Effects of the increased protein level in small intestine on the colonic microbiota, inflammation and barrier function in growing pigs.

BACKGROUND: An increased level of the dietary protein alters the colonic microbial community and metabolic profile of pigs, but it remains unclear whether this leads to colonic inflammation and impairs barrier function in growing pigs. RESULTS: Sixteen pigs (35.2 ± 0.3 kg) were infused with sterile saline (control) or soy protein hydrolysate (SPH) (70 g/day) through a duodenal fistula twice daily during a 15-day experimental period. The SPH treatment did not affect their average daily feed intake and daily weight gain (P > 0.05), but reduced colon index and length (P < 0.05). Illumina MiSeq sequencing revealed that species richness was increased following SPH intervention (P < 0.05). Furthermore, SPH reduced the abundance of butyrate- and propionate-producing bacteria-such as Lachnospiraceae NK4A136 group, Lachnospiraceae_uncultured, Coprococcus 3, Lachnospiraceae UCG-002, and Anaerovibrio-and increased the abundance of potentially pathogenic bacteria and protein-fermenting bacteria, such as Escherichia-Shigella, Dialister, Veillonella, Prevotella, Candidatus Saccharimonas, Erysipelotrichaceae UCG-006, Prevotellaceae_uncultured, and Prevotellaceae UCG-003 (P < 0.05). In addition, a lower content of total short-chain fatty acids, propionate, and butyrate and a higher concentration of cadaverine, putrescine, total biogenic amines, ammonia, and isovalerate were observed following SPH infusion (P < 0.05). Further analysis revealed that SPH increased the concentration of tumour necrosis factor-α, interleukin (IL)-1ß, IL-6, and IL-8 in the colonic mucosa (P < 0.05). Interestingly, SPH intervention increased the expression of occludin, zonula occludens (ZO)-1, and claudin-1 in colonic mucosa (P < 0.05). Correlation analysis showed that different genera were significantly related to the production of metabolites and the concentrations of pro-inflammatory cytokines. CONCLUSION: An increased soy protein level in the small intestine altered the colonic microbial composition and metabolic profile, which resulted in the secretion of colonic proinflammatory cytokines and the increased expression of tight junction proteins.

Supplementation of Moringa oleifera leaf powder orally improved productive performance by enhancing the intestinal health in rabbits under chronic heat stress.

Heat stress jeopardizes animal's growth and health mainly through induction of oxidative stress and inflammation. The current study investigated the effects of Moringa oleifera leaf powder (MOLP) supplementation on productive performance and intestinal health of rabbits under chronic heat stress (HS). Young New Zealand White rabbits (male) at the age of 32 weeks (n = 21, mean body weight of 3318 ± 171 g) for four weeks' period were reared on commercial pelleted diet and divided into three groups: control (CON, 25 °C), HS (35 ± 1 °C) and HS (35 ± 1 °C) with MOLP (HSM) supplemented orally (200 mg/kg body weight). The results demonstrated that rabbits in the HSM group had reduced rectal temperature, respiration rate and improved FCR due to improved daily gain and better crude fiber (NDF) digestibility (P < 0.05) compared with HS group. MOLP improved intestinal integrity and function as indicated by lower serum diamine oxidase level and increased jejunal weight, length, villus height and ratio of villus height to crypt depth than heat-stressed rabbits. MOLP reversed the increased levels of serum cortisol, metabolic indicators i.e. glucose, insulin, and reduced concentrations of serum triiodothyronine. MOLP supplementation also significantly down-regulated the mRNA expression of tumor necrosis factor alpha (α), heat shock protein A2, glutathione peroxidase-1, interleukin (IL)-1α and increased the expression of IL-6. In conclusion, MOLP supplementation could enhance intestinal health along with production and metabolic indicators by alleviating the oxidative stress and inflammatory response in small intestine of hyper-thermic rabbits.

l-Glutamate stimulates cholecystokinin secretion via the T1R1/T1R3 mediated PLC/TRPM5 transduction pathway.

