Effects of dietary leucine supplementation on the gene expression of mammalian target of rapamycin signaling pathway and intestinal development of broilers.
ABSTRACT: This experiment was to investigate the effects of dietary leucine supplementation on the gene expression of mammalian target of rapamycin (mTOR) signaling pathway and intestinal development of broilers. A total of 384 one-day-old broilers were randomly assigned into 4 treatments with 6 replicates (16 broilers per replicate). Broilers in these treatment groups were offered the following diets with 1.37, 1.77, 2.17 and 2.57% of leucine. These diet treatments were named 1.37TM, 1.77TM, 2.17TM, and 2.57TM. The experiment lasted 21 days and all birds had free access to feed and water. Results indicated that there was no significant difference in body weight, average daily gain and average feed intake among all treatments (P > 0.05). The broiler duodenal villus height in 2.57TM was the lowest, but the highest occurred in 1.37TM on d 7 and 14 (P < 0.05). The villus height in the jejunum and ileum increased along with leucine level from 1.37 to 2.17%. The villus height of jejunum was significantly higher in 2.17TM than in 1.37TM on d 7 and 14, and the ratio of villus height to crypt depth (V:C) in the duodenum, jejunum and ileum increased significantly (P < 0.05) on d 21. The gene expression level of mTOR in the duodenum decreased with increasing leucine level and was higher in 1.37TM than in 2.57TM on d 7 and 14 (P < 0.05). On d 14 and 21 of the trial, the expression of S6K1 in the duodenum was higher in 1.37TM than in 2.57TM (P < 0.05), and the expression of mTOR, S6K1 in the jejunum and ileum increased with increasing leucine level form 1.37 to 2.17%, whereas a significant difference occurred between 1.37TM and 2.17TM (P < 0.05). In conclusion, the addition of leucine fails to enhance the growth performance of broilers. However, leucine can improve intestinal development by enhancing villus height and V:C ratio in the jejunum and ileum. Moreover, the expression of mTOR, S6K1 increased as the level of dietary leucine was elevated from 1.37 to 2.17%.
Project description:This study was conducted to evaluate the effect of dietary supplementation with Lactobacillus delbrueckii (LAB) on intestinal morphology, barrier function, immune response, and antioxidant capacity in weaned piglets challenged with lipopolysaccharide (LPS). A total of 36 two-line crossbred (Landrace?×?large Yorkshire) weaned piglets (28 days old) were divided into three groups: (1) nonchallenged control (CON); (2) LPS-challenged control (LPS); and (3) LAB+LPS treatment (0.2% LAB+LPS). Compared to the LPS piglets, the LAB+LPS piglets improved intestinal morphology, indicated by greater (P < 0.05) villus height in the duodenum and ileum; villus height?:?crypt depth ratio in the duodenum, jejunum, and ileum, as well as decreased (P < 0.05) crypt depth in the jejunum and ileum; and better intestinal barrier function, indicated by upregulated (P < 0.05) mRNA expression of tight junction proteins in the intestinal mucosa. Moreover, compared to the LPS piglets, LAB significantly decreased (P < 0.05) concentrations of TNF-? and IL-1? in the small intestine and increased (P < 0.05) IL-10 levels in the jejunum and ileum. Additionally, LAB increased (P < 0.05) T-AOC activities of the colon, GSH concentrations of the jejunum, and mRNA expression of CAT and Cu/Zn-SOD, while reduced (P < 0.05) MDA concentrations in the jejunum and ileum in LPS-changed piglets. Collectively, our results indicate that supplementation of LAB improved intestinal integrity and immune response and alleviated intestinal oxidative damage in LPS-challenged piglets.
