Maternal fiber-rich diet promotes early-life intestinal development in offspring through milk-derived extracellular vesicles carrying miR-146a-5p.

BACKGROUNDS: The intestinal development in early life is profoundly influenced by multiple biological components of breast milk, in which milk-derived extracellular vesicles (mEVs) contain a large amount of vertically transmitted signal from the mother. However, little is known about how maternal fiber-rich diet regulates offspring intestinal development by influencing the mEVs. RESULTS: In this study, we found that maternal resistant starch (RS) consumption during late gestation and lactation improved the growth and intestinal health of offspring. The mEVs in breast milk are the primary factor driving these beneficial effects, especially enhancing intestinal cell proliferation and migration. To be specific, administration of mEVs after maternal RS intake enhanced intestinal cell proliferation and migration in vivo (performed in mice model and indicated by intestinal histological observation, EdU assay, and the quantification of cyclin proteins) and in vitro (indicated by CCK8, MTT, EdU, and wound healing experiments). Noteworthily, miR-146a-5p was found to be highly expressed in the mEVs from maternal RS group, which also promotes intestinal cell proliferation in cells and mice models. Mechanically, miR-146a-5p target to silence the expression of ubiquitin ligase 3 gene NEDD4L, thereby inhibiting DVL2 ubiquitination, activating the Wnt pathway, and promoting intestinal development. CONCLUSION: These findings demonstrated the beneficial role of mEVs in the connection between maternal fiber rich diet and offspring intestinal growth. In addition, we identified a novel miRNA-146a-5p-NEDD4L-ß-catenin/Wnt signaling axis in regulating early intestinal development. This work provided a new perspective for studying the influence of maternal diet on offspring development.

Living, Heat-Killed Limosilactobacillus mucosae and Its Cell-Free Supernatant Differentially Regulate Colonic Serotonin Receptors and Immune Response in Experimental Colitis.

Lactobacillus species have been shown to alleviate gut inflammation and oxidative stress. However, the effect of different lactobacilli components on gut inflammation has not been well studied. This study aims to identify the differences in the effect and mechanisms of different forms and components of Limosilactobacillus mucosae (LM) treatment in the alleviation of gut inflammation using a colitis mouse model that is induced by dextran sodium sulfate (DSS). Seventy-two C57BL/6 mice were divided into six groups: control, DSS, live LM+DSS (LM+DSS), heat-killed LM+DSS (HKLM+DSS), LM cell-free supernatant + DSS (LMCS+DSS), and MRS medium + DSS (MRS+DSS). The mice were treated with different forms and components of LM for two weeks before DSS treatment. After that, the mice were sacrificed for an assessment of their levels of inflammatory cytokines, serotonin (5-HT) receptors (HTRs), and tryptophan metabolites. The results showed that, compared to other treatments, LMCS was more effective (p < 0.05) in the alleviation of DSS-induced body weight loss and led to an increase in the disease activity index score. All three forms and components of LM increased (p < 0.05) the levels of indole-3-acetic acid but reduced (p < 0.05) the levels of 5-HT in the colon. HKLM or LMCS reduced (p < 0.05) the percentages of CD3+CD8+ cytotoxic T cells but increased (p < 0.05) the percentages of CD3+CD4+ T helper cells in the spleen. LM or HKLM increased (p < 0.05) abundances of CD4+Foxp3+ regulatory T cells in the spleen. The LM and LMCS treatments reduced (p < 0.05) the expression of the pro-inflammatory cytokines Il6 and Il17a. The mice in the HKLM+DSS group had higher (p < 0.05) mRNA levels of the anti-inflammatory cytokine Il10, the cell differentiation and proliferation markers Lgr5 and Ki67, the 5-HT degradation enzyme Maoa, and HTRs (Htr1a, Htr2a, and Htr2b) in the colon. All three forms and components of LM reduced the phosphorylation of STAT3. The above findings can help to optimize the functionality of probiotics and develop new dietary strategies that aid in the maintenance of a healthy gut.

Lactobacillus amylovorus Promotes Lactose Utilization in Small Intestine and Enhances Intestinal Barrier Function in Intrauterine Growth Restricted Piglets.

