Corneal epithelial proliferation and thickness in a mouse model of dry eye

Abstract

Although several studies have previously focused on the conjunctival epithelial response to surface dryness, little is known about the effect of a dry environment on corneal epithelium, which is the most clinically significant tissue affected in dry eye. The aim of this study was to quantitatively evaluate the effect of desiccating stress on the number of proliferating corneal epithelial cells and corneal epithelial thickness in mice placed in a controlled-environment chamber (CEC) that induces dry eye.Corneal epithelial cell proliferation and thickness were studied in 8-12 week old female BALB/c mice placed in the CEC (temperature: 22.3 ± 0.7°C; relative humidity: 22.5 ± 4.5%; airflow: 15 L/ min) for 7 days and compared to a control group of mice with no dry eye. Actively proliferating cells were identified by immunofluorescence using a FITC-conjugated antibody against the Ki-67 protein, a cell proliferation marker expressed during active phases of the cell cycle. To detect the spatial distribution of proliferative cells, Ki-67 + cells were counted in three areas of the epithelium: center, periphery, and limbus. Corneal epithelial thickness was evaluated in the central cornea after staining with hematoxylin-eosin. Results from each experimental group were compared using the MannWhitney test.The number of Ki-67 + cells observed in the corneal epithelium of mice exposed to the CEC was significantly higher in each area (center: 32.1 ± 1.1; periphery: 94.2 ± 5.3; limbus: 4.0 ± 1.5) than in the control group (center: 13.2 ± 1.0, p = 0.02; periphery: 42.9 ± 2.3, p = 0.02; limbus: 0.0, p = 0.01). In mice subjected to desiccating stress, a significant number of Ki-67 + positive cells were detected in the basal and suprabasal cell layers (central area 46%; periphery 30.8%: limbus 0%), whereas in the control group the cells were exclusively distributed through the basal cell layer. Ki-67 + cells were not found in the corneal stroma or endothelium in any group. The corneal epithelium was found to be significantly thicker in dry eye mice (54.94 ± 6.09 μm) as compared to the controls (43.9 ± 6.23 μm: p < 0.0001) by a mean of 25%.These results demonstrate that desiccating stress increases corneal epithelial turnover and thickness, similar to what is observed in other chronic inflammatory states of other epithelialized surfaces. The CEC can facilitate the study of the regulation of epithelial cell function and turnover at the molecular and cellular levels under desiccating stress conditions.

In Vivo Confocal Microscopy of Meibomian Glands in Sjögren's Syndrome

“…19,20 MGD patients were characterized by extensive MG dropout, MG secretion that could not be expressed, tear film instability, and ocular surface epithelial damage. According to recent literature, [12][13][14] LSCM in these patients showed increased diameters of acinar units and orifices and highreflective secretion.…”

Assessment of Corneal Epithelial Thickness in Dry Eye Patients

“…12 With the benefit of noncontact, accuracy, stability, and repeatability, OCT was widely used in various studies. 28–31 Fabiani et al 9 figured out that the CET became thicker in 7 days after the establishment of DED in a rat model, indicating that epithelial proliferation has a significant impact on the inflammatory process. Kanellopoulos and Asimellis 32 observed augmented epithelial thickness in dry eye patients, whereas Erdelyi et al 33 examined clinical patients and revealed that the CET tends to be thinner in dry eyes, and attributed it to the destruction of stem cells at the limbus.…”

“…7 To figure out the morphological evidence of epithelial damage, various studies have been conducted to map the thickness of corneal epithelium. 8,9 Brush cytology, 10 impression cytology, 11 ultrasound, 12 and in vivo confocal microscopy 13 are classical techniques in assessing the epithelial thickness. However, these methods require direct or indirect contact between the instruments and the patient's ocular surface.…”

Dry Eye Disease

“…Dry eye disease increases epithelial cell density and thickness, decreases epithelial cell size, and increases epithelial cell turnover. 15– 16 Inflammation of the ocular surface is intimately linked to this epithelial dysfunction. The proinflammatory cytokines IL-1 and IFN-γ cause squamous metaplasia of ocular surface epithelial cells, and IFN-γ decreases goblet cell differentiation.…”