bims-mideyd Biomed News
on Mitochondrial dysfunction in eye diseases
Issue of 2023–08–06
seven papers selected by
Rajalekshmy “Raji” Shyam, Indiana University Bloomington
  1. Hydrogen peroxide-induced oxidative damage and protective role of peroxiredoxin 6 protein via EGFR/ERK signaling pathway in RPE cells.
  2. Ex vivo cultivated retinal pigment epithelial cell transplantation for the treatment of rabbit corneal endothelial dysfunction.
  3. LncRNA NORAD defects deteriorate the formation of age-related macular degeneration.
  4. LOXL1-AS1 lncRNA differentially regulates gene and protein expression, signaling, and morphology of human ocular cells.
  5. Ligand-independent activation of platelet-derived growth factor receptor β promotes vitreous-induced contraction of retinal pigment epithelial cells.
  6. The efficient induction of human retinal ganglion-like cells provides a platform for studying optic neuropathies.
  7. Cardiolipin-mediated temporal response to hydroquinone toxicity in human retinal pigmented epithelial cell line.
  1. Front Aging Neurosci. 2023 ;15 1169211
    Hydrogen peroxide-induced oxidative damage and protective role of peroxiredoxin 6 protein via EGFR/ERK signaling pathway in RPE cells.
    Xiaodong Chen, Radouil Tzekov, Mingyang Su, Yusheng Zhu, Aidong Han, Wensheng Li.
       Introduction: Damage to retinal pigment epithelium (RPE) cells caused by oxidative stress is closely related to the pathogenesis of several blinding retinal diseases, such as age-related macular degeneration (AMD), retinitis pigmentosa, and other inherited retinal degenerative conditions. However, the mechanisms of this process are poorly understood. Hence, the goal of this study was to investigate hydrogen peroxide (H2O2)-induced oxidative damage and protective role of peroxiredoxin 6 (PRDX6) protein via EGFR/ERK signaling pathway in RPE cells.
    Methods: Cells from a human RPE cell line (ARPE-19 cells) were treated with H2O2, and then cell viability was assessed using the methyl thiazolyl tetrazolium assay. Cell death and reactive oxygen species (ROS) were detected by flow cytometry. The levels of PRDX6, epidermal growth factor receptor (EGFR), P38 mitogen-activated protein kinase (P38MAPK), c-Jun N-terminal kinase (JNK), and extracellular signal-regulated kinase (ERK) were detected by Western blot assay. PRDX6 and EGFR were also detected via immunofluorescence staining.
    Results: Our results show that H2O2 inhibited cell viability, induced cell death, and increased ROS levels in ARPE-19 cells. It was also found that H2O2 decreased the levels of PRDX6, EGFR, and phosphorylated ERK but increased the levels of phosphorylated P38MAPK and JNK. PRDX6 overexpression was found to attenuate H2O2-induced inhibition of cell viability and increased cell death and ROS production in ARPE-19 cells. PRDX6 overexpression also increased the expression of EGFR and alleviated the H2O2-induced decrease in EGFR and phosphorylated ERK. Moreover, inhibition of epidermal growth factor-induced EGFR and ERK signaling in oxidative stress was partially blocked by PRDX6 overexpression.
    Discussion: Our findings indicate that PRDX6 overexpression protects RPE cells from oxidative stress damage caused by decreasing ROS production and partially blocking the inhibition of the EGFR/ERK signaling pathway induced by oxidative stress. Therefore, PRDX6 shows promise as a therapeutic target for the prevention of RPE cell damage caused by oxidative stress associated with retinal diseases.
    Keywords:  Peroxiredoxin 6; epidermal growth factor receptor; extracellular signal-regulated kinase; oxidative stress; retinal pigment epithelium cell
    DOI:  https://doi.org/10.3389/fnagi.2023.1169211
  2. Eye Vis (Lond). 2023 Aug 02. 10(1): 34
    Ex vivo cultivated retinal pigment epithelial cell transplantation for the treatment of rabbit corneal endothelial dysfunction.
    Chunxiao Dong, Dulei Zou, Haoyun Duan, Xiangyue Hu, Qingjun Zhou, Weiyun Shi, Zongyi Li.
       OBJECTIVE: Stem cell therapy is a promising strategy for the treatment of corneal endothelial dysfunction, and the need to find functional alternative seed cells of corneal endothelial cells (CECs) is urgent. Here, we determined the feasibility of using the retinal pigment epithelium (RPE) as an equivalent substitute for the treatment of corneal endothelial dysfunction.
