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The study aims to assess the safety and effectiveness of BoneTape™, a new resorbable bone fixation device, using a zygomatic fracture model in rabbits. The study followed BoneTape™ samples and control (sham) groups over 2-, 6-, and 12-week periods post-zygomaticomaxillary (ZM) osteotomy and zygomaticofrontal (ZF) disarticulation. The osteotomized segments were analyzed for bone healing, inflammatory response, and tissue healing. µCT imaging and histological analysis were used to examine the axial alignment, offset, and quality of new bone formation. BoneTape™ samples demonstrated enhanced maintenance of the initial intraoperative positioning, reduced axial offset, and better alignment when compared with the control group, enabling stable bone healing under physiological loading conditions. Complete union was observed at 12-weeks in both groups. The BoneTape™ group experienced minimal immune and tissue reactions, classically associated with wound healing, and showed an increased number of giant cells at 6 and 12-weeks. BoneTape™ represents a promising advancement in osteosynthesis, demonstrating efficacy in maintaining stable zygomatic reconstruction and eliciting minimal immune response in a rabbit model. This study introduces BoneTape™ as a disruptive solution specifically designed for clinical application in cranio-maxillofacial fracture fixation, with the potential to eliminate the use of over-engineered solutions while offering benefits such as ease of application and fewer biologically disruptive steps. [ABSTRACT FROM AUTHOR]

The ex vivo endothelialization of small diameter vascular prostheses can prolong their patency. Here, we demonstrate that heterotypic interactions between human adipose tissue-derived endothelial cells and perivascular cells can be exploited to accelerate the endothelialization of an electrospun ionomeric polyurethane scaffold. The scaffold was used to physically separate endothelial cells from perivascular cells to prevent their diffuse neo-intimal hyperplasia and spontaneous tubulogenesis, yet enable their paracrine cross-talk to accelerate the integration of the endothelial cells into a temporally stable endothelial lining of a continuous, elongated, and aligned morphology. Perivascular cells stimulated endothelial basement membrane protein production and suppressed their angiogenic and inflammatory activation to accelerate this biomimetic morphogenesis of the endothelium. These findings demonstrate the feasibility and underscore the value of exploiting heterotypic interactions between endothelial cells and perivascular cells for the fabrication of an endothelial lining intended for small diameter arterial reconstruction. Adipose tissue is an abundant, accessible, and uniquely dispensable source of endothelial cells and perivascular cells for vascular tissue engineering. While their spontaneous self-assembly into microvascular networks is routinely exploited for the vascularization of engineered tissues, it threatens the temporal stability of an endothelial lining intended for small diameter arterial reconstruction. Here, we demonstrate that an electrospun polyurethane scaffold can be used to physically separate endothelial cells from perivascular cells to prevent their spontaneous capillary morphogenesis, yet enable their cross-talk to promote the formation of a stable endothelium. Our findings demonstrate the feasibility of engineering an endothelial lining from human adipose tissue, poising it for the rapid ex vivo endothelialization of small diameter vascular prostheses in an autologous, patient-specific manner. [Display omitted] [ABSTRACT FROM AUTHOR]

New functional materials for engineering gingival tissue are still in the early stages of development. Materials for such applications must maintain volume and have advantageous mechanical and biological characteristics for tissue regeneration, to be an alternative to autografts, which are the current benchmark of care. In this work, methacrylated gelatin (GelMa) was photocrosslinked with synthetic immunomodulatory methacrylated divinyl urethanes and defined monomers to generate composite scaffolds. Using a factorial design, with the synthetic monomers of a degradable polar/hydrophobic/ionic polyurethane (D-PHI) and GelMa, composite materials were electrospun with polycarbonate urethane (PCNU) and light-cured in-flight. The materials had significantly different relative hydrophilicities, with unique biodegradation profiles associated with specific formulations, thereby providing good guidance to achieving desired mechanical characteristics and scaffold resorption for gingival tissue regeneration. In