Effects of dietary chlorogenic acid on cecal microbiota and metabolites in broilers during lipopolysaccharide-induced immune stress.

Aims: The aim of this study was to investigate the effects of chlorogenic acid (CGA) on the intestinal microorganisms and metabolites in broilers during lipopolysaccharide (LPS)-induced immune stress. Methods: A total of 312 one-day-old Arbor Acres (AA) broilers were randomly allocated to four groups with six replicates per group and 13 broilers per replicate: (1) MS group (injected with saline and fed the basal diet); (2) ML group (injected with 0.5 mg LPS/kg and fed the basal diet); (3) MA group (injected with 0.5 mg LPS/kg and fed the basal diet supplemented with 1,000 mg/kg CGA); and (4) MB group (injected with saline and fed the basal diet supplemented with 1,000 mg/kg CGA). Results: The results showed that the abundance of beneficial bacteria such as Bacteroidetes in the MB group was significantly higher than that in MS group, while the abundance of pathogenic bacteria such as Streptococcaceae was significantly decreased in the MB group. The addition of CGA significantly inhibited the increase of the abundance of harmful bacteria such as Streptococcaceae, Proteobacteria and Pseudomonas caused by LPS stress. The population of butyric acid-producing bacteria such as Lachnospiraceae and Coprococcus and beneficial bacteria such as Coriobacteriaceae in the MA group increased significantly. Non-targeted metabonomic analysis showed that LPS stress significantly upregulated the 12-keto-tetrahydroleukotriene B4, riboflavin and mannitol. Indole-3-acetate, xanthurenic acid, L-formylkynurenine, pyrrole-2-carboxylic acid and L-glutamic acid were significantly down-regulated, indicating that LPS activated inflammation and oxidation in broilers, resulting in intestinal barrier damage. The addition of CGA to the diet of LPS-stimulated broilers significantly decreased 12-keto-tetrahydro-leukotriene B4 and leukotriene F4 in arachidonic acid metabolism and riboflavin and mannitol in ABC transporters, and significantly increased N-acetyl-L-glutamate 5-semialdehyde in the biosynthesis of amino acids and arginine, The presence of pyrrole-2-carboxylic acid in D-amino acid metabolism and the cecal metabolites, indolelactic acid, xanthurenic acid and L-kynurenine, indicated that CGA could reduce the inflammatory response induced by immune stress, enhance intestinal barrier function, and boost antioxidant capacity. Conclusion: We conclude that CGA can have a beneficial effect on broilers by positively altering the balance of intestinal microorganisms and their metabolites to inhibit intestinal inflammation and barrier damage caused by immune stress.

Effects of quercetin and genistein on egg quality, lipid profiles, and immunity in laying hens.

BACKGROUND: After the peak laying stage, laying hens become susceptible to lipid accumulation and inflammatory reactions. The objective of this experiment was to examine the impact of quercetin and genistein on egg quality and lipid profiles in laying hens. A total of 240 Hy-Line Brown hens were randomly assigned to three dietary treatments. Each treatment had eight replicates, with ten hens in each replicate, and the hens were aged between 46 and 56 weeks. The test diets consisted of a corn-soybean meal-based basal diet, a basal diet supplemented with 300 mg kg-1 quercetin, and a basal diet supplemented with 300 mg kg-1 quercetin and 40 mg kg-1 genistein. RESULTS: Results showed that, separately, supplemental quercetin significantly improved egg quality (eggshell strength, albumen height, and Haugh unit, P < 0.05) and reduced the deposition of abdominal fat (P < 0.05). Our findings also showed that, separately or as a combination, supplemental quercetin and genistein significantly increased eggshell thickness (P < 0.05), decreased the levels of lipids in serum (low-density lipoprotein cholesterol, total cholesterol, total triglycerides, and non-esterified fatty acids, P < 0.05) and significantly increased serum immunoglobulins A and G (P < 0.05), and promoted the expression of splenic immune-related genes (IgA and IL-4, P < 0.05). CONCLUSION: This study confirmed that supplemental quercetin and genistein, either separately or in combination, can enhance eggshell thickness, lipid profiles, and immune function in aging hens. Moreover, both quercetin alone and quercetin + genistein exhibited similar abilities to lower lipid levels and improve immune function. © 2023 Society of Chemical Industry.

The ionome and proteome landscape of aging in laying hens and relation to egg white quality.