BACKGROUND: Intrauterine growth restriction (IUGR) resulted in high mortality and many physiological defects of piglets, causing huge economic loss in the swine industry. Lactobacillus amylovorus (L. amylovorus) was identified as one of the main differential bacteria between IUGR and normal piglets. However, the effects of L. amylovorus on the growth performance and intestinal health in IUGR piglets remained unclear. OBJECTIVES: This study aimed to investigate the promoting effects of L. amylovorus Mafic1501, a new strain isolated from normal piglets, on the growth performance and intestinal barrier functions in IUGR piglets. METHODS: Newborn mice or piglets were assigned into 3 groups: CON (normal birth weight, control), IUGR (low birth weight), and IUGR+L. amy (low birth weight), administered with sterile saline or L. amylovorus Mafic1501, respectively. Growth performance, lactose content in the digesta, intestinal lactose transporter, and barrier function parameters were profiled. IPEC-J2 cells were cultured to verify the effects of L. amylovorus Mafic1501 on lactose utilization and intestinal barrier functions. RESULTS: L. amylovorus Mafic1501 elevated body weight and average daily gain of IUGR mice and piglets (P < 0.05). The lactose content in the ileum was decreased, whereas gene expression of glucose transporter 2 (GLUT2) was increased by L. amylovorus Mafic1501 in IUGR piglets during suckling period (P < 0.05). Besides, L. amylovorus Mafic1501 promoted intestinal barrier functions by increasing the villus height and relative gene expressions of tight junctions (P < 0.05). L. amylovorus Mafic1501 and its culture supernatant decreased the lactose level in the medium and upregulated gene expressions of transporter GLUT2 and tight junction protein Claudin-1 of IPEC-J2 cells (P < 0.05). CONCLUSION: L. amylovorus Mafic1501 improved the growth performance of IUGR piglets by promoting the lactose utilization in small intestine and enhancing intestinal barrier functions. Our results provided the new evidence of L. amylovorus Mafic1501 for its application in the swine industry.

Cinnamaldehyde alleviates zearalenone-induced LS174T cell apoptosis, barrier dysfunction and mucin reduction through JNK/NF-κB signaling pathway.

As a natural aldehyde organic compound, cinnamaldehyde (CA) is one of the main components of cinnamon essential oil with multiple bioactivities. In this study, we investigated the protective effects of CA on zearalenone (ZEA)-induced apoptosis, barrier dysfunction and mucin reduction, as well as underlying mechanisms in LS174T cells. In the present study, cells pre-treated with or without CA for 24 h were left untreated or subjected to ZEA for indicated time points Our results showed that 10 µM CA significantly prevented ZEA-induced cell viability decline, reversed ZEA-induced increase of the LDH level, cell cycle disruption and apoptosis in LS174T cells. Periodic acid-schiff (PAS) staining analysis showed that CA significantly alleviated the reduction of mucin secretion in LS174T cells caused by ZEA exposure. Western blot analysis showed that CA significantly reversed ZEA-induced reduction of the expression of mucin 2 (MUC2) and tight junction (TJ) proteins (claudin-1, claudin-3, ZO-1 and ZO-2) in LS174T cells. Notably, CA can significantly reduce the upregulation of the main effector of MAPK and NF-κB signaling pathways in LS174T cells. Further study showed that CA protects cells against ZEA-induced cellular damage through JNK/NF-κB signaling pathway in LS174T cells. Supplementation with CA might be an potential strategy to alleviate the damaging effect of ZEA on epithelial cells.

Limosilactobacillus mucosae and Lactobacillus amylovorus Protect Against Experimental Colitis via Upregulation of Colonic 5-Hydroxytryptamine Receptor 4 and Transforming Growth Factor-ß2.

BACKGROUND: Limosilactobacillusmucosae (LM) exerts anti-inflammatory and health-promoting effects. However, its role in the modulation of gut serotonin or 5-hydroxytryptamine (5-HT) metabolism and 5-HT receptors (HTRs) in inflammation requires further investigation. OBJECTIVES: We compared LM with Lactobacillus amylovorus (LA) for the regulation of 5-HT, HTRs, inflammatory mediators, and their correlations in the colon of mice with experimental colitis. METHODS: Male C57BL/6 mice were randomly assigned to 6 groups: control (Con), LM, LA, dextran sodium sulfate (DSS), and DSS with pre-administration of LM (+LM) or LA (+LA). After 7 d of DSS treatment, mice were killed to analyze the expression of inflammatory mediators, HTRs, and concentrations of 5-HT and microbial metabolites in the colon. RESULTS: LM was more effective than LA in alleviating DSS-induced colonic inflammation. Compared with mice in the DSS group, mice receiving DSS + LM or DSS + LA treatment had lower (P < 0.05) colonic mRNA expression of proinflammatory cytokines. DSS + LM treatment had lower mRNA expression of Il1b, Tnfa, and Ccl3, an abundance of p-STAT3, and greater expression of Tgfb2 and Htr4 in the colon (P < 0.05). The expression of inflammatory mediators (including Tgfb-1) was positively correlated (P < 0.05) with 5-HT and Htr2a and negatively correlated (P < 0.05) with Htr4. However, the expression of Tgfb-2 showed reversed correlations with the 5-HT and HTRs described above. Patterns for these correlations were different for LM and LA. Mice receiving the DSS + LM treatment had greater (P < 0.05) concentrations of acetate and valerate and lower (P < 0.05) concentrations of indole-3-acetic acid in the cecal and colonic contents. CONCLUSIONS: LM showed greater efficacy than LA in alleviating DSS-induced colonic inflammation. The coordinated regulation of transforming growth factor-ß subtypes and serotonin receptors in the colon may be one of the most important mechanisms underlying the probiotic effects of lactobacilli in gut inflammation.

