A 3-dimensional fibre scaffold as an investigative tool for studying the morphogenesis of isolated plant pells

Cell culture

The 3D culture within the fibre scaffolds permits cells to grow freely within a matrix containing both large and small spaces, a technique that is expected to add to current lithographic technologies, where growth is carefully controlled and constricted. The cells, once seeded in the scaffolds, can adopt a variety of morphologies, demonstrating that they do not need to be part of a tightly packed tissue to form complex shapes. This points to a role of the immediate nano- and micro-topography in plant cell morphogenesis. This work defines a new suite of techniques for exploring cell-environment interactions.

We have developed a method of 3D cell culture for plants, which mimics the plant tissue environment, using biocompatible scaffolds similar to those used in mammalian tissue engineering. The scaffolds provide both developmental cues and structural stability to isolated callus-derived cells grown in liquid culture. The protocol is rapid, compared to the growth and preparation of whole plants for microscopy, and provides detailed subcellular information on cells interacting with their local environment. We observe cell shapes never observed for individual cultured cells. Rather than exhibiting only spheroid or ellipsoidal shapes, the cells adapt their shape to fit the local space and are capable of growing past each other, taking on growth and morphological characteristics with greater complexity than observed even in whole plants. Confocal imaging of transgenic Arabidopsis thaliana lines containing fluorescent microtubule and actin reporters enables further study of the effects of interactions and complex morphologies upon cytoskeletal organisation both in 3D and in time (4D). Conclusions: The 3D culture within the fibre scaffolds permits cells to grow freely within a matrix containing both large and small spaces, a technique that is expected to add to current lithographic technologies, where growth is carefully controlled and constricted. The cells, once seeded in the scaffolds, can adopt a variety of morphologies, demonstrating that they do not need to be part of a tightly packed tissue to form complex shapes. This points to a role of the immediate nano- and micro-topography in plant cell morphogenesis. This work defines a new suite of techniques for exploring cell-environment interactions.