Organoid
An organoid is a miniaturised and simplified version of an organ produced in vitro in three dimensions that mimics the key functional, structural, and biological complexity of that organ. It is derived from one or a few cells from a tissue, embryonic stem cells, or induced pluripotent stem cells, which can self-organize in three-dimensional culture owing to their self-renewal and differentiation capacities. The technique for growing organoids has rapidly improved since the early 2010s, and The Scientist named it one of the biggest scientific advancements of 2013. Scientists and engineers use organoids to study development and disease in the laboratory, for drug discovery and development in industry, personalized diagnostics and medicine, gene and cell therapies, tissue engineering, and regenerative medicine.
History
Attempts to create organs in vitro started with one of the first dissociation-reaggregation experiments where Henry Van Peters Wilson demonstrated that mechanically dissociated sponge cells can reaggregate and self-organize to generate a whole organism. In the subsequent decades, multiple labs were able to generate different types of organs in vitro through the dissociation and reaggregation of organ tissues obtained from amphibians and embryonic chicks. The formation of first tissue-like colonies in vitro was observed for the first time by co-culturing keratinocytes and 3T3 fibroblasts. The phenomena of mechanically dissociated cells aggregating and reorganizing to reform the tissue they were obtained from subsequently led to the development of the differential adhesion hypothesis by Malcolm Steinberg.With the advent of the field of stem cell biology, the potential of stem cells to form organs in vitro was realized early on with the observation that when stem cells form teratomas or embryoid bodies, the differentiated cells can organize into different structures resembling those found in multiple tissue types. The advent of the field of organoids started with a shift from culturing and differentiating stem cells in two dimensional media, to three dimensional media to allow for the development of the complex 3-dimensional structures of organs. Utilization of 3D media culture media methods for the structural organization was made possible with the development of extracellular matrices. In the late 1980s, Bissell and colleagues showed that a laminin rich gel can be used as a basement membrane for differentiation and morphogenesis in cell cultures of mammary epithelial cells.
Since 1987, researchers have devised different methods for 3D culturing, and were able to utilize different types of stem cells to generate organoids resembling a multitude of organs. In the 1990s, in addition to their role in physical support, the role of ECM components in gene expression by their interaction with integrin-based focal adhesion pathways was reported. In 2006, Yaakov Nahmias and David Odde showed the self-assembly of vascular liver organoid maintained for over 50 days in vitro. In 2008, Yoshiki Sasai and his team at RIKEN institute demonstrated that stem cells can be coaxed into balls of neural cells that self-organize into distinctive layers.
In 2009 the Laboratory of Hans Clevers at Hubrecht Institute and University Medical Center Utrecht, Netherlands, showed that single LGR5-expressing intestinal stem cells self-organize to crypt-villus structures in vitro without necessity of a mesenchymal niche, making them the first organoids. In 2010, Mathieu Unbekandt & Jamie A. Davies demonstrated the production of renal organoids from murine fetus-derived renogenic stem cells. In 2014, Qun Wang and co-workers engineered collagen-I and laminin based gels and synthetic foam biomaterials for the culture and delivery of intestinal organoids and encapsulated DNA-functionalized gold nanoparticles into intestinal organoids to form an intestinal Trojan horse for drug delivery and gene therapy. Subsequent reports showed significant physiological function of these organoids in vitro and in vivo.
Other significant early advancements included in 2013, Madeline Lancaster at the Institute of Molecular Biotechnology of the Austrian Academy of Sciences established a protocol starting from pluripotent stem cells to generate cerebral organoids that mimic the developing human brain's cellular organization. Meritxell Huch and Craig Dorrell at Hubrecht Institute and University Medical Center Utrecht demonstrated that single Lgr5+ cells from damaged mouse liver can be clonally expanded as liver organoids in Rspo1-based culture medium over several months. In 2014, Artem Shkumatov et al. at the University of Illinois at Urbana-Champaign demonstrated that cardiovascular organoids can be formed from ES cells through modulation of the substrate stiffness, to which they adhere. Physiological stiffness promoted three-dimensionality of EBs and cardiomyogenic differentiation.
Properties
Lancaster and Knoblich define an organoid as a collection of organ-specific cell types that develops from stem cells or organ progenitors, self-organizes through cell sorting and spatially restricted lineage commitment in a manner similar to in vivo, and exhibits the following properties:- it has multiple organ-specific cell types;
- it is capable of recapitulating some specific function of the organ ;
- its cells are grouped together and spatially organized, similar to an organ.
