An investigation of 2D and 3D substrates for embryonic facial process culture

Abstract

This work aims to highlight the differences between the use of 2D and 3D cell culture systems for the growth of samples of primary embryonic mesenchymal tissue derived from facial processes. In the developing embryo, these regions of tissue go on to form the cartilaginous template of the craniofacial skeleton. The tissue used in these experiments was derived from the frontonasal mass and mandible of chick embryos, which form parts of the upper and lower beak respectively. I have shown that explanted pieces of mesenchymal tissue from the frontonasal mass and the mandible of chick embryos display different cell behaviours in both traditional 2D cell culture and when grown in alvetex®, a 3D cell culture system that uses a polymer scaffold, developed by Reinnervate Ltd.
In 2D culture, as previously described in the literature, populations that expand from explanted frontonasal mass form sheets of cartilaginous material, whereas mandible cultures seem to expand into two different populations of cells, a sheet-like population as seen in frontonasal mass cultures as well as a population that is fibrous in appearance.
3D counterparts invaded and populated the alvetex®, and again there appeared to be one population of cells expanding from frontonasal mass explants and two populations in the case of explanted mandible tissue. 3D populations had dramatically higher levels of Alcian blue staining (indicative of the presence of cartilaginous material) than 2D cultures, suggesting that alvetex® provides a more permissive environment for the production of cartilage than traditional 2D substrates used in cell and tissue culture. It was also shown that the alvetex® appears to separate the two different populations of cells observed in mandible cultures, in that the two populations occupy the internal surface of the polymer at 7 days culture, but by 14 days one population remains within the internal surface whilst the other has grown along the top surface of the polymer.
These results suggest that alvetex® provides an environment that is suitable for the growth of primary embryonic tissue and may be suitable for the culture of other samples of primary tissue from which cell lines that have been established in a 3D environment could be derived.