| In modern production,it is a popular trend to feed dairy animals with high-concentration diets to meet the energy requirements of lactation.However,excessive intake of high concentrate,ruminants can lead to accumulation of volatile fatty acids in the rumen and increasing release of harmful substances such as lipopolysaccharides(LPS).The accumulation of volatile fatty acids increases the incidence of subacute ruminal acidosis(SARA).LPS enters the liver through the portal vein,affecting liver morphology and metabolic function,and inducing hepatocyte apoptosis,which weakens liver function.LPS entering the spleen through systemic circulation can also cause damage to the spleen,thereby affecting the health of ruminants.In this study,Guan zhong dairy goat was used as an animal model to simulate the feeding mode in actual production.The SARA model was induced by adding a high proportion of concentrate to the diet to study the effect of SARA on inflammatory factors and hepatocyte and splenic cell apoptosis in the lactating period.The results can accumulate data for the research of the relationship between SARA and the inflammation of liver and spleen and the cell apoptosis of milk goat,and it also can give guide for the use of high-concentrate diet in the breeding of ruminants such as cows and milk goats in the actual production.The research contents and results of this experiment are as follows:Sixteen dairy goats with similar body weight,good health,2-3 foetus age milk goats were randomly divided into the control group and the experimental group,with 8 goats in each group.The control goats were fed a daily with concentrate to coarse 35:65,and the experimental goats were fed a diet with concentrate to coarse 65:35(high concentrate).The trial period is 19 weeks,and all dairy goats were free to access water during the period.1.At the 7th week,the 9th week,the 11 th week,the 13 th week,the 15 th week,the 17 th week,and the 19 th week of the test,blood samples were taken from the jugular vein of all the goats,and blood was biochemically analyzed for 19 weeks.analysis.The results showed that compared with the control group,the white blood cells and lymphocytes of the experimental group had no significant changes during the whole experiment,while the red blood cells increased significantly at the 19 th week of the experiment,and the red blood cell volume distribution width increased significantly during the middle of the experiment(p < 0.05).There was no significant change in other blood routine indicators and the measured biochemical indicators(p > 0.05).The results indicated that feeding high concentrate had no significant effect on blood routine and biochemical indicators of dairy goats,but can increase the number of red blood cells in the blood.2.Liver and spleen tissues were collected at the end of the experiment to make HE sections.The results showed that compared with the control group,the liver cells of the experimental group were swollen,with granular degeneration and vacuolar degeneration,and a large amount of lymphocytes infiltrated in the central vein.The content of hemosiderin in the red pulp of the spleen decreased,but the structure showed no visible changes with naked eye;suggesting that SARA caused inflammatory damage to the liver of lactating dairy goats,and the content of hemosiderin in the spleen of lactating dairy goats decreased.3.Collect liver and spleen samples from all dairy goats,the expression of important proteins of inflammatory factors and regulatory protein of TLRs pathway were detected with RT-qPCR.The results showed that the expression of TLR4,NF-κB2 and IL-6 in the liver tissue of high-concentration dairy goats was significantly increased(p < 0.05),and other genes had no significant change(p > 0.05);TLR4 gene expression in spleen tissue increased significantly(p < 0.05)and the expression of NF-κB2 gene in spleen decreased significantly(p < 0.05).The expression of anti-inflammatory factor IL-10 and pro-inflammatory factor TNF-β gene increased significantly(p < 0.05).4.The expression of key proteins of TLR4 signaling pathway was analyzed using Western blot.The results showed that there were no significant changes in the expression of JNK,ERK and P-38 in the MAPK pathway in the liver and spleen(p > 0.05),and there was no significant change in NF-κB protein in the NF-κB pathway(p > 0.05).5.RT-qPCR was used to detect the expression of important apoptosis-regulating proteins Bcl-2 and Bax and apoptosis-executing proteins caspase3,caspase8 and caspase9.The results showed that the expression of caspase9 gene in liver and spleen of dairy goats in the experimental group all increased significantly(p < 0.05).Western blot was used to detect the expression of caspase3 protein in liver tissues.The results showed that there was no significant change in the expression of caspase3 protein in liver tissues of dairy goats in the experimental group compared with the control group.The results of this study showed that feeding lactating dairy goats with high-concentrate leads to the occurrence of SARA in dairy goats.When milk goats suffer from SARA,the number of red blood cells in the blood increases,granular degeneration and vacuolar degeneration occur in hepatocytes,and lymphocytes infiltrate in hepatic lobules.The expression of IL-6 gene increases in the liver,the expression of TNF-β gene increases in the spleen,the hemosiderin decreases in the spleen,and the expression of NF-κB gene in the spleen decreases.The results indicated that feeding lactating dairy goats with high-concentrate led to the occurrence of SARA in milk goats,and the produced LPS activates the TLR4 receptor pathway and gives rise to the elevated expression of IL-6,TNF-β in the liver and spleen of milk goats,thus causing inflammatory damage to the liver and spleen of milk goats. |
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