Process
Biochemical cues have been incorporated in 3D organoid cultures and with exposure of morphogenes, morphogen inhibitors, or growth factors, organoid models can be developed using embryonic stem cells or adult stem cells. Vascularization techniques can be utilized to embody microenvironments that are close to their counterparts, physiologically. Vasculature systems that can facilitate oxygen or nutrients to the inner mass of organoids can be achieved through microfluidic systems, vascular endothelial growth factor delivery systems, and endothelial cell-coated modules. With patient-derived induced pluripotent stem cells and CRISPR/Cas-based genome editing technologies, genome-edited or mutated pluripotent stem cells with altered signaling cues can be generated to control intrinsic cues within organoids.
Types
A multitude of organ structures have been recapitulated using organoids. This section aims to outline the state of the field as of now through providing an abridged list of the organoids that have been successfully created, along with a brief outline based on the most recent literature for each organoid, and examples of how it has been utilized in research.Cerebral organoid
A cerebral organoid describes artificially grown, in vitro, miniature organs resembling the brain. Cerebral organoids are created by culturing human pluripotent stem cells in a three-dimensional structure using rotational bioreactor and develop over the course of months. The procedure has potential applications in the study of brain development, physiology and function. Cerebral organoids may experience "simple sensations" in response to external stimulation and neuroscientists are among those expressing concern that such organs could develop sentience. They propose that further evolution of the technique needs to be subject to a rigorous oversight procedure. In 2023, researchers have built a hybrid biocomputer that combines laboratory-grown human brain organoids with conventional circuits, and can complete tasks such as voice recognition. Cerebral Organoids are currently being used to research and develop Organoid Intelligence technologies.Gastrointestinal organoid
Gastrointestinal organoids refer to organoids that recapitulate structures of the gastrointestinal tract. The gastrointestinal tract arises from the endoderm, which during development forms a tube that can be divided in three distinct regions, which give rise to, along with other organs, the following sections of the gastrointestinal tract:Organoids have been created for the following structures of the gastrointestinal tract:
Intestinal organoid
Intestinal organoids have thus far been among the gut organoids generated directly from intestinal tissues or pluripotent stem cells. One way human pluripotent stem cells can be driven to form intestinal organoids is through first the application of activin A to drive the cells into a mesoendodermal identity, followed by the pharmacological upregulation of Wnt3a and Fgf4 signaling pathways as they have been demonstrated to promote posterior gut fate. Intestinal organoids have also been generated from intestinal stem cells, extracted from adult tissue and cultured in 3D media. These adult stem cell-derived organoids are often referred to as enteroids or colonoids, depending on their segment of origin, and have been established from both the human and murine intestine.Intestinal organoids consist of a single layer of polarized intestinal epithelial cells surrounding a central lumen. As such, recapitulate the crypt-villus structure of the intestine, by recapitulating its function, physiology and organization, and maintaining all the cell types found normally in the structure including intestinal stem cells. Thus, intestinal organoids are a valuable model to study intestinal nutrient transport, drug absorption and delivery, nanomaterials and nanomedicine, incretin hormone secretion, and infection by various enteropathogens.
For example, Qun Wang's team rationally designed artificial virus nanoparticles as oral drug delivery vehicles with gut organoid-derived mucosal models and demonstrated a new concept of using newly established colon organoids as tools for high-throughput drug screening, toxicity testing, and oral drug development. Or recently, Sakib, S., and Zou, S. developed graphene oxide nanoparticles for delivering siRNA regulating expression of tumor necrosis factor-α, that aimed to treat intestinal organoids exhibiting an inflammatory phenotype.
Intestinal organoids also recapitulate the crypt-Villus structure to such a high degree of fidelity that they have been successfully transplanted to mouse intestines, and are hence highly regarded as a valuable model for research. One of the fields of research that intestinal organoids have been utilized is that of stem cell niche. Intestinal organoids were used to study the nature of the intestinal stem cell niche, and research done with them demonstrated the positive role IL-22 has in maintaining in intestinal stem cells, along with demonstrating the roles of other cell types like neurons and fibroblasts in maintenance of intestinal stem cells.
In the field of infection biology, different intestinal organoid-based model systems have been explored. On one hand, organoids can be infected in bulk by simply mixing them with the enteropathogen of interest. However, to model infection via a more natural route starting from the intestinal lumen, microinjection of the pathogen is required. In addition, the polarity of intestinal organoids can be inverted, and they can even be dissociated into single cells and cultured as 2D monolayers in order to make both the apical and basolateral sides of the epithelium more easily accessible. Intestinal organoids have also demonstrated therapeutic potential. In order to more accurately recapitulate the intestine in vivo, co-cultures of intestinal organoids and immune cells have been developed. Furthermore, organ-on-a-chip models combine intestinal organoids with other cell types such as endothelial or immune cells as well as peristaltic flow.