| Yak is the main livestock on the Qinghai-Tibet Plateau.Because of the special climatic and environmental conditions,the nutritional intake of grazing yaks was inhibited by the fluctuations of grass production and nutritional values according to the season changes,induced to the high occurrence rate of growth retarded yaks.Because the growth performance and feed efficiency were much lower than the normal yaks with the same breed and age,it reduces the economic income of herdsman.Rumen is the special digestive organ of ruminants.Rumen health is the key point of improving the feed efficiency of ruminants and also is the basic of healthy and efficient ruminants farming.Rumen epithelium barrier is comprised of epithelial cells and cell junction complex,which plays an important role on regulating epithelial immune and efficiency nutrients absorption.The cysteamine(CS)and yeast are the important rumen regulating agent.However,if they could stimulate the efficiency growth of growth retarded yaks and its mechanism are still unclear.Therefore,this study compared the differences of rumen development and rumen epithelium transcriptome between growth retarded and normal yaks,and investigated the effects of CS and yeast on the growth performance and rumen microflora,and also investigated their effects on the key gene expressions in the different functional pathway.This study aimed to reveal the different effects of CS and yeast regulating on the gene expressions of rumen epithelial cell junctions through altering the rumen flora and its fermented productions,to provide theoretical and experimental evidence for nutritional regulation of rumen health and stimulating healthy and efficiency growth of yaks.Experiment 1: The comparison of rumen development and rumen epithelial transcriptome between growth retarded and normal yaksAccording to the previous study of our team on the body weight and growth axis hormone secretion of yaks(Hu et al.,2016),ten 15-month-old Qinghai plateau yaks with growth retardation were chosen referred to the criterion that the yak's body weight was below the lowest 10% of yak colony(Jones et al.,2012).Ten normal growth yaks with the same breed and age were chosen,which had the body weight closest to the average weight of yak colony.After blood collected,yaks were slaughtered and rumen epithelium transcriptome were sequenced.The results were shown as following:(1)The digestive organ index of rumen,reticulum,abomasum and jejunum of growth retarded yaks were significantly lower than those of normal yaks(P <0.05)and its rumen epithelial papilla height and muscle layer thickness were significantly lower than the normal yaks(P <0.05).The rumen acetate,butyrate and microbial protein(MCP)concentrations of growth retarded yaks were significantly lower and the ammoniacal nitrogen was significantly higher than those of growth normal yaks,respectively(P <0.05).(2)The serum glucose(GLU),total protein(TP)and albumin(ALB)concentrations of growth retarded yaks were significantly higher than those of normal yaks(P <0.05),blood urea nitrogen(BUN)concentration of growth retarded yaks was significantly lower than normal yaks(P <0.05).The serum LPS and proinflammatory factors including TNF-? and IL-2 concentrations of growth retarded yaks were significantly higher than those of normal yaks(P <0.05).(3)The up-regulated genes in the rumen epithelium of growth retarded yaks were mainly enriched in the functional classification of GO analysis including extracellular region,extracellular matrix,plasma membrane,defense response,immune system process and inflammatory response.The down-regulated genes were mainly enriched in the functional classification of activity of substance transmembrane transporter.This suggested the epithelial cell structure,immune function and substance transport were difference between growth retarded and normal yaks.(4)The rumen epithelium of growth retarded yaks had higher gene expressions in the cell matrix including filamentins,fibronectin,collagen and the key negative regulator of epithelial cell junction myosin light-chain kinase(MYLK)and myosin light 9(MYL9),these genes were significantly enriched in the Focal adhesion and ECM-receptor interaction pathways.The genes related to inflammatory response including complement component C2,C3 and C7,major histocompatibility antigens(MHC),Serum amyloid(SAA)and chemokine were higher expressed in the rumen epithelium of growth retarded yaks,which were enriched in the Complement and coagulation cascades and Antigen processing and presentation pathways.The genes related to ion and VFA transport including SLC40A1,SLC31A1,SLC26A3 and CLCN2 were lower expressed in the rumen epithelium of growth retarded yaks,which were enriched in the Mineral absorption pathway.These results suggested that growth retarded yaks had the maldevelopment and inefficient fermentation of rumen.The different expressed genes were enriched in the focal adhesion,complement and coagulation cascades,antigen processing and presentation and mineral absorption,indicated the cell junction damage,inflammatory response and inefficient nutrients absorption,which inhibited the body growth.Experiment 2: Effects of CS and yeast on the growth performance and rumen microflora of growth retarded yaksBased on the experiment 1.thirty 15-month-old Qinghai plateau growth retarded yaks were chosen according to the criterion in experiment 1 and randomly divided into three groups with 5 repeats and 2 yaks in each repeat.Three groups were randomly assigned to basal ration group(GRB),basal ration addition CS group(GRBC)or basal ration addition yeast(GRBY)group.The dosages of CS and yeast were according to the previous simulated rumen fermentation experiment and referred to the Yang et al.,2005 and Zhang et al.,2014,respectively.After 60-day feeding following 4-day digestion experiment,yaks were slaughtered.(1)Addition CS and yeast both significantly increased the ADG of growth retarded yaks(P <0.05)and reduced the ratio of feed to grain of growth retarded yaks(P <0.05).Addition CS significantly increased the digestibility of ration energy,NDF and ADF(P <0.05),whereas yeast significantly increased the digestibility of ration energy and crud protein(P <0.05),of which addition yeast had the optimal growth promoting effects.