Project description:This research investigated effects of dietary ?-sitosterol addition at different levels on serum lipid levels, immune function, oxidative status, and intestinal morphology in broilers. One-day-old broiler chicks were allocated to 5 groups of 6 replicates. Chickens in the 5 groups were fed a basal diet supplemented with 0 (control group), 40, 60, 80, and 100 mg/kg of ?-sitosterol for 42 D, respectively. ?-Sitosterol linearly decreased (P < 0.05) concentrations of serum total cholesterol, jejunal tumor necrosis factor ? (TNF-?), and ileal interleukin 1? (IL-1?) and mRNA relative expressions levels of jejunal TLR4 and ileal MyD88, whereas it linearly increased (P < 0.05) contents of jejunal immunoglobulin G (IgG), ileal secreted IgA and glutathione, jejunal catalase activity and Nrf2 mRNA relative expression level, villus height (VH), and VH-to-crypt depth (CD) ratio (VH:CD) in the jejunum and ileum. Linear and quadratic increases (P < 0.05) in absolute and relative spleen weight were observed by dietary ?-sitosterol, whereas malondialdehyde (MDA) concentration in the jejunum and ileum followed the opposite trend (P < 0.05). Compared with the control group, dietary ?-sitosterol at higher than or equal to 60 mg/kg level decreased (P < 0.05) contents of serum total cholesterol, ileal MDA, and jejunal TLR4 mRNA relative expression level, whereas it increased (P < 0.05) absolute spleen weight and ileal glutathione content. Higher than or equal to 80 mg/kg level of ?-sitosterol enhanced (P < 0.05) jejunal IgG concentration, VH, catalase activity, and Nrf2 relative expression level and ileal secreted IgA content, but reduced (P < 0.05) ileal IL-1? content and MyD88 mRNA relative expression level. ?-Sitosterol addition at 60 and 80 mg/kg levels increased (P < 0.05) relative spleen weight, whereas it decreased (P < 0.05) jejunal MDA accumulation. Moreover, 100 mg/kg level of ?-sitosterol reduced (P < 0.05) jejunal TNF-? level, but it increased (P < 0.05) VH in the jejunum and VH:CD in the jejunum and ileum. Accordingly, dietary ?-sitosterol supplementation could regulate serum cholesterol level, promote immune function, and improve intestinal oxidative status and morphology in broilers.
Project description:The present study determined the effects of in ovo feeding (IOF) of N-acetyl-L-glutamate (NAG) on early intestinal development and growth performance of broilers. A total of 702 fertile broiler eggs were randomly divided into 3 treatments: 1) non-punctured control group, 2) saline-injected control group, and 3) NAG solution-injected group (1.5 mg/egg). At 17.5 D of incubation, 300 ?L of each solution was injected into each egg of injected groups. Results indicated that the hatchability and healthy chicken rate were not affected by NAG injection (P > 0.05). Chicks from NAG solution-injected group had significantly decreased average daily feed intake and feed conversion ratio during 1-14 D than those in the non-punctured control group (P < 0.05). Compared with the non-punctured control group, IOF of NAG significantly increased the density of goblet cells in jejunum at hatch, duodenum at 7 D, and ileum at 14 D; decreased crypt depth in jejunum at hatch; and increased villus height in duodenum and jejunum and villus height:crypt depth ratio in duodenum at 7 D (P < 0.05). The intestinal mRNA expression of Na+-dependent neutral amino acid transporter, peptide transporter, and excitatory amino acid transporter 3 did not differ between groups at 7 or 14 D. However, the mRNA expression level of rBAT in jejunum significantly increased in the NAG solution-injected group than in the non-punctured control group at 7 D (P < 0.05). In conclusion, IOF of NAG (1.5 mg/egg) accelerated the early intestinal development by enhancing intestinal immune and absorption function, thereby positively affecting the feed efficiency for the first 2 wk post-hatch.
Project description:The objective of this study is to investigate the expression and distribution of heat shock protein 70 (Hsp70) in the intestine of intrauterine growth retardation (IUGR) piglets. Samples from the duodenum, prejejunum, distal jejunum, ileum, and colon of IUGR and normal-body-weight (NBW) piglets were collected at birth. The results indicated that the body and intestine weight of IUGR piglets were significantly lower than NBW piglets. The villus height and villus/crypt ratio in jejunum and ileum of IUGR piglets were significantly reduced compared to NBW piglets. These results indicated that IUGR causes abnormal gastrointestinal morphologies and gastrointestinal dysfunction. The mRNA of hsp70 was increased in prejejunum (P < 0.05), distal jejunum (P < 0.05), and colon in IUGR piglets. However, the hsp70 mRNA in ileum of piglets with IUGR was decreased. Similar to hsp70 mRNA, the protein levels of Hsp70 in prejejunum (P < 0.05), distal jejunum, and colon (P < 0.05) in IUGR piglets were higher than those in NBW piglets. These results indicated that the expression of Hsp70 in the intestinal piglets was upregulated by IUGR, and different intestinal sites had different responses to stress. Meanwhile, the localization of Hsp70 in the epithelial cells of the whole villi and intestinal gland rather than in the lamina propria and myenteron suggested that Hsp70 has a cytoprotective role in epithelial cell function and structure.
