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Zhirong Lin, Tong Zhou, Huan He, Xiaochen Liu, Hui He, Zuguo Liu; A Mouse Model of Limbal Stem Cell Deficiency Induced by Topical Medication of Benzalkonium Chloride. Invest. Ophthalmol. Vis. Sci. 2011;52(14):2018.
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To develop a mouse model of limbal stem cell deficieny induced by topical administration of benzalkonium chloride and investigate the possible mechanisms.
Benzalkonium Chloride (BAC) at concentrations ranging from 0% to 0.5% was applied to the mouse ocular surface for 28 days. Scoring of length of new vessels (NV), corneal inflammatory index and fluorescent staining test were performed to evaluate the toxic effects of BAC on the ocular surface. Global specimens were collected on day (D) 28 and labeled with a series of antibodies including cytokeratin 12 (K12), cytokeratin 19 (K19), connexin 43 (Cx43), P63, ABCG2, EGF-receptor, β-catenin, CD4, cytokeratin 10 (K10), Ki67. In situ TUNEL assay and transmission electron microscopy (TEM) were also applied. TKE2 cells was cultured for MTT assay.
BAC at concentration of 0.5% four times per day successfully induced typical manifestations of limbal stem cell deficiency, including corneal neovascularization, severe stromal inflammation, and diffuse epithelial staining. Conjunctiva-specdific K19 and Cx43 was positive in the corneal surface after BAC treatment. The stem cell associated markers, P63, ABCG2, EGF-receptor, and β-catenin were absent in the limbal basal epithelium. CD4 was present in the corneal stroma. K10 emerged in the corneal and conjunctival surface. TUNEL assay revealed extensive apoptosis in the entire cornea and limbal zone. TEM showed the irregular basement membrane and the loss of stem cell-specific ultrastructure at the limbus. MTT revealed the apparent decrease of proliferative capacity in TKE2 cells after BAC treatment.
Topical administration of 0.5% BAC at high frequency in mouse induces changes resembling that of limbal stem cell deficiency in humans, and thus, represents a novel model of limbal stem cell deficiency.
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