BACKGROUND: It is known that cholecystokinin (CCK) plays an essential role in reducing food intake and driving weight loss. Previous studies demonstrated that amino acids were capable of triggering CCK release through G protein-coupled receptors, but the sensing mechanism remains obscure, especially the intracellular signaling pathway. RESULTS: l-Glu, rather than its d-isomer, robustly stimulated CCK secretion in a porcine duodenal model, and the secretory response was augmented by incubation with the allosteric ligand of T1R1, while T1R3 antagonist attenuated it. Upon inhibiting phospholipase C (PLC) or transient receptor potential M5 (TRPM5) activity, l-Glu failed to increase CCK release. Oral administration of monosodium glutamate in rats also suppressed food intake and increased plasma CCK levels, accompanied by elevated expression of T1R1, PLCß2 and TRPM5 in the duodenum. CONCLUSION: These data demonstrated that l-Glu stimulated CCK secretion through the activation of T1R1/T1R3 in a PLC/TRPM5-dependent manner. © 2020 Society of Chemical Industry.

Effects of low-protein diet on the intestinal morphology, digestive enzyme activity, blood urea nitrogen, and gut microbiota and metabolites in weaned pigs.

This study investigated the effects of low-protein diet supplemented with Lysine (Lys), Methionine (Met), Threonine (Thr), and Tryptophan (Trp) on small intestine morphology, enzyme activity, blood urea nitrogen, and gut microbiota and metabolites in weaned piglets. Eighteen weaned pigs weighing an average of 9.57 kg received one of three treatments: a normal protein diet with 20% crude protein (CP, diet [NP]), a moderately reduced protein diet with 17% CP (MP), or a low-protein diet with 14% CP (LP). All three diets were supplemented with Lys, Met, Thr and Trp to meet essential amino acid requirements for post-weaned piglets according to the NRC (2012). Following a 45 d study period, piglets on the LP and MP diets demonstrated atrophic small intestinal morphology, with decreased villus heights and lower ratios of villus height to crypt depth (p < 0.05); pepsin activity in the stomach was also reduced in these two groups (p < 0.05). Increased plasma cholesterol and decreased blood urea nitrogen presented in the MP and LP groups compared with the NP group (p < 0.05). Overall, gastrointestinal hormones were not affected by dietary protein levels with the exception of reduced somatostatin levels in the MP and LP groups. Jejunum and colon microbiota were not affected at either the phyla or genera level in any of the diets. Colonic ammonia nitrogen concentration was reduced in MP and LP groups. Dietary protein level had no effect on short chain fatty acids or biogenic amines. Our data suggest that reducing dietary protein levels by 3% (MP) or 6% (LP) in weaned pigs has the potential to decrease nitrogen emissions and impaired digestive capacity. Therefore, dietary protein level cannot be reduced by more than 3% in consideration of maladaptive changes to small intestinal morphology and pepsin activity in weaned piglets.

Phenylalanine and tryptophan stimulate gastrin and somatostatin secretion and H+-K+-ATPase activity in pigs through calcium-sensing receptor.

In rodents and humans, aromatic amino acids increase gut hormone secretion and H+-K+-ATPase activity by modulating calcium-sensing receptor (CaSR). However, the role of CaSR and its related signaling molecules in amino acid-induced gut hormone secretion in swine has not been investigated. Here, we examined whether a CaSR-dependent pathway modulated gastrin and somatostatin (SS) secretion and H+-K+-ATPase activity in pigs. Perfusion of pig stomach tissues in the presence of extracellular 80 mM l-phenylalanine (Phe) or 20 mM l-tryptophan (Trp) and a CaSR agonist cinacalcet triggered gastrin and SS secretion and H+-K+-ATPase activity (P < 0.05) and increased CaSR expression (P < 0.05). This effect of Phe and Trp was dependent on Ca2+ (P < 0.05) and was abolished after treatment with NPS 2143, an inhibitor of CaSR, and 2-aminoethyl diphenyl borinate, an inhibitor of CaSR downstream signaling molecule inositol 1,4,5-triphosphate receptor (IP3R). These findings indicate that Phe and Trp induce Ca2+-dependent gastrin and SS secretion and H+-K+-ATPase activity through CaSR and its downstream signaling molecule IP3R.

Progressive response of large intestinal bacterial community and fermentation to the stepwise decrease of dietary crude protein level in growing pigs.

