PEG-Coated Large Mesoporous Silicas as Smart Platform for Protein Delivery and Their Use in a Collagen-Based Formulation for 3D Printing

Abstract

Silica-based mesoporous systems have gained great interest in drug delivery applications due to their excellent biocompatibility and high loading capability. However, these materials face challenges in terms of pore-size limitations since they are characterized by nanopores ranging between 6-8 nm and thus unsuitable to host large molecular weight molecules such as proteins, enzymes and growth factors (GFs). In this work, for an application in the field of bone regeneration, large-pore mesoporous silicas (LPMSs) were developed to vehicle large biomolecules and release them under a pH stimulus. Considering bone remodeling, the proposed pH-triggered mechanism aims to mimic the release of GFs encased in the bone matrix due to bone resorption by osteoclasts (OCs) and the associated pH drop. To this aim, LPMSs were prepared by using 1,3,5-trimethyl benzene (TMB) as a swelling agent and the synthesis solution was hydrothermally treated and the influence of different process temperatures and durations on the resulting mesostructure was investigated. The synthesized particles exhibited a cage-like mesoporous structure with accessible pores of diameter up to 23 nm. LPMSs produced at 140 °C for 24 h showed the best compromise in terms of specific surface area, pores size and shape and hence, were selected for further experiments. Horseradish peroxidase (HRP) was used as model protein to evaluate the ability of the LPMSs to adsorb and release large biomolecules. After HRP-loading, LPMSs were coated with a pH-responsive polymer, poly(ethylene glycol) (PEG), allowing the release of the incorporated biomolecules in response to a pH decrease, in an attempt to mimic GFs release in bone under the acidic pH generated by the resorption activity of OCs. The reported results proved that PEG-coated carriers released HRP more quickly in an acidic environment, due to the protonation of PEG at low pH that catalyzes polymer hydrolysis reaction. Our findings indicate that LPMSs could be used as carriers to deliver large biomolecules and prove the effectiveness of PEG as pH-responsive coating. Finally, as proof of concept, a collagen-based suspension was obtained by incorporating PEG-coated LPMS carriers into a type I collagen matrix with the aim of designing a hybrid formulation for 3D-printing of bone scaffolds.

Keywords: 3D printing; growth factor; hydrothermal treatment; large pores; mesoporous silica particles; pH-triggered release; type I collagen.

Figure 1

Schematic representation of the coating of mPEG-silane on LPMS_HRP obtained through PEGylation method.

Figure 2

N₂ adsorption-desorption isotherms and pore size distributions of synthesized LPMSs using different temperatures and durations of the hydrothermal treatment: LPMSs obtained with 2 h (A,B) and 24 h (C,D) of process, respectively. The classification of hysteresis loop according to pore types is reported to the left (according to reference [86]).

Figure 3

FE-SEM images of LPMS_140_24 at different magnifications: single particles (A), surface cage-like mesoporous network (B,C).

Figure 4

ATR-FTIR spectrum (A) and thermogravimetric analysis (TGA) curves (B) with relative derivative TGA curve (dotted line) of LPMS_140_24 (green), LPMS-HRP (grey), and LPMS-HRP_PEG (purple) particles.

Figure 5

Horseradish peroxidase (HRP) release profiles from LPMS-HRP particles under physiological conditions (grey), PEG-coated samples at pH 7.4 (light blue) and pH 5.5 (light red).

Figure 6

FE-SEM images of LPMS-HRP_PEG material at low (A) and high (B,C) magnifications.

Figure 7

Shear thinning (A) and stability of the storage (G′) and loss (G″) modulus values of the Coll/LPMS-HRP_PEG_TG suspension over time (B) at 10 °C. Sol-gel transition of the composite system at 37 °C (C).

Figure 8

3D mesh-like scaffolds of Coll/LPMS-HRP_PEG_TG (A) and FE-SEM images representing their micro-structures at different magnifications (B,C).