Assistant Professor Boston University, United States
Introduction: Disruption of healthy central nervous system (CNS) tissue by biomaterial implantation stimulates adaptive reprogramming of local astrocytes. As part of this adaptive reprogramming, astrocytes deprioritize regular neuronal interactions and support functions to proliferate, migrate, and spatially reorganize to address the tissue disruption and limit its spread1. Biologically-inspired materials offer a unique opportunity to study and direct context-specific astrocyte adaptive reprogramming by presenting diverse contact-mediated cues2. We hypothesize that biomaterials with variably immunostimulatory chemomechanical properties can be used to direct the extent and dynamics of astrocyte border reprogramming. In this study, we use validated post-polymerization modification synthesis approaches to develop a cellulose-based biomaterial platform that can present material interfaces with varied chemical functionality, surface charge, hydrophilicity, and stiffness. Presentation of these stimuli in vivo results in the formation of context-specific astroglial borders, allowing for insight into biomaterial-property dependent effects on astrocyte adaptive reprogramming.
Materials and Methods: 2-Hydroxyethyl cellulose (HEC, MW 380kDa), 3-(Methylthio)propyl isothiocyanate (MTPI), N,N-Diisopropylethylamine (DIEA), iodomethane, and tert-Butyl hydroperoxide solution (TBHP) were all purchased from Sigma-Aldrich. Iodoacetic acid was purchased from ThermoFisher. C57BL/6J mice were purchased from Jackson Laboratories. HEC was modified with MTPI via a DIEA-catalyzed nucleophilic addition. Secondary modifications of the resulting HECMTP polymer were performed using iodomethane, TBHP, and iodoacetic acid in either water (iodomethane, TBHP) or 0.1M formic acid (iodoacetic acid) for 24hrs (TBHP) or 96hrs (iodomethane, iodoacetic acid). The modified polymers were dialyzed to remove impurities. All synthesized materials were characterized by NMR and FTIR. Hydrogels were formed by solubilizing the modified materials in PBS and hydrogel properties were characterized by dynamic mechanical rheology. Select hydrogels were tested with neural progenitor cells in vitro for cytocompatibility and injected into mouse forebrain to evaluate CNS FBRs using immunohistochemistry (IHC).
Results, Conclusions, and Discussions: MTPI was reacted with primary hydroxyls on HEC to generate HECMTP at high yields and with stoichiometric control. Secondary oxidation or alkylation reactions modified thioether groups on HECMTP to generate sulfoxide (HECSO), sulfonium (HECS+), or zwitterionic (HECzwit) –functionalized materials using a common HECMTP template (Fig. 2a, b). These chemical modifications converted a viscous HEC polymer sol into shear thinning hydrogels with concentration- and chemical functionality-dependent control of stiffness and viscoelastic properties (Fig. 2c). Preliminary investigations show that this HEC material system can be injected using pulled glass micropipettes to form stable, non-resorbable biomaterial deposits in vivo in the CNS. Using a standardized IHC-based FBR evaluation framework3 (Fig. 2d), we are dissecting the unique phenotypes of the astrocyte adaptive reprogramming response stimulated by these HEC-based biomaterials.
Acknowledgements: We would like to thank the Boston University Core Facilities for the use of their rheometer and Olympus IX83. We are grateful to the National Science Foundation for the purchase of the NMR (CHE0619339). This work was supported by funding from Boston University (T.M.O), Craig H. Neilsen Foundation (T.M.O.) and the Bryon Riesch Paralysis Foundation (T.M.O).
References: [1] O’Shea, T. J. Clin. Invest. (2017). 127 (9). 3259–3270. [2] Tam, R. Neuropsychopharmacol. (2014). 39 (1). 169–188. [3] O’Shea, T. M. et al. Nat Commun. (2020). 11(1). 6203.