Proinflammatory Polarization of Macrophages Causes Intestinal Inflammation in Low-Birth-Weight Piglets and Mice.

BACKGROUND: Low-birth-weight (LBW) animals suffer from intestinal damage and inflammation in their early life. OBJECTIVES: The aim of this study was to investigate the role of macrophages in intestinal inflammation in LBW piglets and mice. METHODS: Major genes involved in intestinal barrier function such as claudin-1, zonula occludens-1 (ZO-1), occludin, and mucin 2 and inflammatory cytokines such as IL-1ß, TNF-α, IL-10, and IL-13 were evaluated in 21-day-old, normal-birth-weight (NBW) and LBW piglets and mice. Macrophage markers such as CD16/32, CD163, and CD206 were also assessed by immunofluorescence and flow cytometry. Polarized and unpolarized macrophages were further transferred into NBW and LBW mice, followed by an evaluation of intestinal permeability and inflammation. RESULTS: Claudin-1 mRNA in LBW piglets as well as claudin-1, occludin, ZO-1, and mucin 2 mRNAs in LBW mice, was significantly downregulated. IL-1ß and TNF-α were significantly upregulated in LBW piglets (P < 0.05). LBW mice showed a reduced expression of IL-10 and IL-13 (P < 0.05), with a heightened IL-6 level (P < 0.01) in the jejunum. CD16, a marker for M1 macrophages, was significantly elevated in the jejunum of LBW piglets, whereas CD163, a marker for M2 macrophages, was significantly decreased (P < 0.05). Similarly, LBW mice had more CD11b+CD16/32+ M1 macrophages (P < 0.05) and fewer CD206+ M2 macrophages (P < 0.01) than NBW mice. Moreover, the transfer of M1 macrophages exacerbated intestinal inflammation in LBW mice. Furthermore, 2 major glycolysis-associated genes, hexokinase 2 (HK2) and lactate dehydrogenase A (LDHA), were significantly upregulated in LBW piglets and mice (P < 0.05). CONCLUSIONS: This study revealed for the first time that the intestinal macrophages are polarized toward a proinflammatory phenotype in LBW piglets and mice, contributing to intestinal inflammation. The findings of this study provide new options for the management of intestinal inflammation in LBW animals.

Amuc Prevents Liver Inflammation and Oxidative Stress in Mice Challenged with Salmonella Typhimurium.

BACKGROUND: Salmonella typhimurium is a pathogen that causes gastroenteritis in humans and animals. Amuc_1100 (hereafter called Amuc), the outer membrane protein of Akkermansia muciniphila, alleviates metabolic disorders and maintains immune homeostasis. OBJECTIVE: This study was conducted to determine whether there is a protective effect of Amuc administration. METHODS: Male 6-wk-old C57BL6J mice were randomly allocated into 4 groups: CON (control), Amuc (gavaged with Amuc, 100 µg/d for 14 d), ST (oral administration of 1.0 × 106 CFU S. typhimurium on day 7), and ST + Amuc (Amuc supplementation for 14 d, S. typhimurium administration on day 7). Serum and tissue samples were collected 14 d after treatment. Histological damage, inflammatory cell infiltration, apoptosis, and protein levels of genes associated with inflammation and antioxidant stress were analyzed. Data were analyzed by 2-way ANOVA and Duncan's multiple comparisons using SPSS software. RESULTS: The ST group mice had 17.1% lower body weight, 1.3-3.6-fold greater organ index (organ weight/body weight for organs including the liver and spleen), 10-fold greater liver damage score, and 3.4-10.1-fold enhanced aspartate transaminase, alanine transaminase, and myeloperoxidase activities, and malondialdehyde and hydrogen peroxide concentrations compared with controls (P < 0.05). The S. typhimurium-induced abnormalities were prevented by Amuc supplementation. Furthermore, the ST + Amuc group mice had 1.44-1.89-fold lower mRNA levels of proinflammatory cytokines (interleukin [Il]6, Il1b, and tumor necrosis factor-α) and chemokines (chemokine ligand [Ccl]2, Ccl3, and Ccl8) and 27.1%-68.5% lower levels of inflammation-related proteins in the liver than ST group mice (P < 0.05). CONCLUSIONS: Amuc treatment prevents S. typhimurium-induced liver damage partly through the toll-like receptor (TLR)2/TLR4/myeloid differentiation factor 88 and nuclear factor-κB signaling as well as nuclear factor erythroid-2 related factor signaling pathways. Thus, Amuc supplementation may be effective in treating liver injury in S. typhimurium-challenged mice.

