Keeping an Eye on Molecular Imaging: Assessment of Drug Toxicity in Small Ocular Structure using Mass Spectrometry Imaging

Abstract

 

Purpose

 

Mass spectrometry Imaging (MSI) applications to ophthalmic drug discovery have recently gained growing interest especially for preclinical studies in pharmacology or toxicology. In our study, MSI was applied to assess the distribution of Benzalkonium chloride (BAK) compound (antiglaucoma eye drops preservative) in specific areas of the eye after instillation in animal model and human tissues. They have been reported to cause ocular surface disorders with tear film alteration, eye irritation and to promote dry eye. The distribution of BAK compoundwas investigated in small specific histological regions of the eye (such as iridocorneal angle or sclera, choroid, retina regions) in order to estimate efficiency of action or adverse effects of the treatment.

 

Methods

 

The eyes of New Zealand rabbits were instilled with different BAK solutions at 0.01%, 2 drops BID for 1 month or 5 months or 0.2% at 1 drop/day for 1 month. After sacrifice, eyes were quickly enucleated and embedded in tragacanth gum and frozen at -80°C. Serial cryosections were deposited on glass slides for Hematoxylin-eosin staining or immunohistochemistry (IHC) and conductive glass slide for mass spectrometry imaging.

 

Results

 

High spatial resolution images were performed at cells level (30 µm). Local Drug concentration differences were observed according to histological area and position on the eye section (anterior, posterior, temporal or nasal side).The figure below shows an optical image of H&E staining of rabbit eye 1 month after start of treatment (left) and the distribution of the two benzalkonium ions (Red & Green images, right). MSI and IHC results were put side by side to correlate inflammatory areas (degradation of CSE or apoptosis phenomena within cornea/conjonctiva region) with BAK localization. Moreover, a high accumulation of BAKs were observed at the sclerocorneal junction and near trabecular meshwork involved in aqueous humor outflow. Moreover, differential analysis was carried out to find disease state biomarkers of each ocular structure.

 

Conclusions

 

MSI offers new insight in ocular therapeutic/pharmaceutical research, especially to give a better understanding of the drug candidate migration from the front to the back of the eye to assist drug efficiency or toxicity studies for specific tissue targeting eye diseases.