| In this thesis,the pancreatic tissues of cattle,sheep,pigs and chickens were cultured in vitro under different concentrations of glucose,and the secretion of digestive enzymes and gene expression in the tissues and culture medium were compared.The differences of pancreatic digestive enzymes secretion between ruminants and monogastric animals were studied,and the changes of pancreatic digestive enzymes in ruminants and monogastric animals were compared from the molecular level The specific research content is divided into the following four parts:Experiment 1: the effect of glucose concentration on the secretion of digestive enzymes in the pancreas of cattle,sheep,pigs and chickens in vitroIn this experiment,the pancreatic tissues of cattle,sheep,pigs and chickens were cultured in vitro.Four treatment groups were set up with different gradients of glucose medium,which were control group(glucose concentration was 3.3 m M/L),treatment group(glucose concentration was 6.6 m M/L,9.9 m M/L and 13.2 m M/L).Each treatment was repeated for 5 times,and the culture time was 1 hour and 2 hours respectively.The results showed that glucose could increase the secretion of digestive enzymes in pancreatic tissues of cattle,sheep,pigs and chickens by linear or quadratic curve.Glucose linearly increased the secretion of α-amylase(P < 0.05),and quadratic curve increased the secretion of lipase,trypsin and chymotrypsin(P < 0.05).Compared with the culture time of 2 hours,the sensitivity of tissue activity and enzyme activity of pancreatic tissue cultured for 1 hour was better.Experiment 2: Interaction of glucose and animal species on pancreatic digestive enzyme secretion in vitroIn this experiment,the glucose concentration of 3.3 m M/Lwas selected as the control group,and the glucose concentration of 9.9 m M/L was selected as the experimental group,and the pancreas was cultured in vitro for 1 h.After culture,it was revealed whether there were interspecific differences in pancreatic digestive enzyme secretion between monogastric animals and ruminants.The results showed that high concentration glucose could promote the exocrine quantity of digestive enzymes(P < 0.01);Glucose had no significant effect on the secretion of amylase,trypsin and lipase(P > 0.05),that is,the secretion of amylase,trypsin and lipase was not affected by species;Except amylase in pancreatic tissue and trypsin in culture medium(P > 0.05),other digestive enzymes had interaction with different species(P < 0.01).Experiment 3: Interaction of glucose and animal species on m TOR signaling pathway in pancreasThe purpose of this study is to explore the expression changes of key genes and proteins of m TOR signaling pathway in pancreatic tissue,and to compare whether there are differences in the secretion of digestive enzymes between bovine and pig pancreatic tissue from the molecular level.The test materials are the same as those in test 2.By detecting the expression and phosphorylation level of key genes and proteins in m TOR signaling pathway in pancreatic tissue,the mechanism of glucose regulating digestive enzyme synthesis in bovine and pig pancreatic tissue was revealed.The results showed that the expression of PKB and S6K1 gene was significantly affected by species(P < 0.01),but the m RNA expression of 4E-BP1 gene was not affected by species(P > 0.05).When glucose was increased,the expression of m TOR pathway related genes in pancreas tissue of cattle and pigs was not affected(P > 0.05);The protein expression of PKB,S6K1 and4E-BP1 were significantly affected by species(P < 0.01),but the phosphorylation level was not affected by species(P > 0.05);Except for S6K1 protein expression,there was interaction between species(P < 0.01),and there was no interaction between other genes,protein expression and phosphorylation level(P > 0.05).This indicated that glucose could regulate the secretion of digestive enzymes in pancreas of cattle and pigs through m TOR signaling pathway,and there were some differences in the regulation mechanism of glucose.Experiment 4: Interaction of glucose and animal species on pancreatic ISR signal networkThe purpose of this study is to explore the expression changes of key genes and proteins of ISR signal network in pancreatic tissue,and to compare the differences of glucose secretion of digestive enzymes between monogastric animals and ruminants.The experimental animals and methods were the same as those in Experiment 3.The effects of glucose on the expression and phosphorylation of key genes and proteins in ISR signal network of bovine and pig pancreas were investigated to reveal whether there are interspecific differences in the mechanism of glucose regulating digestive enzyme synthesis in bovine and pig pancreas.The results showed that there were interspecific differences in the m RNA expression of ATF4 gene(P < 0.01),but e IF-2 was not α There was no significant difference in m RNA expression among species(P > 0.05);High concentration of glucose could promote the expression of ATF4 m RNA(P < 0.05);The expression and phosphorylation of ATF4 and e IF-2 were detected α The protein expression of e IF-2 was not significantly different between cattle and pigs(P > 0.05)α The protein phosphorylation levels of the two groups were significantly different(P < 0.01);The increase of glucose concentration can promote the protein expression of ATF4 and e IF-2 in cattle and pigs α The protein phosphorylation level was significantly higher than that of control group(P < 0.01);Except e IF-2 α There was no significant interaction between other genes,protein expression and phosphorylation level(P > 0.05).This suggests that glucose can regulate the secretion of digestive enzymes in bovine and pig pancreas through ISR signaling pathway.The regulation mechanism of glucose on pancreatic digestive enzyme secretion in ruminant and monogastric animals was different.In conclusion,under the experimental conditions,glucose can promote the secretion of digestive enzymes in vitro pancreatic tissue of cattle,sheep,pigs and chickens,and the secretion of digestive enzymes in vitro pancreatic tissue shows a linear increasing trend,without interspecific difference,but the secretion of digestive enzymes is different among species.From the molecular level,glucose can regulate the secretion of digestive enzymes in bovine and pig pancreas through m TOR and ISR signal network,and the regulation mechanism is different among species. |
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