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International Journal of Bioprinting 3D cell culture model for neural cell analysis

particles predominantly exhibited a globular shape, with of 100 μm³ per cell, and ZTA and CoCrMo particles at
a mean size of 5.86 ± 1.20 μm, aligning with findings from 50 μm³ per cell. In all model particle groups, 95% of the
Germain et al., who reported a mean size of 9.87 ± 5.67 particles ranged from 0.1 to 8 μm in size. We assessed cell
μm for commercially available CoCr particles. However, viability, ROS production, and DNA damage over 7 days to
when comparing these to clinically relevant CoCr particles capture both immediate and cumulative cellular responses
generated via a pin-on-plate wear simulator, Germain to model particles. Initial testing of CoCrMo particles at
et al. and Lee et al. observed aggregates and granular concentrations of 0.5, 5, and 50 μm³ per cell showed no
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shapes, with some polygonal shards. Their reported mode significant differences in biological responses, prompting
size ranges for CoCr particles were 10–20 and 30–39 nm, the selection of 50 μm³ per cell for further testing. No
respectively, while the mode size range in our study was significant changes in cell viability or ROS production
0.8–8 μm. This significant discrepancy in particle size were observed between the cell-only controls and model
may help explain the differences in biological responses particle groups for both cell types, likely due to the short
observed in the current study. culture duration. However, the study revealed significant
In this study, bioprinting was used to create a 3D model variations in cell viability and ROS production, which
incorporating neural cells and biomaterial particles for the were influenced by culture time, biomaterial type, and the
first time. This platform offers significant advantages over interactions between particles and cells.
traditional hydrogel casting methods, such as enhanced The literature on neural cell responses to different
spatial precision for cellular placement and the ability to model particles, particularly in studies comparing the
better replicate the in vivo-like organization of tissues. 43,44 biological impact of various particles using 3D in vitro
By combining 5% (w/v) GelMA hydrogels and neural cells models, remains limited. One notable study by Hallab
in bioprinted hydrogels, we established a promising 3D et al. investigated macrophage reactivity to PEEK-
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cell culture model for investigating neural cell responses to OPTIMA™ particles versus ultra-high-molecular-weight
wear particles. The successful printing of model particles polyethylene (UHMWPE) particles, using differentiated
integrated with hydrogels and cells using extrusion-based human macrophages and primary human monocytes. They
bioprinting techniques demonstrated the potential of this found that PEEK-OPTIMA™ particles (0.7 and 2.4 μm) did
approach. Printing parameters were adapted from previous not significantly affect cell viability after 24 or 48 h. This
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studies, such as Rad et al., who successfully used GelMA finding is especially relevant, as our study is the first to
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hydrogels and neural cells in 3D-bioprinted constructs examine neural cell responses to PEEK-OPTIMA™ particles
for spinal cord injury models. The temperature-sensitive in vitro. Hallab et al. also reported that PEEK-OPTIMA™
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properties of GelMA were key, as it undergoes reversible particles induced significant increases in proinflammatory
gelation with temperature change. Rad et al. found that cytokine levels compared to the control, though their
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higher concentrations of GelMA (15% w/v) were needed inflammatory response was less pronounced than that of
at temperatures ≥32°C, while lower concentrations UHMWPE particles. Although our current study did not
(2.5% w/v) were more effective at ≤22°C. Additionally, focus on inflammation, future research should investigate
the current study confirmed that extrusion pressure was the impact of PEEK-OPTIMA™ particles on cytokine
crucial for maintaining the structural integrity of GelMA production in neural cells. Additionally, Hallab et al.
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constructs—too much pressure caused overflow, while conducted an in vivo study using a rabbit model to explore
insufficient pressure hindered extrusion. The nozzle size the effects of PEEK-OPTIMA™ particles via epidural and
also played a critical role in successful extrusion, especially intradiscal injections, which resulted in neurological
when printing hydrogels containing particles of metals, damage. This highlights the critical need to understand the
polymers, and ceramics. Smaller nozzles (25G, 0.25 mm; biological effects of PEEK-OPTIMA™ particles on neural
and 27G, 0.20 mm) were more likely to clog, whereas cells and tissues. 30,45,46 In line with these findings, our
larger nozzles (20G, 0.58 mm; and 22G, 0.41 mm) enabled study observed no major negative impact on cell viability,
consistent, successful extrusion of cell-laden hydrogels with similar results between the cell-only and 3D model
with particles. Our results showed comparable growth particle groups. However, a noteworthy observation was
and proliferation of cells in both 2D- and 3D-bioprinted the significantly higher production of ROS by NG108-15
cultures over 7 days. These findings were consistent with cells exposed to PEEK-OPTIMA™ particles. This increase
Rad et al., who reported similar results using 5% (w/v) in ROS production warrants further investigation into the
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GelMA hydrogel for the same cell lines. potential long-term effects of PEEK-OPTIMA™ particles
In this study, C6 astrocyte-like and NG108-15 cells were in spinal environments. Understanding how PEEK-
exposed to model particles, with PEEK-OPTIMA™ and OPTIMA™ interacts with neural tissues is essential for
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Ceridust 3615 particles administered at a concentration evaluating its safety and clinical outcomes. Such knowledge

Volume 11 Issue 4 (2025) 271 doi: 10.36922/IJB025180174