Corneal protective effects of novel tear substitutes containing sodium hyaluronate and dodecahydrosqualene, squalane, in a porcine dry eye model

Abstract

To develop a novel tear substitute (TS) containing sodium hyaluronate (SH) and dodecahydrosqualene (DHS, squalane), we improved the prescription of a previously developed TS containing saline, 0.5% SH and 1% castor oil (CO), which had corneal protective effects against 60-min desiccation in a porcine dry eye model and viscosity of 106.8 mPa·S. Fresh porcine eyes were treated with a TS containing saline, 0.1%, 0.25%, 0.3% or 0.5% SH, and 1% CO or 1%, 2.5% or 5% DHS, and TS-treated eyes were desiccated for up to 180 min. The corneal damage was evaluated by the staining score of methylene blue (MB), absorbance of MB extracted from the cornea, the staining density of lissamine green (LG) and histopathology. The viscosities of the examined TS were also measured. A saline/0.5% SH/1% DHS solution had corneal protective effects for 90 min under desiccation and a viscosity of 110.0 mPa·s. A TS with saline, 0.1%, 0.25% or 0.3% SH and 1% or 2.5% DHS did not have better protective effects than a saline/0.5% SH/1% DHS solution, although a saline/0.3% SH/5% DHS solution exhibited greater corneal protection against 180-min desiccation on MB and LG staining and histopathological examination, and its viscosity was 34.5 mPa·s, which was similar to the 29.5 mPa·s of 0.3% SH. The saline/0.3% SH/5% DHS solution is available as a novel 3-hr long-lasting TS containing mucinomimetic and liquid oil components to treat and relieve dry eye symptoms in animals and humans.

Keywords: corneal protective effects; dodecahydrosqualene (squalane); porcine dry eye model; sodium hyaluronate; tear substitutes.

Fig. 1.

Typical histopathology of the cornea administered major tear substitutes containing saline, sodium hyaluronate and dodecahydrosqualene after 180-min desiccation using the porcine dry eye model [A; immediately after harvest, B; continuous treatment with saline for 180 min, C; no saline or tear substitute treatment, D; treatment with saline/0.5% sodium hyaluronate (SH)/1% castor oil (CO), E; treatment with saline/0.5% SH/1% dodecahydrosqualene (DHS, squalane), F; treatment with saline/0.3% SH/5% DHS]. There are no histopathological abnormalities in the cornea of the eye immediately after harvest (A). Minimal histopathological alterations are detected on the basal and wing cells in the corneal epithelial layer in the eye continuously treated with saline (B), whereas the untreated eye (no treatment eye) has significant histopathological alterations consisting of marked swelling of basal and wing cells, with cytoplasmic vacuolation and nuclear swelling (C). The abnormalities of the corneal epithelium in the examined eye treated with saline/0.5% SH/1% CO solution are milder than those in the no treatment eye (D). The saline/0.5% SH/1% DHS solution-treated eye has milder histopathological changes than the eye administered saline/0.5% SH/1% CO solution (E). There are minimal histopathological abnormalities in the corneal basal and wing cells, with mild cytoplasmic vacuolation compared with the eye with continuous saline treatment (F). Bar=10 μm.