    METHODS: RPE cells and CECs in situ were obtained from healthy New Zealand male rabbits, and the similarities and differences between them were analyzed by electron microscopy, immunofluorescent staining, and quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR). Rabbit primary RPE cells and CECs were isolated and cultivated ex vivo, and Na+/K+-ATPase activity and cellular permeability were detected at passage 2. The injection of cultivated rabbit primary RPE cells, CECs and human embryonic stem cell (hESC)-derived RPE cells was performed on rabbits with corneal endothelial dysfunction. Then, the therapeutic effects were evaluated by corneal transparency, central corneal thickness, enzyme linked immunosorbent assay (ELISA), qRT-PCR and immunofluorescent staining.
    RESULTS: The rabbit RPE cells were similar in form to CECs in situ and ex vivo, showing a larger regular hexagonal shape and a lower cell density, with numerous tightly formed cell junctions and hemidesmosomes. Moreover, RPE cells presented a stronger barrier and ionic pumping capacity than CECs. When intracamerally injected into the rabbits, the transplanted primary RPE cells could dissolve corneal edema and decrease corneal thickness, with effects similar to those of CECs. In addition, the transplantation of hESC-derived RPE cells exhibited a similar therapeutic effect and restored corneal transparency and thickness within seven days. qRT-PCR results showed that the expressions of CEC markers, like CD200 and S100A4, increased, and the RPE markers OTX2, BEST1 and MITF significantly decreased in the transplanted RPE cells. Furthermore, we have demonstrated that rabbits transplanted with hESC-derived RPE cells maintained normal corneal thickness and exhibited slight pigmentation in the central cornea one month after surgery. Immunostaining results showed that the HuNu-positive transplanted cells survived and expressed ZO1, ATP1A1 and MITF.
    CONCLUSION: RPE cells and CECs showed high structural and functional similarities in barrier and pump characteristics. Intracameral injection of primary RPE cells and hESC-derived RPE cells can effectively restore rabbit corneal clarity and thickness and maintain normal corneal function. This study is the first to report the effectiveness of RPE cells for corneal endothelial dysfunction, suggesting the feasibility of hESC-derived RPE cells as an equivalent substitute for CECs.
    Keywords:  Cell transplantation; Corneal endothelial dysfunction; Corneal endothelium; Retinal pigment epithelium; Stem cell therapy
    DOI:  https://doi.org/10.1186/s40662-023-00351-4
  3. Aging (Albany NY). 2023 Jul 29. 15
    LncRNA NORAD defects deteriorate the formation of age-related macular degeneration.
    Jinfeng Zhang, Jing Jiang, Hongyu Zhou, Shenjun Li, Weihua Bian, Lifu Hu, Daolai Zhang, Cong Xu, Yeying Sun.
      Long noncoding RNAs (lncRNAs) play important roles in the development of age-related macular degeneration (AMD). However, the effect of long non-coding RNA activated by DNA damage (NORAD) on AMD remains unknown. This study aimed to investigate the effect of NORAD on RPE cell senescence and degeneration. Irradiated adult retinal pigment epithelial cell line-19 (ARPE-19) and sodium iodate-treated mice were used as in vitro and in vivo AMD models. Results showed that irradiation-induced AMD characteristics of ARPE-19 and NORAD-knockdown aggravated cell cycle arrest in the G2/M phase, cell apoptosis and cell senescence along with the increased expression of phosphorylated P53 (p-P53) and P21. AMD factors C3, ICAM-1, APP, APOE, and VEGF-A were also increased by NORAD-knockdown. Moreover, NORAD-knockdown increased irradiation-induced reduction of mitochondrial homeostasis factors, (i.e., TFAM and POLG) and mitochondrial respiratory chain complex genes (i.e., ND1 and ND5) along with mitochondrial reactive oxygen species (ROS). We also identified a strong interaction of NORAD and PGC-1α and sirtuin 1 (SIRT1) in ARPE-19; that is, NORAD knockdown increases the acetylation of PGC-1α. In NORAD knockout mice, NORAD-knockout accelerated the sodium iodate-reduced retinal thickness reduction, function impairment and loss of retinal pigment in the fundus. Therefore, NORAD-knockdown accelerates retinal cell senescence, apoptosis, and AMD markers via PGC-1α acetylation, mitochondrial ROS, and the p-P53-P21signaling pathway, in which NORAD-mediated effect on PGC-1α acetylation might occur through the direct interaction with PGC-1α and SIRT1.
    Keywords:  LncRNA NORAD; PGC-1α; SIRT1; age-related macular degeneration; senescence
    DOI:  https://doi.org/10.18632/aging.204917
  4. Hum Mol Genet. 2023 Aug 04. pii: ddad128. [Epub ahead of print]
    LOXL1-AS1 lncRNA differentially regulates gene and protein expression, signaling, and morphology of human ocular cells.
