bims-mideyd 2025-08-17 papers

Int J Mol Sci. 2025 Aug 06. pii: 7622. [Epub ahead of print]26(15):

Role of Endogenous Galectin-3 on Cell Biology of Immortalized Retinal Pigment Epithelial Cells In Vitro.

Caspar Liesenhoff, Marlene Hillenmayer, Caroline Havertz, Arie Geerlof, Daniela Hartmann, Siegfried G Priglinger, Claudia S Priglinger, Andreas Ohlmann.

Galectin-3 is a multifunctional protein that is associated with diseases of the chorioretinal interface, in which the retinal pigment epithelium (RPE) plays a central role in disease development and progression. Since galectin-3 can function extracellularly as well as intracellularly via different mechanisms, we developed an immortalized human RPE cell line (ARPE-19) with a knockdown for galectin-3 expression (ARPE-19/LGALS3+/-) using a sgRNA/Cas9 all-in-one expression vector. By Western blot analysis, a reduced galectin-3 expression of approximately 48 to 60% in heterozygous ARPE-19/LGALS3+/- cells was observed when compared to native controls. Furthermore, ARPE-19/LGALS3+/- cells displayed a flattened, elongated phenotype with decreased E-cadherin as well as enhanced N-cadherin and α-smooth muscle actin mRNA expression, indicating an epithelial-mesenchymal transition of the cells. Compared to wildtype controls, ARPE-19/LGALS3+/- cells had significantly reduced metabolic activity to 86% and a substantially decreased proliferation to 73%. Furthermore, an enhanced cell adhesion and a diminished migration of immortalized galectin-3 knockdown RPE cells was observed compared to native ARPE-19 cells. Finally, by Western blot analysis, reduced pAKT, pERK1/2, and β-catenin signaling were detected in ARPE-19/LGALS3+/- cells when compared to wildtype controls. In summary, in RPE cells, endogenous galectin-3 appears to be essential for maintaining the epithelial phenotype as well as cell biological functions such as metabolism, proliferation, or migration, effects that might be mediated via a decreased activity of the AKT, ERK1/2, and β-catenin signaling pathways.

Keywords: AKT signaling; ARPE-19; EMT; ERK signaling; cell attachment; cell proliferation; galectin-3; galectin-3 knockdown; retinal pigment epithelium cells; β-catenin signaling

DOI: https://doi.org/10.3390/ijms26157622

Free Radic Biol Med. 2025 Aug 13. pii: S0891-5849(25)00876-7. [Epub ahead of print]

HDAC Inhibition Protects RPE Cells from Oxidative Stress via Enhanced Mitochondrial Fusion, Cytoskeletal Repair, and Nrf-2 Activation.

Yarine Lugassy, Eva Berent, Lotan Tarnoy, Sandra Jeries, Tamar Ziv, Naphtali Savion, Hagit Eldar-Finkelman.

Oxidative stress is a key driver of retinal pigment epithelium (RPE) damage and the development of age-related macular degeneration (AMD). Here, we demonstrate that the histone deacetylase (HDAC) inhibitors vorinostat and trichostatin A (TSA) elicit a coordinated cytoprotective response in RPE cells exposed to rotenone. Both compounds significantly reduced reactive oxygen species (ROS) levels, enhanced mitochondrial fusion, increased mitochondrial ATP production, and improved cell morphology and cell survival in the rotenone-treated cells. In addition, the compounds activated Nrf-2 as evidenced by Keap1 downregulation, increased p62/SQSTM1 expression, and induction of Nrf-2 targets, including heme oxygenase 1 (HO-1). Proteomic analysis of drug-treated cells revealed a significant enrichment of proteins involved in cytoskeletal organization and dynamics. Consistently, specific staining for actin filaments confirmed that vorinostat and TSA preserved cytoskeletal architecture and increased levels of the tight junction protein TJP3 in cells exposed to rotenone. Finally, inhibition of the vorinostat/TSA target HDAC6, or blockade of α-tubulin acetyltransferase, demonstrated that modulation of α-tubulin acetylation could influence ROS levels. Similarly, enhanced mitochondrial fusion by Mdivi-1 reduced ROS accumulation in the rotenone-treated cells. However, these last two interventions did not fully recapitulate the antioxidant effects observed with vorinostat or TSA. Our results identify a multifaceted protective mechanism triggered by HDAC inhibition in oxidatively stressed RPE cells and support the therapeutic repurposing of vorinostat in oxidative stress-driven RPE or retinal degeneration.

