Organoides en Odontología. Avances en este Nuevo Enfoque de Investigación: Revisión Sistemática Exploratoria

Los Organoides (O) son un tipo de cultivo celular 3D, que reproducen las características morfológicas y funcionales de diversos órganos o tejidos en un entorno in vivo. Se logran a través de la proliferación y diferenciación de Células Madres (CM) en distintas líneas celulares con capacidad de autoorganizarse. Son capaces de reproducir forma, función, expresión génica o repuesta a estímulos de la misma forma que el órgano original. Esto le ha permitido servir de base para múltiples investigaciones en el ámbito médico y odontológico. En los últimos años, se ha podido recrear con éxito, prácticamente, todos los órganos de nuestro cuerpo, como pulmones, hígado, tracto reproductivo, cerebro y muchos otros (Bartfeld, 2021). De la misma forma, son varias las líneas de investigación odontológicas desarrolladas. En específico, la creación de O de órganos orales como dientes y glándulas salivales, son las más reportadas (Oshima et al., 2017). Sin embargo, no son del común conocimiento del odontólogo general. Esta revisión sistemática exploratoria, tiene como objetivo presentar una visión general de la evidencia acumulada, determinado las áreas odontológicas de investigación, así como sus resultados. La investigación odontológica, en base al uso de O, es de alta calidad y de vanguardia, mostrando resultados prometedores, que auguran un gran futuro, tanto para la odontología como para los pacientes.

Organoids (O) are a type of 3D cell culture, which reproduce the morphological and functional characteristics of various organs or tissues in an in vivo environment. They are achieved through the proliferation and differentiation of Stem Cells (SC) into different cell lines with the ability to self-organize. They are capable of reproducing form, function, gene expression, or responses to stimuli in the same way as the original organ. This has allowed it to serve as the basis for multiple investigations in the medical and dental field. In recent years, it has been possible to successfully recreate practically all human organs, such as the lungs, liver, reproductive tract, brain and many others (Bartfeld, 2021). In the same way, there are several lines of dental research developed, specifically, the creation of O from oral organs such as teeth and salivary glands, are the most reported (Oshima et al., 2017). However, they are not common knowledge of the general dentist. This exploratory systematic review aims to present an overview of the accumulated evidence, determining the dental research areas, as well as their results. Dental research, based on the use of O, is of high quality and cutting-edge, showing promising results and a favorable future, both for dentistry and for patients.

[Alternative methods to animal testing, present and future]. / Les méthodes alternatives à l'expérimentation animale, présent et futur.

Alternative methods to animal testing are used in fundamental and clinical research, for the realization of studies for regulatory purposes, and also screening operations in the development of new molecules. They are based on in vitro (cell models) or in silico (mathematical models) replacement methods. They have been largely promoted by the 3Rs rule (Replace, Reduce, Refine) which aims at regulating animal experimentation. For biomedical research, these different methods are valuable tools for better understanding the physiology of organisms and the mechanisms of the effects of chemicals and physical agents on them.

Title: Les méthodes alternatives à l'expérimentation animale, présent et futur. Abstract: Les méthodes alternatives à l'expérimentation animale sont utilisées en recherche fondamentale et clinique, pour la réalisation d'études à visée réglementaire et d'opérations de criblage en matière de développement de nouvelles molécules. Elles reposent sur des procédures de remplacement in vitro (modèles cellulaires) ou in silico (modèles mathématiques). Les méthodes alternatives ont été largement promues par la règle des 3R (Remplacer, Réduire, Raffiner) qui vise à encadrer l'expérimentation animale. Dans le domaine de la recherche, ces différentes méthodes sont des outils précieux qui permettent de mieux comprendre la physiologie des organismes et les mécanismes d'action des agents chimiques et physiques sur ces derniers.

Organoides de origen humano como modelo fisiopatológico de evaluación terapéutica de fármacos en COVID-19 / Organoids as a physiopathology model and therapeutic drug evaluation tool in COVID-19