accelerated esterase/collagenase degradation models, the new materials exhibited an initial rapid weight loss followed by a more gradual rate of degradation. The degradation profile allowed for the early infiltration of human adipose-derived stromal/stem cells, while still enabling the graft's structural integrity to be maintained. In conclusion, the materials provide a promising candidate platform for the regeneration of oral soft tissues, addressing the requirement of viable tissue infiltration while maintaining volume and mechanical integrity. There is a need for the development of more functional and efficacious materials for the treatment of gingival recession. To address significant limitations in current material formulations, we sought to investigate the development of methacrylated gelatin (GelMa) and oligo-urethane/methacrylate monomer composite materials. A factorial design was used to electrospin four new formulations containing four to five monomers. Synthetic immunomodulatory monomers were crosslinked with GelMa and electrospun with a polycarbonate urethane resulting in unique mechanical properties, and resorption rates which align with the original design criteria for gingival tissue engineering. The materials may have applications in tissue engineering and can be readily manufactured. The findings of this work may help better direct the efforts of tissue engineering and material manufacturing. [Display omitted] [ABSTRACT FROM AUTHOR]

Biodegradable hydrogels are growing in demand to enable the delivery of biomolecules (e.g. growth factors) for regenerative medicine. This research investigated the resorption of an oligourethane/polyacrylic acid hydrogel, a biodegradable hydrogel which supports tissue regeneration. The Arrhenius model was used to characterize the resorption of the polymeric gels in relevant in vitro conditions, and the Flory-Rehner equation was used to correlate the volumetric swelling ratio with the extent of degradation. The study found that the swelling rate of the hydrogel follows the Arrhenius model at elevated temperatures, estimating degradation time in saline solution at 37°C to be between 5 and 13 months, serving as a preliminary approximation of degradation in vivo. The degradation products had low cytotoxicity towards endothelial cells, and the hydrogel supported stromal cell proliferation. Additionally, the hydrogels were able to release growth factors and maintain the biomolecules' bioactivity towards cell proliferation. The study of the vascular endothelial growth factor (VEGF) release from the hydrogel used a diffusion process model, showing that the electrostatic attraction between VEGF and the anionic hydrogel allowed for controlled and sustained VEGF release over three weeks. In a rat subcutaneous implant model, a selected hydrogel with desired degradation rates exhibited minimal foreign body response and supported M2a macrophage phenotype, and vascularization. The low M1 and high M2a macrophage phenotypes within the implants were associated with tissue integration. This research supports the use of oligourethane/polyacrylic acid hydrogels as a promising material for delivering growth factors and supporting tissue regeneration. There is a need for degradable elastomeric hydrogels that can support the formation of soft tissues and minimize long-term foreign body responses. An Arrhenius model was used to estimate the relative breakdown of hydrogels, in-vitro. The results demonstrate that hydrogels made from a combination of poly(acrylic acid) and oligo-urethane diacrylates can be designed to resorb over defined periods ranging from months to years depending on the chemical formulation prescribed by the model. The hydrogel formulations also provided for different release profiles of growth factors, relevant to tissue regeneration. In-vivo , these hydrogels had minimal inflammatory effects and showed evidence of integration into the surrounding tissue. The hydrogel approach can help the field design a broader range of biomaterials for tissue regeneration. [Display omitted] [ABSTRACT FROM AUTHOR]

