The Effect Of IL-8on The Responsiveness Of Voltage-gated Calcium Channels For SH-SY5Y Human Neuroblastoma Cells On Quasi-three-dimensional Micropatterns

Quasi-three-dimensional microwell patterns were fabricated with poly (l-lactic acid)(PLLA) for the development of cell-based assays targeting voltage-gated calciumchannels (VGCCs). SH-SY5Y human neuroblastoma cells were interfaced with themicrowell patterns and cell growth characteristics were categorized as two dimensional(2D), three dimensional (3D) and near two dimensional (N2D) according to the locationof cells within the pattern: on the top surface of the pattern, on the side wall or on thebottom surface of the microwells. The capability of the microwell patterns to support3D cell growth was evaluated in terms of the percentage of the cells in each category.Cell spreading was analyzed in terms of projection areas under light microscopy. VGCCresponsiveness of SH-SY5Y cells before and after differentiation was evaluated withconfocal microscopy and a calcium fluorescent indicator, Calcium Green-1. Thesecretion of IL-8on flat substrat or microwell patterns and the effect of IL-8on VGCCresponse on flat substrate were evaluated. It was found that cells within the microwells,either N2D or3D, showed a more rounded shape and less projection areas than2D cellson flat PLLA substrates. Upon stimulation with50mM K+, cells in microwells showeda significant lower VGCC responsiveness than cells on flat substrates, in terms of eitherresponding magnitudes or percentages of cells responding. For either projection areas orVGCC responsiveness, N2D cells had an intermediate value between those for2D and3D cells, suggesting a correlative relation between cell morphology and VGCCresponsiveness. The secretion of IL-8on microwell patterns was higher than that ofcells on flat substrates. And exerting gradient concerntration of IL-8on SH-SY5Y cellsshows that high concertration IL-8will significant decline VGCC responsiveness.Theseresults indicate that the pattern format and thus the cell growth characteristics werecritical factors in determining cell VGCC responsiveness and that IL-8play animportant role in the functionality establishment of SH-SY5Y cells on microwellpatterns. It also provides an approach for engineering cell functionality in cell-basedassay system establishment.