Dietary Rumen Degradable Starch Regulates Milk Synthesis Of Dairy Goat:Effects And Potential Mechanisms

How to improve milk quality while increasing milk production? This has been a major issue for the development of the dairy industry.Milk is rich in nutrients.However,the main nutrient content(milk fat,milk protein,lactose)in milk is susceptible to factors such as heredity,physiological stages,nutrition,feeding process,and environment.The type and proportion of dietary carbohydrates significantly affect milk yield and milk quality.Studying and evaluating the effective nutritional indexes of diet nutrition is important for optimizing diet formula to improve milk yield and quality.Starch is the main component of non-structural carbohydrates and an important energy source in ruminant diets.The starch present in ruminant diets could be divided into rumen-degradable starch(RDS)and rumen escaped starch(RES)according to their different degradation sites(rumen or intestine).With the rapid development of high-throughput sequencing technology and bioinformatics,the limitations of traditional nutrition research methods have been broken.This study combines traditional nutrition research methods with omics sequencing and bioinformatics analysis.Reveals the effects of different amounts of rumen degradable starch on lactation performance and the transcription and metabolic layers of mammary gland and liver tissues in dairy goats.Experiment 1 Effect of rumen degradable starch on lactation performance and milk precursor synthesis in goatDietary total starch content is the same but different RDS content has different effects on lactation performance.To clarify the effects of RDS on lactation performance and milk precursor synthesis in dairy goats,Eighteen Guanzhong dairy goats(day in milk = 185 ± 12d)with similar parity,weight,and milk yield were selected and randomly assigned to one of three groups(n = 6),which were fed an LRDS diet(Low RDS = 20.52%),MRDS diet(Medium RDS = 22.15%),or HRDS diet(High RDS = 24.88%)for 5 weeks.No effect of RDS on dry matter intake was identified in the present study.The lactation efficiency(FPCM/DMI)tended to increase in the MRDS group(P < 0.1),while the milk yield tended to decrease in the HRDS group(P < 0.1).Moreover,the milk fat content and yield significantly decreased in the HRDS group compared with those in the LRDS group(P <0.01).Although the milk fat content was significantly lower in the MRDS group than in the LRDS group,the milk fat yield was not significantly different from that of the LRDS group.Furthermore,increased RDS significantly decreased ruminal B.fibrisolvens and Pseudobutyrivibrio abundances and increased the trans-10,cis-12 conjugated linoleic acid(CLA)content in the rumen fluid.Each group had no significant effect on milk protein content(P > 0.05),but milk protein yield in the HRDS group was significantly lower than that in the MRDS group(P < 0.05).The supply of leucine,isoleucine,and methionine was significantly higher in the MRDS than HRDS group.And the supply of EAA in the MRDS group had a trend higher than that in the HRDS group(P < 0.01).There was no significant difference in lactose content and yield among the three groups.In summary,when the total starch and NDF contents are similar in diet,appropriately increasing the RDS content will help improve the lactation performance of dairy goats,However,excessive RDS will cause negative effects such as decreased milk fat and milk protein production.Experiment 2 Effect and mechanism of rumen degradable starch on mammary gland based on transcriptomicsThe mammary gland is an exocrine organ with a synthetic function.It can ingest a large amount of nutrients from the blood and synthesize milk fat,milk protein and lactose under the action of a series of enzymes and transport carriers.Based on the results of the first experiment,this experiment will reveal the changes in mammary gland tissue transcription and its related mechanisms under the conditions of feeding different RDS levels.Experiment design is the same as experiment 1.The mammary gland tissues of dairy goats in each treatment group were collected for transcriptomics sequencing.To gain a better understanding of the functional roles of these differentially expressed genes among the 3dietary groups,further GO and KEGG analyses were performed.Most of the top 10 GO terms were related to lipid metabolism,of which only lipid biosynthetic process was significantly enriched(FDR < 0.05).The significant DEGs involved in the lipid biosynthetic processncluding ACSS2,MVD,INSIG1,AGPS,SCD5,FADS2,CERCAM,SC5 D,HSD17B7,HSD17B12,ATM,TP53 RK,GDF1 and LOC102177400.The interaction between DEGs in the mammary tissue of goats was predicted by the R package “WGCNA”.Based on the GO database,most of the top 20 nodes of the blue module's gene coexpression network were associated with lipid metabolism.Among them,INSIG1 had the highest degree of connectivity.Hence,the inhibition of fatty acid synthesis was likely due to a decrease in SREBP activity caused by downregulation of INSIG1,although there