Abby R. Whittington

Calcium phosphate ceramics in bone tissue engineering: A review of properties and their influence on cell behavior

Calcium phosphate ceramics (CPCs) have been widely used as biomaterials for the regeneration of bone tissue because of their ability to induce osteoblastic differentiation in progenitor cells. Despite the progress made towards fabricating CPCs possessing a range of surface features and chemistries, the influence of material properties in orchestrating cellular events such as adhesion and differentiation is still poorly understood. Specifically, questions such as why certain CPCs may be more osteoinductive than others, and how material properties contribute to osteoinductivity/osteoconductivity remain unanswered. Therefore, this review article systematically discusses the effects of the physical (e.g. surface roughness) and chemical properties (e.g. solubility) of CPCs on protein adsorption, cell adhesion and osteoblastic differentiation in vitro. The review also provides a summary of possible signaling pathways involved in osteoblastic differentiation in the presence of CPCs. In summary, these insights on the contribution of material properties towards osteoinductivity and the role of signaling molecules involved in osteoblastic differentiation can potentially aid the design of CPC-based biomaterials that support bone regeneration without the need for additional biochemical supplements.

Fabrication of a model continuously graded co-electrospun mesh for regeneration of the ligament–bone interface

Current scaffolds for the regeneration of anterior cruciate ligament injuries are unable to capture intricate mechanical and chemical gradients present in the natural ligament-bone interface. As a result, stress concentrations can develop at the scaffold-bone interface, leading to poor osseointegration. Hence, scaffolds that possess appropriate mechano-chemical gradients would help establish normal loading properties at the interface, while promoting scaffold integration with bone. With the long-term goal of investigating regeneration of the ligament-bone interface, this feasibility study aimed to fabricate a continuously graded mesh. Specifically, graded meshes were fabricated by co-electrospinning nanohydroxyapatite/polycaprolactone (nHAP-PCL) and poly(ester urethane) urea elastomer solutions from offset spinnerets. Next, mineral crystallites were selectively deposited on the nHAP-PCL fibers by treatment with a 5× simulated body fluid (5× SBF). X-ray diffraction and energy-dispersive spectroscopy indicated calcium-deficient hydroxyapatite-like mineral crystallites with an average Ca/P ratio of 1.48. Tensile testing demonstrated the presence of a mechanical gradient, which became more pronounced upon treatment with 5× SBF. Finally, biocompatibility of the graded meshes was verified using an MC3T3-E1 osteoprogenitor cell line. The study demonstrates that graded meshes, for potential application in interfacial tissue engineering, can be fabricated by co-electrospinning.

Response of bone marrow stromal cells to graded co-electrospun scaffolds and its implications for engineering the ligament-bone interface

Biomaterial scaffolds with gradients in architecture, mechanical and chemical properties have the potential to improve the osseointegration of ligament grafts by recapitulating phenotypic gradients that exist at the natural ligament-bone (L-B) interface. Towards the larger goal of regenerating the L-B interface, this in vitro study was performed to investigate the potential of two scaffolds with mineral gradients in promoting a spatial gradient of osteoblastic differentiation. Specifically, the first graded scaffold was fabricated by co-electrospinning two polymer solutions (one doped with nano-hydroxyapatite particles) from offset spinnerets, while the second was created by immersing the first scaffold in a 5 × simulated body fluid. Rat bone marrow stromal cells, cultured in the presence of osteogenic supplements, were found to be metabolically active on all regions of both scaffolds after 1 and 7 days of culture. Gene expression of bone morphogenic protein-2 and osteopontin was elevated on mineral-containing regions as compared to regions without mineral, while the expression of alkaline phosphatase mRNA revealed the opposite trend. Finally, the presence of osteopontin and bone sialoprotein confirmed osteoblastic phenotypic maturation by day 28. This study indicates that co-electrospun scaffolds with gradients in mineral content can guide the formation of phenotypic gradients and may thus promote the regeneration of the L-B interface.

