| This study consists of two parts:1.Effects of dietary on abundance and mechanism of Na+/H+ exchangers in omasum epithelium of goats.2. Effects of dietary on abundance and mechanism of ammonia transporters in rumen and omasum epithelium of goats.Series 1 Effects of dietary on abundance and mechanism of Na+/H+ exchangers in omasum epithelium of goats.This series is divided into two experiments,experiment 1 Effects of dietary on abundance of Na+/H+ exchangers in omasum epithelium of goats in vivo. 34 crossbred young goats were selected for this study,they were randomly assigned to two dietary groups:PNSC group and PNS group. All goats were feed with peanut stems and water were avaliable at any time. The goats in PNSC group were fed with 100 g concentrate for four times every day. After 42 days feeding, all the goats were slaughtered.Goats in experiment 1 were slaughtered after 16h fasting.goats in experiment 2 were slaughtered after 2h with feeding.It was found that whether fasting or continuous feeding, The pH of omasum fluid in the PNSC group were significantly lower than those in PNS group; The concentration of propionate, acetate and butyrate of omasum fluid in the PNSC group were significantly higher than those in PNS group;Using Real-time PCR and Western-blot methods, it was found that the Na+/H+ exchanger NHE1 mRNA and protein expression level of omasum epithelial in the PNSC group is significantly higher than those in the PNS group in experiment l,and the expression level of NHE2 and NHE3 mRNA and protein had no difference between the two groups in the omasum of goats; in experiment 2,goats were slaughtered after 2h with feeding, it was found that the NHE1 mRNA and protein expression level of omasum epithelial in the PNSC group is significantly higher than those in the PNS group, and the expression level of NHE2 and NHE3 mRNA and protein had no difference between the two groups,the same like experiment 1.Experiment 2 Effect of pH and SCFA on abundance of Na+/H+exchanger in omasum epithelial of goats in vitroOmasum epithelial were taken from the healthy young goats (n=4) after slaughter, and were digested by fractional trypsinization then incubated at 37℃ with 5% CO2 for 24 h to allow attachment. The attached cells were treated by low pH and high SCFA for 24 h.After collecting the cells, using Real-time PCR and Western-blot methods to test the genes and proteins expression. In vitro study, it was found that SCFA can increase Na+/H+ exchanger NHE1 mRNA and protein expression level in omasum epithelial of goats,low pH have no influence on NHE1 expression. Both low pH and high SCFA can reduce NHE2 and NHE3 genes and protein expression in omasum epithelial of goats.Series 2 Effects of dietary on abundance and mechanism of ammonia transporter in rumen and omasum epithelium of goats.This series is divided into two experiments,experiment 1 Effects of dietary nutrition level on mRNA abundance of ammonia transporter proteins in rumen and omasum epithelium of goats in vivo.15 boer goats were divided into high nutrition level group (HLgroup) and low nutrition level group (LLgroup).Goats in HLgroup were feed with concentrate 327g, grass 78.48g, alfalfa 30.52g; goats in LL group were feed with grass 348.8g, alfalfa 69.76g, premix 17.44g. Two times one day. After a period of concentrate added, Finally,goats in LL group were feed with grass 390.47g, alfalfa 78.09g, premix 17.44g, goats in HL group were feed with concentrate 700g, grass 171 g, alfalfa 68g. After 2 weeks feeding, all the goats were slaughtered for sampling. In vivo study, it was found that the pH of rumen and omasum fluid in the HL group were significantly lower than those in LL group; The concentration of propionate, acetate and butyrate of rumen and omasum fluid in the HL group were significantly higher than those in LL group; ammonia concentration in HL group were significantly higher than those in LL group in rumen of goats; Rhbg and Rhcg mRNA expression level in the HL group is significantly higher than those in the LL group in rumen and omasum epithelial of goats.Experiment 2 Effect of pH,SCFA and Nitrogen Compounds on mRNA abundance of ammonia transporters in rumen and omasum epithelial of goats in vitroRumen and omasum epithelial were taken from the healthy young goats (n=4) after slaughter, and were digested by fractional trypsinization then incubated at 37℃ with 5% CO2 for 24 h to allow attachment. The attached cells were treated by pH, SCFA,NH4C1 and Urea for 24 h.After collecting the cells, using Real-time PCR method to test the genes expression. In vitro study, it was found that low pH can increase Rhbg and Rhcg mRNA expression in rumen and omasumepithelial of goats; SCFA,Urea and SCFA+NH4Cl can decreace Rhbg and Rhcg mRNA expression in rumen epithelial of goats; low pH and NH4Cl can increase Rhbg and Rhcg mRNA expression in omasum epithelial of goats; both SCFA and Urea can reduce Rhbg and Rhcg genes expression in omasum epithelial of goats. |
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