Biophysical Regulations of Epigenetic State and Notch Signaling in Neural Development Using Microgroove Substrates

Abstract

A number of studies have recently shown how surface topography can alter the behavior and differentiation patterns of different types of stem cells. Although the exact mechanisms and molecular pathways involved remain unclear, a consistent portion of the literature points to epigenetic changes induced by nuclear remodeling. In this study, we investigate the behavior of clinically relevant neural populations derived from human pluripotent stem cells when cultured on polydimethylsiloxane microgrooves (3 and 10 μm depth grooves) to investigate what mechanisms are responsible for their differentiation capacity and functional behavior. Our results show that microgrooves enhance cell alignment, modify nuclear geometry, and significantly increase cellular stiffness, which we were able to measure at high resolution with a combination of light and electron microscopy, scanning ion conductance microscopy (SICM), and atomic force microscopy (AFM) coupled with quantitative image analysis. The microgrooves promoted significant changes in the epigenetic landscape, as revealed by the expression of key histone modification markers. The main behavioral change of neural stem cells on microgrooves was an increase of neuronal differentiation under basal conditions on the microgrooves. Through measurements of cleaved Notch1 levels, we found that microgrooves downregulate Notch signaling. We in fact propose that microgroove topography affects the differentiation potential of neural stem cells by indirectly altering Notch signaling through geometric segregation and that this mechanism in parallel with topography-dependent epigenetic modulations acts in concert to enhance stem cell neuronal differentiation.

Bioengineered niches that recreate physiological extracellular matrix organisation to support long-term haematopoietic stem cells

“…The subcellular localization and activity of TFEB are regulated by mechanistic target of rapamycin (mTOR)-mediated phosphorylation, which occurs at the lysosomal surface 70 . Materials platforms have also been implicated in changes in chromatin organisation [71][72][73][74] and, for LT-HSCs, understanding chromatin accessibility to suppress CCCTC-binding factor (CTCF), restrains LT-HSCs from transitioning to activated ST-HSCs 75 . These new biological insights guide us for future bioengineered niche developments.…”

Direct deep UV lithography to micropattern PMMA for stem cell culture

“…During neurogenesis and regeneration, neurite growth occurs by a combination of mechanical and chemical signals which are sensed by the tips of growing neurites [ 35 ]. Microgrooves have been previously used to modulate neuronal differentiation of pluripotent stem cells [ 37 , 38 ]. The ability to control and study neuronal growth in vitro can support the design and fabrication of implantable nerve grafts [ 35 ].…”

Recent progresses in neural tissue engineering using topographic scaffolds

“…Grooves are the key determinants of neural cell alignment and orientation. As Hsu and coworkers reported that human pluripotent stem cells (hPSCs) could response to the groove topography, and cell alignment, stiffness, and nerve differentiation are regulated by grooves depth (Figure 1A) [33]. These features could be quantitatively analyzed by means of optical microscope, scanning ion conductivity microscope and atomic force microscope.…”