Nanofibrillar scaffolds induce preferential activation of Rho GTPases in cerebral cortical astrocytes

Volkan Mujdat Tiryaki,¹ Virginia M Ayres,¹ Adeel A Khan,² Ijaz Ahmed,³ David I Shreiber,³ Sally Meiners<sup>4

1</sup>Electronic and Biological Nanostructures Laboratory, Department of Electrical and Computer Engineering, Michigan State University, East Lansing, MI, USA; ²Department of Paper Engineering, Chemical Engineering, Imaging, Western Michigan University, Kalamazoo, MI, USA; ³Department of Biomedical Engineering, Rutgers, The State University of New Jersey, Piscataway, NJ, USA; ⁴Nanoculture, LLC, Piscataway, NJ, USA

Abstract: Cerebral cortical astrocyte responses to polyamide nanofibrillar scaffolds versus poly-L-lysine (PLL)-functionalized planar glass, unfunctionalized planar Aclar coverslips, and PLL-functionalized planar Aclar surfaces were investigated by atomic force microscopy and immunocytochemistry. The physical properties of the cell culture environments were evaluated using contact angle and surface roughness measurements and compared. Astrocyte morphological responses, including filopodia, lamellipodia, and stress fiber formation, and stellation were imaged using atomic force microscopy and phalloidin staining for F-actin. Activation of the corresponding Rho GTPase regulators was investigated using immunolabeling with Cdc42, Rac1, and RhoA. Astrocytes cultured on the nanofibrillar scaffolds showed a unique response that included stellation, cell–cell interactions by stellate processes, and evidence of depression of RhoA. The results support the hypothesis that the extracellular environment can trigger preferential activation of members of the Rho GTPase family, with demonstrable morphological consequences for cerebral cortical astrocytes.

Keywords: stellation, nanofiber, RhoA, atomic force microscopy