June 2013
Volume 54, Issue 15
Free
ARVO Annual Meeting Abstract  |   June 2013
The M33 gene of murine cytomegalovirus (MCMV) is involved in the stimulation of VEGF-A production by mouse macrophages
Author Affiliations & Notes
  • Moon Han
    Viral Immunology Center, Georgia State University, Atlanta, GA
  • Hsin Chien
    Viral Immunology Center, Georgia State University, Atlanta, GA
  • Hamed Laroui
    Center for Inflammation, Immunity, and Infection, Georgia State University, Atlanta, GA
  • Didier Merlin
    Center for Inflammation, Immunity, and Infection, Georgia State University, Atlanta, GA
  • Scott Cousins
    Ophthalmology, Duke University, Durham, NC
  • Richard Dix
    Viral Immunology Center, Georgia State University, Atlanta, GA
    Ophthalmology, Emory University School of Medicine, Atlanta, GA
  • Footnotes
    Commercial Relationships Moon Han, None; Hsin Chien, None; Hamed Laroui, None; Didier Merlin, None; Scott Cousins, Alcon (F), Alcon (C), Heidelberg Engineering (C), Narrow River (C), Nordic Biotech (C), PanOptica (C), Pfizer (C), Salutaris Medical Devices (C), Sanofi-Fovea (C), Valeant Ophthalmics (C), Imagen Biotech (I); Richard Dix, None
  • Footnotes
    Support None
Investigative Ophthalmology & Visual Science June 2013, Vol.54, 129. doi:
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      Moon Han, Hsin Chien, Hamed Laroui, Didier Merlin, Scott Cousins, Richard Dix; The M33 gene of murine cytomegalovirus (MCMV) is involved in the stimulation of VEGF-A production by mouse macrophages. Invest. Ophthalmol. Vis. Sci. 2013;54(15):129.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose: Our laboratory recently demonstrated that chronic MCMV infection of healthy C57BL/6 mice results in a more severe experimental choroidal neovascularization (CNV) (PLoS Pathogens 8:e1002671, 2012). Because the human cytomegalovirus (HCMV)-encoded chemokine receptor US28 has been shown to play a multifunctional role in angiogenesis, we hypothesized that the M33 gene of MCMV (the MCMV homologue to HCMV US28) plays a similar role in experimental CNV. The purpose of this study is to test the hypothesis that the M33 gene of MCMV is involved in the stimulation of VEGF production by mouse macrophages.

Methods: Monolayers of IC-21 mouse macrophages were treated for 24 hours with either M33 siRNAs (single or combined M33-1 siRNA and M33-2 siRNA) or scrambled siRNAs (control) using either HiPerFect transfection reagent (Qiagen) or Lipofectamine RNAiMAX transfection reagent (Invitrogen) and inoculated with MCMV (moi=3 pfu/cell). All siRNA-treated and control MCMV-infected monolayers were harvested at 24 and 48 hours post-infection and subjected to western blot analysis for the comparison of VEGF-A protein production.

Results: Although no significant differences were observed when comparing relative band densities of VEGF-A production by siRNA-treated MCMV-infected macrophages at 24-hour post-infection, treatment using a combination of M33-1/M33-2 siRNAs resulted in a decrease in VEGF-A protein production by ~51%. Use of Lipofectamine RNAiMAX transfection reagent was superior to HiPerFect transfection reagent in reducing VEGF-A protein production by the M33 siRNAs.

Conclusions: The M33 gene of MCMV is involved in the stimulation of VEGF-A production by IC-21 mouse macrophages, but the downregulation of M33 gene by siRNAs did not result in a greater reduction in VEGF-A protein production as expected. We therefore postulate that one or more genes in addition to the M33 gene of MCMV contribute to the stimulation of VEGF-A production by mouse macrophages.

Keywords: 492 cytomegalovirus • 748 vascular endothelial growth factor • 412 age-related macular degeneration  
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