June 2022
Volume 63, Issue 7
Open Access
ARVO Annual Meeting Abstract  |   June 2022
Hybrid molecule SA-2 improve both mitochondrial respiration and glycolysis in primary human trabecular meshwork cells
Author Affiliations & Notes
  • Charles Enyaah Amankwa
    Pharmacology and Neuroscience, University of North Texas Health Science Center, Fort Worth, Texas, United States
  • Biddut Debnath
    Pharmacology and Neuroscience, University of North Texas Health Science Center, Fort Worth, Texas, United States
  • Sudershan Reddy Gondi
    The University of Texas Southwestern Medical Center, Dallas, Texas, United States
  • Dorota Luiza Stankowska
    Pharmacology and Neuroscience, University of North Texas Health Science Center, Fort Worth, Texas, United States
  • Suchismita Acharya
    Pharmacology and Neuroscience, University of North Texas Health Science Center, Fort Worth, Texas, United States
  • Footnotes
    Commercial Relationships   Charles Amankwa None; Biddut Debnath None; Sudershan Gondi None; Dorota Stankowska None; Suchismita Acharya None
  • Footnotes
    Support  NIH/NEI R01EY029823
Investigative Ophthalmology & Visual Science June 2022, Vol.63, 3296 – A0396. doi:
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      Charles Enyaah Amankwa, Biddut Debnath, Sudershan Reddy Gondi, Dorota Luiza Stankowska, Suchismita Acharya; Hybrid molecule SA-2 improve both mitochondrial respiration and glycolysis in primary human trabecular meshwork cells. Invest. Ophthalmol. Vis. Sci. 2022;63(7):3296 – A0396.

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

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Abstract

Purpose : Oxidative stress (OS) caused by hypoxia/hyperoxia environment results in progressive loss of trabecular meshwork (TM) cells in primary open angle glaucoma (POAG). Our previous report demonstrated; a hybrid nitric oxide (NO) donor-antioxidant molecule SA-2 protect primary human (h) TM cells against t-butyl hydrogen peroxide (TBHP) induced cell death and increased superoxide dismutase enzyme level. Here we investigated the effect of SA-2 on mitochondrial energy metabolism by measuring the respiration status, glycolysis rate and energy production.

Methods : Primary hTM cells obtained from human donor eyes (IRB approved at UNTHSC) were seeded in 24-well culture plates (Seahorse XFe 24 Cell Mito Stress kit, Agilent), and starved for 24h before treatment with SA-2 (1 µM,10µM,100µM, and 1mM). In a separate experiment, the cells were pretreated with TBHP (150µM) for 30 minutes, followed by the addition of SA-2 (10µM,100µM). After 24h, the mitochondrial complex inhibitors and uncoupling reagents (oligomycin, FCCP, rotenone/antimycin A) were added. The plate was analyzed for changes in oxygen consumption rate (OCR) and extracellular acidification rate (ECAR) using the Seahorse XFe24 analyzer following the manufacturer instructions.

Results : The mean OCR was significantly decreased (>70%) followed by increase in the mean ECAR (~3-fold) after treatment with TBHP compared to oligo/FCCP/rot treated cells, hereafter called as negative control. Treatment with SA-2 at 1 µM,10µM,100µM and 1mM concentrations increased both oligomycin/FCCP induced decrease in ATP production and maximal mitochondrial respiration followed by an increase in the mean ECAR compared to negative control. The mean OCR was higher in SA-2 (100µM) +TBHP treated cells followed by an increase in ECAR in SA-2 (10µM or 100µM) +TBHP treated cells than TBHP and negative control treated cells. N =2-3.

Conclusions : Mitochondrial respiration was impaired after TBHP treatment to hTM cells following cell death. While most of the mitochondrial targeting anti-oxidant compounds increase OCR but not ECAR, we found the hybrid NO donor-anti-oxidant compound SA-2 increases ATP production, maximal mitochondrial respiration and increases glycolytic energy production in hTM cells. This finding provides a novel direction for further investigation into the effect of SA-2 and mitochondrial bioenergetics during OS-induced cell death.

This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.

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