was no difference in the m RNA abundance of SREBF1.Experiment 3 Effect of rumen degradable starch on arteriovenous plasma metabolites in dairy goatsExperiment 2 revealed that feeding HRDS diet significantly down-regulated genes related to lipid biosynthetic in mammary tissue.However,it is still unclear how the different RDS diets affect the metabolic profile of mammary gland tissue and the mechanism is still unclear.This experiment uses metabolomics to reveals the effects and mechanisms of different RDS diets on the metabolic profile of mammary artery and venous vein plasma in dairy goats.Experiment design is the same as experiment 1.Three hours after morning feeding,the goat arteriovenous plasma was collected for metabolomics analysis.KEGG analysis of differential metabolic pathways in mammary vein blood showed that fatty acid biosynthesis(map00061)was significantly enriched in each comparison group(MRDS vs.LRDS,HRDS vs.LRDS and HRDS vs.MRDS).Differential metabolites related to fatty acids include stearic acid,myristic acid,oleic acid,palmitic acid,dodecanoic acid,cis-9-palmitoleic acid.Dodecanoic acid and myristic acid were the common metabolites of the three comparison groups which indicated that dodecanoic acid and myristic acid are most likely biomarkers related to milk fat synthesis.However,most of the above fatty acids were not significantly different in the mammary arterial plasma,which indicated that the difference in fatty acids in venous plasma was caused by the metabolic activity of mammary epithelial cells.In addition,the content of primary bile acid was significantly different in the mammary artery and vein plasma of each group.Bile acids play an important role in regulating lipid absorption,transport,and metabolism.Therefore,the reduction of primary bile acid concentration in arteriovenous plasma in the HRDS group was likely to be another key factor that interferes with milk fat synthesis.The relationship between bile acids and milk fat synthesis in mammary tissue needs further study.Experiment 4 Effect and mechanism of rumen degradable starch on liver bile acid metabolism based on transcriptomicsBile acid can promote the absorption of fat and fat-soluble vitamins in the small intestine,and can activate SREBPs and PPARs signaling pathways in the form of signal molecules,which plays an important role in the body's lipid metabolism.Based on the results of Experiment 3,this experiment will reveal the effects of dietary RDS on liver bile acid metabolism and provide new insights into abnormal lipid metabolism in mammary tissue induced by diets with high content of RDS diet.The experimental design was the same as Experiment 1.Because the bile acid concentration decreased significantly with the increase of RDS level,liver samples from the LRDS and HRDS groups were selected for transcriptome sequencing.The results showed that the expression of lipopolysaccharide-binding protein gene(LBP)in liver tissues of dairy goats in the HRDSgroup was significantly increased.The white blood cell count of plasma in the HRDS group was significantly increased.The acute phase and acute inflammation response were significantly enriched by GO enrichment analysis.The above results prove that the liver tissues of dairy goats in the HRDS group are affected by LPS and have inflammatory reactions.KEGG enrichment analysis showed that there are 4 down-regulated genes related to the bile acid secretion pathway(MDR1,RXR?,AE2 and SULT2A1).The WGCNA analysis was performed on all the differential genes.Most genes in gene module that are significantly positively related to white blood cell count are related to the immune response.The three genes with higher connectivity(C1QB,C1 QC,CD48)may be become a potential marker of liver inflammation.In summary,high RDS diets can trigger LPS-mediated liver inflammation and down regulated the key genes related to bile acid secretion,which can lead to abnormal bile acid metabolism and affect milk fat synthesis.In summary,when the total starch and NDF contents of the diet are similar,an appropriate increase in the amount of RDS will help improve the lactation performance of dairy goats,but an excessively high amount of RDS will have a negative impact on the lactation performance of dairy goats.High RDS diets can reduce the abundance of B.fibrisolvens + Pseudobutyrvibrio,cause abnormal fatty acid hydrogenation,and produce trans-10,cis-12 CLA,which can inhibit milk fat synthesis.trans-10,cis-12 CLA is circulated to the mammary glands via blood,and INSIG1 mediates down-regulation of genes related to fatty acid and cholesterol regulation by nuclear transcriptional regulator SREBF1,thereby reducing milk fat content and milk fat production.At the same time,feeding high RDS diets reduced the supply of amino acids to the mammary glands and the uptake of the mammary glands,resulting in lower milk protein yield.In addition,excessive RDS can cause liver Inflammatory response,causing abnormal bile acid metabolism,which may be one of the reasons that cause milk goat milk fat content and yield to decrease.