The ionome is essential for maintaining body function and health status by participating in diverse key biological processes. Nevertheless, the distribution and utilization of ionome among different organs and how aging impacts the ionome leading to a decline in egg white quality remain unknown. Thus, we used inductively coupled plasma mass spectrometry (ICP-MS) to analyze 35 elements and their isotopic contents in eight organs of laying hens at 35, 72, and 100 weeks. Moreover, the magnum proteome, amino acids in egg white, and egg white quality were analyzed in laying hens at three different ages using 4D proteomics techniques, an amino acid analyzer, and an egg quality analyzer. Across the organs, we identified varying distribution patterns among macroelements (Mg24, Ca43/44, K39, and P31), transition metals (Zn64/66, Cu63/65, Fe56/57, and Mn55), and toxic elements (Pb208, Ba137, and Sr86). We observed an organ-specific aging pattern characterized by the accumulation of toxic elements (Pb208, Ba137, and Sr86) and calcification in the small intestine. Additionally, a decrease in the utilization of essential trace elements selenium (Se78/82) and manganese (Mn55) was noted in the oviduct. By analyzing ionome in tandem with egg quality, egg white amino acids, and proteome, we unveiled that the reduction of selenium and manganese concentrations in the magnum during the aging process affected amino acid metabolism, particularly tryptophan metabolism, thereby inhibiting the amino acid synthesis in the magnum. Furthermore, it accelerated the senescence of magnum cells through necroptosis activation, leading to a decline in the albumen secretion function of the magnum and subsequently reducing egg white quality. Overall, this study provides insights into the evolution of 35 elements and their isotopes across 8 organs of laying hens with age. It also reveals the elemental composition, interactions, and utilization patterns of these organs, as well as their correlation with egg white quality. The present study highlights the significance of ionome and offers a comprehensive perspective on the selection of ionome for regulating the aging of laying hens.

Effects of Bacillus subtilis Natto NB205 and Its Mutant NBMK308 on Egg Quality in Aging Laying Hens.

In aging laying hens, reproductive changes reduce egg quality. Bacillus subtilis natto (B. subtilis) is a versatile bacterium with high vitamin K2 content, providing health benefits for animals and humans. This study investigated the effect of B. subtilis natto NB205 and its mutant NBMK308 on egg quality in aging laying hens. Results showed that NB205 and NBMK308 supplementation significantly improved albumen height (p < 0.001), Haugh units (p < 0.05), and eggshell thickness (p < 0.001) compared to the control group. Supplementation also increased ovalbumin expression, regulated tight junction (TJ) proteins, reduced pro-inflammatory cytokine levels, and improved the health and productivity of aging laying hens by regulating key apoptosis-related genes in the magnum part of the oviduct. There were differences in the expression of vitamin K-dependent proteins (VKDPs) in the magnum between NB205 and NBMK308, but no significant differences in the improvement of egg quality. Supplementation with NB205 and NBMK308 can improve egg quality in aging laying hens.

Effects of Selenium Yeast on Egg Quality, Plasma Antioxidants, Selenium Deposition and Eggshell Formation in Aged Laying Hens.

Internal egg and eggshell quality are often deteriorated in aging laying hens, which causes huge economic losses in the poultry industry. Selenium yeast (SY), as an organic food additive, is utilized to enhance laying performance and egg quality. To extend the egg production cycle, effects of selenium yeast supplementation on egg quality, plasma antioxidants and selenium deposition in aged laying hens were evaluated. In this study, five hundred and twenty-five 76-week-old Jing Hong laying hens were fed a selenium-deficient (SD) diet for 6 weeks. After Se depletion, the hens were randomly divided into seven treatments, which included an SD diet, and dietary supplementation of SY and sodium selenite (SS) at 0.15, 0.30, and 0.45 mg/kg to investigate the effect on egg quality, plasma antioxidant capacity, and selenium content in reproductive organs. After 12 weeks of feeding, dietary SY supplementation resulted in higher eggshell strength (SY0.45) (p < 0.05) and lower shell translucence. Moreover, organs Se levels and plasma antioxidant capacity (T-AOC, T-SOD, and GSH-Px activity) were significantly higher with Se supplementation (p < 0.05). Transcriptomic analysis identified some key candidate genes including cell migration inducing hyaluronidase 1 (CEMIP), ovalbumin (OVAL), solute carrier family 6 member 17 (SLC6A17), proopiomelanocortin (POMC), and proenkephalin (PENK), and potential molecular processes (eggshell mineralization, ion transport, and eggshell formation) involved in selenium yeast's effects on eggshell formation. In conclusion, SY has beneficial functions for eggshell and we recommend the supplementation of 0.45 mg/kg SY to alleviate the decrease in eggshell quality in aged laying hens.

Effects of dietary chlorogenic acid on intestinal barrier function and the inflammatory response in broilers during lipopolysaccharide-induced immune stress.

