A bioengineering perspective on modelling the intestinal epithelial physiology in vitro

Abstract

The small intestine is a specialised organ, essential for nutrient digestion and absorption. It is lined with a complex epithelial cell layer. Intestinal epithelial cells can be cultured in three-dimensional (3D) scaffolds as self-organising entities with distinct domains containing stem cells and differentiated cells. Recent developments in bioengineering provide new possibilities for directing the organisation of cells in vitro. In this Perspective, focusing on the small intestine, we discuss how studies at the interface between bioengineering and intestinal biology provide new insights into organ function. Specifically, we focus on engineered biomaterials, complex 3D structures resembling the intestinal architecture, and micro-physiological systems.

Fig. 1. Structure of the small intestine.

The intestinal topography includes villi, where the absorptive enterocytes, secretory enteroendocrine and goblet cells reside, and crypt domains harbouring the intestinal stem cell niche including stem cells and Paneth cells. Underlying mesenchymal cells like myofibroblasts support the epithelium by secreting important growth factors such as WNTs, R-spondins and BMP antagonists. Gradients are believed to exist for the factors promoting stem cell self-renewal, including WNT and ligands for the ErbB pathway, whereas opposing gradients exist for differentiation-promoting factors such as BMP. The gradient, indicated here for EGF, represents all the ligands of the family, including Nrg1, Areg, Hbegf and Ereg^–, which support intestinal stem cell function. Many immune cells also reside in the intestine, constantly monitoring microbe and food antigens. Created with BioRender.com.

Fig. 2. Different types of organoids.

Intestinal organoid architecture comes in different flavours ranging from spherical structures to budding organoids (brown enterocytes, turquois intestinal stem cells, pink Paneth cells, purple enteroendocrine cells and green goblet cells) with distinct localisation of different cellular domains containing stem cells and differentiated cells, and organoids that include a supporting mesenchyme (illustrated with pink cells surrounding the organoid). The cells are arranged with the apical membrane inwards and with appropriate cellular junctions that normally support the formation of an intact barrier including tight junctions (purple). Created with BioRender.com.

Fig. 3. Synthetic matrices for stem cell research.

Synthetic matrix assembly using, e.g. click-chemistry, resembling the use of toy building blocks where cell-instructive building blocks of choice, e.g. adhesion peptides or growth factors, are combined with a polymer backbone to form a functionalised scaffold where cells can adhere and be cultured. Created with BioRender.com.

Fig. 4. Establishing complex 3D structures for organ mimicry and dynamic micro-physiological systems.

a Either single cells or organoid fragments can be used to seed these systems. b Complex 3D structures, e.g. crypt-like or villus-like, have been used to guide the cellular organisation. These structures can be fabricated from, e.g. biomaterials, PDMS, silicon or plastics such as polystyrene. c Intestine-on-a-chip devices have been used to create dynamic culture conditions including shear stress from fluid flow as well as cyclic mechanical deformation that resemble the natural movements associated with peristalsis. Intestine-on-a-chip device seen from above with an upper (purple) and a lower (blue) channel. Cross section of the device which is commonly composed of an upper (purple) and a lower (blue) channel separated by a porous membrane. Epithelial cells can be seeded into the upper channel, e.g. as a flat 2D layer or on a 3D construct that provides topographic features such as villi (as shown in the figure). These devices are most often made of PDMS, a transparent polymeric organosilicon, also making them ideal for live-cell imaging. d The intestinal barrier can be assessed by introducing various compounds to the upper surface of the 3D constructs, or through the inlet of the intestine-on-a-chip with access to the apical side of the epithelium. Medium can subsequently be collected from below the 3D construct or through both channel outlets of the intestine-on-a-chip for further analysis. In addition, cells seeded on these structures can later be isolated for, e.g. gene or protein expression analysis. Moreover, cells can be directly monitored to follow cell behaviour, using live-cell imaging. Created with BioRender.com.