The Mechanism Of MiRNA-155 On Apoptosis Of Lymphocytes In Selenium-deficient Broilers

Selenium is an indispensable nutrient element in animals,it participates in multiple biological functions.Selenium mainly involved in neurobiology,muscle metabolism,reproduction,redox reaction,anti-aging,viral infection and other biological functions.Selenium plays an important role in enhancing organism immunity and preventing disease,and mainly emerge its biological function via keep the integrity of the immune cell membrane to maintain the physiological function of the immune system.Selenium deficiency can reduce the activity of immune cells,inhibit the inherent immunity,cellular immunity and humoral immune function.Previous studies have reported that selenium deficiency caused immune tissue damage or immune response abnormalities in human and animals,such as rat,laying hen,pig,and so on.Apoptosis is one of the mechanisms of immunosuppression,and studies have reported that selenium could induce the apoptosis of immune cells.In addition,studies showed that selenium levels could change the expression profile of micro RNAs?mi RNAs?and lead the expressions of related m RNAs also changed,then affected the normal physiological function of cells.Mi RNA-155 is a multifunctional mi RNA,which has the most target genes.Mi RNA-155 plays an important role in immune system,not only participates in the development and differentiation of immune cells,but also affects the function of immune cells.However,the effect of mi RNA-155 on apoptosis in the immune tissue of selenium-deficient broilers is not clear at present.In this study,we analyzed the correlation between mi RNA-155 and apoptosis in immune tissue based on immune tissue apoptosis model of selenium-deficient broiler and the mi RNA-155 over-expression/knock-down model.We clarify the signal transduction pathway of mi RNA-155 in lymphocytes apoptosis of selenium-deficient broiler.The results showed as follws:1. The broilers were fed with low selenium?0.03 mg/kg?diet for 30 days,then we found the immune function was reduced,and the apoptotic phenomenon were appeared by detections of the ultrastructure and histopathological structure.Therefore,immune cells apoptosis model induced by selenium deficiency replicated successfully.2.The results of antioxidant enzyme related genes?Gpx and Mn SOD?,apoptosis related genes?JNK,Bcl-2,Bax,Bak,Cyt-c,caspase 9,caspase 3?,revealed that selenium deficiency induced oxidative stress-related apoptosis in broiler immune tissues.3.The results of mi RNA genomics in spleen tissues,from the 657 entities included on the mi RNAs chip,205 mi RNAs were found to up-regulate and 181 mi RNAs were found to down-regulate by selenium.Mi RNA-155 was showed as one of the down-regulated mi RNAs induced by selenium deficiency.The results of q PCR showed that the expression of mi RNA-155 in the selenium deficient group was reduced to 43%by the control group in broiler spleen tissue.These results suggested selenium deficiency altered the mi RNA profile and down-regulated the expression of mi RNA-155 in spleen tissues.4.The mi RNA-155 nucleic acid chain was successfully transfected into lymphocytes cell by cell transfection.The expression of mi RNA-155 was significantly increased by 7.9 times in over-expression group,it was reduced to about 47%in knock-down group when compared with the control group.The results indicated that the mi RNA-155 over-expression/knock-down of lymphocytes were established successfully.5.Both the recombinant plasmid of TNFRSF1B?wild type and mutant type?and mi RNA-155was co-transfected into lymphocytes.The results of double-luciferase reporter gene assay,q PCR and Western bloting showed that TNFRSF1B is really one of the target genes of mi RNA-155 in broiler lymphocytes.6.The lymphocytes of mi RNA-155 over-expression/knock-down treated by H₂O₂,and detected the indexes AO-EB,Tunnel,antioxidant related genes?Gpx and Mn SOD?,apoptosis related genes?JNK,Bcl-2,Bax,Bak,Cyt-c,caspase 9,caspase 3?,the results showed that mi RNA-155 could regulate the oxidative stress-related apoptosis induced by H₂O₂ through targeting TNFRSF1B in lymphocytes.7.The expressions of Ca²⁺related genes?Ry R1,Ry R3 and SERCA1s?and endoplasmic reticulum stress related genes?GRP78,GRP94,IRE1,e IF2?,ATF6 and caspase 12?were detected in broiler selenium deficient immune tissue,and results showed that selenium deficiency induced the endoplasmic reticulum stress by imbalance of Ca²⁺in broilers immune tissues.The results of experiments in vitro showed that mi RNA-155 effectively alleviated the endoplasmic reticulum stress in lymphocytes.In conclusions,the over-expression of mi RNA-155 could inhibit the occurrence of oxidative stress mediated-apoptosis induced by selenium deficiency via negatively regulating TNFRSF1B.It was also demonstrated that mi RNA-155 could affect endoplasmic reticulum stress and immunity in immune tissues of selenium deficient-broilers.This study laid a foundation for further study of the reaction mechanism between mi RNA-155 and selenium deficiency,and provided a theoretical basis for the prevention and treatment of selenium deficiency immune dysfunction.This work also provided reference for comparative medicine.