Engineered brain organoids: Developing an improved and larger human brain model in vitro

• Autores: Theresa Sarah Petra Rothenbücher
• Directores de la Tesis: Alberto Martínez Serrano (dir. tes.), Marta Pérez Pereira (dir. tes.)
• Lectura: En la Universidad Autónoma de Madrid ( España ) en 2020
• Idioma: inglés
• Número de páginas: 143
• Títulos paralelos:
  • Organoides cerebrales modificados con técnicas de ingeniería: Desarrollo de un modelo de cerebro humano de mayor tamaño in vitro.
• Tribunal Calificador de la Tesis: Francisco Zafra (presid.), Josep Maria Canals Coll (secret.), Isabel Liste Noya (voc.)
• Programa de doctorado: Programa de Doctorado en Biociencias Moleculares por la Universidad Autónoma de Madrid
• Materias:
  • Ciencias de la vida
    • Biología molecular
• Enlaces
  • Tesis en acceso abierto en: Biblos-e Archivo
• Resumen

  • Brain organoids are neural tissues, that are generated in vitro by aggregating human pluripotent stem cells (hPSCs) and inducing them to self-organize towards brain-like tissue. They are considered to be a highly promising model for the human brain and have already been used in developmental and disease studies. Nevertheless, there are still several hurdles that need to be overcome in order to consider brain organoids as a robust and reliable model. Shortcomings of current brain organoids include a lack of reproducibility, the development of a necrotic tissue core due to exceedance of the oxygen- and nutrient diffusion limit, and the absence of some crucial features of the human brain, e.g. immune system, vascularization and developmental characteristics like gyrification.

    This doctoral thesis project focuses on addressing several of these issues. First of all for enhancing reproducibility and quality of the starting material we changed the culture condition of hPSCs to defined and xenogen-free. We furthermore worked on alternative hydrogel scaffolds (microenvironments) with brain-specific properties to promote the development of brain organoids. The key element of this thesis was the development of the engineered flat brain organoid (efBO) protocol, in which a 3D-printed polycaprolactone (PCL) scaffold was included into the culture of brain organoids as macroenvironment. In this way, we were able to freely tune their size and modify their shape into a flat morphology. efBOs possess advantageous diffusion conditions and thus are well supplied with oxygen and nutrients.

    Therefore, efBOs do not show necrotic tissue cores, but develop into a healthy neuronal tissue with a thick continuous neuroepithelial layer, when compared to regular, brain organoids.

    Moreover, the scaffold allows to tune the size of brain organoids directly from the start. By seeding cells onto 1 cm2 scaffolds, we were able to increase the size of brain organoids considerably. In addition, we observed that the very much enhanced surface-to-volume ratio leads to gyrification initiated by self-organization of the tissue around day 25. The efBO methodology provides hydrogel inclusion of the cells directly from day zero, which not only simplifies their culture, but means that beside medium changes, organoids do not need to be handled or manipulated at a later time point, allowing them to grow undisturbed. We consider the herein developed efBO technology as a next step towards the generation of a stable and reliable human brain model for drug screening applications, developmental and patterning studies. In the future the scaffold itself could act as factor-release system to induce spatial patterning, and even degrade with time if desired (4D culture, time being the 4th dimension)