Trends in bioactivity: inducing and detecting mineralization of regenerative polymeric scaffolds.

Due to limitations of biological and alloplastic grafts, regenerative engineering has emerged as a promising alternative to treat bone defects. Bioactive polymeric scaffolds are an integral part of such an approach. Bioactivity importantly induces hydroxyapatite mineralization that promotes osteoinductivity and osseointegration with surrounding bone tissue. Strategies to confer bioactivity to polymeric scaffolds utilize bioceramic fillers, coatings and surface treatments, and additives. These approaches can also favorably impact mechanical and degradation properties. A variety of fabrication methods are utilized to prepare scaffolds with requisite morphological features. The bioactivity of scaffolds may be evaluated with a broad set of techniques, including in vitro (acellular and cellular) and in vivo methods. Herein, we highlight contemporary and emerging approaches to prepare and assess scaffold bioactivity, as well as existing challenges.

Comparative evaluation of mesenchymal stromal cell growth and osteogenic differentiation on a shape memory polymer scaffold.

Trauma-induced, critical-size bone defects pose a clinical challenge to heal. Albeit autografts are the standard-of-care, they are limited by their inability to be shaped to various defect geometries and often incur donor site complications. Herein, the combination of a "self-fitting" shape memory polymer (SMP) scaffold and seeded mesenchymal stromal cells (MSCs) was investigated as an alternative. The porous SMP scaffold, prepared from poly(ε-caprolactone) diacrylate (PCL-DA) and coated with polydopamine, provided conformal shaping and cell adhesion. MSCs from five tissues, amniotic (AMSCs), chorionic tissue (CHSCs), umbilical cord (UCSCs), adipose (ADSCs), and bone marrow (BMSCs) were evaluated for viability, density, and osteogenic differentiation on the SMP scaffold. BMSCs exhibited the fastest increase in cell density by day 3, but after day 10, CHSCs, UCSCs, and ADSCs approached similar cell density. BMSCs also showed the greatest calcification among the cell types, followed closely by ADSCs, CHSCs and AMSCs. Alkaline phosphatase (ALP) activity peaked at day 7 for AMSCs, UCSCs, ADSCs and BMSCs, and at day 14 for CHSCs, which had the greatest overall ALP activity. Of all the cell types, only scaffolds cultured with ADSCs in osteogenic media had increased hardness and local modulus as compared to blank scaffolds after 21 days of cell culture and osteogenic differentiation. Overall, ADSCs performed most favorably on the SMP scaffold. The SMP scaffold was able to support key cellular behaviors of MSCs and could potentially be a viable, regenerative alternative to autograft.

Suitability of EtO Sterilization for Polydopamine-coated, Self-fitting Bone Scaffolds.

Irregularly shaped craniomaxillofacial (CMF) defects may be advantageously treated by "self-fitting" shape memory polymer (SMP) scaffolds, namely those prepared from poly(ε-caprolactone)diacrylate (PCL-DA) networks and PCL-DA/poly(L-lactic acid) (PLLA) (75:25 wt%) semi-interpenetrating polymer networks (semi-IPNs). In addition to achieving good scaffold-tissue contact, a polydopamine (PD) coating can be leveraged to enhance bioactivity for improved osseointegration. Sterilization with ethylene oxide (EtO) represents a logical choice due to its low operating temperature and humidity. Herein, for the first time, the impact of EtO sterilization on the material properties of PD-coated SMP scaffolds was systematically assessed. Morphological features (i.e., pore size and pore interconnectivity), and in vitro bioactivity were preserved as were PCL crystallinity, PLLA crystallinity, and crosslinking. These latter features led to sustained shape memory properties, and compressive modulus. EtO-sterilized, PD-coated scaffolds displayed similar in vitro degradation behaviors versus analogous non-sterilized scaffolds. This included maintenance of compression modulus following 28 days of exposure to non-accelerated degradation conditions.

Evaluation of a self-fitting, shape memory polymer scaffold in a rabbit calvarial defect model.

