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Acute systemic inflammatory response to lipopolysaccharide stimulation in pigs divergently selected for residual feed intake.

ABSTRACT: BACKGROUND:It is unclear whether improving feed efficiency by selection for low residual feed intake (RFI) compromises pigs' immunocompetence. Here, we aimed at investigating whether pig lines divergently selected for RFI had different inflammatory responses to lipopolysaccharide (LPS) exposure, regarding to clinical presentations and transcriptomic changes in peripheral blood cells. RESULTS:LPS injection induced acute systemic inflammation in both the low-RFI and high-RFI line (n?=?8 per line). At 4?h post injection (hpi), the low-RFI line had a significantly lower (p?= 0.0075) mean rectal temperature compared to the high-RFI line. However, no significant differences in complete blood count or levels of several plasma cytokines were detected between the two lines. Profiling blood transcriptomes at 0, 2, 6, and 24 hpi by RNA-sequencing revealed that LPS induced dramatic transcriptional changes, with 6296 genes differentially expressed at at least one time point post injection relative to baseline in at least one line (n?=?4 per line) (|log2(fold change)|???log2(1.2); q?

SUBMITTER: Liu H

PROVIDER: S-EPMC6792331 | biostudies-literature | 2019 Oct

REPOSITORIES: biostudies-literature

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Project description:Microbes and microbial components potentially impact the performance of pigs through immune stimulation and altered metabolism. These immune modulating factors can include endotoxin from gram negative bacterial outer membrane component, commonly referred to as lipopolysaccharide (LPS). In this study, our objective was to examine the relationship between intestinal barrier integrity, endotoxin and inflammation with feed efficiency (FE), using pig lines divergently selected for residual feed intake (RFI) as a model. Twelve gilts (62 ± 3 kg BW) from the low RFI (LRFI, more efficient) and 12 from the high RFI (HRFI, less efficient) were used. Individual performance data was recorded for 5 wk. At the end of the experimental period, ADFI of LRFI pigs was less (P < 0.001), ADG not different between the 2 lines (P = 0.72) but the G:F of LRFI pigs was greater than for HRFI pigs (P = 0.019). Serum endotoxin concentration (P < 0.01) and the acute phase protein haptoglobin (P < 0.05) were greater in HRFI pigs. Transepithelial resistance of the ileum, transport of fluorescein isothiocyanate labeled-Dextran and-LPS in ileum and colon, as well as tight junction protein mRNA expression in ileum, did not differ between the lines, indicating the 2 lines did not differ in transport characteristics at the intestinal level. Ileum inflammatory markers, myeloperoxidase (P < 0.05) and IL-8 (P < 0.10), were found to be greater in HRFI pigs. Alkaline phosphatase (ALP) activity was significantly increased in the LRFI pigs in ileum and liver tissues and negatively correlated with blood endotoxin (P < 0.05). Lysozyme activity in the liver was not different between the lines; however, the LRFI pigs had a twofold greater lysozyme activity in ileum (P < 0.05). Despite the difference in their activity, ALP or lysozyme mRNA expression was not different between the lines in either tissue. Decreased endotoxin and inflammatory markers and the enhanced activities of antimicrobial enzymes in the LRFI line may not fully explain the difference in the FE between the lines, but they have the potential to prevent the growth potential in HRFI pigs. Further studies are needed to identify the other mechanisms that may contribute to the greater endotoxin and acute phase proteins in the HRFI pigs and the greater FE in the LRFI pigs.

