Matrices with Compliance Comparable to that of Brain Tissue Select Neuronal over Glial Growth in Mixed Cortical Cultures

Abstract

Cortical neurons and astrocytes respond strongly to changes in matrix rigidity when cultured on flexible substrates. In this study, existing polyacrylamide gel polymerization methods were modified into a novel method for making substrates capable of engaging specific cell-adhesion receptors. Embryonic cortical dissociations were cultured on polyacrylamide or fibrin gel scaffolds of varying compliance. On soft gels, astrocytes do not spread and have disorganized F-actin compared to the cytoskeletons of astrocytes on hard surfaces. Neurons, however, extend long neurites and polymerize actin filaments on both soft and hard gels. Compared to tissue culture plastic or stiff gel substrates coated with laminin, on which astrocytes overgrow neurons in mixed cultures, laminin-coated soft gels encourage attachment and growth of neurons while suppressing astrocyte growth. The number of astrocytes on soft gels is lower than on hard even in the absence of mitotic inhibitors normally used to temper the astrocyte population. Dissociated embryonic rat cortices grown on flexible fibrin gels, a biomaterial with potential use as an implant material, display a similar mechano-dependent difference in cell population. The stiffness of materials required for optimal neuronal growth, characterized by an elastic modulus of several hundred Pa, is in the range measured for intact rat brain. Together, these data emphasize the potential importance of material substrate stiffness as a design feature in the next generation of biomaterials intended to promote neuronal regeneration across a lesion in the central nervous system while simultaneously minimizing the ingrowth of astrocytes into the lesion area.

Nanoscale Surface Topography Reshapes Neuronal Growth in Culture

“…. Although this value is much lower than any bulk values, this rigidity remains in a range known to have no effect on cortical neuron development 25 . Therefore, even by considering the extreme situation where the nano-pillars might display a linear elastic behavior in a large range of deformations, the changes we observed in neuronal elongation do probably not result from the flexibility of nano-pillars.…”

Tissue Stiffness Dictates Development, Homeostasis, and Disease Progression

“…During the normal wound healing process, it is very common for the ECM of damaged tissues to be replaced by fibrous tissue that is stiffer than the original ECM. 57 The newly formed tissue of increased stiffness is generally referred to as scar tissue. Matrix stiffening in scar tissue tends to be the product of abnormal and/or increased ECM deposition, increased ECM crosslinking, and reduced or abnormal matrix degradation.…”

RPTPα is required for rigidity-dependent inhibition of extension and differentiation of hippocampal neurons

“…Several recent studies have shown that soft substrates stimulated neurite extension and branching, but inhibited glial cell growth (Balgude et al, 2001;Engler et al, 2004;Flanagan et al, 2002;Georges et al, 2006;Lamoureux et al, 2002;Strassman et al, 1973). However, mechanisms of this regulation are not clear.…”