bims-mideyd 2022-05-15 papers

Bioengineered. 2022 May;13(5): 11767-11781

Recombinant human klotho protects against hydrogen peroxide-mediated injury in human retinal pigment epithelial cells via the PI3K/Akt-Nrf2/HO-1 signaling pathway.

Xuewei Wen, Song Li, Yanfei Zhang, Liang Zhu, Xiaoting Xi, Shuyuan Zhang, Yan Li.

Globally, age-related macular degeneration (AMD) is a common irreversible ophthalmopathy. Oxidative stress of retinal pigment epithelial cells is involved in AMD occurrence and development. Klotho is an anti-aging protein with antioxidant properties. We investigated the protective properties of Klotho on hydrogen peroxide (H2O2)-induced injury of retinal pigment epithelial cells (ARPE-19 cells) and its associated pathomechanisms. We found that Klotho pretreatment for 24 h could up-regulate Bcl-2 levels, decrease the cleaved-caspase-3 and Bax levels, inhibit H2O2-induced ARPE-19 cell apoptosis, and promote cell proliferation. Klotho pretreatment inhibited the H2O2-mediated elevations of reactive oxygen species (ROS) in ARPE-19 cells. It enhanced antioxidant activities of the cells and restored the glutathione peroxidase (GPX), superoxide dismutase (SOD2), catalase (CAT), as well as malondialdehyde (MDA) levels to close to the normal level. N-acetylcysteine (NAC), a reactive oxygen scavenger, could reverse the harmful effects of H2O2 on proliferation, apoptosis, and oxidative stress of ARPE-19 cells. Further, Klotho pretreatment enhanced Akt phosphorylation and expression as well as nuclear translocation of Nrf2 in H2O2-treated ARPE-19 cells. This indicates that Klotho protects cells from oxidative stress by activating phosphatidylinositol 3 kinase (PI3K)/protein kinase B (Akt)-nuclear factor E2-related factor 2 (Nrf2)/heme oxygenase 1 (HO-1) signaling pathway. Klotho is, therefore, a potential preventive or treatment option for AMD.

Keywords: Klotho; PI3K/akt-nrf2/HO-1 signaling pathway; apoptosis; oxidative stress

DOI: https://doi.org/10.1080/21655979.2022.2071023

FASEB J. 2022 May;36 Suppl 1

Mitochondrial Transplantation as a Novel Therapeutic for Retinal Metabolic Dysfunction-Driven Disease.

Erik R Butcher, Daisy Y Shu, Scott Frank, Magali Saint-Geniez.

Metabolic dysfunction and mitochondria defect are implicated in several age-associated diseases including age-related macular degeneration (AMD), a leading cause of blindness in the elderly. In AMD, mitochondrial oxidative stress in the retinal pigment epithelium (RPE) drives disease progression and growth of atrophic lesions. Mitochondria release and uptake has been recently identified as a novel mechanism of intercellular communication but its implication in ocular diseases such as AMD has never been investigated. Here, we examined the role of mitochondrial transfer as a new mechanism of metabolic crosstalk between RPE cells. Diseased mitochondria were purified from RPE cells treated with the AMD-associated cytokine TNFα (10 ng/mL), administered to host RPE cells and the effects of MitoTNFA compared to MitoCtrl , isolated from control RPE, or exposure to exogenous TNFα. We showed that treatment of healthy RPE with MitoTNFA , and not MitoCtrl , triggers mitochondrial network fragmentation and transcriptional upregulation of inflammatory factors RelB, IL6, IL8, and repression of PGC1α, mirroring the effect of direct TNFα treatment. ELISA assay confirmed that the effects observed were not caused by presence of soluble TNFα within the mitochondrial fraction. Metabolic profiling using the Seahorse XFe96 bioanalyzer further validated the ability of MitoTNFA to phenocopy TNFα-induced metabolic reprograming in RPE. Finally, we demonstrated that transfer of MitoCtrl improved the bioenergetic functions of both healthy and diseased RPE with significant increases in basal, maximal, and spare respiratory functions. Our results showed that mitochondria transfer recapitulates the context-specific phenotype from donor to host cells in RPE. This new paradigm in RPE biology may not only explain the centrifugal expansion of RPE lesions in AMD but also presents a promising therapeutic avenue for mitochondria-driven disorders such as AMD.

DOI: https://doi.org/10.1096/fasebj.2022.36.S1.R2669

Front Cell Dev Biol. 2022 ;10 878395

Mitochondrial ROS in Slc4a11 KO Corneal Endothelial Cells Lead to ER Stress.

Rajalekshmy Shyam, Diego G Ogando, Joseph A Bonanno.

Recent studies from Slc4a11 -/- mice have identified glutamine-induced mitochondrial dysfunction as a significant contributor toward oxidative stress, impaired lysosomal function, aberrant autophagy, and cell death in this Congenital Hereditary Endothelial Dystrophy (CHED) model. Because lysosomes are derived from endoplasmic reticulum (ER)-Golgi, we asked whether ER function is affected by mitochondrial ROS in Slc4a11 KO corneal endothelial cells. In mouse Slc4a11 -/- corneal endothelial tissue, we observed the presence of dilated ER and elevated expression of ER stress markers BIP and CHOP. Slc4a11 KO mouse corneal endothelial cells incubated with glutamine showed increased aggresome formation, BIP and GADD153, as well as reduced ER Ca2+ release as compared to WT. Induction of mitoROS by ETC inhibition also led to ER stress in WT cells. Treatment with the mitochondrial ROS quencher MitoQ, restored ER Ca2+ release and relieved ER stress markers in Slc4a11 KO cells in vitro. Systemic MitoQ also reduced BIP expression in Slc4a11 KO endothelium. We conclude that mitochondrial ROS can induce ER stress in corneal endothelial cells.

Keywords: ERAD (ER associated protein degradation); MitoQ; ROS—reactive oxygen species; SLC4A11 ammonia transporter; corneal endothelial cells; er stress

DOI: https://doi.org/10.3389/fcell.2022.878395

FASEB J. 2022 May;36 Suppl 1

PP2A and LRP-1 targeted protection of human retinal pigment epithelial cells.

Micah T Starghill, Malia Batiste, Samantha Bowden, Ian Davenport, Partha Bhattacharjee.

Retinal pigment epithelial (RPE) cell disorders contribute to the leading causes of vision loss in the United States. Diabetic retinopathy and Age-related macular degeneration are RPE disorders that develop as a consequence to the accumulation of toxic retinal lipids. However, the mechanism of how RPE regulates lipid homeostasis in response to intra-retinal dysfunction of lipid metabolism is unknown. The purpose of this project is to provide new proof of principle information, toward the understanding, of how RPE regulates oxidative stress induced alteration of intracellular phosphatase/kinase signaling leading to retinal disorders. Through confocal immunofluorescence imaging, transepithelial resistance (TEER) measurement and Western Blot analysis of protein expression of primary human RPE(hRPE) cells are investigated after the treatment of lipid peroxidation products. Protein phosphatase 2A (PP2A), appears to have a pertinent role in the regulation of kinases (p-Akt and p-ERK ½) associated to RPE loss and TEER maintenance in response to oxidative stress induced by lipid peroxidation products. Altered PP2A signaling was found to be modulated by low density lipoprotein related receptor protein-1 (LRP-1) targeted peptidomimetic treatment. This project also provides potential clinical application to the treatment of retinal degeneration associated with intraretinal lipid homeostasis using a patented peptidomimetic targeting LRP-1 and PP2A signaling.

DOI: https://doi.org/10.1096/fasebj.2022.36.S1.00R51