Limited Azithromycin Localization to Rabbit Meibomian Glands Revealed by LC-MS-Based Bioanalysis and DESI Imaging

Abstract

Meibomian gland dysfunction (MGD) is the leading cause of dry eye, and although it affects approximately 4% of the population, treatment options remain limited. Topical azithromycin is one of the most promising pharmacological agents because of its multiple mechanisms of action and long sustainability. Azithromycin is frequently used as an off-label medication in the U.S. However, although azithromycin is presumed to act directly on meibomian gland cells, the mechanisms of action that contribute to its clinical efficacy remain unclear because no studies using a pharmacokinetic approach have been performed. Therefore, we aimed to clarify whether topical azithromycin reaches the meibomian glands sufficiently to generate a biological effect. We measured azithromycin concentrations in rabbit meibomian glands collected using a recently developed method. Moreover, we also visualized the azithromycin micro-distribution using desorption electrospray ionization (DESI) imaging. Azithromycin concentration in the meibomian glands reached only 0.8 µg/g tissue following a single application of a 1% azithromycin ophthalmic solution and was 1000-fold lower than the concentration in conjunctival epithelium. Similarly, no signal was observed in the meibomian glands on DESI images. Our results clearly demonstrated that topical azithromycin had limited access to the meibomian glands and was predominantly distributed in ocular surface tissues such as the palpebral conjunctiva and lid margins. These findings provide new insight into the clinical responses to topical azithromycin therapy and will aid in the further development of effective drugs with more suitable pharmacokinetic properties.Key words meibomian gland; azithromycin; distribution; MS imaging; eye drop Meibomian glands are sebaceous glands embedded within specific types of dense connective tissue in the eyelid. They are primarily responsible for producing meibomian oil (meibum), which plays a physiologically important role in promoting tear film stability and preventing aqueous tear evaporation on the ocular surface.1) Meibomian gland blockages are clinically defined as meibomian gland dysfunction (MGD), which is the leading cause of dry eye disease. However, there is no pharmacologically approved drug useful in treating MGD. Azithromycin ophthalmic solution (AzaSite ® ; InSite Vision, a SUN PHARMA company, Alameda, CA, U.S.A.) is one of the most promising candidates for MGD treatment. 2)Although it has been approved only for the treatment of bacterial conjunctivitis, off-label application of topical azithromycin is the most commonly used pharmaceutical treatment in the U.S. for the management of MGD.3) In addition to its broad anti-bacterial spectrum, azithromycin inhibits pro-inflammatory cytokines and restores lipid properties. Indeed, several clinical studies have demonstrated that topical azithromycin ophthalmic solution significantly relieved the clinical signs and symptoms of MGD.4-6) Although significant progress has been made in human clinical studies to better u...

Experimental Models, Induction Protocols, and Measured Parameters in Dry Eye Disease: Focusing on Practical Implications for Experimental Research

“…Approximately, the upper eye lid tarsus contains about 30-40 glands, and the lower eye lid tarsus contains 20-40 MBG glands [21,22]. MBG primarily secretes lipid meibum, which is an oily substance that forms the lipid layer of the tear film and plays a pivotal physiological role in maintaining tear film stability, preventing evaporation of the ocular surface, preventing tear film contamination, and reducing surface tension [23,24]. In addition, lubricating molecules such as mucinous glycoprotein and lubricin synthesized by the MBG have been reported [24].…”

“…The disturbances of MBG trigger abnormalities in the lipid layer of the tear film, leading to evaporative dry eye. This situation is defined as MBG dysfunction (MGD), which is one of the major causes of DED [22,23,25].…”

MALDI imaging mass spectrometry revealed atropine distribution in the ocular tissues and its transit from anterior to posterior regions in the whole-eye of rabbit after topical administration

“…Since then, its use has rapidly expanded and its application optimized [2,3]. IMS has been used in pharmaceutical research [4,5], drug discovery [6], observation of endogenous molecules (proteins [7], lipids [8], peptides [9], and metabolites [10]), and histopathology [11]. IMS has also been used to analyze the distribution of topically administered benzalkonium chloride in rabbit eye [12].…”