Three-Dimensional Culture Systems for Dissecting Notch Signalling in Health and Disease

Abstract

Three-dimensional (3D) culture systems opened up new horizons in studying the biology of tissues and organs, modelling various diseases, and screening drugs. Producing accurate in vitro models increases the possibilities for studying molecular control of cell-cell and cell-microenvironment interactions in detail. The Notch signalling is linked to cell fate determination, tissue definition, and maintenance in both physiological and pathological conditions. Hence, 3D cultures provide new accessible platforms for studying activation and modulation of the Notch pathway. In this review, we provide an overview of the recent advances in different 3D culture systems, including spheroids, organoids, and "organ-on-a-chip" models, and their use in analysing the crucial role of Notch signalling in the maintenance of tissue homeostasis, pathology, and regeneration.

Keywords: 3D culture systems; Notch signalling; cancer; drug screening; microfluidics; organ-on-a-chip; organoids; regeneration; spheroids; therapy.

Figure 1

Schematic representation of the Notch signalling function. Notch pathway modulation strongly depends on the Notch receptor—DLL/JAG ligand interaction established between neighbouring cells. (a) Lateral induction triggers the expression of the Jag or Dll in the receiving cell. (b) Lateral inhibition reduces the expression of Dll or Jag in the neighbouring cell. (c) Cis-inhibition occurs when receptor and ligand expressed by the same cell, activates the pathway and sequesters active molecules on a single cell surface. Abbreviations: DLL, Delta-like; JAG, Jagged.

Figure 2

3D culture systems used to recapitulate the in vivo tissue organisation and complexity. Simplified overview of three different culture models: spheroids (a), organoids (b), and “organ-on-a-chip” (c). (a) Spheroids represent the simplest 3D culture system. Spheroids constitute efficient model systems for studying cell–cell communication, cellular metabolism, oxygen gradients, and nutrients distribution. Their complexity can be scaled up by integrating into the spheroids different cell types, such as stem cells and endothelial cells, thus providing the ideal platform for biochemical and molecular testing, as well as rapid drug screening. (b) Organoids reproduce the complexity of the native tissue, where cells at various cytodifferentiation states, ranging from stem cells and progenitors to fully differentiated cells, coexist. (c) “Organ-on-a-chip” closely mimics several aspects of the native organ. Integrated into a microfluidic system, 2D cultures, spheroids and organoids can be exposed to a dynamic microenvironment that allows controlled fluid exchange and interactions between different tissues, such as vessels and nerves (adapted from [64,65]).

Figure 3

Summary of 3D systems and their applications. Several aspects of tissue organisation can be modelled in 3D cultures. For all applications, spheroids, organoids, and “organ-on-a-chip” represent more faithfully the main features of native tissues when compared to 2D cultures. 3D systems allow characterisation of specific stem cell populations and could be used for regenerative purposes. Furthermore, 3D structures can reproduce a disease environment and serve for drug testing.