To clarify the influence of tumor necrosis factor (TNF)-alpha on fibrotic phenotypes induced by transforming growth factor (TGF)-beta in retinal pigment epithelial cells (RPECs) by epigenetic regulation. Human primary retinal pigment epithelial cells (RPECs including ARPE19) were used in cultures in the presence or absence of TNF-alpha and/or TGF-beta 2. RT2 Profller (TM) (Qiagen) was used for PCR Array for fibrosis and epithelial mesenchymal transition (EMT). Microarray analysis by 3D gene DNA chip was outsourced to Toray Industries Inc. Quantification of histone acetyl transferase (HAT)-related and histone deacetylase (HDAC) related gene expression were also analyzed. HDAC and HAT activity was measured using an EpiQuik HDAC and HAT Activity/Inhibition Assay Kit (Epigentek). CD44, MMP-9, HAT, and HDAC in RPECs were analyzed by western blotting. Analysis of expression of the EMT/fibrosis related CD44 and MMP-9 phenotypes induced by TNF-alpha+TGF-beta 2 revealed four alterations in RPECs: 1) abolition of TGF-beta 2-induced alpha-SMA by TNF-alpha; 2) synergy between TNF-alpha+TGF-beta 2 for induction of CD44 and MMP-9 phenotypes 3) no inhibition of HDAC activity by either TNF-alpha or TGF-beta 2; and 4) significant inhibition of HAT activity by either TNF-alpha or TGF-beta 2, but no synergy. The HDAC activation through HAT inhibition by TNF-alpha+TGF-beta was counteracted by HDAC inhibitors, leading to the inhibition of EMT/fibrosis. EMT/fibrotic CD44 and MMP-9 phenotypes were epigenetically upregulated by concerted action of TNF-alpha and TGF-beta in RPECs. The intervention in epigenetic regulation may hold potential in preventing intraocular proliferative diseases. (C) 2021 Published by Elsevier Inc.