Immune stress exerts detrimental effects on growth performance and intestinal barrier function during intensive animal production with ensuing serious economic consequences. Chlorogenic acid (CGA) is used widely as a feed additive to improve the growth performance and intestinal health of poultry. However, the effects of dietary CGA supplementation on amelioration of the intestinal barrier impairment caused by immune stress in broilers are unknown. This study investigated the effects of CGA on growth performance, intestinal barrier function, and the inflammatory response in lipopolysaccharide (LPS) mediated immune-stressed broilers. Three hundred and twelve 1-day-old male Arbor Acres broilers were divided randomly into 4 groups with 6 replicates of thirteen broilers. The treatments included: i) saline group: broilers injected with saline and fed with basal diet; ii) LPS group: broilers injected with LPS and fed with basal diet; iii) CGA group: broilers injected with saline and feed supplemented with CGA; and iv) LPS+CGA group: broilers injected with LPS and feed supplemented with CGA. Animals in the LPS and LPS+CGA groups were injected intraperitoneally with an LPS solution prepared with saline from 14 d of age for 7 consecutive days, whereas broilers in the other groups were injected only with saline. LPS induced a decrease in feed intake of broilers during the stress period, but CGA effectively alleviated this decrease. Moreover, CGA inhibited the reduction of villus height and improved the ratio of villus height to crypt depth in the duodenum of broilers 24 and 72 h after LPS injection. In addition, dietary CGA supplementation significantly restored the expression of cation-selective and channel-forming Claudin2 protein 2 h after LPS injection in the ileum. LPS enhanced the expression of tumor necrosis factor-α (TNF-α) and interleukin-1ß (IL-1ß) in the small intestine, but this enhancement was blocked by CGA supplementation. The expression of interleukin-10 (IL-10) increased with LPS injection and CGA promoted the production of IL-10. CGA addition downregulated the expression of intestinal interleukin-6 (IL-6) of broilers under normal rearing conditions. However, CGA supplementation upregulated the expression of IL-6 of broilers 72 h after LPS injection. The data demonstrate that dietary supplementation with CGA alleviates intestinal barrier damage and intestinal inflammation induced by LPS injection during immune stress thereby improving growth performance of broilers.

Energy values evaluation and improvement of soybean meal in broiler chickens through supplemental mutienzyme.

This study measured the metabolizable energy of soybean meal (SBM) and evaluated effects of soybean meal specific enzymes supplementation in corn-soybean diets on growth performance, intestinal digestion properties and energy values of 28-day-old broilers. A total of 336 one-day-old male AA broiler chickens were distributed to 7 groups in a completely random design. The birds were given 7 diets containing 6 diets with different combined soybean meals and a fasting treatment, 8 replicates per treatment and 6 birds per replicate (Trial 1). A total of 672 one-day-old male AA broiler chickens were randomly allocated to 7 dietary treatments including a control diet and 6 diets supplemented with 300 mg/kg α-galactosidase, 200 mg/kg ß-mannanase, and 300 mg/kg protease individually or in combination (Trial 2). Apparent metabolizable energy (AME) of broilers was measured from d 25 to 27 in both trial 1 and trial 2. The results showed that AME values of combined soybean meals averaged 2,894 kcal/kg. Dietary ß-mannanase and protease supplementation increased body weight gain (P < 0.05) during d 0 to 14, whereas did not affect the growth performance (P > 0.05) during d 14 to 28. Addition of ß-mannanase in combination with other enzymes significantly increased lipase and trypsin content (P < 0.05) in ileum. In addition, dietary ß-mannanase and protease supplementation individually or in combination enhanced trypsin enzyme content in jejunum (P < 0.05). The ß-mannanase enzyme enhanced villus height and villus height to crypt depth ratio (P < 0.05) of ileum compared with control diet. Moreover, supplementation of enzyme except for protease enhanced raffinose and stachyose degradation ratio (P < 0.05). Dietary ß-mannanase supplementation individually or in combination enhanced AME and AMEn values (P < 0.05). This study demonstrated that dietary enzyme supplementation especially ß-mannanase improved intestinal digestion properties and contributed to high energy values.

Microbiome and ileum transcriptome revealed the boosting effects of selenium yeast on egg production in aged laying hens.

