Evaluation of a human 3D multi-spheroid model derived from SH-SY5Y cells for cytotoxicity testing

3D culture models have been extensively used in cell testing. Thus, we aimed to develop a novel cellular model of multiple spheroids derived from neuronal SH-SY5Y cells. We observed that poly-D-lysine coating can induce formation of a population of spheroids, i.e. the density of 20 × 10³ of SH-SY5Y cells per well led to formation of approximately 300 and 150 spheroids after 4 and 7 days of incubation, respectively. In the spheroids, we found induced expressions of ICAM-1, connexin 26 and neuron-specific enolase proteins demonstrating increased differentiation of SH-SY5Y cells compared to the cells grown as a monolayer. Finally, we aimed to utilize the new culture model in toxicity testing. Thus, we incubated the cells with CdCl₂, HgCl₂ and PbCl₂ at doses up to 1000 µM for 24-48 h. After the treatment, we measured glutathione levels, dehydrogenase activity, and ATP levels in SH-SY5Y cells grown in both 2D and 3D arrangements showing that the cells grown as monolayers were significantly more susceptible to the toxic effects of heavy metals. In conclusion, we developed a novel cell culture model of multiple spheroids derived from SH-SY5Y cells which can be very useful tool in cellular testing in future.