H₂o₂-Induced Oxidative Stress Modulates Methotrexate and Dexamethasone-Dependent Regulation of Nlrp3-Associated Transcription and Hla Class I Expression in Primary Human Rpe Cells Publisher Pubmed

Summary: Do anti-inflammatory drugs reshape retinal immunity? Lab tests found methotrexate and dexamethasone altered RPE gene signals, without matching cytokine changes. #Retina #Immunology

Souri Z ; Malekahmadi M
Source: European Journal of Pharmaceutical Sciences Published:2026

Abstract

Purpose: Oxidative stress–driven inflammation is a central mechanism in retinal diseases. This study investigated how methotrexate (MTX) and dexamethasone (DEX), alone or in combination, modulate NLRP3-related gene expression and HLA class I expression in primary human retinal pigment epithelial (RPE) cells under basal and oxidative stress conditions. Methods: Primary human RPE cells were isolated from cadaveric donor eyes and treated with MTX (100 µg/mL), DEX (50 µg/mL), or their combination for 24 h under normal culture conditions or following H₂O₂-induced oxidative stress (100 µM). mRNA expression of NLRP3, PYCARD, CASPASE-1, HLA-A, and HLA-B was quantified by qPCR. ELISA measured IL-1β and IL-18 secretion. Results: Under basal conditions, MTX and DEX downregulated NLRP3 and HLA-A/B mRNA expression while increasing PYCARD and CASPASE-1 transcripts. Combination treatment reduced IL-1β secretion, whereas MTX and combination therapy decreased IL-18 levels. In oxidative stress conditions, MTX and combination therapy upregulated NLRP3 and CASPASE-1 mRNA, while HLA-A/B and PYCARD transcripts were primarily increased by combination treatment. IL-1β levels remained unchanged, and IL-18 was reduced only in the combination group. Conclusions: MTX and DEX modulate NLRP3-associated and HLA class I transcriptional responses in RPE cells in a redox-dependent manner. Although oxidative stress modulated drug-associated gene expression profiles, these transcriptional changes were not paralleled by corresponding alterations in secreted inflammatory cytokine levels, suggesting that the context-dependent nature of inflammatory signaling and cytokine release in RPE cells should be taken into consideration. © 2026 The Author(s). Published by Elsevier B.V. This is an open access article under the CC BY-NC-ND license. http://creativecommons.org/licenses/by-nc-nd/4.0/