Ontogenesis Of Cationic Amino Acid Transporter MRNA Expression In Chicken Intestine And The Nutritional Regulation

In order to investigate the ontogenetic change of cationic amino acid transporter mRNA expression in chicken intestine, the difference between Lingnan-yellow chicken and Arbor Acre (AA) chicken and the effect of nutritional factor on the gene expression of cationic amino acid transporters in the chicken intestine, 2 animal trials were designed to compare the growth performance and amino acid digestibility between Lingnan-yellow chicken and AA chicken, the tissue specific and ontogenetic change of cationic amino acid transporter mRNA expression in chicken intestine were studied and the effect of xylanase on the animo acid digestibility and cationic amino acid transporter mRNA expression in the intestine of Lingnan-yellow chicken fed wheat based diet was studied.In trial 1, 160 healthy 0-day-old male parent AA broiler chicks and 160 healthy 0-day-old male parent Lingnan-yellow chicks were divided randomly into 4 replicates respectively. They were fed under the same condition. The average daily feed intake (ADFI) and average daily weight gain (ADG) of every week were recorded. On 16 and 30d, two birds (closest to the mean pen weight) selected from each replicate pen were slaughtered to collected serum of mesentery vein and chyme in ileum for the determination of amino acids concentration in serum and amino acid apparent digestibilities in ileum chyme. The samples of duodenum, jejunum, ileum and colorectum of Lingnan-yellow chickens were collected on 30d for the determination of relative abundance of cationic amino acid transporters rBAT,y+LAT2,CAT1 and CAT4 mRNA expression by reverse transcription PCR(RT-PCR); On 2d, 16d, 30d, 44d and 58d, the samples of duodenum, jejunum and ileum were collected to determination the ontogenetic change of rBAT,y+LAT2,CAT1 and CAT4 mRNA expression in different intestinal segments by RT-PCR. The results were shown as follows:(1) From 1 to 8 w, the ADFI and ADG of AA chicken were significantly higher than that of Lingnan-yellow chicken (P<0.01) and the feed/gain (F/G) of AA chicken was significantly lower than that of Lingnan-yellow chicken(P<0.05); The every week ADFI curves of AA chicken and Lingnan-yellow chicken increased linearly from 1 to 4 w (before changing the phase 1 diet to phase 2 diet) , showed a platform phase from 5 to 6 w, and went on increasing from 7 to 8 w. The change of diet influenced the ADFI; The ADG curve of AA chicken had the same trend with that of Lingnan-yellow chicken in the prophase. From 1 to 3w, ADG increased linearly and the ADG value of AA chicken was 2.21 times of that of Lingnan-yellow chicken. From 4 to 8 w, the ADG increased slowly, and the value of AA chicken was 1.75 times of that of Lingnan-yellow chicken; At the end of trail, the ADFI , ADG and F/G of AA chicken was 1.48 times, 1.75 times and 0.844 times of that of Lingnan-yellow chicken, respectively.(2) The values of ileum amino acid apparent digestibilities of AA chicken were higher than that of Lingnan-yellow chicken( Met and Cys excepted ) on 16d and 30d(P>0.05), and the ileum apparent digestibility of total amino acids of AA chicken was higher than that of Lingnan-yellow chicken by 4.17%(P=0.09) and 4.63%(P>0.05) respectively; The total amino acids in the mesentery vein serum of AA chicken was higher than that of Lingnan-yellow chicken on 16d (P<0.05) while it was reverse on 44d (P<0.05). There was no significant difference between AA chicken and Lingnan-yellow chicken on 30d, 58d. From 16d to 58d, the total amino acids in the mesentery vein serum of the chicken was decreased gradually, and the difference was significant between the different time points.(3) Expression of rBAT mRNA had the same tissue specificity with that of y+LAT2 mRNA in Lingnan-yellow chicken intestine. The abundance of rBAT and y+LAT2 mRNA in colorectum were significant lower than that in duodenum, jejunum and ileum(P<0.01), respectively. which in ileum were higher than that in duodenum and jejunum(P>0.05). CAT1 mRNA expression had the similar tissue specificity with CAT4, but they were distinct from rBAT and y+LAT2. The abundance of CAT4 and CAT1 mRNA in colorectum were significant higher than that in duodenum, jejunum and ileum(P<0.01). The expression abundance of CAT1 mRNA in ileum was significant higher than that in duodenum and jejunum(P<0.01), respectively. The abundance of CAT4 mRNA decreased gradually from duodenum, jejunum to ileum; It indicated that the cationic amino acid transporter system b0,+ located in the apical membrane and system y+L located in the basolateral membrane of epithelial cell would be the main transporter systems for cationic amino acids absorption in intestine of chicken.