Electrospun meshes possessing region‐wise differences in fiber orientation, diameter, chemistry and mechanical properties for engineering bone‐ligament‐bone tissues

Although bone‐patellar tendon‐bone (B‐PT‐B) autografts are the gold standard for repair of anterior cruciate ligament ruptures, they suffer from drawbacks such as donor site morbidity and limited supply. Engineered tissues modeled after B‐PT‐B autografts are promising alternatives because they have the potential to regenerate connective tissue and facilitate osseointegration. Towards the long‐term goal of regenerating ligaments and their bony insertions, the objective of this study was to construct 2D meshes and 3D cylindrical composite scaffolds – possessing simultaneous region‐wise differences in fiber orientation, diameter, chemistry and mechanical properties – by electrospinning two different polymers from off‐set spinnerets. Using a dual drum collector, 2D meshes consisting of an aligned polycaprolactone (PCL) fiber region, randomly oriented poly(lactide‐co‐glycolide) (PLGA) fiber region and a transition region (comprised of both PCL and PLGA fibers) were prepared, and region‐wise differences were confirmed by microscopy and tensile testing. Bone marrow stromal cells (BMSCs) cultured on these meshes exhibited random orientations and low aspect ratios on the random PLGA regions, and high aspect ratios and alignment on the aligned PCL regions. Next, meshes containing an aligned PCL region flanked by two transition regions and two randomly oriented PLGA regions were prepared and processed into 3D cylindrical composite scaffolds using an interpenetrating photo‐crosslinkable polyethylene glycol diacrylate hydrogel to recapitulate the shape of B‐PT‐B autografts. Tensile testing indicated that cylindrical composites were mechanically robust, and eventually failed due to stress concentration in the aligned PCL region. In summary, this study demonstrates a process to fabricate electrospun meshes possessing region‐wise differences in properties that can elicit region‐dependent cell responses, and be readily processed into scaffolds with the shape of B‐PT‐B autografts. Biotechnol. Bioeng. 2014;111: 2549–2559.

Using startup of steady shear flow in a sliding plate rheometer to determine material parameters for the purpose of predicting long fiber orientation

The properties of long glass fiber reinforced parts, such as those manufactured by means of injection molding and compression molding, are highly dependent on the fiber orientation generated during processing. A sliding plate rheometer was used to understand the transient stress and orientation development of concentrated long glass fibers during the startup of steady shear flow. An orientation model and stress tensor combination, based on semiflexible fibers, was assessed in its ability to predict fiber orientation when using model parameters obtained from the fits of the stress responses. Specifically, samples of different initial fiber orientations was subjected to the startup of steady shear flow, and an orientation model based on bead and rod theory was coupled with a derived stress tensor that accounts for the semiflexibility of the fibers to obtain the corresponding model parameters. The results showed the semiflexible orientation model and stress tensor combination, overall, provided improved rheo...

Influence of therapeutic radiation on polycaprolactone and polyurethane biomaterials

Biomedical polymers are exposed in vivo to ionizing radiation as implants, coatings and bystander materials. High levels of ionizing radiation (e.g. X-ray and gamma) have been reported to cause degradation and/or cross-linking in many polymers. This pilot study sought to determine causes of failure, by investigating how therapeutic radiation affects two different porous polymeric scaffolds: polycaprolactone (PCL) and polyurethane (PU). PCL is a bioresorbable material used in biomedical devices (e.g., dentistry, internal fixation devices and targeted drug delivery capsules). PU is commonly used in medical applications (e.g., coatings for pacemakers, tissue expanders, catheter tubing and wound dressings). PU was specifically fabricated to be a non-degradable polymer in this study. Porous scaffolds, fabricated using solvent casting and/or salt leeching techniques, were placed in phosphate buffered saline (PBS, pH=7.4) and exposed to typical cancer radiotherapy. A total dose of 50 Gy was broken into 25 doses over an eleven-week period. Collected PBS was tested for polymer leachants and degradation products using Gas Chromatography Mass Spectroscopy (GC-MS), results revealed no analyzable leachants from either polymer. Scaffolds were characterized using Environmental Scanning Electron Microscopy, Size-exclusion chromatography (SEC), Differential Scanning Calorimetry (DSC) and Fourier Transform Infrared Spectroscopy (FTIR). No gross visual changes were observed in either polymer, however PU exhibited microstructure changes after irradiation. Increased number average molecular weight and weight average molecular weight in PCL and PU were observed after irradiation, indicating crosslinking. PU displayed an increase in intrinsic viscosity that further confirms increased crosslinking. PCL and PU showed decreases in crystallinity after irradiation, and PU crystallinity shifted from long-range-order hard segments to short-range-order hard segments after irradiation. Results from both PCL and PU suggest changes in polymer backbones. This preliminary study suggests that therapeutic radiation doses cause both degradation and crosslinking in PCL and PU.

