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Geliang Gan, Shaomin Peng, Veena S. Rao, Ron A. Adelman, Lawrence J. Rizzolo; Synergistic Action of TNFα and IFN on the Tight Junctions of the Human Retinal Pigment Epithelium. Invest. Ophthalmol. Vis. Sci. 2011;52(14):417.
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Chronic, low-grade, subclinical inflammatory processes likely underlie age-related macular degeneration among other ocular diseases. Tumor necrosis factor-alpha (TNFα) and interferon gamma (IFNγ) act synergistically to affect barrier properties in many epithelia, but it is unclear how they affect tight junctions in the retinal pigment epithelium (RPE).
Secondary cultures of human fetal RPE (hfRPE) were incubated for two days in interleukin 1 beta (IL-1β), IFNγ , TNFα, and TNFα + IFNγ. The transepithelial electrical resistance (TER), ion conductance for Na+ and K+, and permeation of methylpolyethylene glycol (mPEG) were used to assess the function of tight junctions. Gene expression of claudins and occludin was examined by quantitative real-time RT-PCR, and protein expression was examined by immunoblotting and confocal, immunofluorescence microscopy. Claudin expression was modulated by siRNA.
Although IL-1β and IFN-γ had small effects on the expression of the minor claudins, only IFN-γ had a small, but variable, effect on TER. TNF-α consistently lowered TER 85% and decreased selectivity between Na+ and K+. TNF-α decreased the expression of claudin-19, the major claudin, and occludin without affecting the expression of their mRNAs. When combined with TNF-α, IFN-γ consistently lowered the TER and increased the permeation of mPEG. This effect was companied by a 5x decrease in the amount of claudin 19 mRNA with little effect on occludin. Although TNFα also increased the expression of claudin-2 in a small subset of cells, this increase could be blocked by siRNA without altering the effect on TER.
TNFα lowered the TER by affecting steady-state protein levels of occludin and claudin-19. IFNγ decreased the expression of claudin-19 mRNA and protein, but only in the presence of TNFα. Besides decreasing TER, this combination increased the permeation of mPEG. Therefore, IFNγ amplifies the effect of TNFα at least in part by regulating the mRNA.
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