(4) Ontogenesis of rBAT,y+LAT2,CAT1,CAT4 mRNA expression in chicken duodenum were closed to that in jejunum, and was different with that in ileum, respectively. The ontogenesis of rBAT mRNA expression was similar with that of y+LAT2 in duodenum and jejunum and which accorded with the curve of ADG and ADFI. The relative expression abundance of rBAT and y+LAT2 mRNA increased from 2 to 30d, descended on 44d and rebound on 58d. The decrease on 44 and 58d maybe come from the stress of feed change and the decrease of protein level in the feed. The change range of rBAT and y+LAT2 mRNA expression abundance of Lingnan-yellow chicken were smaller than that of AA chicken. The rBAT and y+LAT2 mRNA expression abundance of AA chicken were higher or significantly higher than that of Lingnan-yellow chicken; In duodenum and jejunum, the ontogenesis of CAT1, CAT4 mRNA expression of Lingnan-yellow chicken were different with that of AA chicken. The expression abundance of CAT1 mRNA decreased from 2 to 58d in yellow-feather chicken, while it only descended on 44d and kept stable on the other time points in AA chicken. The expression abundance of CAT1 mRNA decreased from 2 to 44d and rebound on 58d in Lingnan-yellow chicken while it reached the lowest point on 58d and kept the similar level on other time points in AA chicken; Ontogenesis of rBAT,y+LAT2,CAT1,CAT4 mRNA expression in ileum of Lingnan-yellow chicken were different with that of AA chicken, respectively. The abundance of rBAT,y+LAT2 mRNA on 30d, 44d and 58d were higher than that on 2d and 16d in Lingnan-yellow chicken, and they were significantly higher on 58d than that on the other time points and reached the lowest point on 16d in AA chicken. In Lingnan-yellow chicken ileum, the level of CAT1 mRNA descended from 2 to 44d and the level on 58d was closed to that on 44d, while in AA chicken ileum, the level of CAT1 mRNA reached the tiptop on 58d and on the other time points the CAT1 mRNA level was nearly same. The ontogenesis of CAT4 mRNA expression in ileum of Lingnan-yellow chicken was similar with that of AA chicken, while on 58d the level of AA chicken went on descending and the level of Lingnan-yellow increased significantly.In trial 2, 720 healthy 0-day-old male parent yellow–feathered chicks were divided randomly into 3 groups signed with A, B, C, and 6 replicates per group. Group A fed the wheat-based diet, Group B fed the wheat-based diet with 0.1% xylanase (the crude enzyme activity≥12000U/g), Group C fed the corn-soybean diet. The nutrients levels were same among the 3 groups. The ADFI and ADG of every week were recorded. On 16 and 30d, chyme in ileum was collected and serum of mesentery vein was collected on 16, 30 and 44d for the determination of amino acids concentration in serum and amino acid apparent digestibilities in ileum chyme. The samples of jejunum and ileum of A and B groups were collected on 16d for the determination of relative abundance of cationic amino acid transporters rBAT,y+LAT2,CAT1 and CAT4 mRNA expression by RT-PCR.The results were shown as follows:(1) Xylanase increased the body weight of Lingnan-yellow chicken fed the wheat-based diet by 4.4%(P<0.05), increased the ADG by 4.7%(P<0.05) and decreased the F/G by 3.2%(P<0.05) at the end of the trial. There was no effect on the feed intake. The productive performance of Lingnan-yellow chicken fed the diet added with xylanase was nearly same with the that of chicken fed the corn-based diet.(2) Supplementation of xylanase in the wheat-based diet increased the apparent digestibilities of amino acids in ileum chyme, and the apparent digestibility of total amino acids increased by 3.5%(P>0.05) on 16d and 3.0%(P>0.05) on 30d. There was no difference of the ileum apparent amino acid digestibilities between wheat-based diet with xylanase and corn-based diet.(3) Supplementation of xylanase in the wheat-based diet increased the concentrations of most amino acids in the mesentery vein serum of chicken on 16d, and the Lys, His, total amino acids reached the significant level (P<0.05). It had no effect on the concentration of amino acids in the mesentery vein serum on 30and 44d.(4) Supplementation of xylanase in the wheat-based diet significantly increased the expression abundance of rBAT and CAT4 mRNA in the jejunum of Lingnan-yellow chicken (P<0.05)and had the trend to increase the expression abundance of y+LAT2 and CAT1 mRNA in jejunum(P>0.05); The supplementation of xylanase in the wheat-based diet had no effect on the expression of rBAT mRNA in ileum of chicken, but had the trend to increased the expression abundance of y+LAT2 , CAT1 and CAT4 mRNA in ileum(P>0.05).