Effects Of Yeast Selenium On LPS-induced Spleen Injury And Egg Quality In Laying Hens

Selenium is an active component of the antioxidant enzyme glutathione peroxidase.It participates in the vivo synthesis of a variety of selenium-containing proteins,and plays an important role in the body's oxidative stress,immune regulation,cell apoptosis and programmed necrosis.In the breeding of laying hens,it is susceptible to be stim ulated by factors such as lipopolysaccharide(LPS),glucocorticoid,heat stress and cold stress,leading to the reduction of immune function and production performance.In laying hens,the effects of different selenium sources and levels on antioxidant ability,immunity and performance are significantly different.As a common organic selenium source,yeast selenium has the advantages of good safety and high absorption rate.This study was conducted to investigate the following objectives: 1)the effects of yeast selenium on egg quality and production performance;2)to study the yeast selenium antagonism with LPS induced laying hens spleen and lymphocyte damage model;3)to observe the spleen histopathological changes,oxidative stress indicators,inflammato ry response indicators,programmed necrosis related genes,heat shock protein m RNA and protein level changes.At last,the overall aim of this study was to illustrate the mechanism of yeast selenium alleviating spleen tissue damages caused by LPS,and to provide a theoretical basis for the application of selenium yeast in laying hens.Experiment 1: A total of 960 healthy hy-line brown laying hens aged 38 weeks were randomly divided into 2 groups with 6 replicates in each group and 80 hens in each replicate.Group 1 was the control group,feeding to the basal diet(sodium selenite group,containing 0.35mg/kg selenium);Group 2 was added yeast selenium on the basis of the basal diet,containing 0.35mg/kg selenium;Prefeeding period was lasted for 7 days,and then the experimental period was 56 days.The day before the experiment started,the hens were divided into the following groups: control group,control+LPS group,0.35mg/kg yeast selenium group,0.35mg/kg yeast selenium+LPS group.The observing indicators were listed as follows:the spleen tissue and serum antioxidant indexes(GPX,SOD,CAT,MDA,NO,i NOS),serum cytokines(INF-?,TNF-?,IL-2),cytokines(IL-1?,INF-?,IL-2,IL-17);splenic lymphocyte necrosis,heat shock protein gene and protein expression in spleen tissue and lymphocyte(HSP27,HSP40,HSP60,HSP70 and HSP90);inflammatory factor gene expressions in spleen tissue and lymphocyte(i NOS,COX-2,NF-?B,PTGE,TNF-?,HO-1);expressions of 22 selenoprotein-related genes in spleen tissue(Dio1,Dio2,Dio3,GPX1,GPX2,GPX3,GPX4,SelH,SelI,Sel15,SelM,SelO,SelK,SelPb,Selpp,SelT,Sepx1,SelU,SelS,TrxR1,TrxR2,TrxR3);programmed necrosis related index expression in spleen tissue and lymphocyte(MLKL,FADD,RIP1,RIP3,Caspase-8)and changes in the MAPK signaling pathways(ERK/p-ERK,JNK/p-JNK,P38/p-P38)in spleen tissue and lymphocytes.These results were listed as follows:1.The results of AO/EB double fluorescence staining and flow cytometry showed that LPS can induce programmed necrosis in chicken splenic lymphocytes,and the addition of yeast selenium can alleviate the necrosis rate of lymphocytes to certain extent.2.The results showed that there were no significant changes in the antioxidant indexes(GPX,SOD,CAT,MDA,NO,iNOS)and cytokine levels(INF-?,TNF-?,IL-2,IL-1?)in the spleen tissue and serum of the yeast selenium group,while showed significant changes in the LPS group,when compared with the control group.The expression levels of antioxidant indexes and cytokines in the spleen tissue and serum of the yeast selenium+LPS group were between the control group and the LPS group,and both showed statistically significant differences.The results showed LPS has toxic effect on hens,and yeast selenium can alleviate the toxici ty of LPS.3.Through the obsevation of the expression levels of 22 selenoproteins,it was found that compared with the control group,the m RNA expression levels of GPX4,SelH,SelI,SelM,SelO,Selpp,SelT,Sepx1,SelU in the spleen tissue of the yeast selenium group were increased significantly.In LPS group,the mRNA expression levels of GPX2,GPX4,SelH,SelI,Sel15,SelM,and SelO were significantly reduced,while the mRNA expression levels of SelK,SelS,TrxR1,TrxR3 increased.In addition,compared with the LPS group andyeastselenium+LPS group,except for SelU,the mRNA levels of the remaining 21 selenoproteins all had significant changes.4.Through the detection of the expression levels of heat shock protein related indicators(HSP27,HSP40,HSP60,HSP70 and HSP90)in the spleen tissue and lymphocytes,differences were found when compared with the control group and the yeast selenium group.The LPS group expression level of each index was significantly increased,and the expression level of each index