| In recent years,the frequent appearance of the global extreme climate phenomenon and longer period of high temperature weather in summer gradually become an important factors that affect the animals' health and production performance.Gastrointestinal mucosal barrier is more sensitive to damage when animals subject to heat stress(HS)and HS negatively impacts animal's physiology and production performances.While supplementation of Selenium(Se)can alleviate these negative impacts from HS.Dietary Se,mainly through its incorporation into selenoproteins,plays important roles in various physiology and pathology processes.Selenoproteins play roles in regulation of antioxidant activities and inflammatory response,immunomodulation and anti-stress.Hitherto,the effect of HS on expression of selenoproteins in the intestinal epithelial cells,the reciprocal metabolic impacts of Se on the prevention of intestinal epithelial cells damage and expression of selenoproteins during HS remain unclear.Therefore,we choose the IPEC-J2 cells as experimental material and conduct this study to determine:(1)the effect of HS on the expression of 25 selenoproteins;(2)the protective effect of different sources of Se on alleviating cellular damage induced by HS and the possible mechanism linked the protective effects of Se to selenoproteins expression profiles.The results are presented as follows.Experiment 1:The effect of HS on the expression of selenoproteins in IPEC-J2 cells.A single factorial experiment was designed to study the effect of HS on the expression of selenoproteins in IPEC-J2 cells.Cells were cultured at 37 ? for 24 h(Control)or exposed to a mild hyperthermia at 41.5 ? for 24 h(HS).The results showed that:(1)HS up-regulated(P<0.05)Hsp70 and one tight junction-related gene(Zo-1)in IPEC-J2 cells;(2)At the same time,HS up-regulated(P<0.05)4 selenoprotein genes(Gpx3,Dio2,Selk,Sels)and 3 inflammation-related genes(Il-6,Icam-1?Tgf-,?),down-regulated(P<0.05 or as indicated)6 selenoprotein genes(Gpx2,Gpx6,Txnrdl,Selh,Selm,Selx)and 3 inflammation-related genes(Ifn-?,Mcp-1,Tnf-a)in the cells;(3)HS also exhibited impacts on protein expressions,which up-regulated Hsp70,down-regulated SeIX and showed no effect on SelP in IPEC-J2 cells.Experiment 2:The protective effect of different sources of Se on alleviating IPEC-J2 cellular damage induced by HS and the possible mechanism.This study was designed to investigate the protective effects of different sources of Se(sodium selenite,SS and selenomethionine,SeMet)on alleviating IPEC-J2 cellular damage induced by HS,and the expression of selenoproteins were also compared.IPEC-J2 cells were grown in complete medium at 37 ? until to 80%confluence,then cells were grown in serum-free medium for another 24 h under different conditions:grown at 37 ?(Control),at 41.5 ?(HS),at 41.5 ? with 0.42 ?mol/L SS supplementation(SS),at 41.5 ? with 0.42 ?mol/L SeMet supplementation(SeMet).The results showed that:(1)HS up-regulated(P<0.05)expression of Hsp70 gene,both SS and SeMet supplementation prevented(P<0.05)the up-regulation of Hsp70 by HS,no differences were found between SS and SeMet group;(2)HS down-regulated(P<0.05)expression of Cldn-1 and Zo-1 genes,up-regulated(P<0.05)expression of Ocln gene;both SS and SeMet supplementation alleviated(P<0.05)the down-regulation or up-regulation effect of HS on these genes in different degree;(3)Cell viability was decreased(P<0.05)along with the prolongation of HS;both SS and SeMet supplementation effectively alleviated(P<0.05)the decreases of cell viability by HS,especially in later stages;(4)The ROS content of HS groups were higher in value than CK groups,SeMet supplementation reduced(P<0.05)ROS content while SS supplementation increased ROS content in values than that of HS groups;(5)HS up-regulated(P<0.05)expression of Il-6,Il-8,Icam-1 and Ifn-? genes;SS and SeMet supplementation prevented(P<0.05)the up-regulated of these genes by HS and SeMet exhibited a better prevention effects than that of SS;HS down-regulated(P<0.05)expression of Tgf-? and Tnf-a genes,and SS and SeMet supplementation showed no further effects on these two genes;(6)HS up-regulated(P<0.05)10 selenoprotein genes(Gpx3,Gpx4,Diol,Dio2,Selk,Sels,Self,Sepwl,Sep 15 and Sephs2),down-regulated(P<0.05)5 selenoprotein genes(Gpx1?Txnrd2,Selh,Seli and Selm);SS and SeMet supplementation alleviated the up-or down-regulation of these selenoprotein genes by HS;the mRNA abundance of aboved mentioned selenoprotein genes in SeMet groups were more similar to that of CK cells compared with SS cells;both SS and SeMet supplementation further up-regulated(P<0.05)expression of Gpx4,Diol,Sepwl and Sels genes compared to CK and HS groups;(7)HS up-regulated(P<0.05)protein expression of Hsp70,SeMet supplementation further up-regulated(P<0.05)Hsp70 compared to HS groups;HS down-regulated(P<0.05)protein expression of Cldn-1 and Zo-l and had no effect on Ocln;SS and SeMet supplementation prevented(P<0.05)the down-regulation of Cldn-1 and Zo-1 by HS,the protein abundance of these three proteins in SeMet groups were even higher than that of CK cells;HS down-regulated(P<0.05)protein expression of Gpxl,SS and SeMet supplementation inhibited(P<0.05)the down-regulation of Gpxl by HS;HS showed no effect on expression of SelP,while heat stressed cells supplied with SS and SeMet further up-regulated(P<0.05)SelP and SeMet group exhibited a higher(P<0.05)expression of SelP.Conclusion:(1)HS affected the expression of inflammation-related genes,up-regulate the gene and protein expression of Hsp70,and many selenoprotein genes were up-or down-regulated after IPEC-J2 cells were subject to HS may imply a potential link between selenoproteins functions and the response of cells to HS.(2)Under the low-Se culture condition,HS exhibited negative impacts on the expression of inflammation-related genes,Hsp70 and tight junction-related genes,and the cell viability,also the expression of Hsp70 and three tight junction-related protein were affected.Both SS or SeMet supplementation exhibited protective effects and alleviated the negative impacts of HS on those investigated measurements;(3)SS and SeMet supplementation affected the expression of 25 selenoprotein genes or protein,and their cells protective effects was associates with regulation expression of selenoproteins under hyperthermia;(4)In general,SeMet exhibited a better protective effects than SS in alleviating the HS-induced IPEC-J2 cells injury. |
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