Organoids and Mini-Organs

Jamie A. Davies, Melanie Lawrence

• Publisher: Academic Press
• Year: 2018
• ISBN:
  9780128126370
• Topic: Biological Sciences
• Subtopic: Cell Biology
• Index: Biological Sciences

Organs and Mini-Organs combines contributions from leading practitioners who work under the editorial control of an acclaimed researcher who also served for eight years as Editor-in-Chief of the journal Organogenesis, the first journal on this topic. The book begins with an introduction, but then delves into chapters that present advice on how to make organoids for many systems. In addition, case studies that illustrate the uses of organioids are presented, along with discussions on future directions and specific problems that need to be solved.

• Collects the best protocols of organoid cultures from diverse tissues
• Covers a wide range of organs
• Includes troubleshooting cases for common, but specific problems for each culture conditions
• Provides an entire section on the application of organoids

Part I

Introduction

Outline

• Chapter 1 Organoids and mini-organs

Chapter 1

Organoids and mini-organs

Introduction, history, and potential

Jamie A. Davies, University of Edinburgh, Edinburgh, United Kingdom

Abstract

Organoids are three-dimensional assemblies that contain multiple cell types, arranged similarly to the cells in a specific tissue, at least at the micro-scale; mini-organs add to this micro-realism a realistic macro-scale anatomy as well. Researchers have been producing organoids for at least 60 years, initially to explore basic mechanisms of development, but more recently as tools for medical research. Organoids made from human cells are particularly valuable for preclinical studies, because they avoid the need to extrapolate results from one species to another. This chapter outlines the history of organoid research, describes briefly how typical organoids are produced, and goes on to describe their use in pharmacology, toxicology, oncology, and microbiology. It includes an indication of how organoids can be turned into mini-organs, and ends with some cautionary notes on the limitations of current techniques.

Keywords

Organoids; mini-organs; research

Chapter Outline

• Introduction 3
  • Origins of Organoids 5
  • Mechanisms of Organoid Formation 7
  • The Path to Human Organoids 9
  • The Construction of Organoids 11
  • Organoids as Tools for Clinical Research 12
  • From Organoids to Mini-Organs 16
  • Limitations of Organoids 17
• Final Remarks 18
• Acknowledgments 18
• References 18
• Further Reading 23

Introduction

It is important to begin this book with a definition of terms, because the word “organoid” has been coined several times in the history of biomedicine, and has been used to convey at least three distinct meanings. In the 20th century, “organoid” was sometimes used as a synonym for “organelle” (e.g., Duryee and Doherty, 1954), a sub-cellular structure: this use is now obsolete. In oncology, “organoid” is sometimes used as an adjective to imply a tumor with a complex, tissue-like structure, for example a gland-like carcinoma or a teratoma: this use remains current (e.g., Nesland et al., 1985; Heller et al., 1991), if somewhat obscure. Neither of these meanings is relevant to this book. Here, “organoid” is being used in its most common modern sense, to refer to a three-dimensional assembly that contains cells of more than one type, arranged with realistic histology, at least at the micro-scale. An organoid might be made from cells of humans or other animals, and these might be differentiated cells, stem cells, or a mixture of the two.

Interest in organoids has increased significantly in the 21st century (Fig. 1.1), fueled on the one hand by rapid developments in stem cell derivation to provide human progenitor cells, and on the other by a strengthening desire to refine, reduce or replace the use of animals in research (reviewed by Davies, 2012). Organoids are being produced for basic research into development and neoplasia, for industrial and medical applications such as toxicology, and—ultimately—for transplantation. At the end of 2013, The Scientist named organoids as one of its “advances of the year” (Grens, 2013), not because the technology was new, but because it had suddenly become much more pervasive and visible thanks to some high-profile research papers. About a year and a half later, and for much the same reason, Nature published an overview of the potential of the technology (Willyard, 2015). Organoids have now been developed to represent many different parts of the body, for many different reasons, with applications expected to grow still further over the next few years.

Figure 1.1 The rising use of organoids in biological research. The graph shows the number of papers returned from a Pubmed search for “Organoid”, and screened manually for “organoid” being used in the sense in which it is used in this book.

Being published in a period of such intense research, this book has two purposes. The first is to help newcomers to the field to set up working organoid systems, whether by using the exact techniques of other researchers, or by using these techniques to inspire the creation of novel organoid systems. The second purpose is to help researchers with working organoid systems find inspiration and advice to help them use their organoids as tools to solve a range of biological and medical problems.

Origins of Organoids

Organoids have been an important area of research for much longer than most recent reviews imply, particularly the annoyingly large number of reviews that appear to have been written for the sole purpose of portraying their authors as founders of a field. The first mammalian organoids were in fact produced over 60 years ago, and organoids have contributed steadily and significantly to developmental cell biology ever since.

