Transcriptomic Profiling Reveals A Role For TREM-1 Activation In Enterovirus D68 Infection-induced Proinflammatory Responses

Enterovirus D68(EV-D68)was first isolated from California in 1962,a highly pathogenic virus,which could cause severe respiratory and neurological diseases.There were few reports about the pathogenicity of EV-D68 before 2011.In 2014,there were outbreaks of EV-D68 infection in multiple states across the United States,1035 people in 47 states and the District of Columbia had reported respiratory diseases until midOctober.Recently,there have been reports of acute flaccid myelitis(AFM)caused by EV-D68 infection in mainland China,Europe and other places.These cases have caused public concern about the pathogenicity of EV-D68.Unfortunately,there is no effective drugs or vaccine against EV-D68 infection,and the molecular mechanism of its infection and pathogenicity is still unclear.This study applied RNA-seq to analyze the transcript profile of rhabdomyosarcoma(RD)cells infected with EV-D68 to reveal the gene regulatory networkbetween the host and EV-D68.First,the results of RNA-seqand subsequent bioinformatic analysis revealed that EV-D68 reshape the transcriptional genes regulatory network of RD cells,including activating antiviral immune responses,interfering with the normal operation of the cell cycle,and affecting cell differentiation and apoptosis.Importantly,triggering receptor expressed on myeloid cells 1(TREM-1)was involved in highly activated TREM-1 signaling processes as an important mediator in EV-D68 infection,related to up-regulated interleukin 8(IL-8),interleukin 6(IL-6),and tumor necrosis factor alpha(TNF-α)expression.TREM1 is a member of the immunoglobulin superfamily of receptors,and it is reportedly involved in systemic inflammatory responses through its ability to amplify activation of host defense signaling pathways.Next,real-time quantitative PCR and ELISA experiments demonstrated that stimulation of RD and HEK293 T cells with EV-D68 up-regulated TREM-1 and its downstream inflammatory cytokinesproduction.Subsequently,inhibition of TREM-1 in EV-D68-infected RDs with a specific peptide LP17 lowered cytokine production,which suggest that TREM-1 is involved in TREM-1 downstream cytokine expression.Notably,LP17 affects the replication of EV-D68 in RD cells by targeting TREM-1,and reduces the apoptosis and death caused by EV-D68 infection.Further analysis showed that NF-κB p65 was essential for EV-D68-induced TREM-1expression.Knockdown of NF-κB p65 significantly reduce EV-D68-induced TREM-1production.A dual-luciferase assay demonstrated that NF-κB p65 binds to promoter region of the endogenous TREM-1 gene and regulates TREM-1 expression at the transcriptional level.Additionally,inhibition of the TREM1 signaling pathway with the specific inhibitor LP17 specificallydampened activation of the p38 MAPK signaling cascade,which suggest TREM-1 mainly transmits activation signals to phosphorylate p38 mitogen-activated protein kinase(MAPK).The study found that TREM-1 is involved in inflammatory response in EV-D68 infected RD cells,providing new insights for the molecular mechanism of EV-D68,and provide new clues and possible molecular targets for the therapeutic intervention of enterovirus diseases.