Review Article (Invited)
Intestinal and optic-cup organoids as tools for unveiling mechanics of self-organizing morphogenesis
2022 Volume 19 Article ID: e190048
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2022 Volume 19 Article ID: e190048
Organoid, an organ-like tissue reproduced in a dish, has specialized, functional structures in three-dimensional (3D) space. Organoid development replicates the self-organizing process of each tissue development during embryogenesis but does not necessarily require external tissues, illustrating the autonomy of multicellular systems. Herein, we review the developmental processes of epithelial organoids, namely, the intestine, and optic-cup, with a focus on their mechanical aspects. Recent organoid studies have advanced our understanding of the mechanisms of 3D tissue deformation, including appropriate modes of deformation and factors controlling them. In addition, the autonomous nature of organoid development has also allowed us to access the stepwise mechanisms of deformation as organoids proceed through distinct stages of development. Altogether, we discuss the potential of organoids in unveiling the autonomy of multicellular self-organization from a mechanical point of view. This review article is an extended version of the Japanese article, Mechanics in Self-organizing Organoid Morphogenesis, published in SEIBUTSU BUTSURI Vol. 60, p.31–36 (2020).
Recent studies have developed a variety of organoids modelling the brain, intestine, lung, kidney, liver, and pancreas. They are widely used in the studies of embryonic development, cancer, and congenital disease progression, as well as regenerative medicine. Though epithelial organoid development occurs in the absence of surrounding tissue unlike embryonic tissue development, this feature is useful in understanding multicellular self-organization. In this review, by summarizing the mechanical processes of 3D tissue formations and their regulatory factors as revealed in recent organoid studies, we provide a mechanical perspective on how organoids can be promising tools in revealing self-organizing tissue morphogenesis.
Cells self-organize to form three-dimensional (3D) structures of tissue during embryogenesis. To form 3D structures, individual cells generate mechanical forces that are integrated into the tissue as a whole through cell-cell interactions. This cellular force generation varies spatiotemporally throughout development that deforms each tissue in a stepwise manner. While individual tissues form autonomously, they can be regulated and influenced by the surrounding microenvironment and neighboring tissues during embryogenesis. In other words, because cell dynamics are influenced by the surrounding microenvironment, interactions between tissue and its surroundings may be crucial in regulating the development of an entire embryo. Even if the individual cell behaviors driving development are autonomous, it is still unclear how autonomous the developmental process of each tissue is.
To address the autonomy of tissue formation, it is necessary to manipulate the environment of each tissue. Over the past two decades, remarkable advances in 3D cell culture technology have enabled the generation of organoids by successfully reproducing organ-like 3D tissues in a dish [1–6]. Particularly, epithelial organoids reproduce not only the process of cell differentiation but also the process of characteristic 3D deformation of organogenesis. Notably, most epithelial organoids can be reproduced without external tissues such as mesenchyme and other epithelial tissues, representing the autonomy of tissue development. Nevertheless, current organoids with varied morphology only partially reproduce the morphology of each tissue in vivo and therefore its developmental process is less robust than that of embryonic tissues.
Although epithelial organoid development differs from embryonic tissue development in that there is no surrounding tissue, this feature is useful for understanding multicellular self-organization. Recent studies have utilized organoids such as the intestine and optic-cup to investigate their mechanisms of morphogenesis [7–9]. These studies revealed several mechanical factors that play a key role in the deformation of organoid development. This indicates the autonomy of organoids during their development by which various mechanical factors are regulated at a precise time and location.
In this paper, we review the recent progress in the research focusing on the mechanics of organoids, especially the intestine and optic-cup. First, we discuss the variety of mechanical processes involved in each organoid development. Next, we categorize elementary deformation processes during organoid development by their types and discuss the main driving forces and factors controlling them. Finally, we affirm the potentiality of organoids in unraveling the autonomy of multicellular self-organization.
Each cell has mechanisms that control its behaviors autonomously (on its own) and non-autonomously (by signals sensed from external sources). This is an analogy that can be applied not only to individual cells but also to each tissue and organ [10,11]. The current techniques for producing organoids exploit the inherent ability of self-organization of cells at the tissue scale under optimal culture conditions.
Organoids can be derived from embryonic stem cells (ESCs), somatic stem cells, induced pluripotent stem cells (iPSCs), and isolated organ progenitors that differentiate to form organ-like tissues. 3D culture technology has made it possible to create a variety of organoids, including the brain [1,12], gut [2,3], retina [4,13], kidney [14–16], liver [17], mammary gland [18], limb bud [19], and skin [20]. These organoids are useful for elucidating multicellular autonomy inherent in tissue development. Recent studies on intestinal and optic-cup organoids have provided mechanical insights into the mechanisms of tissue development. In the following subsections, we present the developmental processes of these two types of organoids.