The declines in laying performance during the late production period have adverse effects on the length of the production cycle. Improving the nutrition of laying hens is a crucial measure to reverse this declination. This study investigated the effect of selenium yeast (SY) on egg production, ileal gene expression and microbiota, as well as elucidating their associations in aged laying hens. A total of 375 Jinghong laying hens at 76 weeks old were randomly assigned into 5 dietary treatments, which included a selenium-deficient basal diet based on corn-soybean meal, and dietary supplementation of SY at 0.15, 0.30 and 0.45 mg/kg, and sodium selenite at 0.45 mg/kg. The results showed that SY ameliorated the depression in aged laying performance in the 0.30 mg/kg group (P < 0.01). Selenium yeast significantly increased ileum selenium concentration (P < 0.05), and SY groups had higher selenium deposition efficiency than the sodium selenite group. Functional enrichment and Short Time-series Expression Miner (STEM) analysis indicated that SY activated metabolic progress (e.g., glycerolipid metabolism, glycerophospholipid metabolism, and fatty acid metabolism), immune response and oxidative stress response. Four hub genes including thioredoxin reductase 1 (TXNRD1), dihydrolipoamide dehydrogenase (DLD), integrin linked kinase (ILK) and leucine zipper tumor suppressor 2 (LZTS2) were involved in intestinal metabolism which was closely associated with selenium deposition/status. Moreover, the relative abundance of Veillonella, Turicibacter and Lactobacillus was significantly increased, but the relative abundance of Stenotrophomonas was significantly decreased by SY supplementation. Multi-omics data integration and Canonical correspondence analysis (CCA) showed that both the ileum selenium content and the laying rate were highly correlated with pathways and bacteria enriched in metabolism and immune response. Meanwhile, the "switched on" gene prostate stem cell antigen (PSCA) had a positive relationship with Veillonella and a negative relationship with the opportunistic pathogens Stenotrophomonas. Overall, our study offered insight for the further exploration of the role of SY on boosting egg production and balancing ileum intestinal flora in aged laying hens.

Mycotoxins binder supplementation alleviates aflatoxin B1 toxic effects on the immune response and intestinal barrier function in broilers.

This experiment was conducted to evaluate whether a commercial mycotoxins-binder, XL, could effectively attenuate the negative effects of Aflatoxin B1 (AFB1) on growth performance, immunological function, and intestinal health in birds. Two hundred forty 1-day-old Arbor Acres broiler chickens were randomly divided into 4 treatments using a 2 × 2 factorial randomized design with 2 levels of dietary mycotoxins binder (0 or 2g /kg) and 2 AFB1 supplemented levels (0 or 200 µg/kg) from 0 to 42 d. Results showed that AFB1 exposure impaired growth performance by decreasing BWG in 1-21 d and 1-42 d, decreasing FI in 1-21 d, increasing FCR in 1-21 d and 1-42 d (P < 0.05). Broilers fed AFB1- contaminated diet impaired the immune function, as evident by decreasing IgA contents, Newcastle disease antibody titers in serum, and sIgA contents of jejunal mucosa at 21 d (P < 0.05). On the other hand, AFB1 challenge significantly increased the gene expression of proinflammatory factors in spleen at 21 d and liver at 42 d, and significantly decreased claudin-1 expression at 42 d and occludin expression at 21 d, and increased claudin-2 at 21 d in jejunum of broiler chickens (P < 0.05) compared to the basal diet group. Dietary XL supplementation significantly decreased the gene expression of IL-6 in spleen at 21 d and IL-1ß in liver at 42 d, cytochrome P450 3A4 (CYP3A4) expression in liver at 21 d of broilers (P < 0.05) compared with the nonsupplemented birds, regardless of AFB1 challenged or not. Inclusion of 2 g/kg XL increased serum ALB at 42 d, IgM and IgA at 42 d, Newcastle disease antibody titer level at 35 d (P < 0.05). Dietary XL addition enhanced intestinal barrier function by increasing the expression of claudin-1 at 21 d and Occludin at 42 d (P < 0.05) in jejunum. Conclusively, 2 g/kg mycotoxins-binder can relieve the toxic effect of AFB1 on broilers.

Hydroxychloride trace elements improved eggshell quality partly by modulating uterus histological structure and inflammatory cytokines expression in aged laying hens.

The objectives of this study were to investigate the effectiveness of dietary zinc, copper, and manganese hydroxychloride (HC) supplementation on performance, minerals deposition, serum parameters, eggshell ultrastructure, uterus histological structure, and inflammatory cytokines in aged hens. A total of 560 Hyline Brown layers at 62 wk of age were randomly allotted into 3 groups (CON, basal diet without extra minerals supplemented; Sulphate and HC, basal diet with sulphate or hydroxychloride zinc, copper, and manganese supplementation at levels of 80, 15, and 80 mg/kg, respectively). The trial lasted for 16 wk consisting of 4 wk depletion period and 12 wk testing period. The results indicated that dietary hydroxychloride trace elements increased egg weight (P < 0.05) when compared with CON group and improved average Haugh unit and albumen height (P < 0.05) when compared with Sulphate group from 70 to 73 wk. Trace element supplementation significantly increased eggshell strength, ceruloplasmin content in serum, and modified crystallographic structure of eggshell (P < 0.05) that included effective layer height, palisade height, mammillary layer width, and mammillary internal area ratio, but the results did not differ regarding the trace mineral sources used. Furthermore, hens fed with hydroxychloride trace element showed the highest mucosal fold height (P < 0.05) and epithelial height (P = 0.053) in eggshell gland, as well as mRNA expression of TNF-α (P < 0.05) and IL-22 (P = 0.094). It is concluded that supplementation of Zn, Cu, and Mn mixture modified eggshell quality partly through enhancing histological structure and immune responses of uterus. Hydroxychloride source of Zn, Cu, and Mn excelled sulphate in its beneficial effects for birds.

