| During the last phase of incubation,dramatic physiological and metabolic changes occur in chick embryos,and the supplies of the nutrients and energy are always insufficient.It is reported that as a regulating technology of early nutrition supply,in ovo feeding(IOF)of exogenous nutrients for the chick embryo could improve the early development of intestine,the hatching status and subsequent the growth performance of broiler chickens.The objective of this study was to investigate the effects of IOF of L-Arginine(Arg)solution into the amniotic fluid of Arbor Acres(AA)broiler eggs at 17.5 d of incubation on the early development of digestive organs,the mucosa morphological structure,immune barrier function and digestive-absorptive function in small intestine and subsequent on the growth performance of broilers,and further elucidate the possible mechanism.1.Effects of in ovo feeding of L-Arginine on the early development of digestive organs and the growth performance of posthatch broilersThis study was to preliminarily investigate the effects of IOF of Arg solution into the amniotic fluid of broiler eggs at 17.5 d of incubation on the early development of digestive organs and the growth performance of broilers,and then selected the appropriate IOF concentration of Arg(Experiment I).The 1600 embryonated eggs with similar weight were randomly allocated to 5 groups,consisting of 8 replicates of 40 eggs each.The 5 treatments were arranged as non-injected control(NC),diluent-injected(0.75%NaCl solution)group(DG),and Arg solution injected groups contained 0.5,1.0,and 2.0%Arg,all dissolved in diluent.At 17.5 d of incubation,0.6 mL of IOF solution was injected to amniotic fluid of each egg of injected groups.The chicks hatched from each treatment were sexed,and male chicks were randomly divided into 8 replicate cages with 15 broilers/replicate.The chicks were raised until 21 d of age.Results showed that,with the increasing IOF concentration of Arg from 0%to 2%,the hatchability of 2%Arg group was lower(linear,P = 0.025)than the other groups;the body weight gain(BWG;linear,P = 0.025)and the feed/gain ratio(F/G;linear,P = 0.007)during 1-7 d,the feed intake(FI;linear,P = 0.004;quadratic,P =0.001)and the BWG(linear,P = 0.038;quadratic,P = 0.007)during 8-21 d,the FI(linear,P = 0.005;quadratic,P = 0.001)and BWG(linear,P = 0.010;quadratic,P = 0.004)during 1-21 d all increased,the greatest values were both observed in the 1%Arg group.The absolute weights of yolk sac(linear,P = 0.043)and duodenum(linear,P = 0.036;quadratic,P = 0.020)of 1-d-old broilers;the absolute weights of yolk sac(linear,P = 0.026;quadratic,P = 0.009),proventriculus(linear,P = 0.013),gizzard(linear,P = 0.010;quadratic,P =0.037)and small intestine(linear,P<0.050)of 3-d-old broilers;the absolute weights of liver(linear,P = 0.013),pancreas(linear,P = 0.015;quadratic,P = 0.049),proventriculus(linear,P = 0.009),gizzard(linear,P = 0.013)and small intestine(linear,P<0.050)of 7-d-old broilers;the absolute weights of liver(linear,P = 0.045),proventriculus(linear,P =0.026;quadratic,P = 0.043)and duodenum(linear,P = 0.013;quadratic,P = 0.024)of 21-d-old broilers all increased with the increasing IOF concentrations of Arg from 0%to 2%,the greatest values were both observed in the 1%Arg group.Therefore,the 1%Arg group obtained the best results and should be chosen as the appropriate IOF concentration of Arg.2.Effects of in ovo feeding of L-Arginine on the gastrointestinal hormone,the jejunal digestive and absorptive capacity of posthatch broilersThis study was to investigate the effects of IOF of 1%Arg solution into the amniotic fluid at 17.5 d of incubation on the release of gastrointestinal hormones,the digestive enzyme activities,the concentrations of plasma free amino acids,the mRNA expressions of sensing receptors and nutrient transporters of post-hatch broilers.This experiment design was the same with the Experiment I.Results showed that,compared with NC and DG groups,the IOF of 1%Arg solution increased(P<0.05)the concentrations of ghrelin in duodenum of 7-d-old broilers,in jejunum and ileum of 21-d-old broilers;the concentrations of ghcagons-like peptide 2(GLP-2)in duodenum and ileum of 7-d-old broilers,in jejunum and ileum of 21-d-old