Intestinal OrganoidThe intestinal epithelial tissue consists of crypts and villi, where stem, progenitor, and differentiated cell types are aligned properly to maintain an epithelial barrier against luminal contents and carry out organ functions such as nutrient absorption and mucus secretion (Figure 1A). During intestinal development, the basal region of the villus extends into the mesenchymal tissue as the crypt. The base of the crypt is also known as the stem cell zone, which has stem cells and Paneth cells. The region above the stem cell zone is known as the transit-amplifying zone that houses highly proliferative cells, while differentiated cells such as the enterocytes reside farther apart.
Generation of intestinal organoid and its morphogenesis in 3D culture. A section of in vivo mouse intestine (A), where the apical surface is facing the lumen (light blue), and the basal surface is facing the mesenchyme (orange). 3D culture of an isolated crypt (B) generates an intestinal organoid with a 3D structure containing a lumen-enclosed epithelium (C). During development, the organoid initially attains a bulged state (D), and later, transforms into a budded state (E). In this process, lumen volume is reduced, and the crypt bud is protruded. The intestinal organoid as shown here houses different cells such as the stem cells (blue), Paneth cells (pink), and enterocytes (yellow).
Crypt cells can be isolated from the mouse intestinal tissue (Figure 1B) that are harvested in 3D culture using extracellular matrix (ECM) gel and defined niche factors. The isolated crypts contain Lgr5+ stem cells, which can produce all the cell types found in the adult mouse intestinal epithelium such as Paneth cells, enterocytes, enteroendocrine cells, and goblet cells [2]. The crypt cells can self-organize into an intestine-like 3D tissue, i.e., an intestinal organoid with a polarized epithelium in the apical-basal direction enclosing a lumen (Figure 1C). Initially, the organoid attains a bulged state around day 3.5 of culture (Figure 1D), and then it is transformed into a budded state around day 4, where a portion of the epithelium projects outward to form crypt bud (Figure 1E). At the initial stage, the cells of the luminal homogeneous organoid show a transient activation of transcription factor Yap1, but the cell-to-cell variability of Yap1 subcellular localization drives heterogenous Notch-Delta activation, which causes symmetry breaking to induce Paneth cell differentiation and subsequent crypt bud formation [21].
During crypt bud formation, actin filaments and myosin accumulate on the apical surface of the bud region to induce apical constriction of the surface [7]. The apical constriction in the crypt and basal tension in the villus changes the spontaneous curvature of the epithelium. Moreover, this process also involves a reduction of lumen volume that accelerates crypt bud formation [7]. Such regulation of the lumen volume is correlated with the expression of osmotic regulators in the enterocytes of the villus region.
Another study reported that the spreading of crypt bud excised from the intestinal organoid on soft gel coated with ECM forms a monolayer with several different compartments including a central stem cell zone, transit-amplifying zone, and a region of differentiated cells [8]. This monolayer recapitulates the open-lumen architecture of the in vivo intestine, with the base of the monolayer facing the ECM substrate. The stem cells located around the center of the crypt region undergo apical constriction, exerting a pushing force and thereby folding the intestinal monolayer [8].
Optic-Cup OrganoidOptic-cup is the primary source of eye tissue that can be derived from both mouse and human ES cells in culture [4,13,22]. The serum-free culture of embryoid body-like aggregates with quick aggregation (SFEBq) was introduced to generate embryoid bodies (Figure 2A and B) [1], where the embryoid body-like tissues first self-assemble to form a neuroepithelium (Figure 2C) and then form a complex 3D structure of optic cup as observed in embryos.
Generation of optic-cup organoid and its morphogenesis in 3D culture. In the SFEBq method, A) initially, the dissociated embryonic stem cells (ESC) are harnessed in a tube. B) The cells autonomously form an aggregate, and C) the aggregate forms a lumen-enclosed neuroepithelium. D) In the optic-cup formation, a part of the neuroepithelium protrudes outward to form a semispherical optic vesicle (OV). The OV exhibits further development (i-iv). The inner and outer surfaces of the OV are apical and basal surfaces, respectively (i). The distal part of the OV differentiates into the neural retina (NR) and the surrounding retinal pigment epithelium (RPE) (ii). Upon differentiation, the NR invaginates in the apically convex manner (iii) by gradually increasing the NR curvature. Later, the tissue attains a two-walled cup-like structure with a hinge-shaped NR-RPE boundary (iv).