Linking serotonin homeostasis to gut function: Nutrition, gut microbiota and beyond.

Serotonin (5-HT) produced by enterochromaffin (EC) cells in the digestive tract is crucial for maintaining gut function and homeostasis. Nutritional and non-nutritional stimuli in the gut lumen can modulate the ability of EC cells to produce 5-HT in a temporal- and spatial-specific manner that toning gut physiology and immune response. Of particular interest, the interactions between dietary factors and the gut microbiota exert distinct impacts on gut 5-HT homeostasis and signaling in metabolism and the gut immune response. However, the underlying mechanisms need to be unraveled. This review aims to summarize and discuss the importance of gut 5-HT homeostasis and its regulation in maintaining gut metabolism and immune function in health and disease with special emphasis on different types of nutrients, dietary supplements, processing, and gut microbiota. Cutting-edge discoveries in this area will provide the basis for the development of new nutritional and pharmaceutical strategies for the prevention and treatment of serotonin homeostasis-related gut and systematic disorders and diseases.

Excessive dietary L-tryptophan regulated amino acids metabolism and serotonin signaling in the colon of weaning piglets with acetate-induced gut inflammation.

L-Tryptophan (Trp) was shown to improve the gut barrier and growth of weaning piglets. However, whether excessive dietary Trp regulates amino acids (AAs) metabolism and gut serotonin (5-HT) homeostasis in piglets with gut inflammation is not clear yet. We hypothesize that excessive dietary Trp alleviates acetate-induced colonic inflammation and gut barrier damage in weaning piglets partially through the regulation of colonic AAs metabolism and 5-HT signaling. Fifty-four 21-day-old weaned piglets were divided into six groups: control, acetate, 0.2%Trp, 0.2%Trp + acetate, 0.4% Trp, and 0.4%Trp + acetate. Piglets were fed a basal diet supplemented with 0%, 0.2%, or 0.4% of Trp throughout the 12-day experiment. During days 0-7, all piglets had free access to diet and drinking water. On day 8, piglets were intrarectal administered with 10 mL of 10% acetate saline solution or 0.9% saline. During days 8-12, all piglets were pair-fed the same amount of feed per kg bodyweight. Results showed that excessive dietary Trp alleviated acetate-induced reductions in daily weight gain and increase in feed/gain ratio. Trp restored (P < 0.05) acetate-induced increase in concentrations of free aspartate, glutamate/glutamine, glycine, 5-HT, and 3-methylindole in the colon, downregulation of zonula occludens-1 and 5-HT reuptake transporter (SERT) expression and upregulation of IL-1ß, IL-8, TLR4, and 5-HT receptor 2A (HTR2A) expression, and the increase in ratios of p-STAT3/ STAT3 and p-p65/p65 in the colon. The above findings suggested that excessive dietary Trp in the proper amount regulated colonic AAs metabolism, 5-HT homeostasis, and signaling that may contribute as important regulators of gut inflammation during the weaning transition.

Hydrophobic modification improves the delivery of cell-penetrating peptides to eliminate intracellular pathogens in animals.

