Evaluation of Eye Irritation Potential of Solid Substance with New 3D Reconstructed Human Cornea Model, MCTT HCE^TM

Abstract

The eye irritation potential of drug candidates or pharmaceutical ingredients should be evaluated if there is a possibility of ocular exposure. Traditionally, the ocular irritation has been evaluated by the rabbit Draize test. However, rabbit eyes are more sensitive to irritants than human eyes, therefore substantial level of false positives are unavoidable. To resolve this species difference, several three-dimensional human corneal epithelial (HCE) models have been developed as alternative eye irritation test methods. Recently, we introduced a new HCE model, MCTT HCETM which is reconstructed with non-transformed human corneal cells from limbal tissues. Here, we examined if MCTT HCETM can be employed to evaluate eye irritation potential of solid substances. Through optimization of washing method and exposure time, treatment time was established as 10 min and washing procedure was set up as 4 times of washing with 10 mL of PBS and shaking in 30 mL of PBS in a beaker. With the established eye irritation test protocol, 11 solid substances (5 non-irritants, 6 irritants) were evaluated which demonstrated an excellent predictive capacity (100% accuracy, 100% specificity and 100% sensitivity). We also compared the performance of our test method with rabbit Draize test results and in vitro cytotoxicity test with 2D human corneal epithelial cell lines.

Proposal of 28 GHz In Vitro Exposure System Based on Field Uniformity for Three‐Dimensional Cell Culture Experiments

“…Permittivity of the DMEM solution with 10% fetal bovine serum and 1% penicillin/streptomycin (P/S; GE Healthcare HyClone Laboratories, Logan, UT) was measured. This DMEM solution will be used as the culture medium for our future in vitro 3D cell culture experiments [Jung et al, ; Jang et al, ] with mmWave 28 GHz exposure. We adopted a Keysight N9952A Handheld Microwave Analyzer and an N1501A dielectric probe kit (Keysight, Santa Rosa, CA) for measurements up to 40 GHz.…”

“…The permittivity property of Dulbecco's modified Eagle's medium (DMEM) solution as a culture medium was measured for the specific absorption rate (SAR) analysis of the skin cell layer. A simulation model of the Petri dish with a cell culture insert for a 3D culture experiment [Jung et al, ; Jang et al, ] was employed. The best Petri dish setup from the field symmetry point of view was provided, and local SAR simulation was carried out to investigate the polarization effect on the SAR results among different Petri dishes with varying polarization angles.…”

Implementation of an in vitro exposure system for 28 GHz

“…Four 35 mm Petri dishes with inserts for a 3D cell culture, as shown in Figure 7, were placed on the holder tray inside the exposure chamber to verify the temperature regulation during exposure. Saline solutions and Micropig Franz cell membranes (Medi Kinetics Co., Ltd.) were used as a practical model for the 3D cell culture experiments, which are usually composed of a culture medium and a reconstructed cell model [20]. The center-to-center distance between the dishes was 39 mm, in accordance with the exposure area [18].…”

“…Compared to other reported in vitro MMW exposure systems [12][13][14][15][16], little effort has been made to practically realize an in vitro experiment system for 5G MMW exposure. The entire system is based on field uniformity [18] to provide adequate exposure area for three-dimensional (3D) cell culture [19,20], which is more efficient in dealing with a shallow skin depth of approximately 1 mm at MMW frequencies [10]. A significant feature of the implemented system is its "all-in-one" characteristics, that is, every system component is incorporated into a single unit for a more accurate exposure experiment.…”

Egr1 Gene Expression as a Potential Biomarker for In Vitro Prediction of Ocular Toxicity

“…The bovine cornea opacity/permeability assay using a by-product (corneas) from freshly slaughtered cattle [9], the isolated chicken eye test [10], and rabbit cornea epithelium (RCE) model [11] were also developed for evaluation of the in vitro eye irritation potential of various chemicals, such as pharmaceuticals, cosmetics, and their raw materials. Various 3D reconstructed human cornea-like epithelium (RhCE) models, including EpiOcular™ (MatTeck, Ashland, MA, USA), SkinEthic™ HCE (SkinEthic, Lyon, France), MCTT HCE™ (Biosolution, Suwon-si, Korea), and Labcyte Cornea model (JTE, Okinawa, Japan), have also been reported as in vitro alternative tests to eliminate species difference [12][13][14][15][16], presenting a high similarity of morphology, histology, and biomarker expression to human corneal epithelium [15].…”