Project description:This study was conducted to explore whether exposure to bisphenol A (BPA) during pregnancy could change intestinal digestion and absorption function in offspring using pigs as a model, and whether methyl donor (MET) could counteract the BPA-induced impacts. Fifty Landrace × Yorkshire sows were divided into four dietary groups throughout gestation: control diet (CON); control diet supplemented with BPA (50 mg/kg); control diet supplemented with MET (3 g/kg betaine, 400 mg/kg choline, 150 ?g/kg vitamin B12, and 15 mg/kg folic acid); and control diet with BPA and MET supplementation (BPA + MET). Intestine samples were collected from pigs' offspring at birth and weaning. Maternal BPA exposure during pregnancy significantly reduced the ratio of jejunum villus height to crypt depth, decreased the jejunum sucrase activity, down-regulated the mRNA expression of jejunum peptide transporter 1 (Pept1) and DNA methyl transferase 3a (DNMT3a), and decreased the DNA methylation level of jejunum Pept1 in offspring (p < 0.05). Maternal MET supplementation significantly raised the ratio of villus height to crypt depth in jejunum and ileum, improved the jejunum lactase activity, up-regulated the mRNA expression of jejunum Pept1, lactase (LCT), DNMT1, DNMT3a, and methylenetetrahydrofolate reductase (MTHFR), and increased the DNA methylation level of jejunum Pept1 in offspring (p < 0.05). However, the ratio of jejunum villus height to crypt depth was higher in BPA + MET treatment compared with CON and BPA treatment (p < 0.05). Meanwhile, there was no difference in the jejunum sucrase activity, the mRNA expression of jejunum Pept1 and DNMT3a, and the DNA methylation level of jejunum Pept1 between CON and BPA + MET treatment. These results indicated that maternal exposure to BPA during gestation might suppress offspring's intestinal digestion and absorption function, whereas supplementation of MET could counteract these damages, which might be associated with DNA methylation.
Project description:Aflatoxin B1 (AFB1) has potent hepatotoxic, carcinogenic, genotoxic, immunotoxic and other adverse effects in human and animals. The aim of this study was to investigate the molecular mechanism of G2/M cell cycle arrest induced by AFB1 in the jejunum of broilers. Broilers, as experimental animals, were fed 0.6 mg/kg AFB1 diet for 3 weeks. Our results showed that AFB1 reduced the jejunal villus height, villus height/crypt ratio and caused G2/M cell cycle arrest. The G2/M cell cycle was accompanied by the increase of ataxia telangiectasia mutated (ATM), p53, Chk2, p21 protein and mRNA expression, and the decrease of Mdm2, cdc25C, cdc2, cyclin B and proliferating cell nuclear antigen protein and mRNA expression. In conclusion, AFB1 blocked G2/M cell cycle by ATM pathway in the jejunum of broilers.
Project description:Poultry coccidiosis is a costly intestinal disease that leads to considerable tissue damage, inefficient nutrient absorption, increased mortality, and predisposition to secondary infections. This study evaluated the effects of a direct feed microbial (DFM) dietary additive on performance, intestinal morphology, and immune response of broilers during a mixed coccidiosis challenge. In total, 840 Cobb500 male broilers were randomly allocated to 3 treatments (7 replicates, 40 birds/pen) including negative control (NC) fed basal diet; positive control (PC) fed basal diet with coccidiosis challenge; and DFM supplemented diet, with coccidiosis challenge. At 15 days of age, all birds except for the NC treatment were orally gavaged with live oocysts of a commercial vaccine. On d 21 (6 days post challenge), 4 birds/pen were randomly selected and euthanized for scoring of coccidia-caused lesions in the duodenum, jejunum, and ceca. Body weight gain (BWG), feed intake (FI), and feed conversion ratio (FCR) were recorded on d 7, 14, 28, and 42. Jejunal and ileal tissue samples were taken for histomorphological assessment from 2 birds/pen on d 21. Ileal samples were also taken for mRNA expression analysis on d 14 and d 21. The DFM birds had significantly greater BWG than PC birds during d 0-21 (P < 0.05). No differences were observed among the treatment groups in terms of FI and FCR. Dietary DFM supplementation significantly reduced lesion scores in the duodenum and jejunum when compared with PC group (P < 0.05). The coccidia challenge significantly reduced (P < 0.05) ileal villus height when compared to the non-challenged group on d 21. Conversely, dietary DFM supplementation alleviated the negative effects of coccidiosis by increasing ileal villus area on d 21 (P < 0.05). The challenged birds had significantly greater expression of IFN-? and IL-1? in the ileum on d 21. Based on these findings, dietary DFM supplementation may help restore broiler performance during the starter and early grower periods during coccidiosis, likely by maintaining gut integrity via improving intestinal morphology and also by reducing disease severity as manifested by lower lesion scores.