Infections induced by intracellular pathogens are difficult to eradicate due to poor penetration of antimicrobials into cell membranes. It is of great importance to develop a new generation of antibacterial agents with dual functions of efficient cell penetration and bacterial inhibition. In this study, the association between hydrophobicity and cell-penetrating peptide delivery efficiency was investigated by fragment interception and hydrophobicity modification of natural porcine antimicrobial peptide PR-39 and the combination of cationic cell-penetrating peptide (R6) with antimicrobial peptide fragments modified with hydrophobic residues. The chimeric peptides P3I7 and P3L7, obtained through biofunctional screening, exhibited potent broad-spectrum antibacterial activity and low cytotoxicity. Moreover, P3I7 and P3L7 can effectively penetrate cells to eliminate intracellular pathogens mainly through endocytosis. The membrane destruction mechanism makes the peptides fast sterilizers and less prone to developing drug resistance. Finally, their good biocompatibility and antibacterial infection effects were verified in mice and piglets. To conclude, the chimeric peptides P3I7 and P3L7 show great potential as affordable and effective antimicrobial agents and may serve as ideal candidates for the treatment of intracellular bacterial infections. STATEMENT OF SIGNIFICANCE: The low permeability of antibacterial drugs makes infections induced by intracellular bacteria extremely difficult to treat. To address this issue, we designed chimeric peptides with dual cell-penetrating and antibacterial functions. The active peptides P3I7 and P3L7, acquired through functional screening have strong broad-spectrum antibacterial activity and powerful bactericidal effects against intracellular Staphylococcus aureus. The membrane permeation mechanism of P3I7 and P3L7 against bacteria endows fast bactericidal activity with low drug resistance. The biosafety and antibacterial activity of P3I7 and P3L7 were also validated by in vivo trials. This study provides an ideal drug candidate against intracellular bacterial infections.

Chitosan oligosaccharide alleviates and removes the toxicological effects of organophosphorus pesticide chlorpyrifos residues.

The abuse of chlorpyrifos (CHP), a commonly used organophosphorus pesticide, has caused many environmental pollution problems, especially its toxicological effects on non-target organisms. First, CHP enriched on the surface of plants enters ecosystem circulation along the food chain. Second, direct inflow of CHP into the water environment under the action of rainwater runoff inevitably causes toxicity to non-target organisms. Therefore, we used rats as a model to establish a CHP exposure toxicity model and studied the effects of CHP in rats. In addition, to alleviate and remove the injuries caused by residual chlorpyrifos in vivo, we explored the alleviation effect of chitosan oligosaccharide (COS) on CHP toxicity in rats by exploiting its high water solubility and natural biological activity. The results showed that CHP can induce the toxicological effects of intestinal antioxidant changes, inflammation, apoptosis, intestinal barrier damage, and metabolic dysfunction in rats, and COS has excellent removal and mitigation effects on the toxic damage caused by residual CHP in the environment. In summary, COS showed significant biological effects in removing and mitigating blood biochemistry, antioxidants, inflammation, apoptosis, gut barrier structure, and metabolic function changes induced by residual CHP in the environment.

L-Tryptophan Differentially Regulated Glucose and Amino Acid Transporters in the Small Intestine of Rat Challenged with Lipopolysaccharide.

Tryptophan (Trp) has been shown to improve the growth and gut function of weaned piglets. Whether the growth-promoting effect of Trp is due to the improvement in nutrient transport and absorption during weaning or under conditions of inflammation has not been fully characterized. The objective of this study was to determine the effects of Trp on lipopolysaccharide (LPS)-induced changes in glucose and amino acid (AA) transport in the rat jejunum. Twenty-four 7-week-old Sprague Dawley rats were randomly divided into one of three groups: control, LPS, and Trp + LPS. Rats were supplemented with 0 or 0.1 mg Trp per gram body weight/d in drinking water for 7 days and were intraperitoneally injected with LPS (5 mg/kg BW) on day 8. After 24 h, rats were sacrificed, and jejunum samples were isolated for the analysis of glucose and AA transport using an Ussing chamber and the expression of glucose and AA transporters. The results showed that Trp alleviated the LPS-induced increase in jejunal permeability (p < 0.05) and decrease in changes in the short-circuit current of glucose, arginine, glutamine, glutamate, glycine, histidine, leucine, lysine, taurine, threonine, and Trp (p < 0.05). Trp reversed (p < 0.05) the LPS-induced downregulation of expression of the glucose transporter SGLT1 and AA transporters solute carrier family 38 member 2 (SNAT2) and solute carrier family 7 member 8 (LAT2), as well as ATPase Na+/K+ transporting subunit alpha 2 (ATP1A2). However, Trp increased (p < 0.01) the LPS-induced upregulation of acidic AA transporter solute carrier family 1 member 1 (EAAT3) expression. The above findings may help to develop nutritional interventions for the differential targeting of gut nutrient transporters, aiming to improve gut function and health in the presence of inflammation in both humans and animals.

