2. Academic Validation
  3. The Mitochondrial-Derived Peptide Humanin Protects RPE Cells From Oxidative Stress, Senescence, and Mitochondrial Dysfunction

The Mitochondrial-Derived Peptide Humanin Protects RPE Cells From Oxidative Stress, Senescence, and Mitochondrial Dysfunction

  • Invest Ophthalmol Vis Sci. 2016 Mar;57(3):1238-53. doi: 10.1167/iovs.15-17053.
Parameswaran G Sreekumar 1,  Keijiro Ishikawa 1,  Chris Spee 2,  Hemal H Mehta 3,  Junxiang Wan 3,  Kelvin Yen 3,  Pinchas Cohen 3,  Ram Kannan 1,  David R Hinton 4
Affiliations

Affiliations

  • 1 Arnold and Mabel Beckman Macular Research Center, Doheny Eye Institute, Los Angeles, California, United States.
  • 2 Department of Ophthalmology, University of Southern California, Los Angeles, California, United States.
  • 3 USC Leonard Davis School of Gerontology, University of Southern California, Los Angeles, California, United States.
  • 4 Department of Ophthalmology, University of Southern California, Los Angeles, California, United States 4Department of Pathology, University of Southern California, Los Angeles, California, United States.
PMID: 26990160 DOI: 10.1167/iovs.15-17053
Abstract

Purpose: To investigate the expression of humanin (HN) in human retinal pigment epithelial (hRPE) cells and its effect on oxidative stress-induced cell death, mitochondrial bioenergetics, and senescence.

Methods: Humanin localization in RPE cells and polarized RPE monolayers was assessed by confocal microscopy. Human RPE cells were treated with 150 μM tert-Butyl hydroperoxide (tBH) in the absence/presence of HN (0.5-10 μg/mL) for 24 hours. Mitochondrial respiration was measured by XF96 analyzer. Retinal pigment epithelial cell death and Caspase-3 activation, mitochondrial biogenesis and senescence were analyzed by TUNEL, immunoblot analysis, mitochondrial DNA copy number, SA-β-Gal staining, and p16INK4a expression and HN levels by ELISA. Oxidative stress-induced changes in transepithelial resistance were studied in RPE monolayers with and without HN cotreatment.

Results: A prominent localization of HN was found in the cytoplasmic and mitochondrial compartments of hRPE. Humanin cotreatment inhibited tBH-induced Reactive Oxygen Species formation and significantly restored mitochondrial bioenergetics in hRPE cells. Exogenous HN was taken up by RPE and colocalized with mitochondria. The oxidative stress-induced decrease in mitochondrial bioenergetics was prevented by HN cotreatment. Humanin treatment increased mitochondrial DNA copy number and upregulated mitochondrial transcription factor A, a key biogenesis regulator protein. Humanin protected RPE cells from oxidative stress-induced cell death by STAT3 phosphorylation and inhibiting Caspase-3 activation. Humanin treatment inhibited oxidant-induced senescence. Polarized RPE demonstrated elevated cellular HN and increased resistance to cell death.

Conclusions: Humanin protected RPE cells against oxidative stress-induced cell death and restored mitochondrial function. Our data suggest a potential role for HN therapy in the prevention of retinal degeneration, including AMD.

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