(2)Both addition CS and yeast significantly increased the serum GLU and TP concentrations and improved the nutritional status of growth retarded yaks(P <0.05).Addition yeast significantly increased the serum globulin concentration(P <0.05),decreased the ration of albumin to globulin(P <0.05),increased the serum Ig A and Ig G concentrations(P <0.05),improving the immunity of growth retarded yaks.Both addition CS and yeast significantly decreased the serum LPS and IL-2 concentrations(P <0.05),and yeast further significantly decreased the serum TNF-? and IL-1? concentrations(P <0.05),alleviating the inflammatory response of growth retarded yaks.(3)Addition CS significantly increased the rumen acetate concentration and the ratio of acetate to propionate(P <0.05),stimulated the acetate type fermentation,whereas addition yeast significantly increased the rumen p H value(P <0.05),decreased the ratio of acetate to propionate(P <0.05),stimulated the propionate type fermentation of growth retarded yaks.Both CS and yeast addition significantly decreased the rumen ammoniacal nitrogen concentration(P <0.05),suggested rumen fermentation improved of growth retarded yaks.(4)Addition CS and yeast significantly changed the rumen microflora of growth retarded yaks.Both addition CS and yeast significantly increased the abundant of Saccharofermentans and Butyrivibrio 2 in the rumen of growth retarded yaks(P <0.05).Addition CS significantly increased the ratio of Firmicutes to Bacteroidetes and increased the abundant of Spirochaetes,Lachnospiraceae XPB1014 group and Treponema 2(P <0.05).Yeast addition significantly increased the abundant of Prevotella 1,Ruminococcaceae UCG-005 and Pseudobutyrivibrio(P <0.05),and significantly decreased the abundant of Eubacterium coprostanoligenes group.The results suggested that both addition CS and yeast significantly promoted the growth of growth retarded yaks and yeast had the optimal growth promoting effects.Addition CS increased the rumen fiber degrading bacteria,increased the acetate fermentation,increased the digestibility of ration energy and fiber,improved the nutritional status,and promoted the growth of growth retarded yaks.Addition yeast increased the abundances of protein and starch degrading bacteria Prevotella 1 and butyrate-produced bacteria Butyrivibrio 2 and Pseudobutyrivibrio,stimulated the propionate fermentation,increased the rumen p H,increased the digestibility of ration energy and crude protein,improved nutritional status,decreased serum LPS and proinflammatory factor concentrations,alleviated inflammatory response,and promoted the growth of growth retarded yaks.Experiment 3: Different effects of CS and yeast on the gene expression of rumen epithelial cell junctions of growth retarded yaksIn this experiment further detected gene expression of epithelial barrier function in rumen and through correlation analysis to explore the nutrition regulation and effects regulation of rumen flora and fermentation products in rumen epithelial barrier function gene different expressed.The results show:(1)Both addition CS and yeast significantly increased the width of epithelial papillae in rumen(P <0.05)of growth retarded yaks,and CS also significantly increased the rumen papillae surface area and muscular thickness(P <0.05)of growth retarded yaks,improved the development of rumen.(2)Yeast significantly decreased the complement factors C2,C3,C7 and histocompatibility antigen MHC1 gene expression(P <0.05),while relieve the inflammatory response induced by invasion of exogenous antigen of epithelial in rumen in growth retarded yaks.Both addition CS and yeast significantly increased the epithelial rumen Ion and VFA carrier CLCN2?SLC26A3?PAT1 and Na KV gene expression(P <0.05),improved ability of nutrient absorption of rumen epithelial in growth retarded yaks.Yeast significantly decreased(P <0.05)the rumen epithelial barrier key negative regulator MYLK gene expression,while significantly increased(P <0.05)cell junction CLDN1,OCLN and CDH1 gene expression,and enhanced the epithelial barrier function in growth retarded yaks.(3)The concentration of acetic acid and butyric acid in rumen was positively correlated with the width and surface area of rumen papillae(P <0.05),And the concentration of propionic acid and butyric acid was significantly positively correlated with the rumen epithelial cell junction OCLN,CLDN1 and CDH1 gene expression(P <0.05).Prevotella 1,Prevotellaceae UCG-001,Ruminococcaceae NK4A214 group,Ruminococcus 1,Butyrivibrio 2 and other flora abundance were significantly positively correlated with cell junction and ion and VFA carrier gene expression(P <0.05),meantime,significantly negatively correlated with complement factors gene expression(P <0.05).In conclusion,the rumen epithelial in growth retarded yaks cell junction were damaged,ions and the ability of VFA absorption was reduced,inflammatory response induced by invasion of exogenous antigen,rumen epithelial barrier function was insufficient or stunted,inanition and produced inflammation response result in growth retarded yaks.addition CS increased the abundance of rumen utilize fiber bacteria,promote rumen acetic-type fermentation,improved rumen epithelial ion and VFA carrier gene expression,increase the width and muscular thickness of rumen epithelial papillae,promoted rumen epithelial development,improve the digestibility of dietary fiber,energy and other nutrients,and promote growth of growth retarded yaks.Addition yeast increased the abundance of rumen Prevotella 1 and other protein and starch degrading bacteria,as well as probiotics like Butyrivibrio 2.Which Improved the balance of microbial system in the rumen of growth retarded yaks,promote propionic-fermentation,furthermore,improved the cell junction,transport of ions and VFA in the rumen epithelial,alleviated inflammatory response in epithelial,improved the rumen epithelial barrier function,reduced the concentration of LPS moved into the blood and serum pro-inflammatory factors,and improve the digestibility of dietary protein and energy.Addition yeast further promoted the growth of growth retarded yaks compared to addition CS. |
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