Project description:BACKGROUND:Improving feed efficiency (FE) of pigs by genetic selection is of economic and environmental significance. An increasingly accepted measure of feed efficiency is residual feed intake (RFI). Currently, the molecular mechanisms underlying RFI are largely unknown. Additionally, to incorporate RFI into animal breeding programs, feed intake must be recorded on individual pigs, which is costly and time-consuming. Thus, convenient and predictive biomarkers for RFI that can be measured at an early age are greatly desired. In this study, we aimed to explore whether differences exist in the global gene expression profiles of peripheral blood of 35 to 42 day-old pigs with extremely low (more efficient) and high RFI (less efficient) values from two lines that were divergently selected for RFI during the grow-finish phase, to use such information to explore the potential molecular basis of RFI differences, and to initiate development of predictive biomarkers for RFI. RESULTS:We identified 1972 differentially expressed genes (DEGs) (q ? 0.15) between the low (n = 15) and high (n = 16) RFI groups of animals by using RNA sequencing technology. We validated 24 of 37 selected DEGs by reverse transcription-quantitative PCR (RT-qPCR) in a joint analysis of 24 (12 per line) of the 31 samples already used for RNA-seq plus 24 (12 per line) novel samples from the same contemporary group of pigs. Using an analysis of the 24 novel samples alone, only nine of the 37 selected DEGs were validated. Genes involved in small molecule biosynthetic process, antigen processing and presentation of peptide antigen via major histocompatibility complex (MHC) class I, and steroid biosynthetic process were overrepresented among DEGs that had higher expression in the low versus high RFI animals. Genes known to function in the proteasome complex or mitochondrion were also significantly enriched among genes with higher expression in the low versus high RFI animals. Alternatively, genes involved in signal transduction, bone mineralization and regulation of phosphorylation were overrepresented among DEGs with lower expression in the low versus high RFI animals. The DEGs significantly overlapped with genes associated with disease, including hyperphagia, eating disorders and mitochondrial diseases (q < 1E-05). A weighted gene co-expression network analysis (WGCNA) identified four co-expression modules that were differentially expressed between the low and high RFI groups. Genes involved in lipid metabolism, regulation of bone mineralization, cellular immunity and response to stimulus were overrepresented within the two modules that were most significantly differentially expressed between the low and high RFI groups. We also found five of the DEGs and one of the co-expression modules were significantly associated with the RFI phenotype of individual animals (q < 0.05). CONCLUSIONS:The post-weaning blood transcriptome was clearly different between the low and high RFI groups. The identified DEGs suggested potential differences in mitochondrial and proteasomal activities, small molecule biosynthetic process, and signal transduction between the two RFI groups and provided potential new insights into the molecular basis of RFI in pigs, although the observed relationship between the post-weaning blood gene expression and RFI phenotype measured during the grow-finish phase was not strong. DEGs and representative genes in co-expression modules that were associated with RFI phenotype provide a preliminary list for developing predictive biomarkers for RFI in pigs.

Project description:Yorkshire pigs were divergently selected for residual feed intake (RFI) for 8 generations. Pigs with low RFI have reduced feed intake. Based on the resource allocation theory, we hypothesize that these low RFI pigs might have compromised immune response to immune stimulation. To test this hypothesis, twenty eight gilts of initial body weight (BW) of 63 4 kg from pigs selected for low RFI and high RFI were randomly selected for the ISU RFI selection project and utilized in two replicates (14 pigs/replicate) for this study. All pigs were housed in individual metabolism pens, had free access to water and were fed a corn-soy-based diet twice daily, and feed restricted (1.5 kg/day) as previously described. After a 9-day adaptation period pigs were randomly assigned within line to either a control (n = 6, three pigs per line) or lipopolysaccharide (LPS) challenge (n = 8, 4 pigs per line) group. Pigs in the challenge group were then repeatedly challenged with an intramuscular injection of 30, 36, 39, and 42 g/kg BW of LPS from E. coli O5:B5 (sigma-Aldrich, St. Louis, MO) dissolved in saline solution at time points 0, 48, 96, and 144 hours post initial injection (hpi) to compensate for LPS tolerance after initial injection, respectively, while animals in the control group were injected with an equivalent volume of saline solution at the equivalent time points. Rectal temperatures of individual pigs were measured immediately before the initial injection (serving as baseline, called 0 hpi for convenience), and 2, 4, 6, and 24 hpi, and 0, 4, 24 hours after each subsequent injection for all pigs. Blood samples were collected from the jugular vein into TempusTM Blood RNA tubes (Life Technologies, Grand Island, NY) for long-term storage at -80 C, EDTA tubes (BD, Franklin Lakes, NJ) for CBC tests and serum tubes for cytokine assays at 0, 2, 6, 24 and 168 hpi. At 168 hpi, all pigs were euthanized via barbiturate overdose, exsanguinated and tissue samples including the longissimus dorsi muscle, liver, spleen, and ileum were isolated, cleaned an frozen for later use. This study only focused on samples and data collected for the first 24 hours. Total RNA was extracted from the 32 blood samples of the treatment group in Replicate 2, collected at 0, 2, 6, and 24 hpi, by using preserved blood RNA purification kit I (Norgen Biotek Corp, Thorold, Ontario) per manufacturers instruction. On-column DNA digestion was performed as described using DNase I (Qiagen, Valencia, CA). Globin transcripts (HBB and HBA) were depleted by following an RNase H-mediated method. The quantity and integrity of the RNA were monitored by using Nanodrop 2000 (Thermo Scientific, Waltham, MA) and Bioanalyzer 2100 (Agilent Technologies, Santa Clara, CA) before and after globin depletion. The efficiency of globin depletion of each sample was checked by conventional RT-qPCR with -actin and GAPDH as the internal controls. Globin depletion efficiencies for all RNA samples were above 85%.