L-leucine supplementation reduces growth performance accompanied by changed profiles of plasma amino acids and expression of jejunal amino acid transporters in breast-fed intra-uterine growth-retarded piglets.

Previously, we provided an evidence that L-leucine supplementation facilitates growth performance in suckling piglets with normal birth weight. However, it remains hitherto obscure weather breast-fed piglets displaying intrauterine growth restriction (IUGR) show a similar effect in response to L-leucine provision. In this study, seven-day-old sow-reared IUGR piglets were orally administrated with L-leucine (0, 0.7 1.4, 2.1 g/kg BW) twice daily for two weeks. Increasing leucine levels hampered the growth performance of suckling IUGR piglets. The average daily gain of IUGR piglets was significantly reduced in 1.4 g/kg BW and 2.1 g/kg BW L-leucine supplementation groups (P < 0.05). Except for ornithine and glutamine, the plasma concentrations of other amino acids were abated as L-leucine levels increased (P < 0.05). Leucine supplementation led to reduction in the levels of urea, blood ammonia, blood glucose, triglyceride, and total cholesterol, as well as an elevation in the level of low density lipoprotein cholesterol in suckling IUGR piglets (P < 0.05). In addition, 1.4g/kg BW of L-leucine enhanced the mRNA expression of ATB 0,+ , whereas decreased the mRNA abundances of CAT1, y+LAT1, ASCT2 and b 0,+ AT in the jejunum (P < 0.05). Concomitantly, the jejunum of IUGR piglets in L-leucine group contains more ATB0,+ and less SNAT2 protein than in the control (P < 0.05). Collectively, L-leucine supplementation impairs growth performance in breast-fed IUGR piglets, which may be associated with depressed nutritional conditions and alterations in the uptake of amino acids and the expression of amino acid transporters in the small intestine.

Hydroxyproline alleviates 4-hydroxy-2-nonenal-induced DNA damage and apoptosis in porcine intestinal epithelial cells.

Oxidative stress has been confirmed in relation to intestinal mucosa damage and multiple bowel diseases. Hydroxyproline (Hyp) is an imino acid abundant in sow's milk. Compelling evidence has been gathered showing the potential antioxidative properties of Hyp. However, the role and mechanism of Hyp in porcine intestinal epithelial cells in response to oxidative stress remains unknown. In this study, small intestinal epithelial cell lines of piglets (IPEC-1) were used to evaluate the protective effects of Hyp on 4-hydroxy-2-nonenal (4-HNE)-induced oxidative DNA damage and apoptosis. IPEC-1 pretreated with 0.5 to 5 mmol/L Hyp were exposed to 4-HNE (40 µmol/L) in the presence or absence of Hyp. Thereafter, the cells were subjected to apoptosis detection by Hoechst staining, flow cytometry, and Western blot or DNA damage analysis by comet assay, immunofluorescence, and reverse-transcription quantitative PCR (RT-qPCR). Cell apoptosis and the upregulation of cleaved-caspase-3 induced by 4-HNE (40 µmol/L) were inhibited by 5 mmol/L of Hyp. In addition, 5 mmol/L Hyp attenuated 4-HNE-induced reactive oxygen species (ROS) accumulation, glutathione (GSH) deprivation and DNA damage. The elevation in transcription of GADD45a (growth arrest and DNA-damage-inducible protein 45 alpha) and GADD45b (growth arrest and DNA-damage-inducible protein 45 beta), as well as the phosphorylation of H2AX (H2A histone family, member X), p38 MAPK (mitogen-activated protein kinase), and JNK (c-Jun N-terminal kinase) in cells treated with 4-HNE were alleviated by 5 mmol/L Hyp. Furthermore, Hyp supplementation increased the protein abundance of Krüppel like factor 4 (KLF4) in cells exposed to 4-HNE. Suppression of KLF4 expression by kenpaulone impeded the resistance of Hyp-treated cells to DNA damage and apoptosis induced by 4-HNE. Collectively, our results indicated that Hyp serves to protect against 4-HNE-induced apoptosis and DNA damage in IPEC-1 cells, which is partially pertinent with the enhanced expression of KLF4. Our data provides an updated explanation for the nutritional values of Hyp-containing animal products.

Tryptophan regulates bile and nitrogen metabolism in two pig gut lactobacilli species in vitro based on metabolomics study.

