The Carbonic Anhydrase Inhibitor Dorzolamide Stimulates the Differentiation of Human Meibomian Gland Epithelial Cells

ABSTRACT

Purpose

Clinical studies have indicated that the long-term use of topical antiglaucoma drugs, such as carbonic anhydrase inhibitors (CAIs), may lead to meibomian gland dysfunction (MGD). We hypothesize that these adverse effects involve a direct influence on human MG epithelial cells (HMGECs). The purpose our present investigation was to test our hypothesis and determine whether exposure to dorzolamide, a CAI, impacts the proliferation, intracellular signaling and differentiation of HMGECs.

Materials and Methods

We cultured immortalized (i) HMGECs with vehicle or various concentrations of dorzolamide for 6 days. Cells were enumerated with a hemocytometer, and examined for their morphology, Akt signaling activity, accumulation of neutral lipids, phospholipids and lysosomes, and the expression of protein biomarkers for lipogenesis regulation, lysosomes and autophagosomes.

Results

Our results show that a high, 500 µg/ml concentration of dorzolamide causes a significant decrease in Akt signaling and the proliferation of iHMGECs. However, the high dose of dorzolamide also promotes the differentiation of iHMGECs. This response features increases in the number of lysosomes, the accumulation of phospholipids, and the expression of the light chain 3A biomarker for autophagosomes. In contrast, the therapeutic amount (50 µg/ml) of dorzolamide has no impact on the proliferative or differentiative abilities of iHMGECs.

Conclusions

Our results support our hypothesis and demonstrate that the CAI dorzolamide does exert a direct influence on the proliferation and differentiation of iHMGECs. However, this effect is elicited only by a high, and not a therapeutic, amount of dorzolamide.

Abbreviations: AKT: phosphoinositide 3-kinase-protein kinase B; BPE: bovine pituitary extract; CAD: cationic amphiphilic drug; DED: dry eye disease; DMEM/F12: 1:1 mixture of Dulbecco’s modified Eagle’s medium and Ham’s F-12; EGF: epidermal growth factor; FBS: fetal bovine serum; iHMGECs: immortalized human meibomian gland epithelial cells; KSFM: keratinocyte serum-free medium; LAMP-1: lysosomal-associated membrane protein 1; LC3A: light chain 3A; MGD: meibomian gland dysfunction; SREBP-1: sterol regulatory element-binding protein 1

KEYWORDS:

• Dorzolamide
• carbonic anhydrase inhibitor
• human meibomian gland epithelial cells
• meibomian gland dysfunction
• dry eye disease

Disclosure statement

A provisional patent has been filed concerning phospholipidosis-inducing (PLD) compounds. It is possible that dorzolamine may be a PLD compound. The intellectual property for this application is owned by the Schepens Eye Research Institute of Massachusetts Eye and Ear.

Additional information

Funding

This work was supported by NIH grant EY028653, the Margaret S. Sinon Scholar in Ocular Surface Research Fund, and the China Scholarship Council.