    Heather M Schmitt, Kristyn M Hake, Kristin M Perkumas, Brandon M Lê, Maria F Suarez, Michele L De Ieso, Rashad S Rahman, William M Johnson, María Gomez-Caraballo, Allison E Ashley-Koch, Michael A Hauser, W Daniel Stamer.
      Pseudoexfoliation glaucoma (PEXG) is characterized by dysregulated extracellular matrix (ECM) homeostasis that disrupts conventional outflow function and increases intraocular pressure (IOP). Prolonged IOP elevation results in optic nerve head damage and vision loss. Uniquely, PEXG is a form of open angle glaucoma that has variable penetrance, is difficult to treat, and does not respond well to common IOP-lowering pharmaceuticals. Therefore, understanding modulators of disease severity will aid in targeted therapies for PEXG. Genome-wide association studies have identified polymorphisms in the long non-coding RNA LOXL1-AS1 as a risk factor for PEXG. Risk alleles, oxidative stress and mechanical stretch all alter LOXL1-AS1 expression. As a long non-coding RNA, LOXL1-AS1 binds hnRNPL and regulates global gene expression. In this study, we focus on the role of LOXL1-AS1 in the ocular cells (trabecular meshwork and Schlemm's canal) that regulate IOP. We show that selective knockdown of LOXL1-AS1 leads to cell type-specific changes in gene expression, ECM homeostasis, signaling, and morphology. These results implicate LOXL1-AS1 as a modulator of cellular homeostasis, altering cell contractility and ECM turnover, both of which are well-known contributors to PEXG. These findings support LOXL1-AS1 as a key target for modifying the disease.
    DOI:  https://doi.org/10.1093/hmg/ddad128
  5. BMC Ophthalmol. 2023 Aug 03. 23(1): 344
    Ligand-independent activation of platelet-derived growth factor receptor β promotes vitreous-induced contraction of retinal pigment epithelial cells.
    Yajian Duan, Wenyi Wu, Jing Cui, Joanne Aiko Matsubara, Andrius Kazlauskas, Gaoen Ma, Xiaorong Li, Hetian Lei.
       BACKGROUND: Epiretinal membranes in patients with proliferative vitreoretinopathy (PVR) consist of extracellular matrix and a number of cell types including retinal pigment epithelial (RPE) cells and fibroblasts, whose contraction causes retinal detachment. In RPE cells depletion of platelet-derived growth factor (PDGF) receptor (PDGFR)β suppresses vitreous-induced Akt activation, whereas in fibroblasts Akt activation through indirect activation of PDGFRα by growth factors outside the PDGF family (non-PDGFs) plays an essential role in experimental PVR. Whether non-PDGFs in the vitreous, however, were also able to activate PDGFRβ in RPE cells remained elusive.
    METHODS: The CRISPR/Cas9 technology was utilized to edit a genomic PDGFRB locus in RPE cells derived from an epiretinal membrane (RPEM) from a patient with PVR, and a retroviral vector was used to express a truncated PDGFRβ short of a PDGF-binding domain in the RPEM cells lacking PDGFRβ. Western blot was employed to analyze expression of PDGFRβ and α-smooth muscle actin, and signaling events (p-PDGFRβ and p-Akt). Cellular assays (proliferation, migration and contraction) were also applied in this study.
    RESULTS: Expression of a truncated PDGFRβ lacking a PDGF-binding domain in the RPEM cells whose PDGFRB gene has been silent using the CRISPR/Cas9 technology restores vitreous-induced Akt activation as well as cell proliferation, epithelial-mesenchymal transition, migration and contraction. In addition, we show that scavenging reactive oxygen species (ROS) with N-acetyl-cysteine and inhibiting Src family kinases (SFKs) with their specific inhibitor SU6656 blunt the vitreous-induced activation of the truncated PDGFRβ and Akt as well as the cellular events related to the PVR pathogenesis. These discoveries suggest that in RPE cells PDGFRβ can be activated indirectly by non-PDGFs in the vitreous via an intracellular pathway of ROS/SFKs to facilitate the development of PVR, thereby providing novel opportunities for PVR therapeutics.
    CONCLUSION: The data shown here will improve our understanding of the mechanism by which PDGFRβ can be activated by non-PDGFs in the vitreous via an intracellular route of ROS/SFKs and provide a conceptual foundation for preventing PVR by inhibiting PDGFRβ transactivation (ligand-independent activation).