Research has demonstrated that tryptophan (Trp) regulated the composition and metabolism of the gut microbiota. However, the detailed mode of action of Trp on the metabolism of intestinal commensal lactobacilli has not been well characterized. This study aimed to compare the effects of Trp concentration (0.2, 0.4, 0.6 mmol/L) in the media on the metabolism of Lactobacillus amylovorus and Limosilactobacillus mucosae isolated from the small intestine of piglets in vitro by high-performance liquid chromatography and metabolomics study. Results showed that increased Trp concentration increased (P < 0.05) net utilization of lysine, methionine, tryptophan, asparagine/aspartate, glutamine/glutamate, however, increased net production of glycine and taurine in Lac. amylovorus. In contrast, increased Trp concentration decreased (P < 0.05) net utilization of leucine, phenylalanine, and serine and increased (P < 0.05) net utilization of arginine and net production of ornithine and glycine in Lim. mucosae. Targeted metabolomics analysis showed that increased Trp concentration promoted (P < 0.05) the production of indole-3-lactic acid and 3-indoleacetic acid in the two lactobacilli strains. Increased concentration of Trp increased (P < 0.01) glycochenodeoxycholic acid metabolism in Lim. mucosae and glycocholic acid and taurocholic acid metabolism in Lac. amylovorus. Untargeted metabolomics analysis showed that metabolic pathways related to phenylalanine and tryptophan metabolism, and nicotinate and nicotinamide metabolism were regulated by Trp in Lim. mucosae. These findings will help develop new biomarkers and dietary strategies to maintain the functionality of the gut microbiota aiming at improving the nutrition and health of both humans and animals.

Insights into diet-associated oxidative pathomechanisms in inflammatory bowel disease and protective effects of functional amino acids.

There has been a substantial rise in the incidence and prevalence of clinical patients presenting with inflammatory bowel disease (IBD), which includes Crohn's disease and ulcerative colitis. Accumulating evidence has corroborated the view that dietary factors (particularly diets with high levels of saturated fat or sugar) are involved in the development and progression of IBD, which is predominately associated with changes in the composition of the gut microbiota and an increase in the generation of reactive oxygen species. Notably, the ecological imbalance of the gut microbiome exacerbates oxidative stress and inflammatory responses, leading to perturbations of the intestinal redox balance and immunity, as well as mucosal integrity. Recent findings have revealed that functional amino acids, including L-glutamine, glycine, L-arginine, L-histidine, L-tryptophan, and hydroxyproline, are effectively implicated in the maintenance of intestinal redox and immune homeostasis. These amino acids and their metabolites have oxygen free-radical scavenging and inflammation-relieving properties, and they participate in modulation of the microbial community and the metabolites in the gut. The principal focus of this article is a review of recent advances in the oxidative pathomechanisms of IBD development and progression in relation to dietary factors, with a particular emphasis on the redox and signal transduction mechanisms of host cells in response to unbalanced diets and enterobacteria. In addition, an update on current understanding of the protective effects of functional amino acids against IBD, together with the underlying mechanisms for this protection, have been provided.

Roles of stress response-related signaling and its contribution to the toxicity of zearalenone in mammals.

Zearalenone (ZEA) is a mycotoxin frequently found in cereal crops and cereal-derived foodstuffs worldwide. It affects plant productivity, and is also a serious hazard to humans and animals if being exposed to food/feed contaminated by ZEA. Studies over the last decade have shown that the toxicity of ZEA in animals is mainly mediated by the various stress responses, such as endoplasmic reticulum (ER) stress, oxidative stress, and others. Accumulating evidence shows that oxidative stress and ER stress signaling are actively implicated in and contributes to the pathophysiology of various diseases. Biochemically, the deleterious effects of ZEA are associated with apoptosis, DNA damage, and lipid peroxidation by regulating the expression of genes implicated in these biological processes. Despite these findings, the underlying mechanisms responsible for these alterations remain unclear. This review summarized the characteristics, metabolism, toxicity and the deleterious effects of ZEA exposure in various tissues of animals. Stress response signaling implicated in the toxicity as well as potential therapeutic options with the ability to reduce the deleterious effects of ZEA in animals were highlighted and discussed.

Glycine represses endoplasmic reticulum stress-related apoptosis and improves intestinal barrier by activating mammalian target of rapamycin complex 1 signaling.