broilers;the concentrations of vasoactive intestinal peptide(VIP)in duodenum of 7-d-old broilers.The IOF of Arg solution increased(P<0.05)the activities of trypsin and lipase in duodenum,lipase in jejunum,amylase and lipase in pancreas of 7-d-old broilers;the activities of lipase in duodenum,the amylase,lipase and trypsin in jejunum,amylase in ileum,trypsin in gizzard of 21-d-old broilers;the activities of alkaline phosphatase(AKP),maltase,sucrase in small intestine of 7 and 21-d-old broilers.The IOF of Arg also increased(P<0.05)the mRNA expressions of sensing receptors of taste receptor type 1 member 1 and 3,calcium-sensing receptor(CaSR),G protein-coupled receptor,class C,group 6,subtype A(GPRC6A)in duodenum and jejunum;the CaSR and GPRC6A in ileum;the mRNA expressions of the nutrient transporters of SLC7A4?SLC7A6?SLC3A1?SGLT-1?GLUT-2?GLUT-5?FABP-1 in duodenum,the SLC7A4?SLC7A6?SLC7A7?SLC3A1?SLC6A19?SLC1A1?SGLT-1?GLUT-2?GLUT-5?FABP-1 in jejunum,the SLC7A1?SLC7A7?SLC6A19?SLC1A1?SGLT-1?GLUT-2?GLUT-5?FABP-1 in ileum;and the concentrations of plasma amino acids of Val,Met,Ile,Leu,Arg,and Pro.3.Effects of in ovo feeding of L-Arginine on the development of digestive organs and growth performance of chick embryos and posthatch broilersThe present study was to investigate the effects of IOF of 1.0%Arg solution into the amniotic fluid of broiler eggs at 17.5 d of incubation on the development of digestive organs and the growth performance of chick embryos at 19 d of incubation and post-hatch broilers(Experiment II).The 960 embryonated eggs with similar weight were randomly allocated to 3 groups,consisting of 8 replicates of 40 eggs each.The 3 treatments were arranged as non-injected control(NC),diluent-injected(0.75%NaCl solution)group(DG),and 1.0%Arg solution injected groups(10 g Arg dissolved in 1 L diluent).At 17.5 d of incubation,0.6 mL of IOF solution was injected to amniotic fluid of each egg of injected groups.The chicks hatched from each treatment were sexed,and male chicks were randomly divided into 8 replicate cages with 15 broilers/replicate.The chicks were raised until42 d of age.Results showed that,there was no significant difference in the hatchability,hatching weight,and the FI and BWG during 22-42 d among all groups(P>0.05).Compared with NC and DG groups,the IOF of 1%Arg solution increased(P<0.05)the weight of chick embryos at 19 d of incubation,the BWG during 1-7 d,the FI and BWG during 8-21 d?1-21 d?1-42 d.The IOF of Arg solution increased(P<0.05)the absolute weights of yolk sac of chick embryos at 19 d of incubation and 1-d-old broilers;the absolute weights of liver,pancreas,proventriculus,gizzard,small intestine and colorectum of 7-d-old broilers;the absolute weights of liver,pancreas,small intestine,cecum and colorectum of 21-d-old broilers;the absolute lengths of duodenum,jejunum and colorectum of 7-d-old broilers;the absolute lengths of duodenum and cecum of 21-d-old broilers.4.Effects of in ovo feeding of L-Arginine on the enterocyte proliferation,morphological structure and mucosal barriers in small intestine posthatch broilersThis study was to investigate the effects of IOF of 1%Arg solution into the amniotic fluid at 17.5 d of incubation on the enterocyte proliferation,morphological structure and mucosal barriers in small intestine of chick embryos at 19 d of incubation and post-hatch broilers.This experiment design was the same with the Experiment II.Results showed that,compared with NC and DG groups,the IOF of 1%Arg solution increased(P<0.05)the villus height(VH)and the ratio of VH to crypt depth(VH/CD)in duodenum,the VH in jejunum,the VH and VH/CD in ileum of chick embryos at 19 d of incubation;the VH,VH/CD and the density of goblet cells in small intestine of 1,7 and 21-d-old broilers;the VH and VH/CD in small intestine of 42-d-old broilers;the percentage of proliferating cell nuclear antigen(PCNA)positive cells of the villus in jejunum of 21-d-old broilers;and decreased(P<0.05)the CD in ileum of chick embryos at 19 d of incubation;the CD in small intestine of 1 and 21-d-old broilers;the CD in duodenum and jejunum of 7-d-old broilers;the CD in jejunum of 42-d-old broilers.The IOF of Arg increased(P<0.05)the mRNA expressions of mucin-1,claudin-1,claudin-3,zonula