The neuroepithelial cells gradually differentiate into multiple cell types while undergoing stepwise 3D deformation. First, a portion of the sheet-like neuroepithelium i.e., optic vesicle (OV) protrudes outward (around Day 7 of culture) (Figure 2D, (i)). Next, the tip of the protruding epithelium differentiates into the neural retina (NR) and surrounding retinal pigment epithelium (RPE) (Figure 2(ii)). The NR invades inward and overlaps with the RPE to form a two-walled cup-like structure (around 12 to 24 hours later) (Figure 2(iii)). Around 48 hours later, the apical end of the boundary tissue takes an anisotropic hinge shape (Figure 2(iv)). This 3D structure and the process of forming such structures are common among a variety of animals, including fishes, birds, and humans.
During embryogenesis, the optic-cup, like the intestine, is formed in the presence of the surrounding tissues such as ectoderm and ECM. Therefore, the influence of external forces was thought to be important for optic cup formation. However, the optic-cup organoids obtained from stem cells are composed only of neuroepithelia and have almost no surrounding tissue. This suggests that the recapitulation of the optic-cup organoid is a result of self-organization that gives rise to the 3D structure of the optic-cup by the force generated by neuroepithelia alone [4].
Further experiments have also revealed that external forces from the surrounding tissue are irrelevant, and that myosin activity and cell proliferation play a vital role, where two types of myosin-dependent force generations are key in optic cup formation [9]. In the initial stages, increased myosin activity generates a contractile force on the apical surface of the neuroepithelium to form the OV. First, as the distal portion of the OV differentiates into NR, myosin activity decreases on the apical surface, as a result, the generated contractile force is reduced, inducing autonomous inward invagination, also known as apical relaxation. Next, cells at the boundary between the NR and the surrounding RPE actively contract along the direction of tissue thickness, pushing the retinal tissue further inward in response to mechanical feedback, called lateral constriction [9]. Furthermore, the NR curvature is also significantly increased by cell proliferation in the NR region and its periphery.
The formation of intestinal and optic-cup organoids in the absence of other surrounding tissues suggests that the 3D morphologies of these tissues are controlled by autonomous mechanisms in both the organoids; one type of cells initially composing the organoids later differentiates into more than one distinct type of cells. Depending on cell differentiation, cells change the manners of force generation accordingly to form specific 3D structures. Therefore, both intestinal and optic-cup organoids could be promising models to study the principles of epithelial morphogenesis.
Actomyosin contractility is known to generate a driving force in morphogenesis. In particular, the regulation of actomyosin contractility can stretch and bend epithelial tissues in 3D space [23–25]. Importantly, actin filaments and myosin are localized within each cell depending on the cell polarity, and the actin filaments can be oriented to generate anisotropic force. In epithelial tissues, actomyosin contractions are generated on apical, basal, and lateral regions depending on the epithelial polarity. The forces thus generated within each cell alter the spontaneous shape of the cell, and the deformation of each cell is integrated into the deformation of the entire tissue. The following subsections introduce several modes of epithelial deformation induced by actomyosin contractility.
Apical ConstrictionApical constriction is one of the key processes of cell shaping that causes epithelial deformation in a variety of developmental contexts [25,26]. During development, actin filaments and myosin that are localized in the apical region of epithelial cells generate contractile force. This force gradually contracts the apical surface area of the cells, causing the epithelium to bend in an apically concave manner, also known as apical constriction (Figure 3A). Apical constriction has been reported to oscillate during embryonic cleavage, suggesting that the constriction process may involve a ratcheting mechanism [27]. Mathematical modeling of the intestinal organoid as a monolayer of cells has shown that apical constriction is a major driving force for crypt budding, which is further confirmed with the analysis of actomyosin dynamics and mechanical perturbations using 2D and 3D organoid cultures [7,8]. During crypt budding, apical constriction pushes the surrounding substrate, especially around the region containing the stem cells.
Modes of actomyosin-dependent epithelial bending. A) Apically concave invagination by apical constriction. Increase in apical actomyosin contractility constricts the apical cell surface, causing the epithelium to bend in an apically concave manner. B) Apically convex invagination by apical relaxation or expansion. Reduction in apical actomyosin contractility or other machineries expands the apical surface, causing the epithelium to bend in an apically convex manner. C) Curvature amplification by lateral constriction. When lateral constriction occurs in an apically or basally curved epithelium, it constricts the thickness of epithelium, amplifying its curvature.
