In Low Protein Diets Supplemented Branched Chain Amino Acid Influenced The Urea Synthesis In Growing Pigs

In the breeding,nitrogen utilization efficiency and nitrogen emissions is closely related to the farming and environmental problems.This study was to research the fed low protein diet and adding branched chain amino acids(BCAA)on the influence of pig growth performance and the urea cycle.MiRN As as an important molecular is known about;however,whether miRNAs regulate the urea synthesis in liver is unknown.Therefore,the purpose of this study was to evaluate the effect on the expression of urea cycle and other relevant miRNAs with feeding low protein diets and adding BCAA.In vivo experiment fourty Large White x Landrace x Duroc growing pigs(57.4±0.2kg of weight)were divided into four groups and were fed a 14 % crude protein diet,12.5 % crude protein diet,11 % crude protein diet or 11 % crude protein supplemented with BCAA diet for 35 days.At the end of the test,five pigs were killed in each group to collect the data of feed intake,weight gain and the sample of blood,urine,liver tissue and duodenum tissue,jejunum tissue,ileum tissue,which were tested to get related indicators.In vitro experiment hunger cell model,1 mM BC AA treated the intestinal epithelial cell for 12 hours,and then testing urea nitrogen concentration and the expression of enzyme in the urea cycle.In vivo experiment results showed that dietary protein restriction reduced pig growth performance,which were restored by BC AA supplementation.In the serum and plasma of the low protein die group and supplementting BCAA group,the concentration of urea nitrogen significantly reduced.Adding BCAA to the 11% crude protein diet,the protein expression level of liver urea cycle key enzymes CPS1 present ed a down trend but in supplementatting BCAA group,OTC protein expression level was significantly lower(P < 0.01)compared to OTC protein expression levels of the 11% CP diet group.These suggested that adding BCAA to the11% CP diet decreased the protein expression level of CPS1 and OTC to affect the process of synthesis of urea urea cycle.In gene expression,the CPS1 gene expression level of supplementing BC AA in 11% CP diet group decreased significantly(P < 0.05)compared with the 14% CP diet group and the 11% CP diet group,but the gene expression level of OTC and Sirt5 in each group had no difference(P > 0.05),which suggested that BCAA may also affect the gene expression of CPS1.In addition,in the 11% CP diet group and 11% CP added BCAA group protein expression level and gene expression level of duodenal urea cycle key enzymes CPS1 and OTC in different degrees of difference.The protein expression levels of CPS1 and OTC in the 11% CP diet supplement BCAA group rise d significantly(P < 0.05)compared to the 12.5% CP diet group and 11% CP diet group,but the protein expression level of Sirt5 had no difference in each group(P > 0.05).In gene expression,gene expression level of the duodenal CPS1,OTC and Sirt5 was not significant change.Above shows that low protein levels may affect the protein expression level of CPS1 in duodenum,and supplementary BCAA to raise protein expression level of CPS1 and OTC,which means BCAA can affect the protein expression level of CPS1 and OTC in urea cycle to affect the process of urea synthesis in urea cycle.Furthermore,in liver the expression level of six miRN As that were forecast to participate in the urea cycle metabolism had significant difference in the low protein diet group and the complementary BC AA group compared with 14% CP diet group.The miR-20 a and miR-199 b were predicted that targeted CPS1,whose expression level were similar between the 11% CP diet group and 11% CP diet supplement of BCAA group but were significantly lower(P < 0.01)compared to the 14% CP diet group.The miR-92 a and miR-204 were predicted target OTC,whose expression level significantly increased(P < 0.01)in supplementting BCAA group than 11% CP diet group but was similar to 14% CP diet group.The miR-19 a and miR-19 b were predicted targeting Sirt5,and expression level of miR-19 a were similar among 12.5% CP diet group,the 11% CP diet group and 11% CP diet supplement BC AA group,which had a significantly lower than the average 14% CP diet group,while the expression level of miR-19 b was similar to 14% CP diet group(P > 0.05)significantly increased after supplementting the BC AA to the 11% CP diet group(P< 0.01)compared to the 11% CP diet.Comprehensive the above results show that low protein levels down-regulated the expression level of miR-19 a,miR-20 a and mi-199 b,conversely,BCAA can cover the expression levels of the miR-19 b,miR-92 a and miR-204.What was said above suggested that low protein levels and BCAA directly affected the expression of six miRNAs.In addition in the duodenum the expression level of miR-19 a and miR-92 a had no significant differences,and the expression level of miR-19 b in the 14% CP diet group,the 11% CP diet group and 11% CP diet supplement BCAA group had no significant differences,while the expression level of miR-20 a decreased significantly in 12.5% CP diet group,the 11% CP diet group and 11% CP diet supplement BC AA group compared to the 14% CP diet group(P < 0.05),which showed that low protein down-regulated the expression level of miR-20 a.In vitro experiment results show that the concentration of 1 mM BCAA treating intestinal epithelial cells after 12 hours in the hunger model,the urea nitrogen concentration of treating group cells significantly reduced,and the expression level of CPS1 significantly decreased.In conclusion,growth performance and nitrogen metabolism were enhanced by BCAA supplementation to low protein diets,and the expressions of key enzymes of urea cycle were reduced in growing pigs,while six miRN As predicated to be negatively regulate urea synthesis were elevated in their expression.Our results provide a new data for further understanding of BC AA as functional AA in animal nutrition,which effects nitrogen metabolism and urea cycle from the low protein diets and the 11 % crude protein + BCAA diet..