Pathology of Ocular Irritation with Acetone, Cyclohexanol, Parafl uoroaniline, and Formaldehyde in the Rabbit Low-Volume Eye Test

Abstract

The ocular irritation responses to 11 different surfactants and two concentration s of acetic acid and sodium hydroxide have been shown to depend on the extent of initial injury, despite marked differences in the processes leading to tissue damage. The purpose of these studies was to determine the extent to which this fundamenta l relationship applies to other nonsurfactants . Ten l l of acetone (ACT), cyclohexano l (CY), para uoroaniline (PF), or 37% formaldehyd e (FA) was directly applied to the cornea of the right eye of each rabbit. Eyes and eyelids were macroscopicall y scored for signs of irritation beginning 3 hours after dosing and periodically until recovery or 35 days. Tissues were obtained for light microscopic examination after 3 hours and on days 1, 3, and 35. Initial corneal injury was characterized quantitatively at 3 hours and 1 day using in vivo confocal microscopy (CM) and by postmortem quantitation of dead corneal epithelial cells and keratocytes using a Live-Dead Assay (L/D, Molecular Probes) and scanning laser CM. Corneal changes over time were characterized quantitatively using in vivo CM performed at 3 hours and 1, 3, 7, 14, and 35 days. The changes with ACT were consistent with mild irritation. Corneal injury was limited to the epithelium and super cial stroma, with the mean normalized depth of injury (NDI) being less than 10% with the majority of regions showing no stromal injury. Changes with CY and PF were consistent with moderate to severe irritation, and FA caused severe irritation. Speci cally, corneal injury by CY and PF tended to involve the epithelium and anterior stroma, with the mean NDI being 10.4% to 23.8%, while injury with FA involved the epithelium, deep stroma, and at times the endothelium. Interestingly, with FA signi cantly less injury was observed at 3 hours with a dramatic increase in injury observed at 1 day and thereafter. In conclusion, these results continue to support and extend our hypothesis that ocular irritation is principally de ned by the extent of initial injury despite clear differences in the means by which irritants cause tissue damage. We believe this approach can be applied to developing alternative assays based on injury to ex vivo eyes or injury to an in vitro corneal equivalent system.

Pathology of Ocular Irritation with Bleaching Agents in the Rabbit Low-Volume Eye Test

“…Using light microscopy, we have characterized the general ocular irritancy of a broad sampling of surfactants and nonsurfactants (acid, alkali, ketone, alcohol, aromatic amine, aldehyde) causing slight to severe irritation (36)(37)(38)(39)(40). Additionally, using in vivo CM, we have characterized quantitatively the corneal changes occurring with these irritants (27-29, 32, 35, 36, 42).…”

“…These bleaching materials are oxidizing agents and their mechanism of injury is different from that of the surfactants and nonsurfactants (acid, alkali, ketone, alcohol, aromatic amine and aldehyde) previously studied (27,29,32,(35)(36)(37)(38)(39)(40)42).…”

“…A lack of success in efforts to validate alternative tests to replace the use of animals for ocular irritation testing (5,7,51) has led us to propose that a better understanding of the critical cellular and molecular changes occurring in in vivo tests is needed (27,(35)(36)(37)(38)(39)42). This opinion has been separately supported by groups of ophthalmologists and ophthalmologic scientists advising the International Life Sciences Institute's Technical Committee on Alternatives to Animal Testing and the European Cosmetics, Toiletries and Perfume Association (COLIPA) (8,51).…”

In vitro eye irritation testing using the open source reconstructed hemicornea – a ring trial_suppl

“…After chemical exposure of these modified models, the epithelium is stripped off the stroma, and cell viability is analyzed separately for each tissue. This procedure takes into account the observation that certain chemicals are known to damage epithelium and stroma differently (Jester et al, 1998a(Jester et al, ,b, 2001(Jester et al, , 2006(Jester et al, , 2010Maurer et al, 2001Maurer et al, , 2002.…”

Role of Nigella sativa seed oil on corneal injury induced by formaldehyde in adult male albino rats

“…Like other epithelia, the epithelium of the cornea is maintained by stem cells. It has no blood supply, unmyelinated nerve endings that are sensitive to chemicals, temperature and touch [24]. Among the promising medicinal plants, Nigella sativa is an amazing herb with a rich historical and religious background; its seeds are the source of the active ingredients of this plant [9].…”