Protective Effect Of Sodium Selenite On DON Induced GPX1 Overexpressing Spleen Lymphocytes In Pigs

In this study,the primary porcine splenic lymphocytes were cultured in vitro,then the GPX1 overexpressing splenic lymphocyte model was established.Then the DON and sodium selenite?Na₂SeO₃?were added alone or combined to explore the antagonistic effect of DON on the oxidative damage of GPX1 lymphocytes and the oxidative damage of DON to GPX1 overexpressing spleen lymphocytes in pigs.And the role of GPX1 in it.Methods:?1?Establishment of GPX1 overexpressing spleen lymphocytes from pigs:construct pEGFP-GPx1 plasmid and transfer it into cells,and identify it by qRT-PCR and Western Blot?WB?technology.?2?Detection of antioxidant indexes:The experiment was divided into 12 groups,namely D1-D4 four-fold concentration gradient group?DON 0.1025 ug/mL-0.820ug/mL?,S?Na₂SeO₃ 2 umol/L?,SD1-SD4 four-fold concentration gradient group?Na₂SeO₃ 2 umol/L+DON 0.1025 ug/mL-0.820 ug/mL?,P?normal cells?,C?overexpression cells?and K?empty carrier cells?,respectively.Antioxidant index and free radical content of GPX1 overexpressing spleen lymphocytes in pigs at 12,24 and 48h.Results:?1?Establishment of GPX1 overexpressing splenic lymphocyte in pigs:in this experiment,the expression of GPX1 gene was detected by qRT-PCR,and the expression of GPX1 mRNA in the overexpressing group was 3.876 times as high as that in the control group.The WB test of GPX1 showed that the expression of GPx1 protein in overexpression group was 1.570 times higher than that in control group,showing significant difference?P<0.05?.These results indicated that GPX1 overexpressing spleen lymphocytes of pigs were successfully established.?2?Detection of antioxidant markers:GPX1 overexpressing splenic lymphocytes?group C?showed no significant changes at all time points?P>0.05?compared with normal pig spleen lymphocytes?group P?and pig spleen lymphocytes?K group?transferred into empty carriers.DON alone?group D?in GPX1 overexpressing splenic lymphocytes,after cultured for 12,24,48 h,ROS,MDA and H₂O₂ contents in D group showed an upward trend.The activities of SOD,CAT and GSH-Px,the content of GSH,T-AOC and the ability to inhibit hydroxyl radicals showed a downward trend.Comparing with each other,with the increase of DON concentration,ROS,MDA and H₂O₂ content showed an upward trend,SOD,CAT and GSH-Px activity,GSH content,T-AOC and inhibition of hydroxyl radicals showed a downward trend at each time point.Compared with group C,the contents of ROS,MDA and H₂O₂ decreased significantly?P<0.05?,the activities of SOD,CAT and GSH-Px,the contents of GSH,T-AOC and the ability to inhibit hydroxyl radicals increased significantly?P<0.05?in group S treated with Na₂SeO₃ alone at 12,24 and 48 hours.DON and Na₂SeO₃?SD group?were added at the same time.At 12,24 and 48 hours,compared with the corresponding D group,ROS,MDA and H₂O₂ contents in SD group cells showed a downward trend.SOD,CAT and GSH-Px activities,GSH content,T-AOC and inhibition of hydroxyl radicals showed an upward trend.Conclusions:?1?Over expression of GPX1 gene,the antioxidant capacity of pig spleen lymphocytes increased.?2?DON can significantly induce the oxidative damage of spleen lymphocytes in pigs,and with the increase of concentration,the more serious the damage is,the concentration dependent effect.Overexpression of GPX1 reduced oxidative damage,suggesting that GPX1 could affect the toxicity of DON to cells.?3?Sodium selenite acts on GPX1 overexpressing spleen lymphocytes,which will alleviate the oxidative damage of cells.It shows that sodium selenite exerts an antioxidant effect in GPX1 overexpressing spleen lymphocytes,and it can also effectively antagonize the oxidative damage of DON to the cells.It fully demonstrates that sodium selenite can antagonize the oxidative damage and toxicity of DON.In combination with point?2?,GPX1 plays an important role in selenium antagonizing DON toxicity.