title: MicroRNA-221 modulates RSV replication in human bronchial epithelium by targeting NGF expression.
abstract: BACKGROUND
  Early-life infection by respiratory syncytial virus (RSV) is associated with aberrant expression of the prototypical neurotrophin nerve growth factor (NGF) and its cognate receptors in human bronchial epithelium. However, the chain of events leading to this outcome, and its functional implications for the progression of the viral infection, has not been elucidated. This study sought to test the hypothesis that RSV infection modulates neurotrophic pathways in human airways by silencing the expression of specific microRNAs (miRNAs), and that this effect favors viral growth by interfering with programmed death of infected cells.
  
  
  METHODS
  Human bronchial epithelial cells infected with green fluorescent protein-expressing RSV (rgRSV) were screened with multiplex qPCR arrays, and miRNAs significantly affected by the virus were analyzed for homology with mRNAs encoding neurotrophic factors or receptors. Mimic sequences of selected miRNAs were transfected into non-infected bronchial cells to confirm the role of each of them in regulating neurotrophins expression at the gene and protein level, and to study their influence on cell cycle and viral replication.
  
  
  RESULTS
  RSV caused downregulation of 24 miRNAs and upregulation of 2 (p<0.01). Homology analysis of microarray data revealed that 6 of those miRNAs exhibited a high degree of complementarity to NGF and/or one of its cognate receptors TrKA and p75(NTR). Among the selected miRNAs, miR-221 was significantly downregulated by RSV and its transfection in bronchial epithelial cells maximally inhibited gene and protein expression of NGF and TrKA, increased apoptotic cell death, and reduced viral replication and infectivity.
  
  
  CONCLUSIONS
  Our data suggest that RSV upregulates the NGF-TrKA axis in human airways by silencing miR-221 expression, and this favors viral replication by interfering with the apoptotic death of infected cells. Consequently, the targeted delivery of exogenous miRNAs to the airways may provide a new strategy for future antiviral therapies based on RNA interference.
authors.affiliation: Department of Pediatrics and Pediatric Research Institute, West Virginia University School of Medicine, Morgantown, West Virginia, United States of America.
authors.forename: Sreekumar
authors.initials: S
authors.lastname: Othumpangat
authors.forename: Cheryl
authors.initials: C
authors.lastname: Walton
authors.forename: Giovanni
authors.initials: G
authors.lastname: Piedimonte
mesh_terms: D000293:Adolescent; D000328:Adult; D000368:Aged; D017209:Apoptosis / Q000235:genetics; D001980:Bronchi / Q000166:cytology; D002478:Cells, Cultured; D004847:Epithelial Cells / Q000378:metabolism* / Q000821:virology; D005260:Female; D020869:Gene Expression Profiling; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D054884:Host-Pathogen Interactions; D006801:Humans; D008297:Male; D035683:MicroRNAs / Q000235:genetics*; D018613:Microscopy, Confocal; D008875:Middle Aged; D020932:Nerve Growth Factor / Q000235:genetics* / Q000378:metabolism; D020411:Oligonucleotide Array Sequence Analysis; D020917:Receptor, trkA / Q000235:genetics / Q000378:metabolism; D012136:Respiratory Syncytial Viruses / Q000235:genetics* / Q000378:metabolism / Q000502:physiology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection; D014779:Virus Replication / Q000235:genetics; D055815:Young Adult
chemical_list: D049452:Green Fluorescent Proteins; D035683:MicroRNAs; D020932:Nerve Growth Factor; D020917:Receptor, trkA; C525709:MIR221, human