Nanotopographic Influence on the In Vitro Behavior of Induced Pluripotent Stem Cells

Abstract

While the influence of nanotopography on stem cell behavior has been extensively investigated on adult stem cells, far fewer studies have investigated the interaction of induced pluripotent stem cells (iPSCs) with various nanotopographical patterns. The purpose of this study was to identify nanopatterns that can influence the stemness and proliferation, as well as the adhesive properties in iPSCs, and thereby explore the feasibility of applying these nano-features for regenerative medicine. Three kinds of nanopatterns were fabricated from polydimethylsiloxane membranes, irregular patterned membrane (IPM), groove patterned membrane (GPM), and postpatterned membrane (PPM), in addition to flat patterned membrane (FPM) which did not have any nanotopographic features and was used as the control pattern. On the surfaces of GPM or PPM, iPSCs showed tendency for aggregation and did not spread out well at passage 1. However, with continued passaging (P6, P10), the tendency to form aggregates was greatly reduced. While iPSCs cultured on GPM and PPM had low population doubling time values compared with FPM and IPM at P1, the differences disappeared in later passages. The expression of the cell proliferation marker Ki67 in iPSCs gradually decreased with continued passaging in cells cultured on FPM and IPM, but not in those cultured on GPM and PPM. The expression of Oct3/4 and Nanog, marker of stemness, was significantly higher on GPM and PPM than on FPM at P6 and P10. At P5, numerous filopodia were demonstrated in the peripheral attachments of iPSC colonies on FPM and IPM, while GPM and PPM generally had globular appearance. The expression of the focal adhesion (FA) molecules -actinin, vinculin, phalloidin, or FA kinase was significantly greater on GPM and PPM than on FPM and IPM at P6 or P10. In conclusion, continued passaging on regular nanopatterns, including groove- and post-forms, was effective in maintaining an undifferentiated state and proliferation of iPSCs and also in increasing the expression of FA molecules.