Molecular mechanism of ocular surface damage: application to an in vitro dry eye model on human corneal epithelium.
OBJECTIVE
The present study was concerned with the development of a new experimental model of dry eye using human reconstructed in vitro corneal epithelium (HCE). The model is based on the use of adapted culture conditions that induce relevant modifications at the cellular and molecular level thus mimicking dry eye.


METHODS
The HCE model was maintained in a controlled environmental setting (relative humidity <40% and 40 °C temperature) for 24 h and up to 72 h to induce dry eye. The evolution of the dry eye condition was assessed by histology, immunohistochemistry staining, scanning electron microscopy, and gene expression by using TaqMan gene assay technology (mucin-4 [MUC4], matrix metallopeptidase-9 [MMP9], tumor necrosis factor-α [TNF-α], and defensin β-2 [DEFB2). The effects of different commercially available tear substitutes on the induced dry eye condition were tested.


RESULTS
This in vitro dry eye HCE model, that was well established within 24 h, has the characteristic features of a dry eye epithelium and could be satisfactorily used for preliminary assessment of the protective activity of some artificial tears. The transcriptional study of selected biomarkers showed an increase in MUC4, MMP9, TNF-α, and hBD-2 (DEFB2) gene expression.


CONCLUSIONS
By using a dynamic approach, we were able to define a biomarker gene signature of dry eye-induced effects that could be predictive of corneal damage in vivo and to discriminate the efficacy among different commercial artificial tears.
Marisa
M
Meloni
Barbara
B
De Servi
Daniela
D
Marasco
Salvatore
S
Del Prete
D015415:Biomarkers / Q000378:metabolism; D002470:Cell Survival; D003470:Culture Media; D015352:Dry Eye Syndromes / Q000378:metabolism / Q000473:pathology*; D019573:Epithelium, Corneal / Q000473:pathology*; D005786:Gene Expression Regulation; D006801:Humans; D007150:Immunohistochemistry / Q000379:methods; D020780:Matrix Metalloproteinase 9 / Q000378:metabolism; D008855:Microscopy, Electron, Scanning / Q000379:methods; D008856:Microscopy, Fluorescence / Q000379:methods; D010078:Oxazines / Q000494:pharmacology; D013997:Time Factors; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism; D014966:Xanthenes / Q000494:pharmacology; D023083:beta-Defensins / Q000378:metabolism
D015415:Biomarkers; D003470:Culture Media; C106641:DEFB4A protein, human; D010078:Oxazines; D014409:Tumor Necrosis Factor-alpha; D014966:Xanthenes; D023083:beta-Defensins; C005843:resazurin; D020780:Matrix Metalloproteinase 9
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