The use of exogenous biomolecules (BM) for the purpose of repairing and regenerating damaged cardiac tissue can yield serious side effects if used for prolonged periods. As well, such strategies can be cost prohibitive depending on the regiment and period of time applied. Alternatively, autologous monocytes/monocyte-derived macrophages (MDM) can provide a viable path towards generating an endogenous source of stimulatory BM. Biomaterials are often considered as delivery vehicles to generate unique profiles of such BM in tissues or to deliver autologous cells, that can influence the nature of BM produced by the cells. MDM cultured on a degradable polar hydrophobic ionic (D-PHI) polyurethane has previously demonstrated a propensity to increase select anti-inflammatory cytokines, and therefore there is good rationale to further investigate a broader spectrum of the cells' BM in order to provide a more complete proteomic analysis of human MDM secretions induced by D-PHI. Further, it is of interest to assess the potential of such BM to influence cells involved in the reparative state of vital tissues such as those that affect cardiac cell function. Hence, this current study examines the proteomic profile of MDM secretions using mass spectrometry for the first time, along with ELISA, following their culture on D-PHI, and compares them to two important reference materials, poly(lactic-co-glycolic acid) (PLGA) and tissue culture polystyrene (TCPS). Secretions collected from D-PHI cultured MDM led to higher levels of regenerative BM, AGRN, TGFBI and ANXA5, but lower levels of pro-fibrotic BM, MMP7, IL-1β, IL-6 and TNFα, when compared to MDM secretions collected from PLGA and TCPS. In the application to cardiac cell function, the secretion collected from D-PHI cultured MDM led to more human cardiac fibroblast (HCFs) migration. A lower collagen gel contraction induced by MDM secretions collected from D-PHI was supported by gene array analysis for human fibrosis-related genes. The implication of these findings is that more tailored biomaterials such as D-PHI, may lead to a lower pro-inflammatory phenotype of macrophages when used in cardiac tissue constructs, thereby enabling the development of vehicles for the delivery of interventional therapies, or be applied as coatings for sensor implants in cardiac tissue that minimize fibrosis. The general approach of using synthetic biomaterials in order to induce MDM secretions in a manner that will guide favorable regeneration will be critical in making the choice of biomaterials for tissue regeneration work in the future. Immune modulation strategies currently applied in cardiac tissue repair are mainly based on the delivery of defined exogenous biomolecules. However, the use of such biomolecules may pose wide ranging systemic effects, thereby rendering them clinically less practical. The chemistry of biomaterials (used as a potential targeted delivery modality to circumvent the broad systemic effects of biomolecules) can not only affect acute and chronic toxicity but also alters the timeframe of the wound healing cascade. In this context, monocytes/monocyte-derive macrophages (MDM) can be harnessed as an immune modulating strategy to promote wound healing by an appropriate choice of the biomaterial. However, there are limited reports on the complete proteome analysis of MDM and their reaction of biomaterial related interventions on cardiac tissues and cells. No studies to date have demonstrated the complete proteome of MDM secretions when these cells were cultured on a non-traditional immune modulatory ionomeric polyurethane D-PHI film. This study demonstrated that MDM cultured on D-PHI expressed significantly higher levels of AGRN, TGFBI and ANXA5 but lower levels of MMP7, IL-1β, IL-6 and TNFα when compared to MDM cultured on a well-established degradable biomaterials in the medical field, e.g. PLGA and TCPS, which are often used as the relative standards for cell culture work in the biomaterials field. The implications of these findings have relevance to the repair of cardiac tissues. In another aspect of the work, human cardiac fibroblasts showed significantly lower contractility (low collagen gel contraction and low levels of ACTA2) when cultured in the presence of MDM secretions collected after culturing them on D-PHI compared to PLGA and TCPS. The findings place emphasis on the importance of making the choice of biomaterials for tissue engineering and regenerative medicine applied to their use in cardiac tissue repair. Image, graphical abstract [ABSTRACT FROM AUTHOR]

In vascular tissue engineering, the ability to obtain a robust and safe vascular tissue cell source (e.g. vascular smooth muscle cells (VSMCs)) and to promote vascular tissue-specific extracellular matrix (ECM) protein production is critically important. Mature blood vessel-derived VSMCs are not practical for in vitro vascular tissue regeneration. The authors have conceived a strategy to differentiate adipose derived stromal cells (ASCs) into VSMC-like cells (ASC-VSMCs) that were similar to mature umbilical artery VSMCs at the transcriptional, protein and contraction function levels. Monocytes/macrophages are known as important regulators of the inflammation and regeneration processes within different tissue