Organotypic modelling as a means of investigating epithelial-stromal interactions during tumourigenesis

Abstract

The advent of co-culture approaches has allowed researchers to more accurately model the behaviour of epithelial cells in cell culture studies. The initial work on epidermal modelling allowed the development of reconstituted epidermis, growing keratinocytes on top of fibroblasts seeded in a collagen gel at an air-liquid interface to generate terminally differentiated 'skin equivalents'. In addition to developing ex vivo skin sheets for the treatment of burns victims, such cultures have also been used as a means of investigating both the development and repair of the epidermis, in more relevant conditions than simple two-dimensional culture, but without the use of animals. More recently, by varying the cell types used and adjusting the composition of the matrix components, this physiological system can be adapted to allow the study of interactions between tumour cells and their surrounding stroma, particularly with regards to how such interactions regulate invasion. Here we provide a summary of the major themes involved in tumour progression and consider the evolution of the approaches used to study cancer cell behaviour. Finally, we review how organotypic models have facilitated the study of several key pathways in cancer development and invasion, and speculate on the exciting future roles for these models in cancer research.

Figure 1

Organotypic culture of skin equivalent. A. Schematic representation of a skin equivalent organotypic. The stroma consists of collagen and human fibroblasts (5 × 10⁵), with keratinocytes (1 × 10⁶) plated on the top. B. Example of immuno-staining for the differentiation markers Keratin14 and Involucrin in a 10 day old culture. C. A timecourse of wound healing in an organotypic culture using HaCaT human keratinocytes. A 5 mm wound was created using a punch biopsy and the epidermis (pink) of the organotypic was removed using forceps. Wound closure is shown at different time points (0, 2, 4, 7 days post wound).

Figure 2

Breast cancer invasion in an organotypic gel. A. Schematic representation of an organotypic cell culture. The stroma consists of collagen:matrigel (70:30) and 5 × 10⁵ human fibroblasts (yellow). 1 × 10⁶ breast cancer cells (blue) are plated on the top of the matrix. The organotypic is then cultures on a grid at the air-liquid interface. B. H & E staining of a section through an organotypic culture 9 days after seeding breast cancer cells (MDA-MB-231) on the top. The image shows breast cancer cells remaining on top of the organotypic culture as well as cells invaded into the stroma.