| Dairy cows and goats are always fed with high-concentrate diet to keep up with the increasing energy requirement meanwhile to increase the milk yield in the dairy-farming industry,which leads to abnormal fermentation of rumen and then induces subacute ruminal acidosis.Lipopolysaccharide(LPS)in the rumen will accumulate and be translocated into circulatory system and then reach to organ and tissues such as mammary gland.LPS can activate NF-κB signal pathway and regulate the cells apoptosis.Sodium butyrate can promote the development of gastrointestinal tract and enchance the barrier function.In our study,Holstein dairy cows and Saanen dairy goats in middle lactation were used to investigate the effect of high-concentrate diet feeding on ruminants mammary tissues apoptosis and the function of dietary sodium butyrate adding on apoptosis.Experiment 1:The effect of high-concentrate diet on dairy cows mammary tissues cells apoptosis:Twelve Holstein dairy cows in middle lactation were used and allocated into two groups(6/group)randomly and one group was fed with a diet composed of 60%forage and 40%concentrate(low-concentrate,LC group)as a control while the other group received a diet composed of 40%forage and 60%concentrate(high-concentrate,HC group)as a treatment.The experiment period was 18 weeks.Rumen fistula was fit for all the cows before the experiment started and ruminal liquid,milk,mammary artery blood,mammary vein blood and mammary tissues were sampled at the end time of the trial to detect the ruminal pH value,milk yield,milk composition,LPS concentration in the rumen fluid and plasma,the apoptosis-related gene such as caspase-3、caspase-8 and caspase-9 expression,the caspase-3 and caspase-8 enzyme activities and NF-κB protein content.The results showed that long-term high-concentrate diet feeding led to a low pH average value and being below 5.6 for 3 hours.The milk yield,milk fat percentage and milk fat content in HC group were significantly(P<0.01)decreased compared to the LC group while the milk protein percentage was remarkably increased compared with the LC group.The LPS concentration in both rumen fluid and plasma of HC group were significantly(P<0.05)higher than LC group.HC group got a significantly(P<0.05)higher caspase-3 gene expression and remarkably(P<0.05)lower bcl-2 gene expression than LC group.Caspaae-3 and caspase-8 enzyme activities in HC group were significantly(P<0.01)higher than LC group.NF-κB p65 and phospho-NF-KB p65 protein content in HC group were significantly(P<0.05)higher than LC group.Conclusion:long-term high-concentrate diet feding induced mammary tissue cells apoptosis in dairy cows.Experiment 2:The effect of high-concentrate diet feeding on mammary tissues cells apoptosis in dairy goats and the function of dietary sodium butyrate addition:Eighteen multiparous Chinese Saanen dairy goats(38.86±2.06kg)were randomly allocated to three groups(6/group).One group was fed with a diet composed of 60%forage and 40%concentrate(low-concentrate,LC group)as a control while the other two groups,one received a diet composed of 40%forage and 60%concentrate(high-concentrate,HC group),another was fed with a diet which contained 40%forage and 60%concentrate with sodium butyrate(SB)buffer in it(BHC group),both as treatments.The experimental period was 20 weeks.Rumen fistula and hepatic vein catheter were fit on the goats before trail started.Ruminal fluid,milk,hepatic vein blood,and mammary tissues were collected.Real-time PCR was used to evaluate the tissues cell apoptosis.The results were as follows:high concentrate diet feeding leaded to ruminal pH value decrease and addition sodium butyrate helped maintain the pH value in a way.LPS concentration in peripheral blood of HC group was significantly increased(P<0.01)compared with LC group while BHC group got a remarkable(P<0.01)LPS concentrate decrease compared to HC group.Caspase-3 mRNA expression in the HC group was significantly increased(P<0.01)compared to the LC group and BHC group.HC group also got a remarkably higher(P<0.01)caspase-8 mRNA expression than LC group.bcl-2 mRNA expression in BHC group was significantly increased(P<0.01)compared to HC group and LC group.Caspase-3 enzyme activity in the HC group was significantly increased(P<0.01)compared to the LC group and BHC group.HC group got a remarkably higher(P<0.01)caspase-8 enzyme activity than LC group while there was no difference between HC group and BHC group for caspase-8 enzyme activity.HC group got remarkably higher(P<0.05)IL-1β,IL-6,TNF-a compared with LC group and BHC group.Conclusion:High-concentration diet feeding induced mammary tissues cells apoptosis and addition sodium butyrate helped alleviate mammary cell apoptosis.Experiment 3:The effects of high-concentrate diet feeding and dietary sodium butyrate addition on the apoptosis pathway-related proteins of dairy goats mammary tissue cells:Eighteen multiparous Chinese Saanen dairy goats(38.86=12.06kg)were randomly allocated to three groups(6/group).One group was fed with a diet composed of 60%forage and 40%concentrate(low-concentrate,LC group)as a control while the other two groups,one received a diet composed of 40%forage and 60%concentrate(high-concentrate,HC group),another was fed with a diet which contained 40%forage and 60%concentrate with sodium butyrate(SB)buffer in it(BHC group),both as treatments.The experimental period was 20 weeks.Mammary tissues were collected at the end time of the trial.Western blot was used to detect the pathway-related NF-κB、bax、bcl-2、cytochrome C and caspase-3 protein expression level.The results showed that HC group got the highest p65 and phosphor-p65 protein expression among the groups and p65 in BHC group was significantly decreased compared to the LC group.As to bax,cytochrome C(cyt-c),caspase-3 and cleaved caspase-3,HC group got a remarkably higher protein expression.than the other two groups.The bcl-2 protein content in BHC group was significantly increased compared to the LC group and BHC group.Conclusion:High-concentrate diet feeding increased the p65 and phospho-p65 as well as pro-apoptotic protein bax,cyt-c,caspase-3 and cleaved caspase-3 protein expression in mammary tissues of dairy goats and a dietary addition of sodium butyrate helped decrease these protein expressions and increase the anti-apoptotic protein bcl-2 expression. |
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