Introducción: La pandemia causada por SARS-CoV-2 ha impactado al mundo gravemente en diversos ámbitos y con ello ha surgido la necesidad de contar con herramientas con mayor relevancia fisiológica para investigar patologías complejas como el COVID-19. Los organoides son un modelo experimental con características únicas como la capacidad de autoformar una estructura tridimensional utilizando células en cultivo. Sobre esta base, surge la siguiente pregunta ¿son los organoides un modelo experimental factible para reflejar la fisiopatología del COVID-19 y evaluar la eficacia de fármacos que limiten su progresión? Metodología: Para abordar esta interrogante, esta revisión plantea el analizar la validez de los organoides como modelo experimental y verificar su utilidad en la evaluación de fármacos para el COVID-19. Para cumplir estos objetivos se realizó una revisión sistemática cualitativa de la literatura, a través de una búsqueda en PubMed con el término 'COVID-19 and stem cells and organoids' y también en un número especial de la revista Cell. Resultados: Se organizaron los resultados relevantes por sistema fisiológico y en la evaluación de fármacos. Los organoides más empleados para estudios de COVID-19 correspondieron a tejido respiratorio, nervioso y digestivo. Algunos resultados encontrados en la revisión fueron similares a aquellos obtenidos a partir de tejidos de pacientes COVID-19 o autopsias, encontrándose hallazgos relevantes como la posible disrupción de la barrera epitelial del sistema nervioso por infección del plexo coroideo. También se logró observar efectividad de fármacos que posteriormente pasaron a ser aprobados y utilizados exitosamente en pacientes. Conclusión: Los organoides se pueden componer a partir de diferentes tipos celulares y bajo diferentes protocolos experimentales, siendo relevante la lectura crítica de los artículos científicos para decidir si sus resultados son extrapolables a la fisiopatología de la enfermedad.

Introduction: The pandemic caused by SARS-CoV-2 has impacted the world severely in several aspects and has created the need for research tools to study the COVID-19 disease. Organoids are experimental models with unique characteristics, like the ability to self-assemble in a tridimensional structure. Based on this, the following question arises: are organoids an experimental model suitable to reflect the physiopathology of COVID-19 and to allow the evaluation of the efficacy of drugs that limit its progression? Methods: To approach this question, this review aimed to analyze the validity of organoids as an experimental model and verify their utility in COVID-19 drug evaluation. To resolve these objectives, a qualitative systematic review was done through a PubMed search with the terms 'COVID-19 and stem cells and organoids' and on a special issue of the Cell Journal. Results: The results were organized by physiologic system and therapeutic drug evaluation. The most utilized tissues for the COVID-19 study were respiratory, nervous, and digestive. Some results found in the review were like those obtained from COVID-19 patient tissue or autopsies, finding some relevant discoveries like the possibility of the choroid plexus disruption in the nervous system caused by the infection. Efficacy was also observed in approved drugs and used later in patients successfully. Conclusion: Organoids might be composed starting with different cell types and under a variety of experimental protocols, being relevant the critical reading of the scientific literature to decide whether their results can be extrapolated to the pathophysiology of the disease

Organoides de cérebro como modelos de doenças de Alzheimer e Parkinson: uma revisão narrativa sobre as perspectivas para medicina regenerativa e personalizada / Brain organoids as models of Alzheimer's and Parkinson's diseases: a narrative review on perspectives for regenerative and personalized medicine

Há muitos anos a cultura celular bidimensional (2D) é utilizada como modelo de estudo de doenças, possuindo grande importância na medicina regenerativa, apesar de ainda conter limitações significativas. A fim de contornar essas limitações, a cultura celular tridimensional (3D) propõe uma organização mais complexa e sustentável que pode ser produzida a partir de células-tronco adultas (ASCs), células-tronco embrionárias (ESCs) ou células-tronco pluripotentes induzidas (iPSCs). A cultura 3D possibilitou o cultivo de células em um ambiente mais próximo do fisiológico, levando à formação de distintos tecidos órgãos-específicos. Em outras palavras, a cultura de células 3D possibilita a criação de estruturas orgânicas muito semelhantes aos órgãos de um ser humano, tanto estruturalmente, quanto funcionalmente. Desse modo, tem-se o que é chamado de organoides. O uso dos organoides tem crescido exponencialmente em ambientes in vitro, permitindo a análise e observação dos diversos fenômenos fisiológicos existentes. Como exemplo, pode-se citar os organoides cerebrais ("mini-brains") reproduzidos in vitro buscando delinear as peculiaridades e complexidades do cérebro humano, com o objetivo de compreender algumas disfunções neurológicas que acometem esse sistema, como as duas principais doenças neurodegenerativas: Doenças de Alzheimer e Parkinson. Portanto, os organoides cerebrais podem permitir notável avanço da medicina regenerativa aplicada a doenças neurodegenerativas, já que esses "mini-brains" podem ser produzidos a partir de células do próprio paciente. Isso permitirá intervenções personalizadas, como testagens farmacológicas, a fim de definir qual seria o melhor tratamento medicamentoso. Consequentemente, essa tecnologia pode permitir terapias mais eficientes e individualizadas - o que é fundamental para a Medicina Personalizada (AU).