Poly(ether ester) Ionomers as Water-Soluble Polymers for Material Extrusion Additive Manufacturing Processes

Water-soluble polymers as sacrificial supports for additive manufacturing (AM) facilitate complex features in printed objects. Few water-soluble polymers beyond poly(vinyl alcohol) enable material extrusion AM. In this work, charged poly(ether ester)s with tailored rheological and mechanical properties serve as novel materials for extrusion-based AM at low temperatures. Melt transesterification of poly(ethylene glycol) (PEG, 8k) and dimethyl 5-sulfoisophthalate afforded poly(ether ester)s of sufficient molecular weight to impart mechanical integrity. Quantitative ion exchange provided a library of poly(ether ester)s with varying counterions, including both monovalent and divalent cations. Dynamic mechanical and tensile analysis revealed an insignificant difference in mechanical properties for these polymers below the melting temperature, suggesting an insignificant change in final part properties. Rheological analysis, however, revealed the advantageous effect of divalent countercations (Ca2+, Mg2+, and Zn2+) in the melt state and exhibited an increase in viscosity of two orders of magnitude. Furthermore, time-temperature superposition identified an elevation in modulus, melt viscosity, and flow activation energy, suggesting intramolecular interactions between polymer chains and a higher apparent molecular weight. In particular, extrusion of poly(PEG8k-co-CaSIP) revealed vast opportunities for extrusion AM of well-defined parts. The unique melt rheological properties highlighted these poly(ether ester) ionomers as ideal candidates for low-temperature material extrusion additive manufacturing of water-soluble parts.

A review on fabricating tissue scaffolds using vat photopolymerization

Vat Photopolymerization (stereolithography, SLA), an Additive Manufacturing (AM) or 3D printing technology, holds particular promise for the fabrication of tissue scaffolds for use in regenerative medicine. Unlike traditional tissue scaffold fabrication techniques, SLA is capable of fabricating designed scaffolds through the selective photopolymerization of a photopolymer resin on the micron scale. SLA offers unprecedented control over scaffold porosity and permeability, as well as pore size, shape, and interconnectivity. Perhaps even more significantly, SLA can be used to fabricate vascular networks that may encourage angio and vasculogenesis. Fulfilling this potential requires the development of new photopolymers, the incorporation of biochemical factors into printed scaffolds, and an understanding of the effects scaffold geometry have on cell viability, proliferation, and differentiation. This review compares SLA to other scaffold fabrication techniques, highlights significant advances in the field, and offers a perspective on the fields challenges and future directions. STATEMENT OF SIGNIFICANCE Engineering de novo tissues continues to be challenging due, in part, to our inability to fabricate complex tissue scaffolds that can support cell proliferation and encourage the formation of developed tissue. The goal of this review is to first introduce the reader to traditional and Additive Manufacturing scaffold fabrication techniques. The bulk of this review will then focus on apprising the reader of current research and provide a perspective on the promising use of vat photopolymerization (stereolithography, SLA) for the fabrication of complex tissue scaffolds.

Investigation into Polyurethane at Varying Dose Rates of Ionizing Radiation for Clinical Application

Polyurethanes (PUs) are commonly used materials for medical devices. These devices are exposed repeatedly to radiation when patients undergo radiotherapy treatments. It has been found that peripherally inserted central catheters (PICCs) and central venous catheters (CVCs) fail at an increased rate (14.7% and 8.8%, respectively) when radiated. Currently, little research is available on increased failure seen in conjunction with radiation, but complex in vivo environments within a human patient make it difficult to isolate effects of individual variables. This research investigated effects of radiation in an aqueous environment to determine whether radiation combined with a mimicked in vivo environment is sufficient to change PU devices. The following dose rates were used in this study: 3.2 Gy·min−1, 4.5 Gy·min−1, 44 Gy·min−1, and 833 Gy·min−1. Samples were characterized in four main ways: cellular response, physical changes, chemical changes, and mechanical changes. Results reveal normal cellular response at all dose rates, indicating dose rate does not alter cellular adhesion or proliferation, and biocompatibility of the material is not being altered. Results from physical, chemical, and mechanical effects confirm that varying dose rates alone do not initiate material changes, which negates the hypothesis that varying dose rates of radiation contribute to the complications in PICC and CVCs.