Project description:Residual feed intake (RFI) is the difference between observed and predicted feed intake of an animal, based on growth and maintenance requirements. In Yorkshire pigs, divergent selection for increased (Low RFI) and decreased (High RFI) RFI was carried out over 10 generations (G) while feeding a corn- and soybean-meal-based, higher-energy, lower-fiber (HELF) diet. In G8 to G10, representing 4 replicates, barrows and gilts (n = 649) of the RFI lines were fed the HELF diet and a diet incorporating coproducts that were lower in energy and higher in dietary fiber (LEHF). The diets differed in ME, 3.32 vs. 2.87 Mcal/kg, and in neutral detergent fiber (NDF), 9.4% vs. 25.9%, respectively. The impact of the LEHF diet on 1) performance and growth, 2) diet digestibility, 3) genetic parameter estimates, and 4) responses to selection for RFI, when fed the HELF, was assessed. In general, the LEHF diet reduced the performance of both lines. When fed the HELF diet, the Low RFI pigs had lower (P < 0.05) ADFI (-12%), energy intake (-12%), ADG (-6%), and backfat depth (-12%); similar (P > 0.05) loin muscle area (LMA; +5%); and greater (P < 0.05) feed efficiency (i.e., 8% higher G:F and 7% lower RFI) than the High RFI line. These patterns of line differences were still present under the LEHF diet but differences for ADFI (-11%), energy intake (-10%), G:F (+2%), and RFI (-6%) were reduced compared to the HELF diet. Apparent total tract digestibility (ATTD) of the HELF and LEHF diets was assessed using 116 barrows and gilts from G8. When fed the HELF diet, ATTD of DM, GE, N, and NDF were similar between lines (P ≥ 0.27), but when fed the LEHF diet, the Low RFI pigs had greater digestibility (7%, 7%, 10%, and 32%) than the High RFI line (P ≤ 0.04). To measure responses to selection for RFI and estimate genetic parameters, data from all 10 generations were used (HELF; n = 2,310; LEHF, n = 317). Heritability estimates of performance traits ranged from 0.19 to 0.63, and genetic correlations of traits between diets were high and positive, ranging from 0.87 (RFI) to 0.99 (LMA). By G10, RFI in the Low RFI line was 3.86 and 1.50 genetic SD lower than in the High RFI line when fed the HELF and LEHF diets, respectively. Taken together, the results of this study demonstrate that responses to selection for RFI when fed a HELF diet are not fully realized when pigs are fed an extremely LEHF diet. Thus, feeding diets that differ from those used for selection may not maximize genetic potential for feed efficiency.

Project description:Weaning is a stress every piglet has to face. It is a main cause of antibiotic uses due to digestive disorders. In this study, response to weaning was analyzed in pigs from two lines divergently selected for residual feed intake (RFI) during growth. A total of 132 pigs from each line, housed per line and diet in conventional postweaning units of 12 castrated males and 12 females, were fed either a conventional control (two successive diets) or a complex (three successive diets) dietary sequence during the postweaning period (4 to 10 wk of age). BWs were recorded at weaning (days 0 and 28 of age), days 1, 2, 6, 12, 19, 26, and 42 (10 wk of age), and at 23 wk of age. Feces texture was examined before weaning (day -1), at day 1, 2, 6, 12, and 19. Feed intake was recorded at pen level from days 0 to 42 after weaning, and individually thereafter. Plasma was collected after blood samplings at days -1, 6, 19, and 42 on half of the piglets: all piglets of a given sex in each pen were sampled, to achieve a balanced number across factors. Pigs of the low RFI (LRFI) line were heavier at weaning, had greater glucose concentration, and lower levels of diarrhea at days 1 and 2 than pigs from the high RFI (HRFI) line (P < 0.01). At day 42, there was no BW difference between lines, and G:F ratio did not differ between lines (P = 0.40). The LRFI pigs had lower feed intake and growth rate from day 0 to day 19 (P < 0.005), and greater plasma concentration of non-esterified fatty acid (P < 0.001), indicating an increased mobilization of body lipids and proteins immediately after weaning compared with HRFI pigs. They also had greater levels of diarrhea at day 6 (22% for LRFI vs. 14% for HRFI, P = 0.002), but the concentration of plasma haptoglobin did not indicate acute inflammation. The complex diet sequence improved feed intake and growth, and reduced diarrhea, mainly in the LRFI line (P < 0.001). To conclude, pigs from the LRFI line were more negatively affected by weaning stress, but managed to recover afterwards. The complex diet sequence ameliorated some of the negative effects that weaning had on the LRFI pigs, but limited effects of nursery period feeding sequence on growth performance were observed during the growing-finishing period.

