Ethambutol-induced optic atrophy: a mitochondrial fission neuropathy mediated by NO?

Abstract

Purpose : Ethambutol (EMB) is used in the treatment of tuberculosis (TB) or mycobacterium avium complex infections. A devastating adverse effect of EMB is toxic optic atrophy, which occurs in 1–2.5% of patients. To date, the pathophysiology of EMB-induced optic atrophy is incompletely understood.

Methods : Human iPSC-derived retinal ganglion cells (iPSC-RGC) or retinal organoids were treated with 2, 20, 200, or 2,000 µg/ml EMB. Time and dose-dependent EMB induced changes were observed by confocal microscopy and gene expression changes by RT-PCR. Mitochondrial function in iPSC-RGC was measured using the Seahorse XFe analyzer.

Results : Treatment of retinal organoids with EMB resulted in a time and dose-dependent loss of RGC, but not other retinal neurons. 24 hours of EMB treatment induced the dose-dependent formation of bubbles in the cytosol of iPSC-derived RGC concordant with increased NOS3 synthesis (P=0.0001). At higher concentrations significantly elevated expression of the apoptotic mediators BAX, APAF1, CASP3, CASP8, and CASP7 was detected (1.5 to 3-fold). We also detected increased expression of MIEF1 (1.8-fold, P<0.0001), suggesting impaired maintenance of mitochondrial DNA. Mitochondrial staining demonstrated decreased mitochondrial length in EMB treated iPSC-RGC (112.1±41.6 µm vs. 212.9±76.3 µm in untreated controls, P<0.0001), indicating mitochondrial fragmentation. Functional mitochondrial measurements demonstrated that spare respiratory capacity in iPSC-RGC is increased by 21%, 20% and 27% treated with 2, 20, or 200 µg/ml EMB, respectively (P<0.05). Furthermore, cells treated with 200 ug/mL EMB displayed a 24% increase in non-mitochondrial respiration (P=0.005).

Conclusions : Our findings suggest that Ethambutol-induced optic atrophy may be caused by nitric oxide stress and dysregulation of mitochondrial dynamics in favor of mitochondrial fission. This, in turn, causes an increase in energy demand that may lead to RGC dysfunction and, eventually, apoptotic cell death.

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