Introduction: Poly(ethylene glycol) (PEG) hydrogels have been demonstrated as effective synthetic matrices for drug delivery and tissue scaffold engineering applications[1].[1] However, only the chain ends of PEG can be functionalized for crosslinking and/or ligand tethering, which limits chemical versatility for non-invasive biomedical applications[2],[3].
In situ gelling poly(oligoethylene glycol methacrylate) (POEGMA) hydrogels were prepared based on hydrazone bond formation to successfully address these limitations. The physical and biological properties of three POEGMA hydrogels with varying phase transition temperatures (VPTT) below (PO0), close to (PO10) and significantly higher (PO100) than physiological temperature are shown by exploiting the tunability of the lower critical solution temperature (LCST) behaviour of the oligo(PEG) side-chains used[4].
Materials and Methods: Hydrazide (POxHy) and aldehyde (POxAy) functionalized POEGMA precursors were synthesized using free-radical polymerization of oligo(ethylene glycol) methacrylate monomers with varying ethylene oxide chain lengths (n=2 or n=8-9). Reactive group functionality (variation of y, mol% of total monomer residues) and LCST (variation of x, the mol% of n=8-9 and n=2 comonomer used) are selected via copolymerization. Reactive precursors are co-extruded to rapidly form hydrogels in situ.
Results and Discussion: The PO10 hydrogel showed a clear discontinuous VPTT at ~32-33°C while PO0 (100 mol% n=2 monomer) and PO100 (100 mol% n=8-9 monomer) hydrogels showed phase transitions at ~24°C and > 60°C respectively (Fig. 1A). The G’ of PO0 was ~1 order of magnitude higher than that of PO100 (Fig. 1B); similarly, gelation varies significantly from PO0 (~5 s) to PO100 (~20 min) (Fig. 1C).
Fig. 1 Thermoresponsive properties of the PO0, PO10 and PO100 hydrogels (1A). Mechanical properties of gels (1B). Physical appearance and gelation times at 37°C (1C). BSA (1D) adsorption onto the PO0, PO10 and PO100 hydrogels at 37°C as function of the protein concentration in the loading solution.
Fig. 2A Fibroblast adhesion after 2 days to a polystyrene control (A1), POEGMA hydrogel (A2) and an RGD-functionalized hydrogel (A3). Fig 2B. Cells recovered following delamination from a PO0 hydrogel interface, following trypsin treatment (B1) and following thermal treatment at 4°C for 15 minutes (B2).
Very low protein adsorption (Fig. 1D) and cell adhesion were observed for PO10 and PO100 gels; however, grafting a small density of RGD peptide to PO100 precursors significantly increased the matrix cell binding capacity (Fig. 2A). Enzyme-free cell delamination from the PO0 gels upon cooling below the gel transition temperature (Fig 2B). In vivo studies on BALB-c mice show mild inflammatory responses at both the acute and chronic time points for PO10 and PO100 but evidence of capsule formation for PO0 (Fig. 3).
Fig 3. Histology following subcutaneous injection (acute = 2 days, chronic = 30 days)
Conclusions: POEGMA-based gels have highly tunable properties that can readily be matched to a range of tissues (e.g. adipose, muscle, or cartilage) by simple copolymerization. By varying the phase transition temperature and inclusion of RGD, matrices can be designed with low protein adsorption (matching reported PEGylated surfaces) and no cell adhesion, or moderate cell adhesion but minimal non-specific protein adsorption.
Funding from NSERC CREATE-IDEM is gratefully acknowledged
References:
[1] Lin, C.-C.; Anseth, K. S. Pharmaceutical research 2009, 26, 631–43.
[2] Nuttelman, C. R.; Rice, M. A.; Rydholm, A. E.; Salinas, C. N.; Shah, D. N.; Anseth, K. S. Progress in Polymer Science 2008, 33, 167–179.
[3] Luzon, M.; Boyer, C.; Peinado, C.; Corrales, T.; Whittaker, M.; Tao, L. E. I.; Davis, T. P. Journal of Polymer Science Part A: Polymer Chemistry 2010, 48, 2783–2792.
[4] Lutz, J.-F. Journal of Polymer Science Part A: Polymer Chemistry 2008, 46, 3459–3470.