Study On The Mechanism Of Selenium-deficiency Or/and Bisphenol A Exposure Induce Pyroptosis Via ROS/NLRP3 Pathway In Broiler Spleen

Selenium（Se）is an essential trace element in animal nutrition,which plays an important role in maintaining redox homeostasis and enhancing immune function.Selenium deficiency can cause damage to various tissues and organs,including immune organs.Bisphenol A（BPA）is chemical widely used in modern agriculture and industry.It has been proved to have immu-notoxicity.The spleen,as an important peripheral immune organ of the body,is also a crucial target organ selenium deficiency and BPA.Pyroptosis is a programmed cell death that plays an important role in inflammation or immunity.However,there is no report on the occurence mechanism of pyroptosis in the process of spleen injury caused by the combination of selenium deficiency and BPA in poultry.Therefore,this study aim to explore the molecular mechanism of spleen pyroptosis co-induced by selenium deficiency and BPA.In vivo experiment,60healthy 1-day-old AA broilers were randomly divided into 4 groups（n=15）:control goup,BPA group,Se D group and Se D+BPA group.In vitro,the Malik’s disease splenic lymphoma cell line MDCC-MSB1（MSB1）was used as research subjects.Cells were treated with selenium deficient medium or/and 100μM BPA for 24 h.Finally,intervened with 8m M NAC（ROS scavenger）.H&E staining,ICP-MS,ELISA,CCK-8 detection,immunofluorescence,q RT-PCR and Western blot and so on were used to detect the pathological changes of spleen,selenium content,BPA content,cell viability,the level of oxidative stress and pyroptosis.The main results of this study are as follows:（1）The histopathological observation results of spleen showed that the control group had a normal and complete structure,with clear boundaries between the red and white pulp.Com-pared with the Control group,the boun dary between the red and white pulp in the BPA group is blurry,and a small amount of inflammatory infiltration cells can be seen in the red pulp.In Se D group,only a small amount of inflammatory cell infiltration can be seen.The pathological changes in the Se D+BPA group were significantly aggravated,manifested as blurred bounda-ries between red and white pulp,infiltration of inflammatory cells in the red pulp,a decrease in the number of lymphocytes in the white pulp,and degeneration and necrosis occur.This indicates that selenium deficiency and BPA co-treatment can exacerbate spleen damage.（2）The detection results of selenium content in spleen showed that the selenium content in the Se D and Se D+BPA group was significantly lower than that of the control group and BPA group（P<0.05）.Moreover,the selenium content in Se D+BPA group was significantly lower than that of the Se D group（P<0.05）,indicating that BPA exposure can reduce the absorption to selenium of spleen.The detection results of BPA content of spleen showed that the BPA content in the BPA and Se D+BPA group was obviously higher than that in the control and Se D group（P<0.05）.Additionly,the BPA content in the Se D+BPA group was significantly higher than that in the BPA group（P<0.05）,indicating that selenium deficiency increased the accu-mulation of BPA in the spleen.（3）Selenium deficiency or/and BPA exposure significantly reduced the activities T-AOC,SOD,CAT and GSH-Px,and increased the levels of MDA,H ₂O₂ and ROS in spleen and MSB1cells（P<0.05）,enhanced the fluorescence signals of NLRP3 and GSDMD,markedly upregu-lated the expression levels of pyroptosis-related genes（NLRP3,ASC,Caspase-1,IL-1β,IL-18and GSDMD）（P<0.05）,and the changes of the above indexes are more obvious after selenium deficiency and BPA co-treatment（P<0.05）,which suggesting that selenium deficiency or/and BPA can induce oxidative stress and pyroptosis.（5）ROS scavenger NAC significantly alleviated oxidative stress and pyroptosis induced by the combination of selenium deficiency and BPA（P<0.05）,which indicating that oxidative stress mediated pyroptosis induced by selenium deficiency and BPA.To sum up,selenium deficiency or/BPA exposure can induce pyroptosis via activating ROS/NLRP3 pathway.This study clarified for the first time the molecular mechanism of brolier spleen injury induced by selenium deficiency or/and BPA exposure,enrich ing the theoretical research on selenium deficiency induced immune damage and the immunotoxicology of BPA in poultry,and provided environmental decision-making basis for healthy livestock and poultry breeding.