Project description:MicroRNAs (miRNAs) are short non-coding RNAs that post-transcriptionally regulate expression of mRNAs in many biological pathways. Liver plays an important role in the feed efficiency of animals and high and low efficient cattle demonstrated different gene expression profiles by microarray. Here we report comprehensive miRNAs profiles by next-gen deep sequencing in Angus cattle divergently selected for residual feed intake (RFI) and identify miRNAs related to feed efficiency in beef cattle. Two microRNA libraries were constructed from pooled RNA extracted from livers of low and high RFI cattle, and sequenced by Illumina genome analyser. In total, 23,628,103 high quality short sequence reads were obtained and more than half of these reads were matched to the bovine genome (UMD 3.1). We identified 305 known bovine miRNAs. Bta-miR-143, bta-miR-30, bta-miR-122, bta-miR-378, and bta-let-7 were the top five most abundant miRNAs families expressed in liver, representing more than 63% of expressed miRNAs. We also identified 52 homologous miRNAs and 10 novel putative bovine-specific miRNAs, based on precursor sequence and the secondary structure and utilizing the miRBase (v. 21). We compared the miRNAs profile between high and low RFI animals and ranked the most differentially expressed bovine known miRNAs. Bovine miR-143 was the most abundant miRNA in the bovine liver and comprised 20% of total expressed mapped miRNAs. The most highly expressed miRNA in liver of mice and humans, miR-122, was the third most abundant in our cattle liver samples. We also identified 10 putative novel bovine-specific miRNA candidates. Differentially expressed miRNAs between high and low RFI cattle were identified with 18 miRNAs being up-regulated and 7 other miRNAs down-regulated in low RFI cattle. Our study has identified comprehensive miRNAs expressed in bovine liver. Some of the expressed miRNAs are novel in cattle. The differentially expressed miRNAs between high and low RFI give some insights into liver miRNAs regulating physiological pathways underlying variation in this measure of feed efficiency in bovines.

Project description:Improving feed efficiency in cattle is important because it increases profitability by reducing costs, and it also shrinks the environmental footprint of cattle production by decreasing manure and greenhouse gas emissions. Residual feed intake (RFI) is 1 measurement of feed efficiency and is the difference between actual and predicted feed intake. Residual feed intake is a complex trait with moderate heritability, but the genes and biological processes associated with its variation still need to be found. We explored the variation in expression of genes using RNA sequencing to find genes whose expression was associated with RFI and then investigated the pathways that are enriched for these genes. In this study, we used samples from growing Angus bulls (muscle and liver tissues) and lactating Holstein cows (liver tissue and white blood cells) divergently selected for low and high RFI. Within each breed-tissue combination, the correlation between the expression of genes and RFI phenotypes, as well as GEBV, was calculated to determine the genes whose expression was correlated with RFI. There were 16,039 genes expressed in more than 25% of samples in 1 or more tissues. The expression of 6,143 genes was significantly associated with RFI phenotypes, and expression of 2,343 genes was significantly associated with GEBV for RFI ( < 0.05) in at least 1 tissue. The genes whose expression was correlated with RFI phenotype (or GEBV) within each breed-tissue combination were enriched for 158 (78) biological processes (Fisher Exact Statistics for gene-enrichment analysis, EASE score < 0.1) and associated with 13 (13) Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways ( < 0.05 and fold enrichment > 2). These biological processes were related to regulation of transcription, translation, energy generation, cell cycling, apoptosis, and proteolysis. However, the direction of the correlation between RFI and gene expression in some cases reversed between tissues. For instance, low levels of proteolysis in muscle were associated with high efficiency in growing bulls, but high levels of proteolysis in white blood cells were associated with efficiency of milk production in lactating cows.

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Acute systemic inflammatory response to lipopolysaccharide stimulation in pigs divergently selected for residual feed intake.

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