For many years, two-dimensional (2D) cell culture has been used as a model to study diseases, having great importance in regenerative medicine, despite still having significant limitations. In order to circumvent these limitations, three-dimensional (3D) cell culture proposes a more complex and sustainable organization that can be produced from adult stem cells (ASCs), embryonic stem cells (ESCs) or induced pluripotent stem cells (iPSCs). The 3D culture enabled the cultivation of cells in an environment closer to the physiological one, leading to the formation of different organ-specific tissues. In other words, 3D cell culture makes it possible to create organic structures very similar to the organs of a human being, both structurally and functionally. In this way, we have what are called organoids. The use of organoids has grown exponentially in in vitro environments, allowing the analysis and observation of the various existing physiological phenomena. As an example, we can mention the brain organoids ("mini-brains") reproduced in vitro, seeking to delineate the peculiarities and complexities of the human brain, in order to understand some neurological dysfunctions that affect this system, such as the two main neurodegenerative diseases: Alzheimer's and Parkinson's Diseases. Therefore, brain organoids may allow a remarkable advance in regenerative medicine applied to neurodegenerative diseases, as these "mini-brains" can be produced from the patient's own cells. This will allow for personalized interventions, such as drug testing, in order to define what would be the best drug treatment. Consequently, this technology can enable more efficient and individualized therapies - which is fundamental for Personalized Medicine (AU).

Alteraciones de la expresión génica en la infección por SARS-CoV-2 en distintos modelos de estudio / Alterations of gene expression in SARS-CoV-2 infection in different study models

Objetivo: La pandemia por SARS-CoV-2 ha supuesto un auténtico reto para el mundo científico debido a la rápida transmisión y elevada mortalidad que produce este nuevo coronavirus. La enfermedad asociada se ha denominado COVID-19 y abarca desde casos asintomáticos hasta graves que evolucionan rápidamente a síndrome de distrés respiratorio agudo, alteraciones multisistémicas y la muerte. La comunidad científica ha aunado esfuerzos para tratar de conocer mejor el proceso fisiopatológico de la infección con la intención de combatir de forma más eficaz la enfermedad. En este trabajo presentamos un estudio para conocer las alteraciones de la expresión génica provocadas por la infección. Metodología: Se han usado tres modelos de estudio distintos: cultivos de células epiteliales bronquiales, organoides de las vías respiratorias y muestras obtenidas de autopsias en pacientes, con y sin infección por SARS-CoV-2. Se han analizado los perfiles de expresión alterados por la infección en cada modelo, así como las categorías funcionales enriquecidas. Resultados: Solo 4 genes son comunes en los tres tipos de modelos de estudio, siendo el modelo de autopsias el más dispar. Dentro de los genes comunes en los modelos de cultivo celular y organoide de pulmón encontramos funciones relacionadas con procesos inflamatorios. Conclusiones: Los estudios in vitro son un buen modelo para tener una foto fija de las alteraciones en los patrones de infección, mientras que las autopsias no son un buen modelo debido al sesgo provocado por la necrosis. (AU)

Short summary: Three different study models have been used to study the gene expression profiles produced by SARS-CoV-2: bronchial epithelial cell cultures, airway organoids, and autopsy samples from patients with and without SARS-infection. CoV-2. Basis: The SARS-CoV-2 pandemic has been a real challenge for the scientific world due to the rapid transmission and high mortality caused by this new coronavirus. The associated disease has been named COVID-19 and ranges from asymptomatic to severe cases that rapidly progress to acute respiratory distress syndrome, multisystem disorders, and death. The scientific community has joined efforts to try to better understand the pathophysiological process of the infection with the intention of combating the disease more effectively. In this work we present a study to determine the alterations in gene expression caused by the infection. Methods: Three different study models have been used: bronchial epithelial cell cultures, airway organoids, and samples obtained from autopsies in patients with and without SARS-CoV-2 infection. The expression profiles altered by the infection in each model have been analyzed, as well as the functional categories enriched. Results: Only 4 genes are common in the three types of study models, the autopsy model being the most disparate. Within the common genes in cell and organoid culture models of the lung, we find functions related to inflammatory processes. Conclusions: In vitro studies are a good model to have a snapshot of alterations in infection patterns, while autopsies are not a good model due to bias caused by necrosis. (AU)

[3D Tumor organoid models produced by cellular capsules technology CCT]. / Production d'organoïdes tumoraux 3D par la technologie des capsules cellulaires TCC.

