Development of 2D and 3D in vitro skin models to investigate inflammation and wound healing
DOI:
https://doi.org/10.21814/jus.6587Keywords:
Cytokeratin expression, Inflammatory skin model, 2D skin-like co-culture model, 3D skin equivalentAbstract
2D cell cultures are widely used in skin research but often fail to accurately replicate the complexity of the in vivo microenvironment. Co-culture systems offer advantages over monocultures since cell communication and interaction are more realistic. Nevertheless, co-cultures are generally considered simplistic models. The development of 3D skin models can more closely resemble in vivo context, minimizing the need for animal models. Most developed 3D skin models result in complete keratinocyte differentiation, evidenced by the expression of specific protein markers like cytokeratin 10 (CK10), and provide a more accurate platform for drug screening and toxicity testing. Here we report on the development of an inflammatory co-culture model, using activated monocytic cells and keratinocytes. Inflammation was induced by incubating the monocytic cells with Escherichia coli bacterial extract (BE) and validated by quantification of TNF-α, IL-6 and -8 cytokines expression. Co-incubation of these two cell lines for 8h is sufficient to maintain the inflammatory state of keratinocytes for at least 24h. Additionally, a co-culture of fibroblasts and keratinocytes, the main cell types that compose the skin, and a 3D skin equivalent were developed and compared for cytokeratin expression. Although the co-culture of fibroblasts and keratinocytes allows for both juxtacrine and paracrine signaling, it did not lead to CK10 expression, indicating that keratinocyte differentiation was not fully attained. On the other hand, CK10 expression was observed already after 11 days in 3D skin equivalents, and was well-defined at 18 and 28 days, demonstrating that 3D models better mimic reality in terms of keratinocyte differentiation.
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