types of the body. However, our understanding of the potential interactions between specific tissue-like cells differentiated from stem/stromal cells (e.g. ASC-VSMCs) and monocytes/macrophages (cued by specific biomaterial scaffolds) is still limited. In this study, indirect and direct ASC-VSMC-monocyte co-cultures were constructed within a porous polyurethane scaffold (D-PHI) previously shown to have an immunomodulatory character. The effects of monocytes/macrophages on the cellularity (cell number detected with DNA quantification assay), ECM (glycosaminoglycan (GAG), collagen, and elastin) accumulation as well as the maintenance of contractile VSMC markers (calponin and smoothelin) of the ASC-VSMCs after a month of co-culture were investigated. It was found that monocyte paracrine signalling in D-PHI positively affected the cellularity and ECM accumulation of ASC-VSMCs in co-culture. Cause-effect relationships were also identified between the release of pro-inflammatory/anti-inflammatory factors (i.e. IL6, TGF-β1) in co-culture and the expression of contractile proteins (calponin and smoothelin) by ASC-VSMCs. This study demonstrated the importance of combining an immune cell strategy with stromal cell derived VSMCs (i.e. ASC-VSMCs) to achieve a practical vascular tissue engineering outcome. Adipose stromal cell derived-vascular smooth muscle cells (ASC-VSMCs) are a promising cell source for vascular tissue engineering. Monocytes/monocyte derived macrophages can be harnessed as an immune-assisted strategy to promote vascular tissue regeneration. This study demonstrated that the co-culture of human ASC-VSMCs with monocytes significantly enhanced the cellularity and extracellular matrix (ECM) accumulation within anionic polyurethane (D-PHI) scaffolds, partially mediated by monocyte paracrine signalling mechanisms. In addition, specific VSMC contractile markers (calponin and smoothelin) were still present in ASC-VSMCs when the cells were exposed to monocytes for a month in vitro. This study corroborated the potential selection of ASC-VSMCs for in vitro engineering of vascular tissue in an immunomodulatory biomaterial scaffold (e.g. D-PHI) based co-culture system containing monocytes. Image, graphical abstract [ABSTRACT FROM AUTHOR]

Tissue scaffolds need to be engineered to be cell compatible, have timely biodegradable character, be functional with respect to providing niche cell support for tissue repair and regeneration, readily accommodate multiple cell types, and have mechanical properties that enable the simulation of the native tissue. In this study, electrospun degradable polar hydrophobic ionic polyurethane (D-PHI) scaffolds were generated in order to yield an extracellular matrix-like structure for tissue engineering applications. D-PHI oligomers were synthesized, blended with a degradable linear polycarbonate polyurethane (PCNU), and electrospun with simultaneous in situ UV cross-linking in order to generate aligned nanofibrous scaffolds in the form of elastomeric composite materials. The D-PHI/PCNU scaffold fibre morphology, cross-linking efficiency, surface nature, mechanical properties, in vivo degradation and integration, as well as in vitro cell compatibility were characterized. The results showed that D-PHI/PCNU scaffolds had a high cross-linking efficiency, stronger polar nature, and lower stiffness relative to PCNU scaffolds. In vivo , the D-PHI/PCNU scaffold degraded relatively slowly, thereby enabling new tissue time to form and yielding very good integration with the latter tissue. Based on a study with A10 vascular smooth muscle cells, the D-PHI/PCNU scaffold was able to support high cell viability, adhesion, and expression of typical smooth muscle cell markers after a 7-day culture period, which was comparable to PCNU scaffolds. These characterization results demonstrate that the unique properties of a D-PHI/PCNU scaffold, combined with the benefits of electrospinning, could allow for the generation of a tissue engineered scaffold that mimics important aspects of the native extracellular matrix and could be used for functional tissue regeneration. Tissue engineered scaffolds should recapitulate native extracellular matrix features. This study investigates the processing of a classical polycarbonate polyurethane (PCNU) with a cross-linked and degradable ionomeric polyurethane (D-PHI), polymerized via in situ rapid light curing to yield a 3-dimensional co-electrospun nanofibre matrix with chemical diversity and low modulus character. This research advances the use of D-PHI for tissue engineering applications by providing a facile means of changing physical and chemical properties in classical PCNUs without the need to adjust spinning viscosities of the base polymer. Further, the in vivo and cell culture findings set the stage for introducing unique elastic materials which inherently support wound healing, repair, and regeneration in tissues, for applications that require the recapitulation of native extracellular matrix physical features. [ABSTRACT FROM AUTHOR]

