Effect of spatial arrangement of substrate topography on neuronal differentiation of stem cells

Tissue engineered substrates can be modified to provide suitable mechanical cues for the manipulation of stem cell responses. Recent studies have shown that topography plays a significant role in stem cell fate determination. We are interested in studying the neuronal differentiaiton of human embryonic stem cells (hESCs) and murine neural progenitor cells (mNPCs) in response to substrate topography. A Multi ARChitectural (MARC) chip containing fields of various geometries and size was developed to investigate the influence of topography spatial arrangement on differentiation. The hESCs and mNPCs were differentiated on the MARC chip with minimal neuronal supplements for 7 and 14 days, respectively. The gene and protein expression analysis of neuronal markers β Tubulin III (Tuj1) and microtubule associated protein 2 (MAP2), and the glial marker, glial fibrillary acidic protein (GFAP) indicated that hESCs and mNPC grown on anisotropic patterns derived a significantly higher percentage of Tuj1 and MAP2 positive cells and lower percentage of GFAP positive cells, thereby supporting neuronal differentiation. The percentage of GFAP positive cells were significantly higher than the neuronal cells when hESCs or mNPCs were grown on the isotropic patterns, thus supporting glial differentiation. By tracking the differentiation and pluripotency markers and applying topography at various periods of culture, we found that the topography contact during the differentiation period was necessary for topography-induced differentiation. This study showed that the topography-induced neuronal differentiation depends on the size and geometry of topographical patterns, and an optimal combination of topography and biochemical cues could shorten the differentiation period.