Histological analysis

For PAS staining, conjunctival tissue was fixed in 4% paraformaldehyde overnight and embedded in paraffin. Serial sections, 6-μm thick, were cut from each sample. The sections were deparaffinized and stained with the 0.5% PAS for identification of goblet cells. Sections from each group were photographed with a microscope equipped with a digital camera. The number of positively stained goblet cells in the superior and inferior conjunctiva was counted in 3 sections from each eye by using image analysis software (Media Cybernetics, Silver Spring, MD). Data are presented as the average number of goblet cells per millimeter.