Graphical abstract Abstract The synthesis of microspheres for tissue regeneration requires good control over the particle size and size distribution. This is particularly important when considering the immune response that may be triggered by the presence of particles in tissue. This report outlines the design of an injectable microsphere system using a low-inflammatory, degradable-polar-hydrophobic-ionic polyurethane, termed D-PHI, and investigates the system's performance in vitro and in vivo. Crosslinked polyurethane microspheres were prepared via a rapid and controlled process based on membrane emulsion and subsequent photopolymerization. The fabrication process efficiently generated microspheres with a narrow size distribution (12 ± 2 μm, PDI = 0.03). The D-PHI microspheres exhibited a slow and controlled degradation and a high capacity for water uptake. Water within the particles existed primarily within the pores of the particles and to a lesser degree within the polymer matrix itself. D-PHI microspheres supported human endothelial and fibroblast cell growth, and they maintained human blood-derived monocytes in a low-inflammatory state. Sub-acute toxicity was assessed for the particles after being administered via intramuscular injection in the gastrocnemius muscle of rats. Cellular infiltration and vascularization into the tissue region where the particles were deposited were observed along with an absence of a fibrous capsule around the particles. The microspheres did not cause elevated human monocyte induced inflammatory character, and supported tissue integration without a prolonged inflammatory response in the rat muscle. These injectable, degradable and low-inflammatory microspheres provide an attractive system for potential drug delivery and tissue regeneration applications in future studies. Statement of Significance Biodegradable, synthetic polymers are attractive candidates for generating tailored drug delivery vehicles and tissue scaffolds owing to their diverse chemical and physical properties that can be customised for delivering defined macromolecules at specific sites in the body. The past two decades have yielded interesting work exploring the fabrication of polymer microspheres with a narrow size distribution. However, the markedly low number of synthetic polymer chemistries currently used for microsphere production exhibit elevated proinflammatory character, both acute and chronic. Furthermore, a limited number of studies have explored the biocompatibility and immune response of polymeric microspheres with human primary cells and in vivo. In the current study, a method was conceived for efficiently generating low-activating polyurethane microspheres with respect to in vitro monocytes and in vivo macrophages. The biodegradable polyurethane, which contained multiple chemistry function and which has previously demonstrated anti-inflammatory properties in film and mm scale scaffold form, was selected as the base material. In this work we undertook the use of a room temperature membrane emulsification photopolymerization approach to avoid the need for high temperature cures and the use of solvents. The response of immune cells to the microspheres was studied with human primary cells and in the rat gastrocnemius muscle. The present work reveals important progress in the design of microspheres, with well-characterized low monocyte-activating properties and the translational advantages of a synthetic polyurethane which could be investigated in future studies for potential macromolecule delivery and tissue regeneration applications. [ABSTRACT FROM AUTHOR]

Abstract Vascular smooth muscle cells (VSMCs) play essential roles in regulating blood vessel form and function and they are required for vascular tissue regeneration. Multipotent adipose derived stromal cells (ASCs) can be differentiated into VSMC-like cells, which can be used as a potential VSMC source for the development of vascular tissue. However, the effects of cryopreservation on the differentiation of ASCs towards VSMCs are poorly studied to date. This study compared fresh ASCs (FA) vs. cryopreserved ASCs (CA) with respect to their differentiation potential towards VSMC-like cells. The expression of contractile VSMC markers (such as smoothelin) and cell contractility were investigated. It was found