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

Jang WH, Jung KM, Yang HR, Lee M, Jung HS, Lee SH, Park M, Lim KM - Biomol Ther (Seoul) (2015)

Bottom Line: Recently, we introduced a new HCE model, MCTT HCE(TM) which is reconstructed with non-transformed human corneal cells from limbal tissues.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.

View Article: PubMed Central - PubMed

Affiliation: Amorepacific Corporation R&D Center, Yongin 446-729.

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 HCE(TM) which is reconstructed with non-transformed human corneal cells from limbal tissues. Here, we examined if MCTT HCE(TM) 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.

No MeSH data available.


Related in: MedlinePlus

Comparison with in vivo MMAS and cytotoxicity on 2D HCE-T cells. (A) Tissue viability versus MMAS (substances with MMAS data available are presented in Table 1). (B) Cytotoxicity of eye irritants on human corneal cell-line, HCE-T. Mean ± SD (n=3).
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f3-bt-23-379: Comparison with in vivo MMAS and cytotoxicity on 2D HCE-T cells. (A) Tissue viability versus MMAS (substances with MMAS data available are presented in Table 1). (B) Cytotoxicity of eye irritants on human corneal cell-line, HCE-T. Mean ± SD (n=3).

Mentions: Additional 7 solid test materials were evaluated with this modified eye irritation test. When 50% of cell viability was set as the criteria for eye irritation potential evaluation according to that for liquid substances, all of the solid chemicals were predicted correctly (Fig. 2A, Table 2). Histological examination also supported the clear manifestation of irritation on the tissue (Fig. 2B). Moreover, repeated runs showed an excellent consistency between runs (Fig. 2A). This approach resulted in excellent sensitivity (100%) and specificity (100%). Consequently, an overall accuracy of 100% was obtained (Table 2). Pearson correlation analysis of tissue viability versus MMAS exhibited a high and significant correlation (Fig. 3A, correlation coefficient=−0.771, p=0.009) but the test appears to overestimate the toxicity of Cat 2 irritants compared to MMAS, which may reflect potential risk of high false negative rates (high sensitivity but low specificity). In addition, we also evaluated the cytotoxicity of several irritants that could be solubilized in culture medium (Fig. 3B). The results showed that cytotoxicity and tissue viability obtained in MCTT HCETM for the solid substances were comparable. However, since test articles are applied as it is (i.e., at 100% concentration) in MCTT HCETM, it was difficult to discriminate Cat 2 from Cat 1. Overall protocol for eye irritation test is described in Fig. 4.


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

Jang WH, Jung KM, Yang HR, Lee M, Jung HS, Lee SH, Park M, Lim KM - Biomol Ther (Seoul) (2015)

Comparison with in vivo MMAS and cytotoxicity on 2D HCE-T cells. (A) Tissue viability versus MMAS (substances with MMAS data available are presented in Table 1). (B) Cytotoxicity of eye irritants on human corneal cell-line, HCE-T. Mean ± SD (n=3).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4489834&req=5

f3-bt-23-379: Comparison with in vivo MMAS and cytotoxicity on 2D HCE-T cells. (A) Tissue viability versus MMAS (substances with MMAS data available are presented in Table 1). (B) Cytotoxicity of eye irritants on human corneal cell-line, HCE-T. Mean ± SD (n=3).
Mentions: Additional 7 solid test materials were evaluated with this modified eye irritation test. When 50% of cell viability was set as the criteria for eye irritation potential evaluation according to that for liquid substances, all of the solid chemicals were predicted correctly (Fig. 2A, Table 2). Histological examination also supported the clear manifestation of irritation on the tissue (Fig. 2B). Moreover, repeated runs showed an excellent consistency between runs (Fig. 2A). This approach resulted in excellent sensitivity (100%) and specificity (100%). Consequently, an overall accuracy of 100% was obtained (Table 2). Pearson correlation analysis of tissue viability versus MMAS exhibited a high and significant correlation (Fig. 3A, correlation coefficient=−0.771, p=0.009) but the test appears to overestimate the toxicity of Cat 2 irritants compared to MMAS, which may reflect potential risk of high false negative rates (high sensitivity but low specificity). In addition, we also evaluated the cytotoxicity of several irritants that could be solubilized in culture medium (Fig. 3B). The results showed that cytotoxicity and tissue viability obtained in MCTT HCETM for the solid substances were comparable. However, since test articles are applied as it is (i.e., at 100% concentration) in MCTT HCETM, it was difficult to discriminate Cat 2 from Cat 1. Overall protocol for eye irritation test is described in Fig. 4.

Bottom Line: Recently, we introduced a new HCE model, MCTT HCE(TM) which is reconstructed with non-transformed human corneal cells from limbal tissues.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.

View Article: PubMed Central - PubMed

Affiliation: Amorepacific Corporation R&D Center, Yongin 446-729.

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 HCE(TM) which is reconstructed with non-transformed human corneal cells from limbal tissues. Here, we examined if MCTT HCE(TM) 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.

No MeSH data available.


Related in: MedlinePlus