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R. H. Nagaraj, M. Mailankot; Interferon-Gamma Induces Indoleamine 2,3-Dioxygenase and Causes Apoptosis in Lens Epithelial Cells. Invest. Ophthalmol. Vis. Sci. 2010;51(13):2618.
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We investigated the effects of interferon-γ (IFN-γ) on IDO-mediated synthesis of kynurenines and on apoptosis in human lens epithelial cells (HLE-B3).
HLE-B3 cells were cultured in the presence of 1-100 units of IFN-γ for 2 days. The effects on IDO expression and its downstream signaling pathways were investigated using an enzyme activity assay and Western blotting. Intracellular production of kynurenines was assessed by HPLC. Apoptosis was determined by Hoechst staining.
IFN-γ promoted the synthesis of IDO and activated the JAK/STAT signaling pathway in HLE-B3 cells in a dose-dependent manner. Fludarabine, a STAT1-specific inhibitor, blocked IFN-γ mediated IDO expression. Nfk, Kyn and 3OHKyn were detected in cells, with 3OHKyn concentrations 2- to 3-fold higher than other kynurenines. The intracellular production of kynurenines could be completely inhibited by 1-methyl D, L-tryptophan (MT), an inhibitor of IDO. All three kynurenines were detected in the cell culture medium from IFN-γ–treated cells. Kyn- and 3OHKyn-modified proteins were detected in IFN-γ-treated cells. Induction of IDO by IFN-γ caused elevations in hydrogen peroxide levels, caspase-3 activity and apoptosis in HLE-B3 cells. At equimolar concentrations, 3OHKyn caused higher levels of apoptosis than the other kynurenines in HLE-B3 cells. MT and a kynurenine 3-hydroxylase inhibitor (Ro61-8048) effectively inhibited IFN-γ-mediated apoptosis in HLE-B3 cells.
Our results show that induction of IDO by IFN-γ is JAK1/2-STAT1 pathway-dependent, and that this induction causes 3OHKyn-mediated apoptosis in HLE-B3 cells. Our study suggests that IDO-mediated kynurenine formation could play a role in cataract formation related to chronic Inflammation.
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