Modulation of TNF-α production by small RNA in dengue virus infected human monocytic cells

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2012-08
Authors
Hammond, Kimberly Elaine
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[Honolulu] : [University of Hawaii at Manoa], [August 2012]
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The immunopathology of dengue virus (DENV) infection is associated with increased TNF-α production. In this study, small RNA-mediated regulation of TNF-α and the effect of TNF-α knockdown during DENV infection were analyzed. This provides insight into the role of TNF-α during DENV infection, both in terms of its contribution to immunopathogenesis and its regulatory mechanism by miRNAs. Utilizing a lentiviral expression system, human monocytic U937 cells that express short hairpin RNAs designed to target TNF-α mRNA were established. TNF-α expression was downregulated in these monocytes, and upon DENV infection they showed decreased endothelial cell activation ability. This demonstrates an overall decrease in the proinflammatory response upon TNF-α knockdown during DENV infection. To analyze the role of microRNAs (miRNAs) in the TNF-α response, miRNAs that potentially target the 3' UTR of TNF-α were predicted. Many of the miRNAs were differentially regulated during DENV infection. miR-320a and miR-592 were among those downregulated, and chosen for further analysis. TNF-α post-transcriptional regulation by miR-320a and miR-592 was confirmed utilizing a TNF-α 3'UTR luciferase reporter. U937 cells transfected with miR-320a and miR-592 mimics followed by DENV infection displayed decreased TNF-α expression and their culture supernatants demonstrated decreased ability to activate endothelial cells. It is concluded that one function of these miRNAs is to negatively regulate TNF-α, and their downregulation contributes to the immunopathogenesis of DENV infection.
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M.S. University of Hawaii at Manoa 2012.
Includes bibliographical references.
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dengue virus
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Theses for the degree of Master of Science (University of Hawaii at Manoa). Microbiology.
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