initial BW = 55 ± 5.6 kg) were allocated into six treatments: 1) thermoneutral (TN) ad libitum control diet (TN-Ctl; n = 6), 2) TN pair-fed (PF) control diet (PF-Ctl; n = 6), 3) HS ad libitum control diet (HS-Ctl; n = 8), 4) TN ad libitum high SBM diet (TN-SBM; n = 6), 5) TN PF high SBM diet (PF-SBM; n = 6), and 6) HS ad libitum high SBM diet (HS-SBM; n = 8). The control diet included 15% crude protein (CP) while the high SBM diet included 23% CP with excess amino acids. The study consisted of two periods: Period 1 (P1; 7 d) in which all pigs were housed in TN environments (26.7 ± 1.4°C) with ad libitum access to respective diets. Period 2 (P2; 21 d), HS groups were exposed to cyclical HS with daytime temperatures reaching 35°C and evening temperatures decreased to 29.4°C. During P2, PF groups were fed to respective HS-Ctl or HS-SBM equivalents in TN conditions. Pigs exposed to HS had an overall increase in rectal temperature and respiration rate (0.91°C and 45 bpm, respectively; P ≤ 0.01) and HS-SBM pigs had increased evening rectal temperature (± 0.13°C; P ≤ 0.01), relative to HS-Ctrl. Overall, HS decreased ADFI (19%; P ≤ 0.01), ADG (26%; P ≤ 0.01) and increased F:G (10%; P ≤ 0.01) irrespective of dietary CP levels. Loin muscle area (P = 0.19) and backfat (P = 0.98) were not different between the TN and HS pigs regardless of diet. Circulating blood urea nitrogen levels did not differ between HS and TN pigs (P = 0.68), but an increase was observed in pigs fed the SBM diet relative to the control diet (P ≤ 0.01). In summary, these results demonstrate that HS decreases growth performance, but the negative effects are not augmented by feeding a high SBM diet.

Transcriptomic regulations of heat stress response in the liver of lactating dairy cows

BackgroundThe global dairy industry is currently facing the challenge of heat stress (HS). Despite the implementation of various measures to mitigate the negative impact of HS on milk production, the cellular response of dairy cows to HS is still not well understood. Our study aims to analyze transcriptomic dynamics and functional changes in the liver of cows subjected to heat stress (HS). To achieve this, a total of 9 Holstein dairy cows were randomly selected from three environmental conditions - heat stress (HS), pair-fed (PF), and thermoneutral (TN) groups - and liver biopsies were obtained for transcriptome analysis.ResultsBoth the dry matter intake (DMI) and milk yield of cows in the HS group exhibited significant reduction compared to the TN group. Through liver transcriptomic analysis, 483 differentially expressed genes (DEGs) were identified among three experimental groups. Especially, we found all the protein coding genes in mitochondria were significantly downregulated under HS and 6 heat shock proteins were significant upregulated after HS exposure, indicating HS may affect mitochondria integrity and jeopardize the metabolic homeostasis in liver. Furthermore, Gene ontology (GO) enrichment of DEGs revealed that the protein folding pathway was upregulated while oxidative phosphorylation was downregulated in the HS group, corresponding to impaired energy production caused by mitochondria dysfunction.ConclusionsThe liver transcriptome analysis generated a comprehensive gene expression regulation network upon HS in lactating dairy cows. Overall, this study provides novel insights into molecular and metabolic changes of cows conditioned under HS. The key genes and pathways identified in this study provided further understanding of transcriptome regulation of HS response and could serve as vital references to mitigate the HS effects on dairy cow health and productivity.