Apical relaxation is conceptually the opposite of apical constriction, i.e., a reduction of actomyosin contractility in the apical region, causing relaxation. This reduced contractility expands the apical surface area of cells, bending the epithelium in an apically convex manner (Figure 3B). Similarly, actin polymerization at the apical region also expands the apical area of cells which causes epithelial bending. These epithelial relaxation and expansion are evident during embryonic development, including the morphogenesis of zebrafish hindbrain [28], rounding of mitotic cells in the Drosophila tracheal placode [29], and internuclear migration in the chick embryo [30].
Apical relaxation occurs at the region undergoing NR invagination during optic-cup organoid formation [9]. Initially, apical myosin activity is high in the OV region, generating contractile force. However, in the region that is differentiated to NR, the apical myosin activity decreases, reducing contractility. The reduction of apical contractility changes the spontaneous curvature, causing NR invagination that autonomously bends the NR in an apically convex manner. The induced NR bending results in the formation of a two-walled cup-like structure with an internal NR and an external RPE. This phenomenon has been observed in the optic-cup organoids derived from both mouse and human cells [4,13] and may also be important for optic cup formation in vivo.
Lateral ConstrictionLateral constriction is the constriction of cells in the apical-basal direction mediated by the actomyosin complexes aligned along the apical-basal axis, which decreases the thickness of the epithelial sheet (Figure 3C). Lateral constriction has a role in increasing the curvature of an already curved epithelium. In other words, if the epithelial sheet is already bent to some degree by apical constriction, lateral constriction causes the sheet to bend further, as if creased. Lateral constriction has been reported in the apoptotic cells of Drosophila [31], in the formation of ascidian progut [32], and during vulval lumen morphogenesis in C. elegans [33].
It was initially thought that the shape of the optic cup was obtained by apical constriction at the NR-RPE boundary. However, it is lateral constriction at the NR-RPE boundary that facilitates NR invagination [9]. Furthermore, apical constriction may distort the epithelial sheet along the NR-RPE boundary, failing to reproduce the smooth cup-like shape, whereas lateral constriction could reproduce the smooth hinge structure of the optic-cup.
Most studies elucidating the mechanics of morphogenesis focus on driving forces. However, tissue deformation is a process driven not only by various forces but also by the mechanical properties and states of the tissue itself as well as the surrounding constraints and force fields. One of the significant mechanical properties of 3D deformation of epithelia is the stiffness with respect to bending. Whereas the surrounding constraints and force fields are influenced by the adjacent lumen and ECM as well as the surrounding tissue. Here, we discuss bending stiffness regulated by the thickness of the epithelial sheet and geometric constraints regulated by lumen volume reported in the studies of intestinal and optic-cup organoids.
Bending Rigidity Regulated by the Thickness of Epithelial SheetThe thickness of epithelial tissue, which varies with cell types and cycle phases, has significant effects on bending stiffness. The thicker the tissue, the higher the bending stiffness, i.e., greater force is required for bending (Figure 4A). On the other hand, the thinner the tissue, the lower the bending stiffness, i.e., less force is required for bending (Figure 4B). As a simple example, if we approximate the epithelial tissue as a uniform plate and consider its small deformation, the bending rigidity, G is proportional to the first order of the elastic modulus, E, and the third order of the thickness, h, of the epithelium, that is, G α Eh3 [34]. This equation indicates that the effect of epithelial thickness on bending rigidity is greater than that of the elastic modulus. Although it is an extreme approximation to consider the epithelial tissue as a uniform plate, this equation provides a more intuitive understanding of morphogenesis.
Bending rigidity regulated by epithelial thickness. A) Thicker epithelium has higher bending rigidity, requiring larger external force to bend the epithelium. B) Thinner epithelium has lower bending rigidity, requiring smaller external force to bend the epithelium.
Let us try to understand the impact of epithelial thickness with the help of an example of optic cup formation [9]. If E and h of the NR are about 146 Pa and 78 μm respectively, and those of the RPE are about 391 Pa and 23 μm respectively, then one might think that NR is more flexible because of the lower elastic modulus value. However, since the NR is more than three times thicker than the RPE, it will be more difficult to bend due to its higher bending rigidity of about fourteen times or more. This implies that epithelial thickness is an important parameter for morphogenesis. Moreover, the higher thickness of the NR as compared to the NR-RPE boundary also suggests that if there were apical constriction at the NR-RPE boundary, the resulting bending moment would be unlikely to deform the NR. Rather, it is more plausible that the NR bends autonomously by generating forces by itself and that the resulting bending moment deforms the NR-RPE boundary.
