Desarrollo de un andamiaje de matriz extracelular para modelos de cáncer óseo.
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Date
2024
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Universidad de Concepción
Abstract
Los tumores y las metástasis en el tejido óseo son un problema sanitario relevante debido a su impacto en la salud y el bienestar. Los tumores óseos metastásicos y tumores malignos con un diagnóstico tardío presentan una tasa de supervivencia reducida, siendo inferior a un año para los pacientes metastásicos. Es urgente la necesidad de desarrollar medidas complementarias para mejorar la eficacia de las terapias farmacológicas actuales y también el reducir la duración del tratamiento para evitar una carga económica creciente en los pacientes y en el sistema sanitario. Los modelos óseos tridimensionales in vitro tienen la capacidad de simular al microambiente óseo en el laboratorio con la posibilidad de introducir células derivadas de pacientes a partir de biopsias de tejido de cáncer óseo para estudiar tratamientos personalizados que puedan conducir a terapias más efectivas y personalizadas para los pacientes. Sin embargo, para lograr una traslación clínica de los resultados in vitro se requiere de un biomaterial que pueda reproducir la fisiopatología del tejido óseo en sus propiedades que permita a las células adherirse, sobrevivir, expandirse y generar procesos de relación con la matriz extracelular.
El objetivo de este trabajo fue el desarrollar un andamio tridimensional que simule la matriz extracelular ósea con la capacidad de biofabricación de modelos de tejido óseo in vitro que simulen patologías tumorales. Para esto se trabajó con células asociadas al cáncer óseo y otras relevantes para conformar modelos más complejos, tales como de cáncer de mama (MCF7), osteosarcoma (Saos2), pre-osteoblastos (MC3T3-E1) y cultivo primario (células madre mesenquimales derivadas de tejido adiposo). El modelo de ingeniería de tejidos se basó en un enfoque de hidrogel de gelatina metacrilada con el empleo de un componente biológico, en este caso micropartículas descelularizadas de hueso humano como parte de un biomaterial compuesto que fue validado mediante caracterización físico-químicas y biológicas.
Se desarrolló un hidrogel compuesto por gelatina metacrilada y micropartículas óseas humanas con comportamiento pseudoplástico. Estas propiedades fueron validadas mediante procedimientos de bioimpresión, mientras que la presencia de micro y nanopartículas óseas se confirmó mediante microscopía electrónica. La biocompatibilidad del hidrogel se evaluó a través de diversos ensayos para la cuantificación de la citocompatibilidad, incluyendo análisis por imágenes y mediciones moleculares. No se registraron efectos tóxicos, lo que indica que las células pudieron adherirse y proliferar dentro del andamiaje, manteniendo su viabilidad durante un período de hasta 30 días en cultivos celulares tridimensionales. Además, las diferentes validaciones demostraron ser compatible con procesos clínicos para distintos tipos de análisis celulares y moleculares. El hidrogel compuesto desarrollado en este trabajo contribuye a la inovación en la ingeniería de tejidos en el área músculo esqueletal, específicamente de biofabricación de tejido óseo y al reemplazo de modelos animales para estudios del tejido óseo por la sustitución de modelos in vitro.
Bone tissue tumours and metastases are a significant healthcare problem due to their impact on health and well-being. Metastatic bone tumours and late-diagnosed malignant tumours have a reduced survival rate, being less than one year for metastatic patients. There is an urgent need to develop complementary measures to improve the efficacy of current drug therapies and also to reduce treatment time to avoid an increasing economic burden on patients and the healthcare system. Three-dimensional in vitro bone models can simulate the bone microenvironment in the laboratory, offering the possibility of introducing patient-derived cells from bone cancer tissue biopsies to study personalized treatments that may lead to more effective and personalized therapies for patients. However, achieving clinical translation of in vitro results requires a biomaterial that can reproduce the pathophysiology of bone tissue in its properties, allowing cells to adhere, survive, expand, and generate processes related to the extracellular matrix. The objective of this work was to develop a three-dimensional scaffold that simulates the bone extracellular matrix and is capable of biofabricating in vitro bone tissue models that mimic tumour pathologies. To this end, we worked with cells associated with bone cancer and other relevant cells to form more complex models, such as breast cancer (MCF7), osteosarcoma (Saos2), pre-osteoblasts (MC3T3-E1), and primary cultures (adipose-derived mesenchymal stem cells). The tissue engineering model was based on a methacrylated gelatin hydrogel approach using a biological component, in this case decellularized human bone microparticles as part of a composite biomaterial that was validated through physicochemical and biological characterization. A hydrogel composed with pseudoplastic behaviour was developed. These properties were validated using bioprinting procedures, while the presence of bone micro- and nanoparticles was confirmed by electron microscopy. The hydrogel's biocompatibility was evaluated through various cytocompatibility quantification assays, including imaging análisis and molecular measurements. No toxic effects were recorded, indicating that cells were able to adhere and proliferate within the scaffold, maintaining their viability for up to 30 days in three dimensional cell cultures. Furthermore, the various validations demonstrated its compatibility with clinical processes for various types of cellular and molecular analyses. The composite hydrogel developed in this work contributes to innovation in musculoskeletal tissue engineering, specifically in the biofabrication of bone tissue, and to the replacement of animal models for bone tissue studies with in vitro models.
Bone tissue tumours and metastases are a significant healthcare problem due to their impact on health and well-being. Metastatic bone tumours and late-diagnosed malignant tumours have a reduced survival rate, being less than one year for metastatic patients. There is an urgent need to develop complementary measures to improve the efficacy of current drug therapies and also to reduce treatment time to avoid an increasing economic burden on patients and the healthcare system. Three-dimensional in vitro bone models can simulate the bone microenvironment in the laboratory, offering the possibility of introducing patient-derived cells from bone cancer tissue biopsies to study personalized treatments that may lead to more effective and personalized therapies for patients. However, achieving clinical translation of in vitro results requires a biomaterial that can reproduce the pathophysiology of bone tissue in its properties, allowing cells to adhere, survive, expand, and generate processes related to the extracellular matrix. The objective of this work was to develop a three-dimensional scaffold that simulates the bone extracellular matrix and is capable of biofabricating in vitro bone tissue models that mimic tumour pathologies. To this end, we worked with cells associated with bone cancer and other relevant cells to form more complex models, such as breast cancer (MCF7), osteosarcoma (Saos2), pre-osteoblasts (MC3T3-E1), and primary cultures (adipose-derived mesenchymal stem cells). The tissue engineering model was based on a methacrylated gelatin hydrogel approach using a biological component, in this case decellularized human bone microparticles as part of a composite biomaterial that was validated through physicochemical and biological characterization. A hydrogel composed with pseudoplastic behaviour was developed. These properties were validated using bioprinting procedures, while the presence of bone micro- and nanoparticles was confirmed by electron microscopy. The hydrogel's biocompatibility was evaluated through various cytocompatibility quantification assays, including imaging análisis and molecular measurements. No toxic effects were recorded, indicating that cells were able to adhere and proliferate within the scaffold, maintaining their viability for up to 30 days in three dimensional cell cultures. Furthermore, the various validations demonstrated its compatibility with clinical processes for various types of cellular and molecular analyses. The composite hydrogel developed in this work contributes to innovation in musculoskeletal tissue engineering, specifically in the biofabrication of bone tissue, and to the replacement of animal models for bone tissue studies with in vitro models.
Description
Tesis presentada para optar al grado de Doctor en Biotecnología Molecular
Keywords
Matriz extracelular, Cáncer de huesos, Materiales biocompatibles