211-OR: Dual Amylin and Calcitonin Receptor Agonists Increase Mitochondrial Respiratory Capacity in Adipose Tissues of Obese Rat

Dual amylin and calcitonin receptor agonists (DACRAs) are potential therapeutic candidates for treatment of obesity. Preclinical studies have shown favorable effects of DACRAs on appetite regulation contributing to a reduction in body weight. Interestingly, the DACRA-induced effects on weight loss are superior to the effects of a suppressed food intake, suggesting an effect on energy expenditure potentially by targeting the mitochondria. We investigated the effects of long-term treatment with a class of DACRAs, namely KBPs, on body weight, food intake and mitochondrial respiratory capacity (MRC) in Sprague-Dawley rats fed a high-fat diet and treated s.c. with KBPs for 8 weeks. Moreover, a pair-fed (PF) group was used to examine food intake-independent effects of KBPs on MRC. At study end, MRC was analyzed in perirenal (pAT) and inguinal (iAT) adipose tissue using high resolution respirometry. Expectedly, KBP treatment significantly reduced body weight compared to PF and vehicle. PF was successfully KBP-matched in terms of accumulated food intake and the food intake was significantly lower than that of vehicle-treated rats. Coupled and uncoupled MRC supported by complex I+II substrates and fatty acids were significantly greater in iAT and pAT after KBP treatment indicating effects on both carbohydrate and lipid metabolism. Additionally, an increased oligomycin-induced leak-respiration was found to be independent of food intake in iAT. Further analysis will shed light on whether these changes in MRC are a result of an increased mitochondrial content or of the respiratory capacity of each mitochondrion. In conclusion, treatment with KBPs in obese rats is associated with an increased MRC in pAT and iAT and elicits effects additional to those obtained by diet-induced weight loss. This highlights DACRAs’ potential as anti-obesity agents with possible benefits on energy expenditure as a contributing cause to the DACRA-induced weight loss. Disclosure E.A.Petersen: Employee; Nordic Bioscience A/S. I.Blom: None. S.A.Melander: None. M.A.Karsdal: Speaker's Bureau; Pfizer Inc. S.Larsen: None. K.Henriksen: Employee; Nordic Bioscience A/S, Stock/Shareholder; Nordic Bioscience A/S. A.T.Larsen: Employee; Nordic Bioscience A/S. Funding Innovation Fund Denmark

Changes in Lipid Metabolism Enzymes in Rat Epididymal Fat after Chronic Central Leptin Infusion Are Related to Alterations in Inflammation and Insulin Signaling.

Leptin inhibits food intake and reduces the size of body fat depots, changing adipocyte sensitivity to insulin to restrain lipid accrual. This adipokine may modulate the production of cytokines that could diminish insulin sensitivity, particularly in visceral adipose tissue. To explore this possibility, we examined the effects of chronic central administration of leptin on the expression of key markers of lipid metabolism and its possible relationship with changes in inflammatory- and insulin-signaling pathways in epididymal adipose tissue. Circulating non-esterified fatty acids and pro- and anti-inflammatory cytokines were also measured. Fifteen male rats were divided into control (C), leptin (L, icv, 12 μg/day for 14 days), and pair-fed (PF) groups. We found a decrease in the activity of glucose-6-phosphate dehydrogenase and malic enzyme in the L group, with no changes in the expression of lipogenic enzymes. A reduction in the expression of lipoprotein lipase and carnitine palmitoyl-transferase-1A, together with a decrease in the phosphorylation of insulin-signaling targets and a low-grade inflammatory pattern, were detected in the epididymal fat of L rats. In conclusion, the decrease in insulin sensitivity and increased pro-inflammatory environment could regulate lipid metabolism, reducing epididymal fat stores in response to central leptin infusion.

