Study Of TLR7Expression Regulated By NRF2/ARE Pathway Under Oxidative Stress Induced By Respiratory Syncytial Virus Infection

Objective To investigate the effect of Toll-like receptor7(TLR7) expressionregulated by Nrf2/ARE pathway under oxidative stress induced by respiratory syncytialvirus infected human lung epithelial cells (A549) in vitro, and its related effect in theTLR7and the main impact of signaling molecules. Accordingly,this study has veryimportant medical significance in the precaution and treatment of RSV infection.Methods A549cells infected with RSV in vitro were pretreated with or withoutInhibitors and agonist of Nrf2, cells and cellular supernatants were harvested at theselected time points after RSV infection (4,8,12and24, respectively). The uninfectedA549cells were as normal controls. The concentrations of NO, hydroxy radical (OH)in cellular supernatants were measured according to the kit reference. The expression ofTLR7, TNF-α, IL-6gene mRNA were evaluated semiquantitatively by RT-PCR. Theexpression levels of TLR7, Nrf2and HO-1protein were detected by Western blot. Theconcentration of TNF alpha and IL-6in cell supernatant was detected by ELISA. TheMTT assays were used to detect the cell proliferation.Results①The group of infection The levels of NO and OH increasedgradually. The RSV infection could markedly up-regulate the expression of TLR7,TNF-α and IL-6mRNA in a time-dependent manner. The cell supernatant concentrationof TNF-α and IL-6increased gradually. The expression levels of TLR7, Nrf2and HO-1protein gone up. It was found that RSV infection could up-regulate the growthinhibition of cell in a time-dependent manner.②The group of TRI The levels of NO and OH were higher than in RSV infectedcells. The expression of TLR7, TNF-α and IL-6mRNA could up-regulate markedly in a time-dependent manner and these were higher than the group of infection. The cellsupernatant concentration of TNF-α and IL-6were higher than in RSV infected cells.The expression levels of TLR7protein was higer than that in the infected cells, but theNrf2and HO-1protein were lower. The growth inhibition of cell ascented less markedlythan that in RSV-infected group.③The group of BHA The levels of NO and OH were lower than in RSVinfected cells. The expression of TLR7, TNF-α and IL-6mRNA could up-regulate in atime-dependent manner but these were lower than the group of infection. The cellsupernatant concentration of TNF-α and IL-6were lower than that in the RSV infectedcells. The expression levels of TLR7protein was lower than the infected cells, but theNrf2and HO-1protein were higher than that in RSV infected cells. The growthinhibition of cell ascented more markedly than that in RSV-infected group.Conclusions: RSV infected A549cells in vitro can induce oxidative stress, RSVinfection could up-regulate the expression of TLR7and its major downstream signalmolecules. Pretreatment with specific agonist BHA to Nrf2, down-regulate theactivation of TLR7and its major downstream signal molecules. Pretreatment withspecific inhibitor Trigonelline to Nrf2, significantly up-regulate the expression of TLR7and its major downstream signal molecules. Pretreatment oxidative stress with specificinhibitor or agonist, respectively, the expression of TLR7and its major downstreamsignal molecules show significant changes. It illustrated that the effect of expression ofTLR7is probably regulated by oxidative stress induced by RSV infection in A549cellsthrough the Nrf2/ARE Pathway.