The effect of Young’s modulus on the neuronal differentiation of mouse embryonic stem cells

Abstract

There is substantial evidence that cells produce a diverse response to changes in ECM stiffness depending on their identity. Our aim was to understand how stiffness impacts neuronal differentiation of embryonic stem cells (ESC's), and how this varies at three specific stages of the differentiation process. In this investigation, three effects of stiffness on cells were considered; attachment, expansion and phenotypic changes during differentiation. Stiffness was varied from 2 kPa to 18 kPa to finally 35 kPa. Attachment was found to decrease with increasing stiffness for both ESC's (with a 95% decrease on 35 kPa compared to 2 kPa) and neural precursors (with a 83% decrease on 35 kPa). The attachment of immature neurons was unaffected by stiffness. Expansion was independent of stiffness for all cell types, implying that the proliferation of cells during this differentiation process was independent of Young's modulus. Stiffness had no effect upon phenotypic changes during differentiation for mESC's and neural precursors. 2 kPa increased the proportion of cells that differentiated from immature into mature neurons. Taken together our findings imply that the impact of Young's modulus on attachment diminishes as neuronal cells become more mature. Conversely, the impact of Young's modulus on changes in phenotype increased as cells became more mature.

The rotation of mouse myoblast nuclei is dependent on substrate elasticity

“…Our results indicate that nuclear rotation is dependent on substrate elasticity. Our results on hard substrates are coherent with previous studies; however, we showed for the first time that a significant reduction in the prevalence, magnitude, and speed of rotation occur on soft substrates designed to mimic the physical properties of resting muscle tissue (Al-Rekabi and Pelling, 2013; Ali et al, 2015;Collinsworth et al, 2002;Mathur et al, 2001;Nyland and Maughan, 2000;Ogneva, 2010). Previous studies on nuclear rotation have been performed on hard substrates that can have a stiffness in the GPa range (Brosig et al, 2010;Ji et al, 2007;Levy and Holzbaur, 2008;Albrecht-Buehler, 1986b, 1988;Wilson and Holzbaur, 2012;Yao and Ellingson, 1969).…”

“…Traditionally, mammalian cells have been cultured on hard glass or plastic substrates. Here, we created “soft” substrates by coating them with a thick GE cross‐linked with glutaraldehyde (GA) gel (GXG gel), as described previously (Al‐Rekabi and Pelling, ; Ali, Wall, Mason, Pelling, & Veraitch, ). The GXG gel stiffness was tuned to be within the resting muscle tissue stiffness regime (Collinsworth et al, ; Mathur et al, ; Nyland and Maughan, ; Ogneva, ).…”

Neurite extension and neuronal differentiation of human induced pluripotent stem cell derived neural stem cells on polyethylene glycol hydrogels containing a continuous Young's Modulus gradient

“…The mechanical properties of the matrix are known to affect the cellular behavior and differentiation of many cell types . However, conflicting reports exist in the literature regarding the sensitivity of pluripotent and neural stem cells during neural differentiation to changes in the Young's Modulus of the culture substrate . It has been suggested that both the lineage and maturation state of the cells play a role in the mechanosensitivity of neural cell types, but the transmittance of mechanical signals to neural cells is complex and not yet fully understood, as multiple pathways play a role .…”

Substrate stiffness affects the morphology and gene expression of epidermal neural crest stem cells in a short term culture

“…In this study, the β‐III tubulin mRNA level reached a 3.2‐fold increase roughly on all hydrogels after 1 day of culture. However, several investigations reported the β‐III tubulin upregulation in other cell types after a longer culture period on the same substrate elasticity (Ali, Wall, Mason, Pelling, & Veraitch, ; Banerjee et al, ; Saha et al, ). In the next part of our study, the gene expression analysis of our panel of growth factors showed that most of them could be modulated by substrate stiffness.…”