occludens-2(ZO-2)in duodenum;the mucin-2,claudin-1,ZO-1 and ZO-2 in jejunum;the mucin-2,claudin-3,occludin and ZO-2 in ileum;as well as the protein abundance of phosphor-mTOR,phosphor-S6K1 and phosphor-4E-BP1 in jejunum of 21-d-old broilers.5.Effects of in ovo feeding of L-Arginine on the development of lymphoid organs and small intestinal immune barrier function in posthatch broilersThis study was to investigate the effects of IOF of 1%Arg solution into the amniotic fluid at 17.5 d of incubation on the development of lymphoid organs,the nitric oxide synthase(NOS)activity and nitric oxide(NO)production,immunological cytokines and toll-like receptors mRNA expression in small intestine as well as the inducible NOS(iNOS)gene promoter methylation status and expression level in jejunum of broilers.This experiment design was the same with the Experiment II.Results showed that,compared with NC and DG groups,the IOF of 1%Arg solution increased(P<0.05)the absolute weights of thymus,bursa of Fabricius and spleen as well as the relative weight of thymus;the contents of secretory immunoglobulin A(slgA),IL-2 and IL-4 in small intestinal mucosa and immunoglobulin A(IgA),IL-2 and IL-4 in serum;the relative mRNA expressions of toll-like receptor-2(TLR-2)and toll-like receptor-4(TLR-4)in jejunum;the mRNA expressions of TLR-4 in duodenum and ileum of 21-d-old broilers.The IOF of Arg increased(P<0.05)the activities of iNOS in duodenum and ileum,the activities of total NOS(TNOS)and iNOS in jejunum of 7-d-old broilers;the activities of TNOS and iNOS in duodenum and jejunum,the activities of iNOS in ileum;the iNOS activity and NO concentration in serum of 21-d-old broilers.Meanwhile,the IOF of Arg increased(P<0.05)the mRNA expression level and protein abundance of iNOS,and decreased(P<0.05)the iNOS gene promoter methylation percentage in jejunum of 21-d-old broilers.As stated above,the conclusions are as follow:(1)With the increasing IOF concentration of Arg from 0%to 2%,the BW,BWG,FI and the early growth development of digestive organs of post-hatch broilers increased firstly and then decreased,and the hatchability of 2%Arg group significantly decreased,however,IOF of 1%Arg solution obtained the best results in the growth performance of broilers without affecting the hatchability.Therefore,the 1%Arg was chosen as the appropriate IOF concentration of Arg.(2)The IOF of 1%Arg solution into the amniotic fluid facilitated affecting the release of gastrointestinal hormones,increasing the activities of digestive enzymes and brush border enzymes,the mRNA expressions of sensing receptors and nutrient transporters in small intestine,and then improving the digestive-absorptive capacity of nutrients as well as the plasma free amino acid levels,and finally improving the development of digestive organs and the growth performance of 21-d-old broilers.(3)The IOF of 1%Arg solution facilitated saving the yolk and supplying the nutrient/energy for the chick embryos,improving the development of digestive organs,the enterocyte proliferation,morphological structure and mucosal barriers in small intestine as well as the finally growth performance of chick embryos at 19 d of incubation and post-hatch broilers,which might be associated with the improvement of digestive-absorptive capacity and the activation of mTOR pathway in small intestine.In addition,the improved intestinal function might explain the improvement in feed intake and consequently on the growth performance of broilers.However,the IOF of Arg might be limited and mainly promoted the growth performance of early stage chicks,and these significantly positive effects could persist during 1-21 d.From 22 to 42 d,these positive effects were attenuated.(4)The IOF of 1%Arg solution could facilitate improving the immunological barrier functions in intestinal mucosa and the intestinal health as well as the overall immunity of 21-d-old broilers by suppressing the iNOS gene methylation and activating the Arg-NO signaling pathway.Therefore,IOF of appropriate Arg solution could be as an effective technology for regulating early nutrition supply in broliers,improving the intestinal development and subsequent the finally growth performance in poultry industry. |
PDF Full Text Request