    Keywords:  Akt; Contraction; Epithelial-mesenchymal transition; Indirect activation; Migration; PDGFRβ; Proliferation; Retinal pigment epithelial cells; Vitreous
    DOI:  https://doi.org/10.1186/s12886-023-03089-8
  6. Cell Mol Life Sci. 2023 Aug 04. 80(8): 239
    The efficient induction of human retinal ganglion-like cells provides a platform for studying optic neuropathies.
    Roxanne Hsiang-Chi Liou, Shih-Wei Chen, Hui-Chen Cheng, Pei-Chun Wu, Yu-Fen Chang, An-Guor Wang, Ming-Ji Fann, Yu-Hui Wong.
      Retinal ganglion cells (RGCs) are essential for vision perception. In glaucoma and other optic neuropathies, RGCs and their optic axons undergo degenerative change and cell death; this can result in irreversible vision loss. Here we developed a rapid protocol for directly inducing RGC differentiation from human induced pluripotent stem cells (hiPSCs) by the overexpression of ATOH7, BRN3B, and SOX4. The hiPSC-derived RGC-like cells (iRGCs) show robust expression of various RGC-specific markers by whole transcriptome profiling. A functional assessment was also carried out and this demonstrated that these iRGCs display stimulus-induced neuronal activity, as well as spontaneous neuronal activity. Ethambutol (EMB), an effective first-line anti-tuberculosis agent, is known to cause serious visual impairment and irreversible vision loss due to the RGC degeneration in a significant number of treated patients. Using our iRGCs, EMB was found to induce significant dose-dependent and time-dependent increases in cell death and neurite degeneration. Western blot analysis revealed that the expression levels of p62 and LC3-II were upregulated, and further investigations revealed that EMB caused a blockade of lysosome-autophagosome fusion; this indicates that impairment of autophagic flux is one of the adverse effects of that EMB has on iRGCs. In addition, EMB was found to elevate intracellular reactive oxygen species (ROS) levels increasing apoptotic cell death. This could be partially rescued by the co-treatment with the ROS scavenger NAC. Taken together, our findings suggest that this iRGC model, which achieves both high yield and high purity, is suitable for investigating optic neuropathies, as well as being useful when searching for potential drugs for therapeutic treatment and/or disease prevention.
    Keywords:  Direct conversion; Human induced pluripotent stem cells (hiPSCs); Induced retinal ganglion cells (iRGCs); Optic neuropathies
    DOI:  https://doi.org/10.1007/s00018-023-04890-w
  7. Biochim Biophys Acta Mol Cell Res. 2023 Jul 29. pii: S0167-4889(23)00127-1. [Epub ahead of print] 119554
    Cardiolipin-mediated temporal response to hydroquinone toxicity in human retinal pigmented epithelial cell line.
    Magdalena Davidescu, Letizia Mezzasoma, Katia Fettucciari, Luisa Pascucci, Marilena Pariano, Alessandro Di Michele, Oxana Bereshchenko, Carlo Cagini, Barbara Cellini, Lanfranco Corazzi, Ilaria Bellezza, Lara Macchioni.
      Hydroquinone, a potent toxic agent of cigarette smoke, damages retinal pigmented epithelial cells by triggering oxidative stress and mitochondrial dysfunction, two events causally related to the development and progression of retinal diseases. The inner mitochondrial membrane is enriched in cardiolipin, a phospholipid susceptible of oxidative modifications which determine cell-fate decision. Using ARPE-19 cell line as a model of retinal pigmented epithelium, we analyzed the potential involvement of cardiolipin in hydroquinone toxicity. Hydroquinone exposure caused an early concentration-dependent increase in mitochondrial reactive oxygen species, decrease in mitochondrial membrane potential, and rise in the rate of oxygen consumption not accompanied by changes in ATP levels. Despite mitochondrial impairment, cell viability was preserved. Hydroquinone induced cardiolipin translocation to the outer mitochondrial membrane, and an increase in the colocalization of the autophagosome adapter protein LC3 with mitochondria, indicating the induction of protective mitophagy. A prolonged hydroquinone treatment induced pyroptotic cell death by cardiolipin-mediated caspase-1 and gasdermin-D activation. Cardiolipin-specific antioxidants counteracted hydroquinone effects pointing out that cardiolipin can act as a mitochondrial "eat-me signal" or as a pyroptotic cell death trigger. Our results indicate that cardiolipin may act as a timer for the mitophagy to pyroptosis switch and propose cardiolipin-targeting compounds as promising approaches for the treatment of oxidative stress-related retinal diseases.
    Keywords:  Cardiolipin; MTP-131; Mitochondria; Mitophagy; Pyroptosis; SkQ1
    DOI:  https://doi.org/10.1016/j.bbamcr.2023.119554
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