The study aimed to determine the effects of reduction of dietary crude protein (CP) level with balanced essential amino acids (EAA) on intestinal bacteria and their metabolites of growing pigs. Forty pigs (initial BW 13.50 ± 0.50 kg, 45 ± 2 days of age) were randomly assigned to four dietary treatments containing CP levels at 20.00% (normal crude protein, NP); 17.16% (medium crude protein, MP); 15.30% (low crude protein, LP); and 13.90% (extremely low crude protein, ELP), respectively. Crystalline AAs were added to meet the EAA requirement of pigs. After 4-week feeding, eight pigs per treatment (n = 8) were randomly selected and slaughtered for sampling of ileal, cecal, and colonic digesta and mucosa. Pigs with moderately reduced CP level had increased bacterial diversity, with the Shannon diversity indices for the colon digesta in the LP group and mucosa in the MP and LP groups significantly (P < 0.05) higher than those in the NP and ELP groups. As the CP level reduces, the Bifidobacterium population were linearly decreased (P < 0.05) both in ileum, cecum, and colon, and the ELP group had the lowest Bifidobacterium population in the cecum and colon, with its value significantly lower than NP and MP groups (P < 0.05). However, the ELP group had the highest population of Escherichia coli in the colon, with its value significantly higher than the LP group (P < 0.05). For bacterial metabolites, as CP level decreased, total short-chain fatty acid (T-SCFA), acetate, and butyrate were linearly increased (linear, P < 0.05) in the ileum, while all SCFAs except formate in the cecum and T-SCFA and acetate in the colon, were linearly decreased (P < 0.05). Reducing CP level led to a linear decrease of microbial crude protein (MCP) in the ileum (P < 0.05) and ammonia in all intestine segments (P < 0.05). The spermidine in cecum and total amines, cadaverine, methylamine, and spermidine in colon were shown a quadratic change (P < 0.05) as dietary CP decreases, with the highest concentration in LP group. These findings suggest that moderate reduction of dietary CP level may benefit large intestinal bacterial community and its fermentation, which was negatively affected by extremely low CP diet.

[Feeding Lactobacillus plantarum and Lactobacillus casei increased microbial diversity and short chain fatty acids production in the gut-intestinal tract of weaning piglets].

Objective: The objective of the study was to study the effects of Lactobacillus plantarum and Lactobacillus casei on the changes in composition and quantity of microbial populations and the concentrations of short chain fatty acids in the gut digesta of piglets before and after weaning, to explore the mechanisms of the tested two strains to relieve weaning stress. Methods: Fifteen litters of piglets (Duroc×Yorkshire×Landrace) at the age of 7 days were randomly allocated to 3 groups (5 each), including the control group with an oral administration of saline, LP group with L. plantarum and and LC group, with L. casei. On day 21, 24 and 35, the piglets were slaughtered, and the ileum and colon digesta were analyzed for microbial populations and short chain fatty acids. Results: In the ileum and colon, the test strains significantly increased the microbial diversity (P<0.05), promoted the growth of Lactobacilli spp. and Bifidobacteria spp. after 2 weeks of post-weaning. The two test strains increased the concentration of acetate, propionate, butyrate and total short chain fatty acids both in ileum and colon before weaning, and the concentration of acetate, total short chain fatty acids after weaning. Correlation analysis showed that the decreased diarrhea rate before weaning in the LP and LC groups was significant associated with the increased short chain fatty acids concentration and total bacteria of ileum and colon, the increased height of the ileum villi. The improved average daily gain presented a correlation with the increased concentration of acetate and total short chain fatty acids of colon. Conclusion: The findings imply that the tested strains are contribute to increase the microbial diversity, the quantities of beneficial bacteria and the production of short chain fatty acids in the gut intestinal tract of weaning piglets.

Amino acid metabolism in intestinal bacteria and its potential implications for mammalian reproduction.

Reproduction is vital for producing offspring and preserving genetic resources. However, incidences of many reproductive disorders (e.g. miscarriage, intrauterine growth restriction, premature delivery and lower sperm quality) have either increased dramatically or remained at high rates over the last decades. Mounting evidence shows a strong correlation between enteral protein nutrition and reproduction. Besides serving as major nutrients in the diet, amino acids (AA) are signaling molecules in the regulation of diverse physiological processes, ranging from spermatogenesis to oocyte fertilization and to embryo implantation. Notably, the numbers of bacteria in the intestine exceed the numbers of host cells by 10 times. Microbes in the small-intestinal lumen actively metabolize large amounts of dietary AA and, therefore, affect the entry of AA into the portal circulation for whole-body utilization. Changes in the composition and abundance of AA-metabolizing bacteria in the gut during pregnancy, as well as their translocation to the uterus, may alter uterine function and epigenetic modifications of maternal physiology and metabolism, which are crucial for pregnancy recognition and fetal development. Thus, the presence of the maternal gut microbiota and AA metabolites in the intrauterine environments (e.g. endometrium and placenta) and breast milk is likely a unique signature for the programming of the whole-body microbiome and metabolism in both the fetus and infant. Dietary intervention with functional AA, probiotics and prebiotics to alter the abundance and activity of intestinal bacteria may ameliorate or prevent the development of metabolic syndrome, while improving reproductive performance in both males and females as well as their offspring.