Enterococcus faecium PNC01 isolated from the intestinal mucosa of chicken as an alternative for antibiotics to reduce feed conversion rate in broiler chickens.

BACKGROUND: The development and utilization of probiotics had many environmental benefits for replacing antibiotics in animal production. Bacteria in the intestinal mucosa have better adhesion to the host intestinal epithelial cells compared to bacteria in the intestinal contents. In this study, lactic acid bacteria were isolated from the intestinal mucosa of broiler chickens and investigated as the substitution to antibiotic in broiler production. RESULTS: In addition to acid resistance, high temperature resistance, antimicrobial sensitivity tests, and intestinal epithelial cell adhesion, Enterococcus faecium PNC01 (E. faecium PNC01) was showed to be non-cytotoxic to epithelial cells. Draft genome sequence of E. faecium PNC01 predicted that it synthesized bacteriocin to perform probiotic functions and bacteriocin activity assay showed it inhibited Salmonella typhimurium from invading intestinal epithelial cells. Diet supplemented with E. faecium PNC01 increased the ileal villus height and crypt depth in broiler chickens, reduced the relative length of the cecum at day 21, and reduced the relative length of jejunum and ileum at day 42. Diet supplemented with E. faecium PNC01 increased the relative abundance of Firmicutes and Lactobacillus, decreased the relative abundance of Bacteroides in the cecal microbiota. CONCLUSION: E. faecium PNC01 replaced antibiotics to reduce the feed conversion rate. Furthermore, E. faecium PNC01 improved intestinal morphology and altered the composition of microbiota in the cecum to reduce feed conversion rate. Thus, it can be used as an alternative for antibiotics in broiler production to avoid the adverse impact of antibiotics by altering the gut microbiota.

Role of Vitamin K in Intestinal Health.

Intestinal diseases, such as inflammatory bowel diseases (IBDs) and colorectal cancer (CRC) generally characterized by clinical symptoms, including malabsorption, intestinal dysfunction, injury, and microbiome imbalance, as well as certain secondary intestinal disease complications, continue to be serious public health problems worldwide. The role of vitamin K (VK) on intestinal health has drawn growing interest in recent years. In addition to its role in blood coagulation and bone health, several investigations continue to explore the role of VK as an emerging novel biological compound with the potential function of improving intestinal health. This study aims to present a thorough review on the bacterial sources, intestinal absorption, uptake of VK, and VK deficiency in patients with intestinal diseases, with emphasis on the effect of VK supplementation on immunity, anti-inflammation, intestinal microbes and its metabolites, antioxidation, and coagulation, and promoting epithelial development. Besides, VK-dependent proteins (VKDPs) are another crucial mechanism for VK to exert a gastroprotection role for their functions of anti-inflammation, immunomodulation, and anti-tumorigenesis. In summary, published studies preliminarily show that VK presents a beneficial effect on intestinal health and may be used as a therapeutic drug to prevent/treat intestinal diseases, but the specific mechanism of VK in intestinal health has yet to be elucidated.

Research Note: Effects of organic zinc on broiler intestinal permeability and integrity in Clostridium perfringens-challenged condition.

This study evaluates the effects of a zinc-amino acid complex on broiler's intestinal permeability and integrity challenged with Clostridium perfringens. A total of 180 Arbor Acres 1-day-old male broilers were assigned to 6 treatments in a completely randomized 2 × 3 factorial design. The experiment investigated the comparative effects of inorganic and organic zinc supplements, that is (ZnSO4 treatment: 80 mg zinc/kg from ZnSO4; iso-dose replacement group. (ISO) treatment:40 mg zinc/kg from ZnSO4 plus 40 mg zinc/kg from a zinc-amino acid complex; and organic treatment: 40 mg zinc/kg from a zinc-amino acid complex) on C. perfringens-challenged broilers. C. perfringens, on the one hand, compromised intestinal mucosal barrier function by increasing the intestinal permeability of fluorescein isothiocyanate-labeled dextran (P < 0.05) and plasma endotoxin level, on the other hand, decreased both the transepithelial electrical resistance and the relative expression of occludin levels in the ileum at day 21 (P < 0.05). However, zinc supplement alleviates C. perfringens-induced pathologic changes in the intestinal permeability. ISO treatment, in particular, enhanced the villus height-to-crypt depth ratio and transepithelial electrical resistance and also reduced plasma endotoxin levels. In addition, zinc supplement relatively enhanced the expression of occludin levels in the ileum compared with the C. perfringens-challenged group. ISO treatment had the highest relative expression of occludin levels in the ileum. Thereby, the results indicated that partial replacement of ZnSO4 with a zinc-amino acid complex in the broiler diet could promote intestinal mucosal barrier function during C. perfringens infection via increased expression of occludin in the ileum.