Monolayer cultures of cell lines and derived-patient cells have long been the in vitro model of choice in oncology. In particular, these models have made it possible to decipher the mechanisms that determine tumor proliferation and invasion. However these 2D models are insufficient because they do not take into account the spatial organization of cells and their interactions with each other or with the extracellular matrix. In the context of cancer, there is a need to develop new 3D (tumoroid) models in order to gain a better understanding of the development of these pathologies but also to assess the penetration of drugs through a tissue and the associated cellular response. We present here the cell capsule technology (CCT), which allows the production of different tumoroid models: simple or more complex 3D culture models including co-culture of tumor cells with components of the microenvironment (fibroblasts, matrix, etc.). The development of these new 3D culture systems now makes it possible to propose refined physiopathological models that will allow the implementation of improved targeted therapeutic strategies.

[Normal organoids and their applications in cancer research]. / Les organoïdes normaux et leurs applications dans la recherche sur le cancer.

Three-dimensional (3D) culture of organoids from primary cells (wild type) or tumoroids from tumor cells, is used to study the physiological mechanisms in vivo, in order to model normal or tumor tissues more accurately than conventional two-dimensional (2D) culture. The features of this 3D culture, such as the three-dimensional structure, the self-renewal capacity and differentiation are preserved and appropriate to cancer study since their cellular characteristics are very similar to in vivo models. Here, we summarize the recent advances in the rapidly evolving field of organoids and their applications to cancer biology, clinical research and personalized medicine.

[Spheroids to organoids: Solid cancer models for anticancer drug discovery]. / Des sphéroïdes aux organoïdes : modèles de cancers solides pour la découverte de molécules anticancéreuses.

Cell culture is an important and necessary technology in oncology research. Currently, two-dimensional (2D) cell culture models are the most widely used, but they cannot reproduce the complexity and pathophysiology of tumors in vivo. This may be a major cause of the high rate of attrition of anticancer drugs entering clinical trials, the rate of new anticancer drugs entering the market being less than 5 %. One way to improve the success of new cancer drugs in the clinic is based on the use of three-dimensional (3D) cell culture models, more able to represent the complex environment and architecture of tumors. These 3D culture systems are also a powerful research tool for modeling the evolution of cancer from early stages to metastasis. Spheroids and organoids, the most adaptable models among 3D culture systems, are beginning to be used in pharmaceutical research and personalized medicine. In this article, we review the use of spheroids and organoids by highlighting their differences, discussing their impact on drug development, and looking at future challenges.

New methodologies for old problems: tridimensional gastrointestinal organoids and guts-on-a-chip / Novas metodologias para velhos problemas: organoides gastrointestinais e guts-on-a-chip tridimensionais

ABSTRACT Objectives: The present review intended to present a critical overview of the methodological and experimental advances concerning tridimensional cell culture models within the scope of gastrointestinal research. Methods: A literature review was performed and some of the main published articles in the area were mentioned. Main results: Classic studies and high impact results were presented, starting from the pioneer works with gastrointestinal organoids, with a small gut organoid, to the achievement of guts-on-a-chip and multi-organ-chips. It was also discussed which implications the construction of such co-cultures bring, as well as future applications arising from these new methodologies. Conclusions: Despite the still discrete number of publications, in quantitative terms, there are qualitative promising and consistent results addressing physiopathological aspects and new therapeutic perspectives of tridimensional in vitro cultures in the gastroenterology field. It is expected, thus, that such new methodological approaches, including organoids and guts-on-a-chip, may contribute decisively to the advance in knowledge on basic aspects, as well as on the translation to new therapeutic approaches in gastrointestinal diseases.

RESUMO Objetivos: A presente revisão visou apresentar uma abordagem crítica dos avanços metodológicos e experimentais referentes a modelos de cultura celular tridimensionais no âmbito do sistema gastrintestinal. Métodos: Foi realizada revisão da literatura com ênfase nos principais artigos publicados na área. Resultados principais: São apresentados trabalhos clássicos e resultados de maior impacto, desde os trabalhos pioneiros com organoides do sistema gastrintestinal, com intestino delgado, até a obtenção de guts-on-a-chip e multi-organ-chips. Discutiu-se, ainda, as implicações decorrentes da elaboração de tais co-culturas, bem como as futuras aplicações decorrentes dessas novas metodologias. Conclusões: Apesar do número ainda discreto de publicações, em termos quantitativos, há, qualitativamente, resultados promissores e consistentes abordando aspectos fisiopatológicos e de novas perspectivas terapêuticas em gastrenterologia decorrentes das culturas tridimensionais in vitro. É esperado, portanto, que essas novas abordagens metodológicas incluindo organoides e guts-on-a-chip possam contribuir decisivamente para o avanço no conhecimento sobre de aspectos básicos, bem como para a translação do conhecimento para novas abordagens terapêuticas em doenças gastrintestinais.