Effects of low dietary protein on the metabolites and microbial communities in the caecal digesta of piglets.

Thirty-six healthy piglets (weighing 10 ± 1 kg; three animals per pen) were randomly allocated to two treatments: (i) a low protein diet (14% crude protein [CP]) supplemented with lysine, methionine, threonine and tryptophan (Group LP) and (ii) a normal protein diet (20% CP, Group NP), resulting in six replicate pens per treatment. One piglet from each pen was slaughtered at days 10, 25 and 45 of the experiment. For the whole experimental period of 45 d, Group LP had lower feed intake and daily gain and a higher feed-to-gain ratio compared with Group NP. At day 10, no effects on measured caecum metabolites were observed, but at days 25 and 45 in Group LP the concentration of ammonia-N, cadaverine, branched chain fatty acids and acetate were reduced. This was also true for the concentration of short chain fatty acids at day 45. The results of denaturing gradient gel electrophoresis showed that microbial diversity in Group LP was less abundant at day 25, but there was no difference at days 10 and 45. An unweighted pair group mean average analysis showed that the similarities were lower between Groups LP and NP at day 10 and higher at days 25 and 45. Quantitation results indicated that the numbers of Firmicutes and Clostridium cluster IV were lower in Group LP than in Group NP at day 25, but there were no differences at days 10 and 45. In conclusion, the low protein diet markedly reduced the metabolites of protein and carbohydrate fermentation and altered microbial communities in the caecal digesta of piglets.

Diversity of butyrivibrio group bacteria in the rumen of goats and its response to the supplementation of garlic oil.

This study aimed to investigate the diversity of the Butyrivibrio group bacteria in goat rumen and its response to garlic oil (GO) supplementation as revealed by molecular analysis of cloned 16S rRNA genes. Six wethers fitted with ruminal fistulas were assigned to two groups for a cross-over design with 28-d experimental period and 14-d interval. Goats were fed a basal diet without (control) or with GO ruminal infusion (0.8 g/d). Ruminal contents were used for DNA extraction collected before morning feeding on d 28. A total bacterial clone library was firstly constructed by nearly full-length 16S rRNA gene cloned sequences using universal primers. The resulting plasmids selected by Butyrivibrio-specific primers were used to construct a Butyrivibrio group-specific bacterial clone library. Butyrivibrio group represented 12.98% and 10.95% of total bacteria in control and GO group, respectively. In libraries, clones were classified to the genus Pseudobutyrivibrio, Butyrivibrio and others within the family Lachnospiraceae. Additionally, some specific clones were observed in GO group, being classified to the genus Ruminococcus and others within the family Ruminococcaceae. Based on the criterion that the similarity was 97% or greater with database sequences, there were 29.73% and 18.42% of clones identified as known isolates (i.e. B. proteoclasticus and Ps. ruminis) in control and GO groups, respectively. Further clones identified as B. fibrisolvens (5.41%) and R. flavefaciens (7.89%) were specifically found in control and GO groups, respectively. The majority of clones resembled Ps. ruminis (98% to 99% similarity), except for Lachnospiraceae bacteria (87% to 92% similarity) in the two libraries. The two clone libraries also appeared different in Shannon diversity index (control 2.47 and GO group 2.91). Our results indicated that the Butyrivibrio group bacteria had a complex community with considerable unknown species in the goat rumen.

Palm Kernel Cake Extracts Obtained from the Combination of Bacterial Fermentation and Enzymic Hydrolysis Promote Swine Small Intestine IPEC-J2 Cell Proliferation and Alleviate LPS-Induced Inflammation In Vitro.