Self-fitting scaffolds prepared from biodegradable poly(ε-caprolactone)-diacrylate (PCL-DA) have been developed for the treatment of craniomaxillofacial (CMF) bone defects. As a thermoresponsive shape memory polymer (SMP), with the mere exposure to warm saline, these porous scaffolds achieve a conformal fit in defects. This behavior was expected to be advantageous to osseointegration and thus bone healing. Herein, for an initial assessment of their regenerative potential, a pilot in vivo study was performed using a rabbit calvarial defect model. Exogenous growth factors and cells were excluded from the scaffolds. Key scaffold material properties were confirmed to be maintained following gamma sterilization. To assess scaffold integration and neotissue infiltration along the defect perimeter, non-critically sized (d = 8 mm) bilateral calvarial defects were created in 12 New Zealand white rabbits. Bone formation was assessed at 4 and 16 weeks using histological analysis and micro-CT, comparing defects treated with an SMP scaffold (d = 9 mm x t = 1 or 2 mm) to untreated defects (i.e. defects able to heal without intervention). To further assess osseointegration, push-out tests were performed at 16 weeks and compared to defects treated with poly(ether ether ketone) (PEEK) discs (d = 8.5 mm x t = 2 mm). The results of this study confirmed that the SMP scaffolds were biocompatible and highly conducive to bone formation and ingrowth at the perimeter. Ultimately, this resulted in similar bone volume and surface area versus untreated defects and superior performance in push-out testing versus defects treated with PEEK discs. STATEMENT OF SIGNIFICANCE: Current treatments of craniomaxillofacial (CMF) bone defects include biologic and synthetic grafts but they are limited in their ability to form good contact with adjacent tissue. A regenerative engineering approach using a biologic-free scaffold able to achieve conformal fitting represents a potential "off-the-shelf" surgical product to heal CMF bone defects. Having not yet been evaluated in vivo, this study provided the preliminary assessment of the bone healing potential of self-fitting PCL scaffolds using a rabbit calvarial defect model. The study was designed to assess scaffold biocompatibility as well as bone formation and ingrowth using histology, micro-CT, and biomechanical push-out tests. The favorable results provide a basis to pursue establishing self-fitting scaffolds as a treatment option for CMF defects.

Bioactive Siloxane-Containing Shape-Memory Polymer (SMP) Scaffolds with Tunable Degradation Rates.

A material-guided, regenerative approach to heal cranial defects requires a scaffold that cannot only achieve conformal fit into irregular geometries but also has bioactivity and suitable resorption rates. We have previously reported "self-fitting" shape-memory polymer (SMP) scaffolds based on poly(ε-caprolactone) diacrylate (PCL-DA) that shape recover to fill irregular defect geometries. However, PCL-DA scaffolds lack innate bioactivity and degrade very slowly. Polydimethylsiloxane (PDMS) has been shown to impart innate bioactivity and modify degradation rates when combined with organic cross-linked networks. Thus, this work reports the introduction of PDMS segments to form PCL/PDMS SMP scaffolds. These were prepared as co-matrices with three types of macromers to systematically alter PDMS content and cross-link density. Specifically, PCL90-DA was combined with linear-PDMS66-dimethacrylate (DMA) or 4-armed star-PDMS66-tetramethacrylate (TMA) macromers at 90:10, 75:25, and 60:40 wt % ratios. Additionally, a triblock macromer (AcO-PCL45-b-PDMS66-b-PCL45-OAc), having a 65:35 wt % ratio PCL/PDMS, was used. Scaffolds exhibited pore interconnectivity and uniform pore sizes and further maintained excellent shape-memory behavior. Degradation rates increased with PDMS content and reduced cross-link density, with phase separation contributing to this effect. Irrespective of PDMS content, all PCL/PDMS scaffolds exhibited the formation of carbonated hydroxyapatite (HAp) following exposure to simulated body fluid (SBF). While inclusion of PDMS expectedly reduced scaffold modulus and strength, mineralization increased these properties and, in some cases, to values exceeding or similar to the PCL-DA, which did not mineralize.