| In recent years, cells cultured in three-dimensional conditions showed a differentgrowth behavior and features than it cultured in the two-dimensional conditions. Thecells cultured in a three-dimentional environment are much closer to the body cells thanit cultured in the two-dimentional environment. Most previous studies focused on themorphology description of cells. Few studies explored the factor of cell markers in thethree-dimensional environment. Therefore, the SH-SY5Y cells have been seeded on thePLLA microwell patterns in the studies, we evaluated the three-dimentionalconfiguration and differentiation status on secretion of VEGF and IL-8of SH-SY5Ycells.Objective The aims of the studies are to compare the differences of growthbehavior of SH-SY5Y on the nine kinds of microwell patterns. Also, we explored theconfiguration of microwell patterns and differentiation status on secretion of VEGF andIL-8of SH-SY5Y cells. This study laid the foundation for establishingthree-dimensional cell-based biosensing platforms.Methods We fistly used silicon etching techniques and the PDMS-based softlithography to fabricate microwell patterns. The diameter of microwell patterns were80μm、100μm and120μm, and the channel of microwell patterns are0μm、20μm and40μm. And then the SH-SY5Y cells integrated with microwell patterns. At the sametime, we used ELISA analysis to detect the secretion of VEGF and IL-8of SH-SY5Ycells on the flat substrates and microwell patterns before and after differentiation5days.Results The results of the integrating experimental show that SH-SY5Y cellsbehaved three types of growth behavior: while some cells in the hole attached to theside-wall and the bottom of the microwell patterns, the other cells lied on the surface ofthe microwell patterns respectively. Quantitative measurements of ELISA showed:compared with the cells on flat substrates, the secretion of VEGF on the100-0μm、100-20μm of microwell patterns which were53.5±11.1pg/10~4、52.4±17.4pg/10~4havevery significant difference（P <0.01）,but the secretion of IL-8on100-40microwellpatterns which was46.8±16.9pg/10~4has significant difference（P <0.05）. Also thesecretion of IL-8on100-0μm、100-20μm and100-40μm were63.4±6.40pg/10~4、74.1±2.95pg/10~4、73.7±6.35pg/10~4.100-0μm compared with the cells on flatsubstrates, has significant difference（P <0.05）.100-20μm and100-40μm have very significant difference（P <0.01）compared with the cells on flat substrates.The secretion of IL-8on the100-0μm、100-20μm of microwell patterns whichwere6.37±2.97pg/10~4、7.16±3.40pg/10~4before differentiation, compared with thesecretion of IL-8on the flat substrates which is4.32±3.33pg/10~4has significantdifference（P <0.05）. Also secretion of IL-8after differentiation5days showed that thesecretion of the sample were18.3±4.22pg/10~4、22.1±3.03pg/10~4、25.7±4.82pg/10~4.compared with the secretion of the sample on the flat substrate have very which is13.7±1.65pg/10~4significant difference（P <0.01）.Conclusion Configuration of microwell patterns can form effectivelythree-dimensional growth mode and boundary two-dimensional growth mode ofSH-SY5Y. Through rearching we found secretion of VEGF and IL-8of SH-SY5Y cellson the microwell patterns after differentiation were significantly higher than the cells onthe microwell patterns before differentiation. The secretion of VEGF and IL-8ofSH-SY5Y cells which were un differentiation and differentiated by5days on themicrowell patterns were higher than cells on the flat substrates, but100-40μmmicrowell patterns was not higher than cells on the flat substrates. In summary,combined with the relation patterns,100-0μm and100-20μm can more effectiveconform cell growth in three dimension in9microwell patterns.The configuration ofmicrowell patterns and differentiation status could be the important factors of affectingSH-SY5Y cells to secrete VEGF and IL-8. Increased secretion of VEGF and IL-8mayas secretion sign of the cells in three-dimensional growth. |
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