Co-fermentation with bacteria and enzymes can reduce sugar content in palm kernel cake (PKC); however, the chemical changes and their effects on cell functionality are unclear. This study investigated the active components in pre-treated PKC extracts and their effects on pig small intestine IPEC-J2 cell proliferation and LPS-induced inflammation. The extracts contained 60.75% sugar, 36.80% mannose, 1.75% polyphenols and 0.59% flavone, as determined by chemical analyses, suggesting that the extracts were palm kernel cake oligosaccharides (PKCOS). Then, we found that 1000 µg/mL PKCOS counteracted the decrease in cell viability (CCK8 kit) caused by LPS induction by 5 µg/mL LPS (p < 0.05). Mechanistic studies conducted by RNA-seq and qPCR analyses suggested PKCOS promoted cell proliferation through the upregulation of TNF-α, PI3KAP1, MAP3K5 and Fos in the PI3K/MAPK signalling pathway; alleviated inflammation caused by LPS via the downregulation of the target genes Casp3 and TNF-α in association with apoptosis; and regulated the expression of the antioxidant genes SOD1, SOD2 and GPX4 to exert positive antioxidant effects (p < 0.05). Furthermore, PKCOS upregulated SLC5A1 (encoding SLGT1), HK and MPI in the glycolytic pathway (p < 0.05), suggesting cell survival. In summary, PKCOS has positive effects on promoting swine intestine cell proliferation against inflammation.

mTOR signaling pathway regulation HIF-1 α effects on LPS induced intestinal mucosal epithelial model damage.

BACKGROUND: Sepsis-induced small-intestinal injury is associated with increased morbidity and mortality. Our previous study and other papers have shown that HIF-1α has a protective effect on intestinal mucosal injury in septic rats. The purpose of this study is to further verify the protective effect of HIF-1α on intestinal mucosa and its molecular mechanism in vitro experiments. METHODS: Caco-2 cells were selected and experiment was divided into 2 parts. Part I: HIF-1α activator and inhibitor were used to treat lipopolysacchrides (LPS)-stimulated Caco-2 cells respectively, to explore the effect of HIF-1α on LPS induced Caco-2 cell epithelial model; Part II: mTOR activator or inhibitor combined with or without HIF-1α activator, inhibitor to treat LPS-stimulated Caco-2 cells respectively, and then the molecular mechanism of HIF-1α reducing LPS induced Caco-2 cell epithelial model damage was detected. RESULTS: The results showed that HIF-1α activator decreased the permeability and up regulated tight junction (TJ) expression, while HIF-1α inhibitor had the opposite effect with the HIF-1α activator. mTOR activation increased, while mTOR inhibition decreased HIF-1α protein and expression of its downstream target molecules, which can be attenuated by HIF-1α activator or inhibitor. CONCLUSION: This study once again confirmed that HIF-1α alleviates LPS-induced mucosal epithelial model damage through P70S6K signalling pathway. It is of great value to explore whether HIF-2α plays crucial roles in the regulation of mucosal epithelial model functions in the future.

Multiple promoters and targeted microRNAs direct the expressions of HMGB3 gene transcripts in dairy cattle.

HMGB3 (high-mobility group box 3) is an X-linked member of a family of sequence-independent chromatin-binding proteins and functions as a universal sentinel for nucleic acid-mediated innate immune responses. The splice variant expression, promoter characterization and targeted microRNAs of the bovine HMGB3 gene were investigated to explore its expression pattern and possible regulatory mechanism. The results revealed that the expression of HMGB3 transcript variants 1 and 2 (HMGB3-TV1 and HMGB3-TV2) mRNA in the mastitis-infected mammary gland tissues was up-regulated by 8.46- and 5.31-fold respectively compared with that in healthy tissues (P < 0.05). HMGB3-TV1 was highly expressed in the mammary gland tissues, whereas HMGB3-TV2 was expressed primarily in liver. Functional analyses indicated that HMGB3 transcription is regulated by three distinct promoters - promoters 1, 2 and 3 (P1, P2 and P3) - resulting in two alternative transcripts with the same 3'-untranslated region. Promoter luciferase activity analysis suggested that the core sequences of P1 and P2 were mapped in the region of g.1535 to ~g.2076 and g.2074 to ~g.2491 respectively. The g.5880C>T SNP in P3 affected its base promoter activity, and different genotypes were associated with the bovine somatic count score. The expression of targets bovine miR-17-5p, miR-20b and miR-93 of the HMGB3 gene was down-regulated 1.56-, 1.72- and 2.94-fold respectively in mammary gland tissues as compared with that in healthy tissues (P < 0.05). The findings suggest that HMGB3 expression is under complex transcriptional and post-transcriptional control by alternate promoter usage, alternative splicing mechanism and microRNAs in dairy cattle.

