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Yangyang Qi, Matthew G J Trese, Y Li, Paul A Edwards, Hua Gao, Xiaoxi Qiao; Inflammatory Insult Aggravates Hydroxychloroquine-Induced Toxicity on Human Retinal Pigment Epithelial Cells. Invest. Ophthalmol. Vis. Sci. 2014;55(13):4880.
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It is well known that HCQ can cause macular toxicity. This study aims to investigate whether inflammation has any impact on HCQ-induced cellular toxicity on proliferation, viability, and cellular morphology of retinal pigment epithelial cell (RPE) in vitro.
Adult human RPE cells, ARPE19 were exposed to various HCQ concentrations (50 µM to 2.5 mM). In addition, each of the groups were also exposed to a sublethal dosage of an inflammatory cytokine mixture (ICM) containing 10 ng/mL interleukin 1-β, 10 ng/mL tumor necrosis factor α, and 100 units/mL interferon γ. After 24-hour treatment, cell survival and viability were quantified using cell counting and an MTS assay. Cell morphology was assessed using a combination of light microscopy, immunocytochemistry and LysoSenor staining. Finally, terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay was used to detect potential apoptosis.
There was a dose-dependent reduction in both ARPE19 cell number and viability after HCQ exposure. A 30% decrease of cell number was seen at 2 mM HCQ immediately after drug application when compared to the control. Morphological study with light microscopy and immunocytochemistry demonstrated increased cell surface area, increased cytoplasmic vacuolization, increased LysoSensor staining intensity, indicating an elevated lysosome pH, and the presence of a prominent cytoskeletal after HCQ exposure. These toxic effects were significantly worsened when the RPE cells were co-treated with ICM, which resulted in a further reduction of cell number by 50% at 2 mM HCQ plus ICM. Co-treatment with ICM at various doses of HCQ always induced additional reductions of viable cell number and proliferation rate, and progressive increase of the morphologic changes and lysosome pH. TUNEL assay was negative for all treatment groups, indicating either HCQ, ICM alone, or co-treatment does not induce apoptosis.
Co-treatment with HCQ and ICM induced more pathologic changes on human RPE cells than HCQ alone. Inflammation enhances HCQ-induced RPE cellular toxicity. However, this cellular toxicity is not associated with apoptosis.
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