OpenTox 2022 Virtual Conference

Skin and ocular co-culture models for in-vitro toxicity  assessment 

Dayan Guerra Flórez (1,2), Angie Castro Montoya 1, Andrés Pareja 1 

1. Unidad de Toxicidad in vitro, Facultad de ciencias y biotecnología,  Universidad CES. Medellín, Colombia. 
2. Maestría en ciencias biológicas, Universidad CES. Medellín, Colombia *E-mail contact: dguerraf@ces.edu.com 

Co-culture models are promising alternative methods to animal experimentation as they recreate a microphysiologic system and enable the study of in-vivo-like interactions. This work aims to generate two co-culture models, one for skin and one for ocular epithelium, as they are two of the most exposed tissues to the external environment, and provide it a useful tool to study in-vitro toxicity effects, with a biological response closet to that of the actual tissue. Cells were cultured in transwells, hanging porous polymeric membranes, to obtain multilayer co-cultures with an in-contact strategy, in which, fibroblast cell line Balb/3T3 was seeded on the bottom of the polymeric insert and incubated for 72 hours, allowing them to attach and proliferate to form a feeder layer. Then Human epithelial cells HEK001 or ocular  cell line SIRC were co-cultured in the upper compartment until reach confluence.  Both, ocular, and skin epithelial co-cultures, were exposed to four concentrations of  MMS (300, 500, 1000 and 1500 �g/mL) for 24 hours, or four concentrations of SDS  (1, 2, 3,2 y 4 mg/mL) for 30 minutes. Then cell viability was determined using MTT  assay. Both tested substances caused a reduction of cell viability above 50% for corneal and skin epithelial models, even for the lowest concentrations evaluated.  Furthermore, dose-dependent effects were observed for both substances, being more prominent than for SDS. These findings suggest that co-cultures here generated are useful models for risk assessment, approaching actual tissue responses to test substance of interest, in addition to being an alternative method to animal testing. Moreover, substance here evaluated can be used as positive control for cytotoxicity assays in co-culture models developed.