Effects of dietary quercetin on the antioxidative status and cecal microbiota in broiler chickens fed with oxidized oil.

This study was conducted to evaluate the effects of quercetin on the antioxidant ability, intestinal barrier functions, and cecal microbiota in broiler chickens fed with oxidized soya oil. Four hundred eighty male Arbor Acres broilers were randomly assigned to 5 treatments, each involving 8 cages (12 birds per cage). The treatment groups were as follows: the control group, birds fed with basal diets containing oxidized oil, and birds fed with basal diets containing oxidized oil and supplemented with 200 ppm of quercetin, 400 ppm of quercetin, and 800 ppm of quercetin. The results showed that dietary supplementation with quercetin at a dose of 400 ppm or 800 ppm alleviated the increased serum malondialdehyde (MDA) level induced by oxidized oil on day 11 (P = 0.005) and reversed the increased MDA level in the mucosa on day 11 (P = 0.021). Quercetin significantly upregulated the transcription of nuclear factor erythroid 2-related factor 2 (Nrf2) and its downstream genes such as catalase (P < 0.001), superoxide dismutase 1 (P < 0.001), glutathione peroxidase 2 (P = 0.018), heme oxygenase-1 (HO-1) (P = 0.0), and thioredoxin (P = 0.002) and reversed the mRNA expression of HO-1 (P = 0.007) in the ileal mucosa. Tight junction protein 1 was only downregulated by oxidized oil (P = 0.013). In addition, quercetin (800 ppm) alleviated the decreased mRNA expression of mucin 2 (MUC2), which contributed to the intestinal chemical barrier (P = 0.039). The supplemental dose of 400 ppm of quercetin was able to promote Lactobacillus in the cecum, which enhanced the gastrointestinal tract health. In summary, these results indicated that quercetin ameliorated the oxidized oil-induced oxidative stress by upregulating the transcription of Nrf2 and its downstream genes to restore redox balance and reinforced the intestinal barrier via higher expression and secretion of MUC2 and facilitating the growth of Lactobacillus in the cecum. Therefore, quercetin could be a potential feed additive that can be applied in poultry production for amelioration of oxidative stress caused by oxidized oil and preventing the potential invasion of exogenous pathogens.

Dietary Quercetin Alleviated DSS-induced Colitis in Mice Through Several Possible Pathways by Transcriptome Analysis.

BACKGROUND: The prevalence of inflammatory bowel disease is rapidly increasing around the world. Quercetin is a flavonoid commonly found in vegetables and fruits and has been reported to exert numerous pharmacological activities such as enhancing antioxidant capacity or suppressing inflammation. OBJECTIVE: We aimed to explore whether quercetin was effective for IBD and the underlying mechanism of quercetin for the ameliorative effects on the DSS-induced colitis in mice. METHODS: Thirty-six mice were randomly assigned to three treatments, including the control group (Ctr), DSS-induced colitis group (DSS) and DSS-induced colitis supplemented with 500 ppm quercetin (DQ500). Colitis was induced by DSS intake, and body weight was recorded every day. After six days administration of DSS, intestinal permeability was measured, and the liver was taken for antioxidant enzyme tests. Colonic tissue was taken for the histopathlogical score and RNA-sequencing analysis. RESULTS: In this experiment, dietary quercetin for 500ppm alleviated the DSS-induced colitis, possibly by strengthening intestinal integrity, liver antioxidant capacity. Based on the results of the transcriptome of colon tissue, several key genes were modulated by quercetin. ERK1/2-FKBP pathway and RXR-STAT3 pathway were involved in the development of IBD, furthermore, in the down-regulation of S100a8/9, FBN2 contributed to lowering the risk of colongenesis. CONCLUSION: We demonstrated that dietary quercetin alleviated the DSS-induced colitis in mice. This is most likely due to its beneficial effects on intestinal integrity and modulation of several key pathways. Based on our research, quercetin was a promising candidate for IBD and its pharmaceutical effects on both IBD and colongenesis need further research.

Targeted Isolation of Antioxidant Constituents from Plantago asiatica L. and In Vitro Activity Assay.