Techniques for constructing organoids have their origins in basic research about the nature of biological organization. A good starting point for this brief history would be the work of H.V. Wilson (1910), who showed that, if a sponge is dissociated into its constituent cells, and these cells are reaggregated randomly, they reorganize to make a realistic and viable new sponge (Wilson, 1910). Though not aimed at making organoids in the modern sense, this experiment was an important demonstration that cells of an adult organism can contain sufficient information to specify a multicellular structure, without any need for outside instructions, and without the need for cells to start from some specific anatomical arrangement contingent on their embryological history. This point is critical to organoid production. In the 1950s, several laboratories began to use the same basic method—disaggregation followed by reaggregation—to determine whether the cells of more complex animals such as vertebrates also had the ability to self-organize (this term was already in use) or whether, for these complex animals, the spatial relationships contingent on past developmental history were critical. Early examples of the disaggregation−reaggregation method of organoid construction being applied to higher vertebrates were provided by Moscona (1952) and by Moscona and Moscona (1952), who made suspensions of chick mesonephric kidney cells. The source tissue included both tubular epithelial cells and mesenchymal cells. On reaggregation and incubation, epithelial cells made small clusters that went on to make tubules surrounded by mesenchyme-derived stroma. This arrangement was reminiscent of the small-scale anatomy of normal mesonephroids, although the overall gross-scale organization of the organ was absent: the structure met the modern definition of an “organoid”. These experiments demonstrated that at least some of the cells of embryonic chicks, like the cells of sponges, carried sufficient information to organize themselves realistically, even when their original spatial relationships had been erased.

The observation that different types of cells in the suspension (e.g., epithelial, mesenchymal) would separate in the reaggregate raised questions about how cells choose their neighbors. It was already known from culture experiments that similar cells tend to unite even if they come from tissues of different species. In the 1940s, Harris (1943), Medawar (1948), and Grobstein and Younger (1949) all tried coculturing fragments of tissues from different species to explore the mechanisms of transplant rejection (this was before the role of the immune system in the process was well understood). They each showed that fragments of the same tissue from different species could join and behave as one: indeed, in the case of heart muscle, heart-beat synchronized across the interspecies boundary. Moscona (1956) built on these studies of interspecific combinations to ask whether the association of cells was controlled more by their being of the same histological type, or by their coming from the animal. He mixed suspended mouse and chick embryonic liver cells, and showed that they cooperated in making an organoid, epithelia associating with epithelia, and stroma with stroma, regardless of the species of origin. He also mixed different tissues, for example chick kidney and mouse cartilage, and observed that the cells sorted out reasonably well, and each resulting organoid was formed only of cells exhibiting nuclear markers of its own species. The species-specific nuclear morphologies ruled out any possible mechanism of cells trans-differentiating according to their surroundings, and supported instead the idea of cells choosing their neighbors. The author concluded that type specificity was stronger than species specificity in arranging cells, and that cells were already determined to make a particular tissue. The paper also showed an early application of organoids to problems of pathology, when its author mixed mouse melanoma cells with normal chick cells, and saw some evidence of invasive growth (“infiltration” in his words) by the sarcoma cells.

Part of the drive behind these early organoid experiments, acknowledged and discussed by Weiss and Taylor (1960), was to provide a counterbalance to the prevailing view that most embryogenesis was driven by inductive signaling, a mechanism that had obsessed many embryologists since its discovery in the 1920s (Spemann and Mangold, 1924). The ability of mixtures of cells, isolated from their normal anatomical relationships with the rest of the embryo, to self-organize was taken by these authors as an indication that much epigenetic information was held by the cells themselves, and did not rely on inductive instructions from elsewhere. This did not, though, rule out signaling taking place between different cell types within the self-organized aggregate and, in the 1950s, Grobstein’s laboratory used the dissociation−reaggregation technique to explore these signals. It was already known, from the experiments of Gruenwald (1937, 1942), that kidney development relies on inductive signaling between the ureteric bud (the progenitor of the urine collecting duct system) and the surrounding metanephrogenic mesenchyme (“Metanephrogenic mesenchyme” is Grobstein’s original term that captures the developmental potential of the tissue.). The mesenchyme induces the ureteric bud to grow and branch, while the bud induces the mesenchyme to make nephrons and stroma. Auerbach and Grobstein (1958) made cell suspensions of metanephrogenic mesenchyme, reaggregated them, and cultured them. On its own this reaggregate did nothing but, when placed in contact with inducing tissue, it made tubules. This showed that the disaggregation and reaggregation process did not itself substitute for induction: the “rules” of development in reaggregates apparently remained the same as in the embryo. The researchers also made suspensions of inducing tissue, mixed them with suspensions of metanephrogenic mesenchyme, and cultured the reaggregate. The cell types segregated and inductive signals passed to the mesenchymal cells, inducing them to make nephrons, and indicating that inductive activity is robust to disaggregation and reaggregation. The inducing tissue used for these experiments was spinal cord, because ureteric buds die. Only decades later, when the Auerbach and Grobstein work was revisited, was a method developed to prevent anoikis in the ureteric bud cells of a disaggregate−reaggregate in culture: in the presence of an inhibitor of Rho-activated kinase (ROCK), the ureteric bud cells survive, segregate from the mesenchyme, and make small collecting duct cysts and treelets that induce the metanephrogenic cells to make nephrons, the whole being a renal organoid (Unbekandt and Davies, 2010).

Mechanisms of Organoid Formation

An important question hanging over the work on organoids in the 1940s and 1950s was the mechanism by which cells of a mixed reaggregate sort out into distinct...

Table of contents

• Cover image
• Title page
• Table of Contents
• Copyright
• List of contributors
• Acknowledgments
• Disclaimer
• Part I: Introduction
• Part II: Constructing Organoids
• Part III: Applications of Organoids
• Index