The thickness of epithelial tissue is known to be regulated by the expansion and contraction of the height of each cell in the apical-basal axis [35], actomyosin contractility, binding of adhesion molecules, and elongation of microtubules along the apical-basal axis.
Geometric Constraint Regulated by Volume of Epithelial LumenA lumen is a space enclosed by the apical surface of the epithelium. This fluid-filled lumen exerts pressure on the apical surface [36,37], which can both function as a driving force that deforms the epithelium and a geometric constraint that regulates deformability (Figure 5). For example, the reduction of lumen volume allows the epithelium to bend freely, in contrast, the expansion of lumen volume prevents the epithelium from bending. The lumen volume can be regulated by cellular fluid transportation in the apical-basal direction.
Epithelial deformation regulated by lumen volume. The decrease in lumen volume allows the epithelium to deform freely (A to B), whereas the increase in lumen volume expands the epithelium and prevents its deformation as a constraint (B to A).
The regulation of lumen volume and its effects have been studied during intestinal organoid formation [7]. The transition from the bulged to the budded state has been shown to decrease the lumen volume, and swell the villi, accelerating the process of crypt budding. The decrease in the lumen volume and swelling of the villus create a negative pressure in the lumen, allowing autonomous bending of the epithelium. This change in lumen volume and villus size is actively regulated by the sodium-glucose co-transporter SGLT-1, which induces water uptake by absorptive epithelial cells to increase cell volume. On the other hand, the development of the optic-cup organoid does not require such active regulation of lumen volume [4].
During morphogenesis, cellular force generation in the appropriate location and time causes the deformations of tissues in a stepwise manner as the tissues progress through stages of development. To robustly regulate the stepwise deformation without overarching control, it is necessary to have not only a bottom-up regulation from microscopic cells to macroscopic tissues but also a top-down regulation. Such multiscale regulation can be explained by individual cells sensing the 3D deformation of the entire tissue [9,38,39].
In the process of optic-cup formation, a cup-like 3D structure is formed by a two-step deformation process driven by apical relaxation and lateral constriction. A mechanical feedback loop explains the regulatory mechanism of stepwise force generations [9]. First, the NR region autonomously bends inward to the surrounding RPE (Figure 6A). This autonomous bending of NR applies a bending moment to the NR-RPE boundary region, generating a mechanical strain at the basal surface of the NR-RPE boundary (Figure 6B). The boundary cells sense the basal strain and are triggered to cause lateral constriction (Figure 6C). The lateral constriction generates a further bending moment to amplify the NR invagination, forming a hinge structure at the boundary. That is, the cells at the boundary adjust the entire-tissue shape by sensing the entire-tissue deformation through mechanical force. This mechanism is referred to as ‘strain-triggered mechanical feedback’ describing how mechanical strain triggers further force generation by cells causing the entire tissue to deform in 3D space [9].
Mechanical feedback from the entire tissue to individual cells. A) In the optic-cup formation, the NR invaginates and the NR-RPE boundary is wedged simultaneously. The NR invagination is autonomously caused to generate a bending moment. B) The bending moment generated in the NR is exerted on the NR-RPE boundary, causing a strain at the basal surface. C) The basal strain triggers lateral constriction at the NR-RPE boundary, constricting the epithelial thickness and facilitating the NR invagination.
While the generation of cellular forces that drive morphogenesis has been thought to be triggered by molecular cues, mechanical feedback in the development of the optic-cup organoid revealed a key role for mechanical forces that feed back 3D tissue deformation to individual cell force generation at different scales. Although the underlying molecular mechanism is still unclear, this finding overturned the conventional train of thought regarding the development of multicellular organisms. Mechanical feedback may play a role in maintaining the robustness of morphogenesis, as it occurs adaptively in response to the deformation of 3D tissues. Especially in optic-cup formation, it plays an important role in the robust formation of smooth hinge structures at the NR-RPE boundary under environmental disturbances since the position of the NR-RPE boundary is highly variable [40]. As the hinge structures are found at the boundaries between two different tissues, the effects of mechanical feedback may often be involved in self-organizing developmental processes.
