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

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

Acknowledgements

The support of National Science Foundation grants PHY-0957776 (VMA and VMT) and ARRA-CBET-0846328 (DIS and IA), the Michigan-Louis Stokes Alliance for Minority Participation Summer Undergraduate Research Academy (AAK), the General Directorate for Higher Education, Ministry of National Education of the Republic of Turkey (VMT), and the New Jersey Commission on Spinal Cord Research Grants 06A-007-SCR1and 06-3058-SCR-E-O (SM) are gratefully acknowledged. The authors thank Dr Melinda Frame (Michigan State University USA) for valuable discussions about colocalization and quantitative immunolabeling analysis, Lars Haubold (Fraunhofer USA) for assistance in contact angle measurements, and Katerina Moloni (nPoint Inc) for assistance in closed-loop multimode AFM investigation.

Disclosure

The authors report no conflicts of interest in this work.