Effects of garlic oil on milk fatty acid profile and lipogenesis-related gene expression in mammary gland of dairy goats.

BACKGROUND: Garlic oil (GO) has blood lipid-lowering effects. Milk fatty acid (FA) originates partly from plasma, and can be affected by the mammary lipogenesis. This study aimed to investigate GO effects on milk FA profile and mammary lipogenesis-related gene expression. Early-lactation goats were randomly allocated to four treatments with six goats each, and offered corn silage ad libitum and fixed amount of 0.79 kg day(-1) dry matter (DM) concentrate mixed with GO (0, 0.57, 1.14, 1.71 g kg(-1) DM) for 30 days consisting of 26-day adaptation. RESULTS: Intake of corn silage reduced (P≤0.05) as GO level increased in the concentrate. Lipase activity and lactose content linearly increased, while non-esterified FA concentration quadratically decreased with increasing GO level (P≤0.05). The proportions of short- and medium-chain (C14:0, C15:0 and C16:0) and saturated FA decreased, whereas C18, cis9 trans11 conjugated linoleic acid (c9t11 CLA), t10c12 CLA, monounsaturated and polyunsaturated FA, and some ≥ C20 FA proportions increased in a linear manner with increasing GO level (P≤0.05). The mRNA abundance of genes remained unchanged (P > 0.1) as GO level increased. CONCLUSION: Garlic oil altered milk FA profile and these effects may not be related to the mammary lipogenesis-related genes expression.

Lactiplantibacillus plantarum L47 and inulin alleviate enterotoxigenic Escherichia coli induced ileal inflammation in piglets by upregulating the levels of α-linolenic acid and 12,13-epoxyoctadecenoic acid.

Alternatives to antibiotics for preventing bacteria-induced inflammation in early-weaned farm animals are sorely needed. Our previous study showed that Lactiplantibacillus plantarum L47 and inulin could alleviate dextran sulfate sodium (DSS)-induced colitis in mice. To explore the protective effects of L. plantarum L47 and inulin on the ileal inflammatory response in weaned piglets challenged with enterotoxigenic Escherichia coli (ETEC), 28 weaned piglets were assigned into four groups, namely, CON group-orally given 10 mL/d phosphate buffer saline (PBS), LI47 group-orally given a mixture of 10 mL/d L. plantarum L47 and inulin, ECON group-orally given 10 mL/d PBS and challenged by ETEC, and ELI47 group-orally given 10 mL/d L. plantarum L47 and inulin mixture and challenged by ETEC. The results demonstrated that the combination of L. plantarum L47 and inulin reduced inflammatory responses and relieved the inflammatory damage caused by ETEC, including ileal morphological damage, reduced protein expression of ileal tight junction, decreased antioxidant capacity, and decreased anti-inflammatory factors. Transcriptome analysis revealed that L. plantarum L47 and inulin up-regulated the gene expression of phospholipase A2 group IIA (PLA2G2A) (P < 0.05) as well as affected alpha-linolenic acid (ALA) metabolism and linoleic acid metabolism. Moreover, L. plantarum L47 and inulin increased the levels of ALA (P < 0.05), lipoteichoic acid (LTA) (P < 0.05), and 12,13-epoxyoctadecenoic acid (12,13-EpOME) (P < 0.05) and the protein expression of Toll-like receptor 2 (TLR2) (P = 0.05) in the ileal mucosa. In conclusion, L. plantarum L47 and inulin together alleviated ETEC-induced ileal inflammation in piglets by up-regulating the levels of ALA and 12,13-EpOME via the LTA/TLR2/PLA2G2A pathway.

Exploring the effects of lysozyme dietary supplementation on laying hens: performance, egg quality, and immune response.

