Modular molecular design of polymerized pro-estrogen materials enables controlled astrocyte response

Abstract

The hormone 17β-estradiol (estrogen or “E2”) has demonstrated robust neuroprotective properties in countering oxidative stress-induced neurotoxicity, as well as strong neurotrophic properties to promote axonal growth, following injury to the central nervous system (CNS). However, oral or injected E2 is a suboptimal drug, as systemic administration fails to achieve a therapeutic dose at the injury site, in addition to being contraindicated in most male patients. Polymerized pro-drug biomaterials can mitigate these issues by locally releasing small quantities of drug over vastly extended timescales. We sought to study the effect of the biomaterial properties of poly(pro-E2) on astrocyte-surface interactions because astrocytes are the most abundant cell type in the central nervous system, play vital roles in neuron support and traumatic injury, and express estrogen receptors. Herein, we sought to study the effect of novel poly(pro-E2) films on astrocyte behavior, as well as to gauge the biomaterial properties that would lead to optimal astrocyte functionality. We synthesized pro-E2 as carbonate and ester derivatives and copolymerized each of these monomers with either oligoethylene glycol dithiol (EG) or hexylene dithiol (Hex) linkers to generate four unique poly(pro-E2) materials with tunable physiochemical and mechanical properties. We found that films of polymer with Hex-linkers supported sustained astrocyte adhesion, whereas the EG-linked analogs did not. To explain this behavior, we investigated the physical and chemical surface properties that may influence cell attachment. SEM images for films incubated in buffer showed marked surface roughness with micro- and nano-scale topography for the polymers with Hex-linkers, whereas those with EG-linkers appeared smooth. This result suggests that astrocytes preferentially adhere to rougher surfaces. Hex-linked polymer surfaces also demonstrated more negative zeta potentials compared to EG-linked polymer surfaces – indicating favorable electrostatic interactions for astrocyte adhesion. Finally, while all polymers exhibited hysteresis during mechanical testing, films with Hex-linkers demonstrated greater dissipation, suggesting more pronounced viscoelasticity. Taken together, these results indicate that a combination of physiochemical surfaces properties, which arise from subtle differences in chemical composition, can exert marked effects on astrocyte adhesion and spreading.