June 2021
Volume 62, Issue 8
Open Access
ARVO Annual Meeting Abstract  |   June 2021
Maturation of retinal pigment epithelium cells in vitro enhances the endogenous antioxidant defense system
Author Affiliations & Notes
  • Jenni J. Hakkarainen
    Research & Development Division, Experimentica Ltd, Kuopio, Finland
  • Olga Vergun
    Research & Development Division, Experimentica Ltd, Kuopio, Finland
  • Anita K. Ghosh
    Research & Development Division, Experimentica Ltd, Kuopio, Finland
    Department of Ophthalmology, Loyola University Chicago, Maywood, Illinois, United States
  • Simon Kaja
    Research & Development Division, Experimentica Ltd, Kuopio, Finland
    Department of Ophthalmology, Loyola University Chicago, Maywood, Illinois, United States
  • Footnotes
    Commercial Relationships   Jenni Hakkarainen, Experimentica Ltd (F), Experimentica Ltd (I), Experimentica Ltd (E), Experimentica Ltd (R), Experimentica Ltd (S); Olga Vergun, Experimentica Ltd (E), Experimentica Ltd (R); Anita Ghosh, Experimentica Ltd (F), Experimentica Ltd (R), Experimentica Ltd (S), eyeNOS Inc. (I), eyeNOS Inc. (P), eyeNOS Inc. (S), K&P Scientific LLC (F), K&P Scientific LLC (C), K&P Scientific LLC (R); Simon Kaja, Experimentica Ltd (F), Experimentica Ltd (I), Experimentica Ltd (P), Experimentica Ltd (R), Experimentica Ltd (S), eyeNOS Inc. (P), K&P Scientific LLC (F), K&P Scientific LLC (I), K&P Scientific LLC (R), K&P Scientific LLC (S)
  • Footnotes
    Support  Experimentica Ltd., Dr. John P. and Therese E. Mulcahy Endowed Professorship in Ophthalmology (SK), Illinois Society for the Prevention of Blindness (AKG).
Investigative Ophthalmology & Visual Science June 2021, Vol.62, 206. doi:
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    • Get Citation

      Jenni J. Hakkarainen, Olga Vergun, Anita K. Ghosh, Simon Kaja; Maturation of retinal pigment epithelium cells in vitro enhances the endogenous antioxidant defense system. Invest. Ophthalmol. Vis. Sci. 2021;62(8):206.

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

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Abstract

Purpose : The aim of the study was to evaluate the endogenous antioxidant defense system in human induced pluripotent stem cell (hiPSC)-derived retinal pigment epithelium (RPE) cells.

Methods : hiPSC-RPE cells were differentiated according to the manufacturer’s instructions for different times in vitro (days in vitro, DIV) on Matrigel®-coated 96-well cell culture plates. Oxidative stress was applied using different concentrations of tert-butyl hydroperoxide (tBHP) for 22 h. Resistance of hiPSC-RPE cells to oxidative stress was measured using the general cell viability assays, resazurin assay and lactate dehydrogenase (LDH) release assay. In addition, production of reactive oxygen species (ROS) in cells was quantified using the ROS indicator, chloromethyl 2',7'-dichlorodihydrofluorescein diacetate (CM-H2DCFDA).

Results : Cell viability as assessed by resazurin assay was similar from DIV13 to DIV19 with IC50 values for tBHP of approx. 0.4 mM. IC50 values started to shift significantly on DIV26 and DIV28, increasing to 0.7 - 0.9 mM. LDH release and CM-H2DCFDA assays confirmed the shift of IC50 value with concomitantly increased levels of LDH release and ROS generation.

Conclusions : hiPSC-RPE cells have a strong endogenous antioxidant defense system, as evident by strong cellular resistance to exogenous oxidative stress insult. Maturation periods longer than 26 days induce sudden, increased resistance against exogenous oxidative stress in hiPSC-RPE cells. Our results necessitate careful and precise characterization of individual hiPSC-RPE cell pools, as rapid changes in the endogenous antioxidant defense pose a significant confounding factor in experimental designs that utilize hiPSC-RPE cells for drug discovery.

This is a 2021 ARVO Annual Meeting abstract.

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