Development and Characterization of a 3-Dimensional Human Corneal Full Thickness Tissue Model

Abstract

 

Purpose

 

The FDA and other regulatory agencies require ocular irritation testing of ophthalmologic pharmaceuticals and many consumer products. Although widely used, animal testing has limitations due to interspecies differences, variability, high cost, and animal welfare issues. A highly reproducible, cost effective, human cell-based in vitro corneal tissue model would be of high utility for ocular irritation testing and ophthalmic drug development.

 

Methods

 

Using primary culture and state of the art tissue engineering techniques, we have developed a human corneal full thickness (CFT) tissue model which includes epithelial, stromal, and endothelial layers. This model has been characterized in terms of histology and by testing a battery of test articles (TA, n=65) spanning the range from ocular corrosives and severe irritants to non-irritants (NI). In vivo irritancy levels were obtained from a published database and in vitro effects were determined using the MTT tissue viability assay and transepithelial electrical resistance (TEER) measurements.

 

Results

 

Using a tissue viability cutoff of 50%, the ocular CFT (OCFT) model was able to detect irritants with 100.0% sensitivity and non-irritants with 75.0% specificity (overall accuracy = 87.7%). In addition, the OCFT model was used to assess tissue recovery following exposure to increasing concentrations of surfactants and TA spanning the range of irritancy. Significant recovery at 24 and 48 hours was observed for NI chemicals. For the same 65 TA, an ex vivo assay, the Bovine Corneal Opacity and Permeability (BCOP), had 87.9%, 68.8%, and 78.5% assay sensitivity, specificity, and accuracy, respectively.

 

Conclusions

 

The OCFT model will address the needs of ophthalmic and other formulators who need to screen their products for irritancy and efficacy. Similar to other in vitro assays, high levels of reproducibility at a low cost are anticipated. The OCFT model will facilitate the development and testing of ophthalmic pharmaceuticals and allow for basic studies related to corneal physiology, wound repair, and pathologies.

 

 

Histological cross-section: Formalin fixed, paraffin embedded, H&E stained cross-section of OCFT tissue model (4X). 1. Corneal epithelium (10X); 2. Stroma with embedded keratocytes; 3. Endothelial layer (20X); 4. Micorporous membrane; 5. Air-liquid interface; 6. Culture medium fed through the membrane.

 

Keywords: 620 ocular irritancy/toxicity testing • 503 drug toxicity/drug effects • 765 wound healing