Vitamin A-Deficient Rats have Only Mild Changes in Jejunal Structure and Function

This study investigated the effect of clinical and subclinical vitamin A deficiency on intestinal structure and function in rats. Weanling male rats fed a vitamin A-deficient diet (VA-) for 40–42 or 60–63 d were compared with rats either pair-fed (PF) or with free access to the same diet supplemented with vitamin A (VA+). A reference (REF) group was fed a standard rat diet. Weight began to plateau in VA- rats after 42 d, becoming significantly different from PF rats at 60–63 d (P < 0.02). Diarrhea did not develop in any study group. VA- rats had clinical signs of vitamin A deficiency in the 60–63 d study, but not in the 40–42 d study. However, serum and liver retinol concentrations were negligible in all VA- rats. VA- rats in the 60–63 d study had significantly reduced villus height (P < 0.02), and sucrase and maltase activities (P < 0.02) compared with PF rats. There were no differences between VA- and PF rats in mucosal wet weights, protein and DNA concentrations, thymidine kinase activity and glucose transport. No differences were detected in the 40–42 d study for any variable measured. Because clinical vitamin A deficiency in rats causes only mild changes in intestinal structure and function, it is unlikely that these alterations alone are responsible for the interactions observed in epidemiological studies between vitamin A deficiency and diarrheal disease.

Butyrate ameliorates chronic alcoholic central nervous damage by suppressing microglia-mediated neuroinflammation and modulating the microbiome-gut-brain axis

BackgroundAlcohol abuse triggers neuroinflammation, leading to neuronal damage and further memory and cognitive impairment. Few satisfactory advances have been made in the management of alcoholic central nervous impairment. Therefore, novel and more practical treatment options are urgently needed. Butyrate, a crucial metabolite of short-chain fatty acids (SCFAs), has been increasingly demonstrated to protect against numerous metabolic diseases. However, the impact of butyrate on chronic alcohol consumption-induced central nervous system (CNS) lesions remains unknown. MethodsIn this study, we assessed the possible effects and underlying mechanisms of butyrate on the attenuation of alcohol-induced CNS injury in mice. Firstly, sixty female C57BL/6 J mice were randomly divided into 4 groups: pair-fed (PF) group (PF/CON), alcohol-fed (AF) group (AF/CON), PF with sodium butyrate (NaB) group (PF/NaB) and AF with NaB group (AF/NaB). Each group was fed a modified Lieber-DeCarli liquid diet with or without alcohol. After six weeks of feeding, the mice were euthanized and the associated indicators were investigated. ResultsAs indicated by the behavioral tests and brain morphology, dietary NaB administration significantly ameliorated aberrant behaviors, including locomotor hypoactivity, anxiety disorder, depressive behavior, impaired learning, spatial recognition memory, and effectively reduced chronic alcoholic central nervous system damage. To further understand the underlying mechanisms, microglia-mediated inflammation and the associated M1/M2 polarization were measured separately. Firstly, pro-inflammatory TNF-α, IL-1β, and IL-6 in brain and peripheral blood circulation were decreased, but IL-10 were increased in the AF/NaB group compared with the AF/CON group. Consistently, the abnormal proportions of activated and resting microglial cells in the hippocampus and cortex regions after excessive alcohol consumption were significantly reduced with NaB treatment. Moreover, the rectification of microglia polarization (M1/M2) imbalance was found after NaB administration via binding GPR109A, up-regulating the expression of PPAR-γ and down-regulating TLR4/NF-κB activation. In addition to the direct suppression of neuroinflammation, intriguingly, dietary NaB intervention remarkably increased the levels of intestinal tight junction protein occludin and gut morphological barrier, attenuated the levels of serum lipopolysaccharide (LPS) and dysbiosis of gut microbiota, suggesting that NaB supplementation effectively improved the integrity and permeability of gut microecology. Finally, the neurotransmitters including differential Tryptophan (Trp) and Kynurenine (Kyn) were found with dietary NaB administration, which showed significantly altered and closely correlated with the gut microbiota composition, demonstrating the complex interactions in the microbiome-gut-brain axis involved in the efficacy of dietary NaB therapy for alcoholic CNS lesions. ConclusionDietary microbial metabolite butyrate supplementation ameliorates chronic alcoholic central nervous damage and improves related memory and cognitive functions through suppressing microglia-mediated neuroinflammation by GPR109A/PPAR-γ/TLR4-NF-κB signaling pathway and modulating microbiota−gut−brain axis.