In vitro 3D regeneration-like growth of human patient brain tissue

Abstract

In vitro culture of primary neurons is widely adapted with embryonic but not mature brain tissue. Here, we extended a previously developed bioengineered three-dimensional (3D) embryonic brain tissue model to resected normal patient brain tissue in an attempt to regenerate human neurons in vitro. Single cells and small sized (diameter < 100 μm) spheroids from dissociated brain tissue were seeded into 3D silk fibroin-based scaffolds, with or without collagen or Matrigel, and compared with two-dimensional cultures and scaffold-free suspension cultures. Changes of cell phenotypes (neuronal, astroglial, neural progenitor, and neuroepithelial) were quantified with flow cytometry and analyzed with a new method of statistical analysis specifically designed for percentage comparison. Compared with a complete lack of viable cells in conventional neuronal cell culture condition, supplements of vascular endothelial growth factor-containing pro-endothelial cell condition led to regenerative growth of neurons and astroglial cells from "normal" human brain tissue of epilepsy surgical patients. This process involved delayed expansion of Nestin+ neural progenitor cells, emergence of TUJ1+ immature neurons, and Vimentin+ neuroepithelium-like cell sheet formation in prolonged cultures (14 weeks). Micro-tissue spheroids, but not single cells, supported the brain tissue growth, suggesting importance of preserving native cell-cell interactions. The presence of 3D scaffold, but not hydrogel, allowed for Vimentin+ cell expansion, indicating a different growth mechanism than pluripotent cell-based brain organoid formation. The slow and delayed process implied an origin of quiescent neural precursors in the neocortex tissue. Further optimization of the 3D tissue model with primary human brain cells could provide personalized brain disease models.

A flexible 3-dimensional microelectrode array for in vitro brain models

“…[3][4][5] Mirroring the 3D architecture of the brain has also been a significant goal in recent years, with a number of model systems developed such as brain organoids [6][7][8][9] or the use of natural and/or synthetic 3D matrices to encapsulate neurons. [10][11][12][13][14] Several of these studies have shown that there are significant differences in the way neurons organize, develop, and communicate when cultured in a 3D matrix as compared to those grown on 2D substrates, and that 3D models more closely resemble the functional behavior of in vivo tissue. [15][16][17] Multi-electrode arrays (MEAs) provide a non-invasive method to monitor the electrophysiological activity of neurons in vitro over time.…”

3d Bioprinted Material That Recapitulates the Perivascular Bone Marrow Structure for Sustained Hematopoietic and Cancer Models

“…3D bioprinting has revolutionized the manner to create 3D in vitro culture systems, allowing for rapid fabrication of constructs with pre-defined geometries. However, despite its success in creating a wide array of tissues in silico (e.g., breast, brain, lung, bladder), faithfully replicating very soft tissues through 3D bioprinting has proven difficult due to their limited printability and low fidelity [67][68][69][70][71][72][73][74][75][76][77]. This is a significant obstacle because the degree of control over the composition, size, and structure of 3D bioprinted scaffolds can meaningfully influence physiological and pathological models, including screening for drug performance.…”

In Vitro 3D Modeling of Neurodegenerative Diseases

“…The scaffold-based method is often prepared using natural or synthetic hydrogels and can be carried out using two techniques: either the culture plate is first coated with a hydrogel, and the cells are then deposited on the surface and cultured under agitation to promote their adhesion and the formation of spheroids while in contact with the matrix (Figure 2C); or the hydrogel and the cells are deposited simultaneously on the plate to induce spheroid formation within the scaffold (Figure 2D) [94]. The goal of using this method is to reproduce the role of ECM in vivo, and, thus, promote cell-ECM interactions thanks to scaffold facilitating cell adhesion and migration [83].…”