Non-invasive redox assessment of mitochondria at the optic nerve head predicting retinal ganglion cell loss

Abstract

Purpose : Eye diseases are currently diagnosed by visualizing often irreversible structural changes which occur late in disease progression, such as retinal ganglion cell (RGC) loss in glaucoma. Early clinical detection of mitochondrial dysfunction could allow intervention before irreversible damage to prevent permanent visual loss. We investigated the value of non-invasive real-time mitochondria and hemoglobin redox state assessment in the mouse eye with elevated intraocular pressure (IOP) for early diseases detection using resonance Raman spectroscopy (RRS)

Methods : We developed a retinal imaging system with an integrated Raman spectrometer to assess mitochondrial and hemoglobin redox states in the mouse eye in vivo. A wavelength-specific library of eye structures (cornea, lens, aqueous humor and vitreous) and porphyrin containing molecules (hemoglobin and mitochondrial cytochromes) was generated to allow regression-based quantification of each component. The anterior chamber of the mouse eye was canulated and the IOP was altered by changing the height of a connected fluid reservoir. The IOP was elevated for 90 min during which the redox states were non-invasively assessed at the optic nerve head rim. The eyes were enucleated 7 days after IOP elevation for immunohistochemical evaluation of RGC loss.

Results : IOP of 100 mmHg resulted in a higher reduced mitochondrial fraction (RMF), and a lower oxygenated hemoglobin fraction (StO₂) consistent with retinal ischemia in a real-time, reversible, and repeatable manner. Retinal ischemia at 100 mmHg was validated by visible retinal blanching on fundoscopy. IOP elevation to 60 mmHg achieved a wide range of RGC degeneration and a significantly higher RMF, lower StO₂ and reduced total hemoglobin signal (rhemo) versus baseline. The RMF value positively correlated with the degree of later RGC loss (7 days post-IOP elevation) (r=0.71, p=0.015 n=11) and the StO₂ value correlated negatively with degree of later RGC loss at day 7 aft IOP elevation (n= 11, r= -0.62, p=0.042).

Conclusions : This technology has the potential to map the metabolic status of eye tissue in vivo before irreversible damage occurs to help guide early intervention based on improving mitochondrial function to prevent permanent visual loss.

This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.

 

In vivo resonance Raman redox state assessment of mitochondrial cytochromes and hemoglobin