June 2021
Volume 62, Issue 8
Open Access
ARVO Annual Meeting Abstract  |   June 2021
Autophagy inhibition-induced alterations of energy metabolism and fluorescence lifetime of the retinal pigment epithelium ex vivo
Author Affiliations & Notes
  • Richard Foerster
    Istitute of biomedical optics, University of Lübeck, Lübeck, Schleswig-Holstein, Germany
    Department of Opthalmology, University of Lübeck, Lübeck, Schleswig-Holstein, Germany
  • Erdal Tan Ishizuka
    Istitute of biomedical optics, University of Lübeck, Lübeck, Schleswig-Holstein, Germany
  • Kensuke Shima
    Department of Infectious Diseases and Microbiology, University of Lübeck, Lübeck, Schleswig-Holstein, Germany
  • Britta Lewke
    Istitute of biomedical optics, University of Lübeck, Lübeck, Schleswig-Holstein, Germany
  • Svenja Rebecca Sonntag
    Department of Opthalmology, University of Lübeck, Lübeck, Schleswig-Holstein, Germany
  • Jan Rupp
    Department of Infectious Diseases and Microbiology, University of Lübeck, Lübeck, Schleswig-Holstein, Germany
  • Ralf Brinkmann
    Istitute of biomedical optics, University of Lübeck, Lübeck, Schleswig-Holstein, Germany
    Medical Laser Center Lübeck, Lübeck, Schleswig-Holstein, Germany
  • Salvatore Grisanti
    Department of Opthalmology, University of Lübeck, Lübeck, Schleswig-Holstein, Germany
  • Yoko Miura
    Istitute of biomedical optics, University of Lübeck, Lübeck, Schleswig-Holstein, Germany
    Department of Opthalmology, University of Lübeck, Lübeck, Schleswig-Holstein, Germany
  • Footnotes
    Commercial Relationships   Richard Foerster, None; Erdal Tan Ishizuka, None; Kensuke Shima, None; Britta Lewke, None; Svenja Sonntag, None; Jan Rupp, None; Ralf Brinkmann, None; Salvatore Grisanti, None; Yoko Miura, None
  • Footnotes
    Support  Eyenovative Förderpreis 2018
Investigative Ophthalmology & Visual Science June 2021, Vol.62, 2219. doi:
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      Richard Foerster, Erdal Tan Ishizuka, Kensuke Shima, Britta Lewke, Svenja Rebecca Sonntag, Jan Rupp, Ralf Brinkmann, Salvatore Grisanti, Yoko Miura; Autophagy inhibition-induced alterations of energy metabolism and fluorescence lifetime of the retinal pigment epithelium ex vivo. Invest. Ophthalmol. Vis. Sci. 2021;62(8):2219.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose : Reduced autophagy function of retinal pigment epithelial (RPE) cells is suggested as one of the key factors in the pathogenesis of degenerative chorioretinal disorders, such as age-related macular degeneration. The purpose of this study is to investigate the energy metabolism of ex vivo RPE under the inhibitory condition of autophagy, and its correlation with its fluorescence lifetime (FLT) using fluorescence lifetime imaging ophthalmoscopy (FLIO).

Methods : Porcine ex-vivo RPE/choroid-explants were exposed to the autophagy inhibitor Bafilomycin A1 (1x10-8 M) for 24 h. The viability of the RPE cells was examined with a calcein test. The inhibitory effect of Bafilomycin A1 on autophagy was verified by measuring intracellular autophagosomes. To understand the influence of the autophagy inhibition on the energy metabolism of the RPE, the oxygen consumption rate (OCR) and extracellular acidification rate (ECAR) were measured by a Seahorse-XF-Analyzer after 24 h of Bafilomycin A1 incubation. The FLT of the RPE was examined by FLIO (excitation: 473 nm, detection in the short spectral channel (SSC): 498-560 nm and in the long spectral channel (LSC): 560-700 nm). In order to enable the FLIO on the tissues placed in a culture plate with culture medium, the horizontal stage and a 45-degree-angled mirror were built in front of the FLIO camera.

Results : Incubation of the explants with 1x10-8 M Bafilomycin A1 for 24 h did not cause increased cell death compared with the non-treated control group, whereas Bafilomycin A1 was proven to inhibit autophagy. The OCR of the treated tissue was significantly reduced (e.g., basal respiration from 220 pmol/min ± 43 pmol/min to 147 pmol/min ± 62 pmol/min, p<0.05). The ECAR, an indicator of the glycolytic activity was not significantly altered. In FLIO, Bafilomycin A1 significantly extended the FLT (mean FLT: τm) of the RPE by about 350 ps in both detection channels.

Conclusions : The results of the metabolic analysis clearly showed that the autophagy inhibition may induce mitochondrial dysfunction in RPE cells. In this study, it was experimentally demonstrated that FLIO may indicate alteration of mitochondrial function in RPE cells and could be utilized to assess it in clinical practice.

This is a 2021 ARVO Annual Meeting abstract.

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