An international research team from the Autonomous University of Madrid and the Technical University of Denmark (TUD) reported that it used 3D printing to create scaffolds for engineered flat brain organoids. The scaffolds allowed the brain organoid size to be significantly increased and, after 20 days, self-generated folding was observed, according to the scientists who published their study, “Next generation human brain models: engineered flat brain organoids featuring gyrification,” in Biofabrication.
“Brain organoids are considered to be a highly promising in vitro model for the study of the human brain and, despite their various shortcomings, have already been used widely in neurobiological studies. Especially for drug screening applications, a highly reproducible protocol with simple tissue culture steps and consistent output, is required,” the investigators wrote.
“Here we present an engineering approach that addresses several existing shortcomings of brain organoids. By culturing brain organoids with a polycaprolactone scaffold, we were able to modify their shape into a flat morphology. Engineered flat brain organoids (efBOs) possess advantageous diffusion conditions and thus their tissue is better supplied with oxygen and nutrients, preventing the formation of a necrotic tissue core.”
“Moreover, the efBO protocol is highly simplified and allows to customize the organoid size directly from the start. By seeding cells onto 12 by 12 mm scaffolds, the brain organoid size can be significantly increased. In addition, we were able to observe folding reminiscent of gyrification around day 20, which was self-generated by the tissue.”
“To our knowledge, this is the first study that reports intrinsically caused gyrification of neuronal tissue in vitro. We consider our efBO protocol as a next step towards the generation of a stable and reliable human brain model for drug screening applications and spatial patterning experiments.”
“The lack of vascularization leads to diffusion limitations for nutrients and oxygen, resulting in a necrotic tissue core for organoids larger than approximately 500 mm,” said one of the lead authors, Theresa Rothenbücher, a PhD student at the University of Madrid, in addressing one of the shortcomings of existing brain organoids. “In an attempt to solve this problem, brain organoids have been vascularized. While including endothelial cells in the culture system increases the complexity of the model, the generated vessel structures show no functionality (blood flow) in vitro. We are able to circumvent this issue by applying bioengineering techniques.”
According to another lead author, Hakan Gürbüz, from the TUD, “By culturing brain organoids with a polycaprolactone (PCL) scaffold, we were able to modify their shape into a flat morphology. efBOs possess advantageous diffusion conditions and thus their tissue is better supplied with oxygen and nutrients, preventing the formation of a necrotic tissue core. The shift from a spherical to a flat shape leads to a significant increase in size and surface-to-volume ratio of the brain organoids.”
eFBOs also offer increased potential to create biologically relevant systems, due to the complexity of the models that they enable, the research team pointed out. Ensuring the long-term viability of these models is a major aim of this branch of research, which has been difficult until now. Flat organoids appear to address the problem of longevity by avoiding the formation of necrotic tissue.
The 3D printing of scaffolds was key to overcoming the shape limitations of the previous spherical models, emphasized contributing TDU author Jenny Emneus, PhD.
“By introducing a 3D-printed scaffold into the culture protocol, the size of the brain organoids and the tissue density and thickness can be tuned,” she said, adding that the resulting model showed consistent formation of neuroepithelial folding resembling gyrification.