In this review, we presented the new findings of tissue deformation elucidated from the studies conducted in two different organoids, the intestine, and the optic-cup. We discussed how a local increase or decrease of myosin activity in cells causes constriction and elongation of the apical and lateral surfaces of epithelial cells, leading to various modes of 3D tissue deformation. In addition, we reviewed that the magnitude of tissue deformation induced by these driving forces is highly dependent on epithelial thickness and lumen volume. Finally, based on the study of the optic-cup organoid, we explained the importance of the mechanisms, by which the mechanical force generated by tissue deformation is fed back to the force generation in individual cells to coordinate the entire-tissue morphology. These findings indicate that organoids are useful in elucidating novel mechanisms of tissue deformation and understanding self-organization in multicellular systems.
Organoids have allowed us to modify the environment of each tissue, leading to the discovery of new features of epithelial deformation as reviewed in this paper. Epithelial organoids are developed in a dish, where each organoid is surrounded by artificial gels and media. The composition of the gels and media in a dish is invariable that are replaced as the organoids attain different stages of development. In contrast, within the embryo, individual epithelial tissues are surrounded by other tissues spanning the ECM and body fluids. Importantly, because multiple tissues develop simultaneously, effects from tissue to tissue, including both biochemical and mechanical effects, also change dynamically. Moreover, because of the presence of several other neighboring tissues in 3D space, influences from the surroundings are spatially non-uniform. Therefore, it is still a challenge to understand how autonomous tissue development is or how much influence from the surroundings is required for each tissue to develop.
To address the autonomy of tissue formation, it is necessary to be able to precisely manipulate the environment of each organoid. Emerging engineering techniques can help us achieve this goal. Examples of engineering techniques that have already been employed on organoids include the addition of appropriate biochemical and mechanical cues to regulate cell-cell and cell-ECM interactions [41], promote vascularization [42], and control apical constriction using optical techniques [43]. Engineered biomaterials that can mimic the tissue geometry with patterned ECM stiffness and cell crowding suggest that tissue geometry has sufficient ability to instruct cells to differentiate as observed in the intestinal organoids, where budding crypts containing stem cells and Paneth cells are formed in the softened and curved regions, and absorptive epithelial cells in the non-budding regions [44]. Synthetic biology approaches would also aid in manipulating the autonomous processes of tissue formation. Artificial genetic circuits can program spatial self-organization of cells by coupling synthetic cell-cell signaling with an output of differential cell-cell adhesion [45]. Current protocols of organoid culture rely on the addition of morphogens and chemicals in culture, which, however, lacks spatiotemporal information about morphogen gradients within. A synthetic morphogen system would provide a new way to precisely program such positional information inside an organoid [46]. If such engineering methods can input spatiotemporally appropriate cues to organoids, it would be possible to exploit the self-organizing ability inherent in cells and manipulate the developmental processes of tissues.
Notably, organoids will open new avenues for understanding the autonomy of multicellular systems and their regulatory mechanisms. Especially, the organoid studies focusing on mechanical aspects will provide a bird's-eye view linking the conventional biological understandings of each gene and cell to organogenesis. This understanding includes multimodal (both biochemical and mechanical) interactions among cells. Furthermore, understanding organoid development, which recapitulates the stepwise tissue deformation of in vivo development will also lead to an understanding of time-dependent regulatory mechanisms revealing how each cellular dynamic event is scheduled to create complex 3D structures of tissues. Moreover, the precise manipulation of the organoid environment through engineering methods may be useful in quantitatively assessing the autonomy of the developmental process of each tissue. Studying the mechanics of organoid development will lead to understanding how each living cell can drive the development of an entire embryo by coordinating the hierarchical dynamics of cells from molecules to the whole body.
The authors declare no conflicts of interest.
SN and SO wrote the first draft of the manuscript. ST provided ideas. All authors reviewed and edited the manuscript and contributed to the final manuscript.
We thank Aya Matsuoka for her help with the illustrations and Keishi Kishimoto of the RIKEN Center for Biosystems Dynamics Research for his valuable comments. This work was supported by the Japan Science and Technology Agency (JST), CREST Grant No. JPMJCR1921 (S.O.) and PRESTO Grant No. JPMJPR2147 (S.T.); the Japan Agency for Medical Research and Development (AMED), Grant No. 21bm0704065h0002 (S.O.) and Grant No. 22bm0704048h0003 (S.T.); the Japan Society for the Promotion of Science (JSPS), KAKENHI Grants No. 21H01209, 22K18749, and 22H05170 (S.O.); and the World Premier International Research Center Initiative, Ministry of Education, Culture, Sports, Science and Technology (MEXT), Japan (S.O. and S.T.).