Keywords: Age-related macular degeneration (AMD); Cytoskeleton; HDAC inhibitors; Mitochondrial dynamics; Nrf-2 signaling; Oxidative stress; Proteomics; Retinal pigment epithelium (RPE); Vorinostat

DOI: https://doi.org/10.1016/j.freeradbiomed.2025.08.007

Cell Death Discov. 2025 Aug 14. 11(1): 380

Targeting hypoxia-inducible factor-1 in a hypoxidative stress model protects retinal pigment epithelium cells from cell death and metabolic dysregulation.

Annika Schubert, Maria Eduarda Lobo Barbosa da Silva, Tabea Ambrock, Orbel Terosian, Anna Malyshkina, Claudia Padberg, Safa Larafa, Johann Matschke, Joachim Fandrey, Yoshiyuki Henning.

Oxidative stress and hypoxia lead to dysfunction of retinal pigment epithelium (RPE) cells and are hallmarks of diseases such as age-related macular degeneration (AMD), the most common blinding disease in the elderly population. We have previously shown that a combination of these two risk factors, i.e. hypoxidative stress, exacerbates RPE cell death by ferroptosis. Hypoxia leads to stabilization of hypoxia-inducible factors (HIFs), key regulators of cellular adaptation to hypoxic conditions. In the present study, we have therefore investigated the roles of HIF-1 and HIF-2 in RPE cell death in a human RPE cell line under hypoxidative stress. For this purpose, we conducted siRNA-mediated knockdowns of the α-subunits of HIF-1 and HIF-2. We found that especially iron metabolism, in particular the expression of transferrin receptor 1 (TFR1) was affected by HIF-1α silencing, resulting in decreased intracellular iron levels and ferroptosis susceptibility. We also found that heme oxygenase 1 (HO-1) contributed to cell death by hypoxidative stress. In addition, we also observed that cell metabolism was improved by HIF-1α silencing under hypoxia, most likely contributing to the protective effect. Furthermore, we identified an FDA-approved small molecule inhibitor, Vorinostat, to downregulate HIF-1α, TFR1, and HO-1 and improve cell metabolism, which eventually resulted in a full rescue of RPE cells from hypoxidative stress-induced cell death. In conclusion, this study highlights the importance of considering targeted HIF inhibition as a promising approach to protect RPE cells from degeneration.

DOI: https://doi.org/10.1038/s41420-025-02675-7

Aging Cell. 2025 Aug 10. e70195

Superoxide Activates Ferroptosis via the Haber-Weiss Reaction and Enhances Age-Related Macular Degeneration.

Ying Huang, Zhenxing Zhou, Mengjia Huan, Qi Guo, Xiaoqian Zhang, Ruiqi Lu, Lushu Chen, Xiumiao Li, Jin Yao, Qin Jiang, Yong Xu.

Antioxidant decline is crucial to driving age-related macular degeneration (AMD). Ferroptosis, a regulated cell death mediated by iron-dependent hydroxyl radical-catalyzed phospholipid peroxidation through the Fenton reaction, is implicated in various chronic degenerative diseases. Here, we show that superoxide activates ferroptosis in retinal pigment epithelium (RPE) cells via the Haber-Weiss reaction, thereby contributing to dry AMD. We silenced manganese superoxide dismutase (MnSOD/SOD2) in RPE cells and exposed the cells to blue light to induce ferroptosis by increasing superoxide anions. Additionally, MnSOD deficiency triggered the Hsp70-linked ubiquitin-dependent degradation of GPX4, further aggravating ferroptosis. We validated blue light-induced ferroptosis in the RPE layer as a driver of the dry AMD phenotype in Sod2+/- mice. Consequently, SOD mimetics efficiently protected RPE against phototoxicity by reducing superoxide-activated ferroptosis. Iron chelators or overexpressing GPX4 sufficiently eradicated ferroptosis. The finding reveals that excessive superoxide contributes to phospholipid peroxidation, providing a promising approach for preventing dry AMD by elevating MnSOD to inhibit RPE cell ferroptosis.

Keywords: GPX4; MnSOD; blue light; dry AMD; ferroptosis; hydroxyl radical; superoxide

DOI: https://doi.org/10.1111/acel.70195