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A. E. Buckner, C. O. Ekworomadu, R. D. Dix; Nicotine Treatment Stimulates NF-B Activity in Both Human Cytomegalovirus and Herpes Simplex Virus Type 1-Infected ARPE-19 Cells. Invest. Ophthalmol. Vis. Sci. 2007;48(13):4734.
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© ARVO (1962-2015); The Authors (2016-present)
Although human cytomegalovirus (HCMV) infection of ARPE-19 cells results in dampening of NF-ΚB activity, nicotine treatment stimulates NF-ΚB activity thereby favoring productive, lytic HCMV replication. We have performed additional studies to test the hypothesis that nicotine-induced stimulation of NF-ΚB activity is not unique to HCMV-infected ARPE-19 cells, but also takes place during herpes simplex virus type 1 (HSV-1) infection of nicotine-treated ARPE-19 cells even though these human herpesviruses exhibit different patterns of virus-host cell interaction.
Monolayers of uninfected ARPE-19 cells were transfected with cationic lipid-mediated NF-ΚB luciferase DNA, treated with nicotine (100 nM) for 1 hr, and nicotine-treated for an additional 4 hrs following inoculation with HCMV [Towne] or HSV-1 [H129] (moi = 1 PFU/cell). Monolayers of untreated cells, mock-infected cells, or cells inoculated with UV-inactivated virus served as controls. All monolayers were lysed, and cell extracts were compared by luminometer analysis for NF-ΚB luciferase activity. Cell extracts were also subjected to western blot analysis for detection of p65 and p50 NF-ΚB subunits and ELISA for determination of VEGF protein levels.
As expected, untreated HCMV-infected ARPE-19 cells showed decreased NF-ΚB expression, but, in contrast, untreated HSV-1-infected ARPE-19 cells showed increased NF-ΚB expression. Nevertheless, both HCMV and HSV-1 exhibited sharp increases in NF-ΚB expression in nicotine-treated cells, although the level of NF-ΚB luciferase activity in HSV-1- infected ARPE-19 cells was ~3-fold higher when compared to HCMV-infected ARPE-19 cells. These findings were confirmed by western blot analysis using antibody to NF-ΚB p65 and p50 subunits. VEGF levels were also elevated in nicotine-treated HCMV and HSV-1-infected cells and at equivalent levels of expression.
Although our findings suggest that HCMV and HSV-1 utilize distinct transcriptional regulatory pathways in ARPE-19 cells, nicotine treatment stimulates NF-ΚB expression in both HCMV and HSV-1-infected ARPE-19 cells. Nicotine treatment of HCMV or HSV-1 infected ARPE-19 cells also upregulates VEGF production possibly in concert with IL-1ß. These findings provide further evidence that smoking of nicotine-containing tobacco products serves as a cofactor to stimulate productive virus replication and may contribute to onset and/or progression of various retinal diseases associated with HCMV or HSV-1 infection.
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