Modulating Bioglass Concentration in 3D Printed Poly(propylene fumarate) Scaffolds for Post-Printing Functionalization with Bioactive Functional Groups.

Poly(propylene fumarate) (PPF) has shown potential for the treatment of bone defects as it can be 3D printed into scaffolds to suit patient-specific needs with strength comparable to that of bone. However, the lack of specific cell attachment and osteogenic signaling moieties have limited their utility as it is necessary to provide these signals to aid in bone tissue formation. To address this issue and provide a platform for functionalization, Bioglass (∼1-2 µm) microparticles have been incorporated into PPF to create a 3D printable resin with concentrations ranging from 0 to 10 wt %. The zero-shear viscosity of PPF-Bioglass resins increased proportionally from 0 to 2.5 wt % Bioglass, with values of 0.22 and 0.34 Pa·s, respectively. At higher Bioglass concentrations, 5 and 10 wt %, the resin viscosity increased to 0.44 and 1.31 Pa·s, exhibiting a 2- and 6-fold increase from the 0 wt % Bioglass resin. Despite this increase in viscosity, all resins remained printable with no print failures. In addition, the surface available Bioglass can tether catechol containing molecules for postprinting functionalization. Analysis of PPF-Bioglass functionalization using a catechol dye analyte shows functionalization increases with Bioglass concentration, up to 157 nmol/cm2, and demonstrates it is possible to modulate functionalization. This presents a versatile and highly translationally relevant strategy to functionalize 3D printed scaffolds post printing with a diverse array of functional species.

Molecular Mass-Dependent Resorption and Bone Regeneration of 3D Printed PPF Scaffolds in a Critical-Sized Rat Cranial Defect Model.

The emergence of additive manufacturing has afforded the ability to fabricate intricate, high resolution, and patient-specific polymeric implants. However, the availability of biocompatible resins with tunable resorption profiles remains a significant hurdle to clinical translation. In this study, 3D scaffolds are fabricated via stereolithographic cDLP printing of poly(propylene fumarate) (PPF) and assessed for bone regeneration in a rat critical-sized cranial defect model. Scaffolds are printed with two different molecular mass resin formulations (1000 and 1900 Da) with narrow molecular mass distributions and implanted to determine if these polymer characteristics influence scaffold resorption and bone regeneration in vivo. X-ray microcomputed tomography (µ-CT) data reveal that at 4 weeks the lower molecular mass polymer degrades faster than the higher molecular mass PPF and thus more new bone is able to infiltrate the defect. However, at 12 weeks, the regenerated bone volume of the 1900 Da formulation is nearly equivalent to the lower molecular mass 1000 Da formulation. Significantly, lamellar bone bridges the defect at 12 weeks with both PPF formulations and there is no indication of an acute inflammatory response.

Optimization of photocrosslinkable resin components and 3D printing process parameters.

The role of 3D printing in the biomedical field is growing. In this context, photocrosslink-based 3D printing procedures for resorbable polymers stand out. Despite much work, more studies are needed on photocuring stereochemistry, new resin additives, new polymers and resin components. As part of these studies it is vital to present the logic used to optimize the amount of each resin constituent and how that effects printing process parameters. The present manuscript aims to analyze the effects of poly(propylene fumarate) (PPF) resin components and their effect on 3D printing process parameters. Diethyl fumarate (DEF), bisacylphosphine oxide (BAPO), Irgacure 784, 2-hydroxy-4-methoxybenzophenone (HMB) and, for the first time, in biomedical 3D printing, ethyl acetate (EA), were the resin components under investigation in this study. Regarding printing process parameters, Exposure Time, Voxel Depth, and Overcuring Depth were the parameters studied. Taguchi Design of Experiments was used to search for the effect of varying these resin constituent concentrations and 3D printing parameters on the curing behavior of 3D printable PPF resins. Our results indicate that resins with higher polymer cross-link density, especially those with a higher content of PPF, are able to be printed at higher voxel depth and with greater success (i.e., high yield). High voxel depth, as long as it does not sacrifice required resolution, is desirable as it speeds printing. Nevertheless, the overall process is governed by the correct setup of the voxel depth in relation to overcuring depth. In regards to resin biocompatibility, it was observed that EA is more effective than DEF, the material we had previously relied on. Our preliminary in vitro cytotoxicity tests indicate that the use of EA does not reduce scaffold biocompatibility as measured by standard cytotoxicity testing (i.e., ISO 10993-5). We demonstrate a workpath for resin constituent concentration optimization through thin film tests and photocrosslinkable process optimization. STATEMENT OF SIGNIFICANCE: We report here the results of a study of photo-crosslinkable polymer resin component optimization for the 3D printing of resorbable poly(propylene fumarate) (PPF) scaffolds. Resin additives are initially optimized for PPF thin film printing. Once those parameters have been optimized the 3D printing process parameters for PPF objects with complex, porous shapes can be optimized. The design of experiments to optimize both polymer thin films and complex porous resorbable polymer scaffolds is important as a guess and check, or in some cases a systematic method, are very likely to be too time consuming to accomplish. Previously unstudied resin components and process parameters are reported.

Magnesium Catalyzed Polymerization of End Functionalized Poly(propylene maleate) and Poly(propylene fumarate) for 3D Printing of Bioactive Scaffolds.

The ring-opening copolymerization of maleic anhydride and propylene oxide, using a functionalized primary alcohol initiator and magnesium 2,6-di-tert-butyl phenoxide as a catalyst, was investigated in order to produce high end-group fidelity poly(propylene maleate). Subsequent isomerization of the material into 3D printable poly(propylene fumarate) was utilized to produce thin films and scaffolds possessing groups that can be modified with bioactive groups postpolymerization and postprinting. The surface concentration of these modifiable groups was determined to be 30.0 ± 3.3 pmol·cm-2, and copper-mediated azide-alkyne cycloaddition was used to attach a small molecule dye and cell adhesive GRGDS peptides to the surface as a model system. The films were then studied for cytotoxicity and found to have high cell viability before and after surface modification.

Effect of Chemical and Physical Properties on the In Vitro Degradation of 3D Printed High Resolution Poly(propylene fumarate) Scaffolds.

Two distinct molecular masses of poly(propylene fumarate) (PPF) are combined with an additive manufacturing process to fabricate highly complex scaffolds possessing controlled chemical properties and porous architecture. Scaffolds were manufactured with two polymer molecular masses and two architecture styles. Degradation was assessed in an accelerated in vitro environment. The purpose of the degradation study is not to model or mimic in vivo degradation, but to efficiently compare the effect of modulating scaffold properties. This is the first study addressing degradation of chain-growth synthesized PPF, a process that allows for considerably more control over molecular mass distribution. It demonstrates that, with greater process control, not only is scaffold fabrication reproducible, but the mechanical properties and degradation kinetics can be tailored by altering the physical properties of the scaffold. This is a clear step forward in using PPF to address unmet medical needs while meeting regulatory demands and ultimately obtaining clinical relevancy.