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International Journal of Bioprinting 3D model of neurogenesis in Alzheimer’s disease

Given that GEL liquefies at 37°C, a hydrogel containing The electrical conductivity of the hydrogel was also
more GEL than ALG would gradually deteriorate evaluated. This parameter is essential for understanding
during incubation and disintegrate within a few days, the physicochemical relationship between biological and
shortening the time for cell culture. Therefore, we utilized artificial systems, mimicking physiological electrical
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a hydrogel with greater ALG content to prevent such processes. In vivo, electrical conductivity facilitates
drawbacks. Although our hydrogel exhibited viscoelastic- neuronal communication 87,88 and directs NSC maturation
liquid behavior throughout our oscillatory rheological into functional neuronal networks. Even without
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measurements, it could still be printed and maintain its externally applied electric fields, conductive hydrogels
original shape post-extrusion, indicating a promising promote neurite outgrowth and NSC differentiation,
profile for use as a bioink for cellular culture in a 3D efficiently contributing to the native CNS support for
environment. neural cells. At room temperature, the hydrogel we
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The chemical structure composition of the bioprinted produced has an electrical conductivity of 27.46 mS/
hydrogel (crosslinked or not), the crosslink agent CaCl , cm. Conductivity measurements for native CNS tissues
2
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and the polymeric components of the hydrogel, ALG, and (matrix and cells) ranged from 2 to 7 mS/cm, indicating a
GEL, were analyzed by ATR-FTIR spectroscopy (Figure closer conductivity of the bioink to the brain tissue. High-
3A). In the ALG spectrum, the transmission bands at 1600 conductive biomaterials for neural applications could reach
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and 1410 cm represent the asymmetric and symmetric up to 17 S/cm to specifically electro-stimulate NSCs and
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vibrations of COO , respectively, while the bands at 1090 up to 7500 S/cm for further biomedical applications. 92
–
and 1030 cm are attributed to C-O and C-O-C bonds. 78,79 3.2. Aβ oligomers incorporated into the bioink
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In GEL, the bands observed at 1630, 1543, and 1238 cm induce a decrease in cell viability and an increase in
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are attributed to the amide I, II, and III vibration peaks, oxidative stress
respectively. The crosslinked hydrogel exhibited all the Neurospheres are free-floating populations derived
bands mentioned above. However, the amide I band of from neural stem and progenitor cells isolated from the
GEL is obscured by the strong absorption band at 1600 neurogenic niches (dentate gyrus of the hippocampus
cm of ALG, attributed to the asymmetric stretching or the SVZ). In the constructs we produced, Aβ was
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of the COO vibration, and shifted to a discretely lower incorporated into the bioink rather than inside the
–
wavelength, suggesting that there was an interaction neurospheres, aiming at mimicking senile plaques
between the positive charges of the amino group of GEL distributed within the neurogenic niches and in the brain
and the negative charges of the terminal COO groups of parenchyma of AD patients as extracellular deposits.
–
ALG. Additionally, as there was more ALG than GEL in The neurosphere structure was maintained in the bioink,
the hydrogel, the characteristic bands of GEL diffused with and Aβ oligomers were retained within the hydrogel,
reducing chain order. Notably, crosslinking with CaCl did simulating one of the pathological features of AD, i.e., the
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not modify the main structures of the hydrogel.
deposition and accumulation of Aβ in the extracellular
As our primary goal is to develop a 3D model for AD, microenvironment.
we investigated whether the presence of Aβ in the hydrogel Simpson et al. demonstrated that Aβ aggregates more
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composition would affect its structure. In Figure 3B and C, readily in a 3D environment of collagen hydrogel than in
we present the characterization of the Aβ structure alone 2D, suggesting that a 3D environment may increase Aβ-
and the structure after incorporation into the hydrogel Aβ interactions, accelerating aggregation and shifting Aβ
with neurospheres as oligomeric/fibrillary structures. The organization from oligomers to fibril. According to the
main amide I band (1700–1600 cm ) of the Aβ peptide authors, Aβ aggregation kinetics is fundamentally different
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backbone is correlated with oligomer size, where larger in 3D structures compared to a 2D environment.
oligomers are associated with lower wavenumbers. The
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amide I band of Aβ was at 1625 cm , representing Aβ A critical issue in modeling the neural tissue is
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oligomeric arrangement. 81–83 This is an important feature optimizing cell density to ensure that they will be properly
once monomeric Aβ is found in healthy brains. However, preserved within the construct and viable after printing.
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oligomers and fibrils are toxic and are the components Moreover, to achieve optimal printability and provide
of amyloid plaques, one of the AD hallmarks. 12,84,85 The a proper 3D matrix, cells must be compatible with the
amyloid or senile plaques are responsible for increasing hydrogel to proliferate and/or differentiate within it. It is
oxidative stress, contributing to neuronal dysfunction, recognized that cytoviability can be reduced by extrusion-
disruption of synaptic function, compromising based bioprinting due to the high shear force exerted on the
neuroplasticity, decreasing memory formation, and cells by the printing nozzles. Using code 1 (Table 2; Figure
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inducing neuroinflammation. 1A), we demonstrate that the total area of the neurospheres

Volume 10 Issue 5 (2024) 513 doi: 10.36922/ijb.3751