An experiment was conducted to evaluate the dietary supplementation with lysozyme's impacts on laying performance, egg quality, biochemical analysis, body immunity, and intestinal morphology. A total of 720 Jingfen No. 1 laying hens (53 weeks old) were randomly assigned into five groups, with six replicates in each group and 24 hens per replicate. The basal diet was administered to the laying hens in the control group, and it was supplemented with 100, 200, 300, or 400 mg/kg of lysozyme (purity of 10% and an enzyme activity of 3,110 U/mg) for other groups. The preliminary observation of the laying rate lasted for 4 weeks, and the experimental period lasted for 8 weeks. The findings demonstrated that lysozyme might enhance production performance by lowering the rate of sand-shelled eggs (P < 0.05), particularly 200 and 300 mg/kg compared with the control group. Lysozyme did not show any negative effect on egg quality or the health of laying hens (P > 0.05). Lysozyme administration in the diet could improve intestinal morphology, immune efficiency, and nutritional digestibility in laying hens when compared with the control group (P < 0.05). These observations showed that lysozyme is safe to use as a feed supplement for the production of laying hens. Dietary supplementation with 200 to 300 mg/kg lysozyme should be suggested to farmers as a proper level of feed additive in laying hens breeding.

One SNP in the 3'-UTR of HMGB1 gene affects the binding of target bta-miR-223 and is involved in mastitis in dairy cattle.

High-mobility group box protein 1 (HMGB1) gene has a universal sentinel function for nucleic acid-mediated innate immune responses and acts as a pathogenic mediator in the inflammatory disease. In an effort to identify the functional single-nucleotide polymorphism (SNP) in the 3'-untranslated region (UTR) of the bovine HMGB1 gene that affects the binding to its target microRNA, first, the expression of HMGB1 mRNA in different genotypes and its candidate bta-miR-223 was investigated. Quantitative real-time polymerase chain reaction results showed that the relative expression of HMGB1 mRNA in cows with the genotype GG is significantly higher than those in cows with the genotype AA (P < 0.05). The expression of bta-miR-223 was significantly upregulated by 1.95-fold (P < 0.05) in the bovine mastitis-infected mammary gland tissues compared with that in the healthy tissues. Subsequently, luciferase assay indicated that the HMGB1 expression was directly targeted by bta-miR-223 in human embryo kidney 293 T (HEK 293T) cells. One novel SNP (g. +2776 A > G) in the HMGB1 3'-UTR, altering the binding of HMGB1 and bta-miR-223, was found to be associated with somatic count scores in cows. Taken together, the g. +2776 A > G-GG was an advantageous genotype which can be used as a candidate functional marker for mastitis resistance breeding program.

Determination of the Effects of Duodenal Infusion Soy Protein Hydrolysate on Hepatic Glucose and Lipid Metabolism in Pigs Through Multi-Omics Analysis.

High animal protein intake increases hepatic lipid deposition and the risk of diabetes. However, the effects of high plant protein (HPP) intake on glycaemic responses and hepatic lipid metabolism in healthy people, as well as the underlying mechanisms, remain unclear. The current study explored the metabolomic and transcriptomic responses in the livers of pigs to assess the effects of HPP intake on host glucose and lipid metabolism. Sixteen pigs were infused with sterile saline or soy protein hydrolysate (SPH; 70 g/day) through a duodenal fistula twice daily during a 15 days experimental period. Hepatic metabolomic and transcriptomic analyses were performed, and the serum and hepatic biochemical parameters were measured. The results revealed that SPH infusion decreased serum glucose, hepatic triglyceride (TG), total cholesterol and low-density lipoprotein cholesterol levels, while it increased serum urea and eight hepatic amino acid levels (P < 0.05). Hepatic metabolomics displayed that SPH treatment produced seven different metabolites, four of which were related to lipid metabolism and one was related to glucose metabolism. In particular, lower (P < 0.05) glycocholic acid and glucose 1-phosphate levels and higher (P < 0.05) phosphatidylethanolamine (PE), arachidonic acid, prostaglandin F2α, l-carnitine and indole-3 acetic acid levels were observed following SPH infusion. A further metabolic pathway enrichment analysis found that these differential metabolites were mainly enriched in pathways related to lipid and glucose metabolism. Hepatic transcriptomics also demonstrated that multiple genes related to glucose and lipid metabolism were affected by SPH (P < 0.05). Together, SPH infusion reduced the hepatic TG levels by accelerating fatty acid ß-oxidation and inhibiting TG synthesis. In addition, SPH infusion reduced the serum glucose levels by promoting hepatic glucose uptake and glycolysis. This study's result demonstrated that HPP intake regulated glycaemic responses and hepatic lipid metabolism in pigs without increasing the risk of hepatic lipid deposition and hyperglycaemia.

