Macroporous heparin-based microcarriers allow long-term 3D culture and differentiation of neural precursor cells

Abstract

Adult neurogenesis and the neurogenic niche in the dentate gyrus are subjects of much research interest. Enhancing our knowledge of this niche process and the role played by this unique microenvironment would further our understanding of plasticity and its relevance for cognition in health and disease. The complex three-dimensional (3D) nature of the niche microenvironment is poorly recapitulated in current cell culture experimental procedures. Neural precursor cells (NPCs) are cultured either on two-dimensional (2D) surfaces, where cells quickly reach confluency and passaging is required, or as 3D neurospheres, with the limitation of poor diffusion of nutrients and thus partial differentiation of cells over time. Herein, we culture NPCs on microscale scaffolds termed microcarriers, composed of poly(ethylene glycol) and heparin, designed to more closely represent the 3D environment of the neurogenic niche. The interconnected macroporous structure of the microcarriers allows NPCs to attach to their pore walls with subsequent continuous proliferation (analyzed up to 28 days) without formation of a necrotic core. Removal of basic fibroblast growth factor and epidermal growth factor from the culture medium results in differentiation of the NPCs. Unlike 2D culture, a high percentage of neurons was achieved on the microcarriers (22% MAP2 positive cells) indicating that these 3D microscale scaffolds give a more conducive environment for neuronal differentiation. Microcarrier culture of NPCs allows long-term cell expansion and better differentiation, which provides superior culture conditions for studying/modelling the neurogenic niche.

Multi-Functionalized Self-Assembling Peptides as Reproducible 3D Cell Culture Systems Enabling Differentiation and Survival of Various Human Neural Stem Cell Lines

“…Also, a possible refinement of the current technique to overcome the progressive increment of cell apoptosis, could be the usage of microfluidic devices (Mofazzal Jahromi et al, 2019) or spinner flasks (Newland et al, 2020) to increase oxygen and nutrients exchange inside the 3D culture system. In turn, this could also ameliorate cell maturation yielding out more entangled, and likely electrically active neuronal networks to be confirmed by electrophysiology tests.…”

Poly(ethylene glycol) based nanotubes for tuneable drug delivery to glioblastoma multiforme

“…Confocal microscope live/dead imaging was performed 3 days postinjection using calcein AM solution (PromoKine) and propidium iodide (Sigma) as previously described. 34,35 Experiments were performed in quadruplicate.…”

Chondroitin Sulfate-Based Cryogels for Biomedical Applications

“…As the synthesized CS-based cryogels are biodegradable and hemocompatible, it is suggested here that CS-based cryogels can be safely used for tissue engineering, or bioreactor, cell separation or scaffolding materials. As cryogels possess interconnected super porous material, they can also be used for neural precursor cell microcarriers [ 47 ], and are useful for nerve guidance conduits, which are designed to guide nerves. The CS-based cryogels provide great potential in not only neurological applications due to porosity, permeability, adhesion of growth factors, multi-channel structure and polymer binding ability [ 48 ], but also metal ions such as Cu(II) and Fe(II) binding capability.…”