Endoplasmic reticulum (ER) stress has been associated with the dysfunction of intestinal barrier in humans and animals. We have previously shown that oral administration of glycine to suckling-piglets improves ER stress-related intestinal mucosal barrier impairment and jejunal epithelial apoptosis. However, the underlying mechanism remains unknown. In this study, the protective effect and the mechanism of glycine on apoptosis and dysfunction in intestinal barrier induced by brefeldin A (BFA), an ER stress inducer, was explored in porcine intestinal epithelial cells (IPEC-1). The results showed that BFA treatment led to enhanced apoptosis and upregulation of proteins involved in ER stress signaling, including inositol-requiring enzyme 1α (IRE1α), activating transcription factor 6α (ATF6α), c-Jun N-terminal kinase (JNK), and C/EBP-homologous protein (CHOP). In addition, BFA induced a dysfunction in intestinal epithelial barrier, as evidenced by the increased paracellular permeability, decreased transepithelial electrical resistance (TEER), and reduced abundance of tight junction proteins (occludin, claudin-1, zonula occludens [ZO]-1, and ZO-2). These alterations triggered by BFA were significantly abolished by glycine treatment (P < 0.05), indicating a protective effect of glycine on barrier function impaired by ER stress. Importantly, we found that the regulatory effect of glycine on intestinal permeability, proteins implicated in ER stress and apoptosis, as well as the morphological alterations of the ER were reversed by rapamycin. In summary, our results indicated that glycine alleviates ER stress-induced apoptosis and intestinal barrier dysfunction in IPEC-1 cells in a mammalian target of rapamycin complex 1 (mTORC1)-dependent manner. The data provides in vitro evidence and a mechanism for the protective effect of glycine against the disruption of intestinal barrier integrity induced by ER stress.

Glycine regulates mucosal immunity and the intestinal microbial composition in weaned piglets.

Glycine is an amino acid with a diverse array of health benefits regarding metabolism, immunity, and development. The aim of this study was to test the hypothesis that glycine supplementation alters the intestinal microbial composition and improves the intestinal mucosal immunity of weaned piglets. One hundred and twenty-eight weaned piglets divided into 4 groups were fed with a corn- and soybean meal-based diet supplemented with 0 (control), 0.5, 1, or 2% glycine for 7 days. The intestinal microbiota and tissue samples from the control and the 2% glycine-supplemented piglets were collected for determination of the composition of microbial community and the intestinal mucosal barrier function. Piglets fed with diet containing 2% glycine, instead of 0.5% or 1% glycine, presented elevated average daily gain and feed conversion ratio, as compared with the control. 2% glycine enhanced the abundance of mucins in the jejunum and ileum and mRNA level of porcine ß-defensin (pBD) 2 and pBD-3, as well as the protein level of secretory immunoglobulin A (sIgA) in the jejunum. The mRNA expression of tumor necrosis factor (TNF)-α, interleukin (IL)-1ß, and IL-6, and the protein level of phosphorylated p38 mitogen-activated protein kinase (MAPK), signal transducer and activator of transcription 3 (STAT3), nuclear factor (NF)-κB p65, and claudin-2 in the jejunum were lower in the 2% glycine group than that in the control. In addition, an elevated ratio of CD4+/CD8+ T lymphocytes was observed in the jejunum of piglets receiving diet supplemented with 2% glycine. The colon content of piglets fed with 2% glycine exhibited a reduction in abundance of pathogenic bacteria (Escherichia-Shigella, Clostridium, and Burkholderiales) and an increase in short-chain fatty acid-producing bacteria (Blautia, Lachnospiraceae, Anaerostipes, and Prevotella) in comparison with the control. We conclude that dietary supplementation with 2% glycine improves the intestinal immunological barrier function and the microbial composition, therefore, contributing to the growth performance of weaned piglets.

Amino Acids in Microbial Metabolism and Function.

Amino acids (AAs) not only serve as building blocks for protein synthesis in microorganisms but also play important roles in their metabolism, survival, inter-species crosstalk, and virulence. Different AAs have their distinct functions in microbes of the digestive tract and this in turn has important impacts on host nutrition and physiology. Deconjugation and re-conjugation of glycine- or taurine- conjugated bile acids in the process of their enterohepatic recycling is a good example of the bacterial adaptation to harsh gut niches, inter-kingdom cross-talk with AA metabolism, and cell signaling as the critical control point. It is also a big challenge for scientists to modulate the homeostasis of the pools of AAs and their metabolites in the digestive tract with the aim to improve nutrition and regulate AA metabolism related to anti-virulence reactions. Diversity of the metabolic pathways of AAs and their multi-functions in modulating bacterial growth and survival in the digestive tract should be taken into consideration in recommending nutrient requirements for animals. Thus, the concept of functional amino acids can guide not only microbiological studies but also nutritional and physiological investigations. Cutting edge discoveries in this research area will help to better understand the mechanisms responsible for host-microbe interactions and develop new strategies for improving the nutrition, health, and well-being of both animals and humans.