Duodenal infusion of soy protein hydrolysate activates cAMP signaling and hypothalamic neurotransmitter synthesis in pigs.

Neurotransmitters in the brain are important for cognition and memory. As bioactive substrates, whether increased soy protein levels in pigs can promote hypothalamic neurotransmitter synthesis remains unclear. The effect of increased soy protein hydrolysate (SPH) levels in the small intestine of pigs on neurotransmitter precursor supply, hypothalamic neurotransmitter synthesis and underlying molecular processes was investigated by using sixteen pigs (35.2 ± 0.3 kg) infused either with SPH (70 g day-1) or sterile saline (control) twice daily for 15 days via a duodenal fistula. It demonstrated that SPH infusion increased the expression of the neutral amino acid transporter B0AT1 in the jejunal mucosa, the serum tyrosine/large neutral amino acid ratio, the concentrations of serum tyrosine, hypothalamic tyrosine, dopamine and brain-derived neurotrophic factor (BDNF) (P < 0.05). It also increased the jejunal and serum choline, hypothalamic choline and acetylcholine levels (P < 0.05). Hypothalamic transcriptome revealed that differential genes were significantly enriched in the cholinergic synapse, dopaminergic synapse and cyclic adenosine monophosphate (cAMP) signalling pathways, and that the expression of key enzyme genes in the synthesis of acetylcholine and dopamine and dopamine receptors 1 (DRD1) was upregulated by SPH (P < 0.05). Western blotting showed that SPH infusion activated the hypothalamic cAMP signalling pathways. Overall, SPH infusion promoted the synthesis of hypothalamic dopamine and acetylcholine, and the synthesised dopamine promoted BDNF production most likely through the activation of the cAMP signalling pathways by the DRD1.

Dietary Moringa oleifera leaf powder improves jejunal permeability and digestive function by modulating the microbiota composition and mucosal immunity in heat stressed rabbits.

Heat stress (HS) has detrimental effects on intestinal health by altering digestive and immune responses in animals. Dietary Moringa oleifera leaf powder (MOLP) has been implicated in ameliorating the impact of HS, but its effects in terms of intestinal function improvement under HS remain poorly characterized. Therefore, the current study investigated the impact of HS and MOLP supplementation on tight junction barriers, intestinal microbiota (jejunal digesta), and differentially expressed genes (DEGs) in jejunal mucosa of heat-stressed rabbits by using the next-generation sequencing techniques. A total of 21 male New Zealand White rabbits (32 weeks old mean body weight of 3318 ± 171 g) were divided into three groups (n = 7/group) as control (CON, 25 °C), heat stress (HS, 35 °C for 7 h daily), and HS with MOLP supplementation (HSM, 35 °C for 7 h daily) gavage at 200 mg/kg body weight per day for 4 weeks. The results indicated that MOLP supplementation increased mRNA expression of tight junction proteins and glutathione transferase activity, while the malonaldehyde concentration was decreased in the jejunal mucosa compared to HS group (P < 0.05). Furthermore, MOLP decreased the concentrations of lipopolysaccharide, pro-inflammatory cytokines, and myeloperoxidase compared with HS group (P < 0.05). Intestinal microbiota analysis revealed that at phyla level, the relative abundance of Bacteroidetes was higher in HSM group compared to CON and HS groups. MOLP supplementation also resulted in higher abundance of putatively health-associated genera such as Christensenellaceae R-7 gut group, Ruminococcaceae NK4A214 group, Ruminococcus 2, Lachnospiraceae NK4A136 group, and Lachnospiraceae unclassified along with higher butyrate levels in HSM group as compared to HS group. The analysis of DEGs revealed that MOLP reversed inflammatory response by downregulation of genes, such as TNFRSF13C, LBP, and COX2 in enriched KEGG pathway of NF-kß pathway. MOLP supplementation also significantly upregulated the expression of genes in protein digestion and absorption pathway, including PRSS2, LOC100349163, CPA1, CPB1, SLC9A3, SLC1A1, and SLC7A9 in HSM group. Three genes of fibrillar collagens, i.e., COL3A1, COL5A3, and COL12A1 in protein digestion were also down-regulated in HSM group. In conclusion, MOLP supplementation could improve jejunal permeability and digestive function, positively modulate microbiota composition and mucosal immunity in heat-stressed rabbits.