Plantago asiatica L. is widely distributed in Eastern Asia and a commonly used drug in China, Korea, and Japan for diuretic and antiphlogistic purposes. In this experiment, the present study was performed to isolate antioxidant molecules based on the DPPH scavenging activity assay and discover the bioactive compounds which contributed to performing the function of Plantago asiatica L. Each faction was chosen for further isolation guided by DPPH scavenging activity assay. Afterwards, two potential bioactive molecules, aesculetin and apigenin, were isolated for in vitro antioxidant activity in cells. Hydrogen-peroxide-induced oxidative stress led to decreased cell viability, impaired intercellular junction, and damage to the cell membrane and DNA. Furthermore, aesculetin ameliorated decreased cell viability induced by hydrogen peroxide via upregulation of antioxidant related genes, and apigenin also protected against H2O2 mainly by improving the glutathione (GSH) antioxidant system, such as increasing the activity of glutathione peroxidase (GPX), glutathione reductase (GR), and the ration of GSH/glutathione disulfide (GSSG). Above all, these findings suggest that aesculetin and apigenin may be bioactive compounds for antioxidant function in Plantago asiatica L.

Effects of Dietary Zinc on Performance, Zinc Transporters Expression, and Immune Response of Aged Laying Hens.

This study was to investigate the effects of dietary zinc (Zn) supplementation on performance, zinc transporter gene expression, and immune function in aged laying hens. In experiment 1, twenty 31-week-old hens (young) and twenty 60-week-old hens (old) with the same genetic background were fed with the same diet for 4 weeks. In experiment 2, a basal diet supplemented with zinc sulfate (ZnS) and zinc glycine chelate (ZnG) at 30, 60, 90, and 120 mg Zn/kg to constitute nine experimental diets. Eight hundred and ten 60-week-old layers were distributed in a completely randomized experimental design with 9 treatments, 6 replicates of 15 birds each, and birds were fed for 10 weeks. In experiment 1, results showed that zinc and metallothionein (MT) concentration in the shell gland of old hens was significantly lower than young layers (P < 0.05). Zinc transporters ZnT1, 4, 5, 6, and 7 messenger RNA (mRNA) abundance in old layers were significantly lower versus the young (P < 0.05). In experiment 2, results indicated that dietary zinc supplementation did not significantly affect the laying rate, average feed intake, egg weight, feed conversion efficiency, broken egg rate, or mortality (P > 0.05). Supplemental ZnG significantly improved eggshell breaking strength than ZnS, with a higher alkaline phosphatase (ALP) activity and more abundant ZnT4 expression in shell gland versus ZnS (P < 0.05). ZnG supplementation at 90 mg Zn/kg affected the duodenal mucus by significantly increasing ZnT1, 6, 7, ZIP13, and MT-4 mRNA level (P < 0.05). Zinc level significantly increased bovine serum albumin (BSA) antibody concentration on 14 day after BSA injection (P < 0.05). Supplementation of ZnG improved eggshell quality of aged layers by upgrading zinc transporter expression in the shell gland and intestine also enhanced humoral immunity.

Dietary sodium butyrate improves intestinal development and function by modulating the microbial community in broilers.

This study investigated the effects of dietary sodium butyrate (SB) supplementation, provided as a specially coated product, on growth performance, intestinal development, morphological structure and function in broilers. In total, 720 one-day-old Arbor Acres male broilers were randomly allocated into six treatment groups with six replicates each and then fed basal diets (control) supplemented with 0, 200, 400, 800 or 1000 mg/kg of SB or with antibiotics (100 mg/kg aureomycin and 20 mg/kg colistin sulfate). The growth trial lasted for 42 days. No differences (P>0.05) in growth performance were detected between groups during the grower period (1-21 d) or over the total (1-42 d) trial period, whereas the addition of SB improved the intestinal structure by stimulating (P<0.05) goblet cells on jejunal and ileal villi accompanied by a trend towards increased (Pdiets<0.10) ileal villus height. In addition, more inerratic leaf-shaped villi and mucus secretion and significantly fewer erosions were demonstrated by scanning electron microscopy. Apart from decreased (P<0.05) malondialdehyde (MDA) in the ileal mucosa at 21 d of age, supplemental SB at higher doses (800 mg/kg) led to greater (P<0.05) total antioxidant capacity and depressed (P<0.05) MDA concentrations in the jejunal mucosa. Birds fed with 400 mg/kg and 800 mg/kg SB had higher (P<0.05) acetic acid concentrations at 42 d and higher butyric acid at 21 d in the jejunum chyme. Morever, chicks fed SB diet were found to have higher concentrations of butyric acid (P<0.05) in the ileal chyme. SB additions at 400 mg/kg displayed higher Firmicutes and Proteobacteria levels, while a higher (P<0.05) relative abundance of Bacteroidetes was observed at 800 mg/kg. Furthermore, we found a striking decrease in Enterobacteriaceae and increases in Lachnospiraceae and Rikenellaceae in the cecal lumen of birds fed 800 mg/kg SB as well as a higher proportion of Ruminococcaceae and a noticeable reduction (P<0.05) of Lactobacillaceae in birds treated with 400 mg/kg SB. Taken together, our results support the importance of SB in improving the intestinal development, morphological structure and biological functions of broilers through modulation of the microbial community, which seems to be optimized for gut health at higher doses (800 mg/kg) of SB.

