Mid-way between in vivo models and in vitro cell cultures, a new tool is emerging that could intensify medical discoveries: the ex vivo organoid model.
Over previous decades, in vitro cultures had been limited by flat, plastic and physiologically aberrant environments. However, research has recently been abandoning them in favor of more lifelike 3D constructions known as organoids. These in vitro culture representations of organs maintained in 3D possess architectures and functionalities similar to those of the tissues from which they are derived. Like miniature organs, they are formed from stem cells, or progenitor cells that are slightly more differentiated, which self-organize within an appropriate 3D environment (hydrogel, porous matrix, etc.). All of which is thanks to growth and differentiation factors, whose nature, quantity and window of exposure orient the future cell type.
"In Petri dishes, the cells stick to the bottom and remain immobile, but in hydrogel – which provides a 3D extracellular matrix – the cells tend to spontaneously aggregate into functional tissues, describes Nathalie Vergnolle, Director of the Digestive Health Research Institute* in Toulouse. There is no magic involved." Between 2D cultures and 3D organoids, only the architecture changes – and yet it makes all the difference. Just as a single cell does not behave in the same way as a group cell, 3D formation reveals functionalities and genetic characteristics that are more representative of living entities. But some scientists are not leaving organoid formation to chance. Jean-Christophe Fricain and the researchers from the BioPrint technology research accelerator (ART)** in Bordeaux have set themselves the challenge of bioprinting tissues. "The spontaneous self-organization of cells in 3D – while representing progress in relation to 2D – is still far from being able to reflect full organ functionality, hence the idea of using bioprinting to direct organoid formation." While the technology is more costly, it does give the mini organs more information and detail.
Tools that are both indispensable...
This quest for the most physiologically relevant representation of an organ in culture is not without reason. It legitimizes the use of organoids in certain scientific and medical discoveries. Originally designed for the study and comprehension of tissue organization and arrangement processes, they have since become excellent research tools. "Capable of being maintained in culture much longer than a biopsy and much more functional than the 2D cultures, organoids have an indisputable place as a model for understanding physiological and pathophysiological mechanisms, specifies Vergnolle, who manufactures human and mouse intestinal and bladder organoids in order to study intestinal and bladder diseases (IBD, colorectal cancer, cystitis, bladder cancer) and their treatments. Indeed, we noticed that the organoids derived from diseased tissues did not have the same morphologies and behaviors as those derived from healthy tissues."
It is also a tool that is becoming essential for understanding the toxicity and action of pharmacological molecules, which is necessary for new drugs to obtain market approval. Prévitox, a network of 35 laboratories and infrastructures dedicated to evaluating drug toxicity, funded by the French National Agency for Medicines and Health Products Safety (ANSM) and steered by Inserm, uses expertise in the field of organoids. The researchers particularly intend to combine different organoids, such as the liver and the kidneys, to model the molecules’ assimilation stages in the body and "obtain an integral view of drug metabolism and toxicity in a complex system", explains Bruno Clément, Director of the Nutrition, Metabolism and Cancer Institute*** in Rennes, who is heading up the adventure. This laboratory is developing pig liver and intestine organoids for the project. The researchers even envisage creating organoids from patient cells in order to test, adapt and optimize therapies to the individual. Still in its infancy, this personalized medicine could become possible, particularly for certain forms of cancer that respond poorly to treatment.
"To me it appears evident that the use of organoids will become commonplace in the field of pharmacotoxicology", assures Jean-Christophe Fricain. "Especially in combination with the new genome editing technologies such as CRISPR-Cas9 which are starting to enable research into the genetic consequences on human organoids, as we currently do on animals", adds Karim Si-Tayeb from the Thorax Institute**** in Nantes, who is using induced pluripotent stem cells from patients with high and low cholesterol differentiated into 3D hepatic structures in order to understand cholesterol regulation. "It is by expanding the work on humans that the organoids will be used to their full potential", adds Robert Barouki, Director of the Inserm Environmental Toxicity, Therapeutic Targets, Cellular Signaling and Biomarkers***** unit in Paris, who is studying pollutants and endocrine disruptors on human liver organoids.
Is the organoid therefore set to replace the other in vitro and in vivo models? The researchers see it more as a tool to supplement the range currently available.
... and complementary
Two-dimensional in vitro cell culture has been around for decades. Despite it reaching the limits of what it can do and despite 3D appearing to outshine it, its use will not necessarily be abandoned. "It remains comparatively simple and inexpensive and so will be useful as an initial approach" assures Si-Tayeb. As for animal research, even if the assembly of mini organs is starting to develop, this system could never be as comprehensive or representative as the animal model. Organoids enable us to improve our understanding of the complexity of living organisms without actually becoming a substitute for them. "These new technologies will not cut down on the need for animal research. However, they will refine our knowledge of living organisms and expand the structure and density of knowledge. It is clear that this will render animal research more effective, precise and targeted, reducing the number of animals used. But organoids will not replace whole animals. The organoid would need to become an animal itself in order to do that", affirms Bruno Verschuere, veterinarian for the French interprofessional animal research discussion and communication group Gircor.
Organoids are therefore set to occupy a crucial position in future research and bring about a technical, preclinical and clinical revolution. Will they be just as groundbreaking as vaccines, antibiotics and chemotherapy in the war against disease? We will need to wait a few more years in order to know for sure.
*unit 1220 Inserm/Inra/Ecole nationale vétérinaire/Université Toulouse 3, IRSD, Toulouse
**unit 1026 Inserm/Université de Bordeaux, Biotis, Bordeaux
***unit 1241 Inserm/Université de Rennes 1, Numecan, Rennes
****unit 1087 Inserm/CNRS/Université de Nantes - Nantes teaching hospital, Thorax Institute, Nantes
*****unit 1124 Inserm/Université Paris Descartes, T3S, Paris