Project description:A total of 75 pigs were used to investigate effects of feeding Two Macrocephala Flavored Powder (TMFP) on small intestinal morphology, intestinal microbiota in weaning pigs. The dietary treatments were: a control diet; control diet + 3 g/kg TMFP; control diet + 0.3 g/kg colistin sulfate (ANT). The results showed that supplementation with TMFP increased (P < 0.05) villus height at duodenum, jejunum at 3 time points, increased (P < 0.05) crypt depth at duodenum, jejunum at day 14, improved villus height: crypt depth ratio (P < 0.05) in jejunum at day 21 as compared with ANT. Supplementation of TMFP and ANT had lower (P < 0.05) E. coli counts in the ileum, cecum and colon at day 7 as compared with control. Supplementation of TMFP had higher (P < 0.05) bifidobacteria counts in the ileum, cecum and colon compared with ANT, except for colon at day 21. No effect (P > 0.05) on lactobacilli in colon has been seen with supplementation of TMFP and ANT at 3 time points, while both of supplementations showed increased the number of lactobacilli in cecum at day 14 and day 21. Analysis of DGGE fingerprints indicated that a highest similarity was observed for profiles from samples taken 14 d, 21 d from TMFP. The diversity of DGGE fingerprints of TMFP was higher than those of ANT and control. The results suggest that TMFP is potential to enhancing intestinal morphology and microbiota of weaning pigs, and can be served as an effective and safe dietary additive for weaning pigs.
Project description:The effects of in ovo injection of raffinose (RFO) as a prebiotic on growth performance, relative weight of proventriculus, gizzard, drumstick and breast muscles, and ileum mucosa morphology were examined in Cobb 500 broilers. A total of 240 fertilized eggs were divided into 4 groups: a non-injected with intact shell and 3 levels of RFO solution (1.5, 3.0, and 4.5 mg in 0.2 mL of an aqueous diluents). The RFO solution was injected into the air sac on d 12 of incubation. In total 144 birds were fed a standard diet and management and sacrificed at d 21 post hatch for collection of samples. Total RNA was extracted from the small intestine, and RT-qPCR was performed to quantify mRNA levels of marker genes of immune cells. Injection of RFO had no significant effect (P > 0.05) on d one body weight of chicks. On d 21, the relative weight of the proventriculus, drumstick, breast, and gizzard was not affected (P > 0.05) by RFO. On hatch d, the villus height increased linearly (P < 0.01) with an increasing dose of RFO. Also, an increasing dose of RFO increased the villus height and villus height:crypt depth ratio (P < 0.05) but did not affect the crypt depth on d 21. The expression levels of CD3 and chB6, which are T cell and B cell marker genes, respectively, were significantly enhanced by high dose RFO (4.5 mg). In conclusion, although an increasing dose of RFO in ovo injection did not significantly influence growth performance or slaughter yield of broilers, RFO has the potential of enhancing ileum mucosa morphology and improving immunity in the small intestine, which are indicators of improved gut health.
Project description:Mycotoxin exposure is common in the poultry industry. Deoxynivalenol (DON) is usually detected at levels below the maximum threshold (5000 ppb), but depending on diet and age, broiler performance can be affected. We evaluated the effects of 900 ppb and 2300 ppb DON on the performance, intestinal morphometry, and lesion scores of broiler chickens. One-day-old male Ross broilers (<i>n</i> = 736) were divided into 4 treatments with 8 replicates each, and a pen containing 23 birds was the experimental unit. The animals were fed diets naturally contaminated with two levels of DON: 900 (Low DON-LD) or 2300 (Moderate DON-MD) ppb, with or without activated charcoal, over 28 days. After this, all birds were fed a marginally DON-contaminated diet without charcoal. During the first 28 days, body weight gain (BWG) and feed conversion ratio (FCR) were significantly impaired when broilers were fed a MD diet without activated charcoal. Even after feeding a marginally contaminated diet from D28-35, birds previously fed the MD diet presented a significantly lower performance. The villus height:crypt depth (VH:CD) ratio was significantly higher in the ileum from 14-day-old broilers fed the MD when compared with the LD diet. At D28, the MD diet caused decreased villus height (VH) and increased crypt depth (CD), affecting VH:CD ratio in both intestinal segments, with higher levels in the jejunum from 28-day-old broilers fed a non-supplemented LD diet. Broiler production was negatively affected by DON, even at moderate levels (2300 ppb).