in the yeast selenium + LPS group was significantly increased;Compared with the LP S group,the expression level of each index in the yeast selenium + LPS group was significantly reduced.5.Through the detection of the expression levels of inflammatory factors(i NOS,COX-2,NF-?B,PTGE,TNF-?,HO-1)in the spleen tissue and lymphocytes,differences were found in the spleen tissue and lymphocytesin LPS group.The expression of i NOS,COX-2,NF-?B,PTGE,and TNF-? were significantly higher than the control group and yeast selenium group,and the expression of HO-1 in the LPS group was significantly decreased.The expression levels of i NOS,COX-2,NF-?B,PTGE,and TNF-? in the yeast selenium+LPS group were significantly higher than those in the control group and the yeast selenium group.The expression levels of i NOS,COX-2,NF-?B,PTGE and TNF-? in the yeast selenium + LPS group were significantly lower than the LPS group.The expression of HO-1 in the yeast selenium+LPS group significant increased when compared with the LPS group.6.Through the observations of the expression levels of MAPK signal ing pathway related indicators(ERK/p-ERK,JNK/p-JNK,p38/p-p38)in spleen tissue and lymphocytes,differences were found in spleen tissue when compared with the control group and the yeast selenium group.The expression of MAPK signaling pathway related indicators in lymphocytes and lymphocytes did not change significantly.The expression level of MAPK signaling pathway related indicators in the LPS group increased when compared with the control group.Compared with the LPS group,the expression levels of MAPK signaling pathway related indicators in the yeast selenium+LPS group were significantly lower.7.Through the observations of the expression levels of the programmed necrosis signal pathway related indicators(MLKL,FADD,RIP1,RIP3,caspase-8)in the spleen tissue and lymphocytes,differences were found when compared with in the spleen tissue and lymphocytes in the control group and the yeast selenium group.The expressions of related indicators of the programmed necrosis signal pathway did not change significantly.The expression levels of MLKL,FADD,RIP1,and RIP3 in the LPS group increased comparing with those in the control group,while the expression levels of caspase-8 decreased.Compared with the LPS group,the expression levels of MLKL,FADD,RIP1,and RIP3 in the yeast selenium+LPS group were significantly lower,while the expression level of caspase-8 increased.Experiment 2: A total of 1440 healthy Hy-line brown laying hens aged 38 weeks were randomly divided into 3 groups with 5 replicates in each group and 96 hens in each replicate.Group 1 was the control group,feeding to the basal diet(sodium selenite group,containing 0.35mg/kg selenium);Group 2 and 3 were added yeast selenium on the basis of basal diet,containing 0.35mg/kg and 0.5mg/kg selenium respectively.The experimental period was lasted for 56 days.The effects of egg laying performance(egg laying rate,egg production,egg weight and feed-to-egg ratio),egg quality(selenium content,lutein content,egg quality,coefficien t of yolk,height of air chamber and haugh unit)and antioxidant indexes(GSH-Px?SOD?MDA?T-AOC)were detected in each group.The results showed that there were no significant differences in laying rate,egg mass,egg weight and feed to egg ratio among these groups.The selenium content in eggs were significantly increased with the increasing levels of yeast selenium content in the diet.At the same time,yeast selenium can significantly increase the content of lutein in eggs and deepen the color of egg yolk.The results also showed that the egg quality loss rate and the air chamber height had a tendency to decrease,while the yolk coefficient and haugh unit increased significantly in the yeast selenium group.The content of glutathione peroxidase(GSH-PX)and the superoxide dismutase(SOD)increased significantly,while the content of malondialdehyde(MDA)in yeast selenium group decreased significantly.In the current study,the addition of yeast selenium to the diet had no significant effects on laying performance of hens,but significantly improved the egg quality,especially the concentrations of selenium in eggs and the antioxidant activity,thereby extending the egg shelf period.In addition,LPS can cause changes in the pathological structure of the spl een of laying hens,inducing oxidative stress in spleen cells,activating the NF-?B/MAPK signaling pathway,causing inflammation and programmed necrosis of spleen cells,and inducing heat shock protein family genes expressions.The yeast selenium can alleviate the occurrence of histopathological changes,oxidative stress,inflammations and programmed necrosis to certain extent,and regulate the expression of 21 kinds of selenoproteins,and enhance the immune regulation ability of laying hens,which plays an active protective role in production.