Zinc Supplementation, via GPR39, Upregulates PKCζ to Protect Intestinal Barrier Integrity in Caco-2 Cells Challenged by Salmonella enterica Serovar Typhimurium.

Background: Zinc has been shown to improve intestinal barrier function against Salmonella enterica serovar Typhimurium (S. typhimurium) infection, but the mechanisms involved in this process remain undefined.Objective: We aimed to explore the roles of G protein-coupled receptor (GPR)39 and protein kinase Cζ (PKCζ) in the regulation by zinc of intestinal barrier function.Methods: A Transwell Caco-2 monolayer was pretreated with 0, 50, or 100 µM Zn and then incubated with S. typhimurium for 0-6 h. Afterward, cells silenced by the small interfering RNA for GPR39 or PKCζ were pretreated with 100 µM Zn and incubated with S. typhimurium for 3 h. Finally, transepithelial electrical resistance (TEER), permeability, tight junction (TJ) proteins, and signaling molecules GPR39 and PKCζ were measured.Results: Compared with controls, S. typhimurium decreased TEER by 62.3-96.2% at 4-6 h (P < 0.001), increased (P < 0.001) permeability at 6 h, and downregulated (P < 0.05) TJ protein zonula occludens (ZO)-1 and occludin by 104-123%, as well as Toll-like receptor 2 and PKCζ by 35.1% and 75.2%, respectively. Compared with S. typhimurium-challenged cells, 50 and 100 µM Zn improved TEER by 26.3-60.9% at 4-6 h (P < 0.001) and decreased (P < 0.001) permeability and bacterial invasion at 6 h. A total of 100 µM Zn increased ZO-1, occludin, GPR39, and PKCζ 0.72- to 1.34-fold (P < 0.05); however, 50 µM Zn did not affect ZO-1 or occludin (P > 0.1). Silencing GPR39 decreased (P < 0.05) zinc-activated PKCζ and blocked (P < 0.05) the promotion of zinc on epithelial integrity. Furthermore, silencing PKCζ counteracted the protective effect of zinc on epithelial integrity but did not inhibit GPR39 (P = 0.138).Conclusion: We demonstrated that zinc upregulates PKCζ by activating GPR39 to enhance the abundance of ZO-1, thereby improving epithelial integrity in S. typhimurium-infected Caco-2 cells.

Zinc enhances intestinal epithelial barrier function through the PI3K/AKT/mTOR signaling pathway in Caco-2 cells.

Zinc plays an important role in maintaining intestinal barrier function as well as modulating cellular signaling recognition and protein kinase activities. The phosphatidylinositol 3-kinase (PI3K) cascade has been demonstrated to affect intercellular integrity and tight junction (TJ) proteins. The current study investigated the hypothesis that zinc regulates intestinal intercellular junction integrity through the PI3K/protein kinase B (AKT)/mammalian target of rapamycin (mTOR) pathway. A transwell model of Caco-2 cell was incubated with 0, 50 and 100 µM of zinc at various time points. Transepithelial electrical resistance (TEER), paracellular permeability, TJ proteins, cell proliferation, differentiation and cell damage were measured. Compared with controls, 50 and 100 µM of zinc increased cell growth at 6, 12 and 24 h and the expression of proliferating cell nuclear antigen at 24 h. Zinc (100 µM) significantly elevated TEER at 6-24 h and reduced TJ permeability at 24 h, accompanied by the up-regulation of alkaline phosphatase (AP) activity and zonula occludens (ZO)-1 expression. In addition, zinc (100 µM) affected the PI3K/AKT/mTOR pathway by stimulating phosphorylation of AKT and the downstream target mTOR. Inhibition of PI3K signaling by LY294002 counteracted zinc promotion, as shown by a decrease in AP activity, TEER, the abundance of ZO-1 and phosphorylation of AKT and mTOR. Additionally, TJ permeability and the expression of caspase-3 and LC3II (markers of cell damage) were increased by addition of PI3K inhibitor. In conclusion, the activation of PI3K/AKT/mTOR signaling by zinc is involved in improving intestinal barrier function by enhancing cell differentiation and expression of TJ protein ZO-1.