| This study aimed to clarify quantitative distributions of free amino acids(FAA)and small peptides(PAA)in gastrointestinal tract of sheep and the mRNA expression of the small peptide transporter PEPT1(SLC15A1),the basic amino acid transporter CAT1(SLC7A1),the acidic amino acid transporter EAAT3(SLC1A3),the neutral amino acid transporters y+LAT2(SLC7A6)and ASCT2(SLC1A5),aminopeptidase APN and dipeptidase DPEP2 in gastrointestinal tract of sheep.This study provides the basic data for protein digestive absorption and the mechanism of amino acid transports in gastrointestinal tract of sheep,and then provide the theoretical basis for feed manufacturing.A total of 6 healthy Han sheeps(non pregnant ewes)at the age of 18 months were butchered to collect chyme and tissues of rumen,duodenum,jejunum,ileum and cecum.Concentrations of FAA and PAA were gauged in chyme supernatant of rumen,duodenum,jejunum,ileum and cecum by LC-MS method to analysis the quantitative distribution of FAA and PAA in gastrointestinal tract.Primers were designed according to the sequences of PEPT1?CAT1?EAAT3?y+LAT2?ASCT2?APN and DPEP2 cDNA of sheep in GenBank.These genes' mRNA were quantified by RT-PCR used double standard curve method to analysis their expression pattern in gastrointestinal tract.The relative abundance of protein of PEPT1 and CAT1 in duodenum,jejunum and ileum were quantified by Western blot.Main results showed that as follows:1.Besides cysteine,concentrations of all PAA in chyme became lower and lower from the duodenum,jejunum,ileum,cecum to rumen.They were significant different(P<0.05).Proportions of PAA in these parts were different.The content of peptide binding lysine was highter in jejunum but lower in ileum;The content of peptide binding glutamate was highter in almost all parts;The content of peptide binding cysteine was lower in almost all parts.Most of concentrations of FAA in chyme became lower and lower from the jejunum,duodenum,ileum,rumen to cecum.Jejunum and duodenum were significant higher than others(p<0.05),while duodenal concentrations of some neutral amino acid such as arginine,tyrosine,tryptophan,methionine,glycine and phenylalanine were higher than that of ileum,and two of them,tyrosine and tryptophan,were significant different(P<0.05).Proportions of FAA in these parts were different.Aspartic acid had lower content in ileum but higher content in cecum;arginine had lower content in rumen but higher content in duodenum;Content of glutamate and lysine were highter in almost all parts;Content of cysteine were lower in all parts except cecum.The proportion of PAA in total amino acids was the significantly lowest in jejunum(P<0.05),and the hightest in cecum(P<0.05).2.CAT1 mRNA in jejunum had the highest abundance,but there were no significant differences in 5 parts.EAAT3,y+LAT2,PEPT1 and APN mRNA had a similar expression pattern that they all had the highest abundance in ileum,followed by jejunum and duodenum,and had the lowest abundance in rumen and cecum.ASCT2 and DPEP2 mRNA had a similar expression pattern that they all had the highest abundance in duodenum and cecum,and had the lowest abundance ileum.3.CAT1 protein had the highest abundance in duodenum,followed by jejunum and ileum,but there were no significant differences in 3 parts(P>0.05).PEPT1 protein had the highest abundance in ileum,followed by duodenum and jejunum,but there were no significant difference in 3 parts(P>0.05).Results indicated that concentrations and proportions of FAA in these chyme of five parts were different,the same as PAA.The major production site of PAA is duodenum.The major digestion site of PAA is jejunum.The major absorption site of PAA in non mesenteric system is ileum.The major production site of FAA is jejunum.The major absorption site of FAA is ileum.But some of the neutral amino acids and basic amino acids are absorbed major in jejunum. |
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