that VSMC-like cells derived from CA expressed smoothelin gene and protein at lower levels and showed compromised contractility in response to vasoconstrictors, when compared with those derived from FA. Moreover, it was demonstrated that this negative effect of cryopreservation could be mediated by MEK1/2-ERK1/2, IFNγ and Smad2/3 associated Signalling pathways. Treatment of CA with MEK1/2-ERK1/2 activator or IFNγ neutralizing antibodies enhanced Smad2/3 phosphorylation and showed a rescue of the negative effect of cryopreservation on the differentiation of ASCs towards VSMC-like cells. These findings are important for defining approaches that may use cryopreserved ASCs for vascular tissue regeneration. Graphical Abstract Unlabelled Image Highlights • VSMC-like cells derived from CA expressed smoothelin gene/protein at lower levels. • CA derived VSMC-like cells showed compromised contractility with vasoconstrictors. • Negative effect of CA could be mediated by MEK1/2-ERK1/2, IFNγ, Smad2/3 Signaling. • Negative effect of CA could possibly be alleviated by intervening molecular agents. [ABSTRACT FROM AUTHOR]

Monocyte interactions with materials that are biofunctionalized with fibronectin (Fn) are of interest because of the documented literature which associates this protein with white blood cell function at implant sites. A degradable-polar hydrophobic ionic polyurethane (D-PHI), has been reported to promote an anti-inflammatory response from human monocytes. The aim of the current work was to study the influence of intrinsic D-PHI material chemistry on Fn adsorption (mono and multi-layer structures), and to investigate the influence of such chemistry on the structural state of the Fn, as well as the latter’s influence on the activity of human monocytes on the protein coated substrates. Significant differences in Fn adsorption, surface hydrophobicity and the availability of defined peptide sequences (N terminal, C terminal or Cell Binding Domain) for the Fn in mono vs multilayer structures were observed as a function of the changes in intrinsic material chemistry. A D-PHI-formulated polyurethane substrate with subtle changes in anionic and hydrophobic domain content relative to the polar non-ionic urethane/carbonate groups within the polymer matrix promoted the lowest activation of monocytes, in the presence of multi-layer Fn constructs. These results highlight the importance of chemical heterogeneity as a design parameter for biomaterial surfaces, and establishes a desired strategy for controlling human monocyte activity at the surface of devices, when these are coated with multi-layer Fn structures. The latter is an important step towards functionalizing the materials with multi-layer protein drug carriers as interventional therapeutic agents. Statement of Significance The control of the behavior of monocytes, especially migration and activation, is of crucial interest to modulate the inflammatory response at the site of implanted biomaterial. Several studies report the influence of adsorbed serum proteins on the behavior of monocytes on biomaterials. However, few studies show the influence of surface chemical group distribution on the controlled adsorption and the subsequent induced conformation- of mono versus multi-layer assembled structures generated from specific proteins implicated in wound repair. The current research considered the role of Fn adsorption and conformation in thin films while interacting with the intrinsic chemistry of segmented block polyurethanes; and the influence of the former on modulation and activation of human monocytes. [ABSTRACT FROM AUTHOR]

Tissue engineering (particularly for the case of load-bearing cardiovascular and connective tissues) requires the ability to promote the production and accumulation of extracellular matrix (ECM) components (e.g., collagen, glycosaminoglycan and elastin). Although different approaches have been attempted in order to enhance ECM accumulation in tissue engineered constructs, studies of underlying signalling mechanisms that influence ECM deposition and degradation during tissue remodelling and regeneration in multi-cellular culture systems have been limited. The current study investigated vascular smooth muscle cell (VSMC)-monocyte co-culture systems using different VSMC:monocyte ratios, within a degradable polyurethane scaffold, to assess their influence on ECM generation and degradation processes, and to elucidate relevant signalling molecules involved in this in vitro vascular tissue engineering system. It was found that a desired release profile of growth factors (e.g. insulin growth factor-1 (IGF-1)) and hydrolytic proteases (e.g. matrix-metalloproteinases 2, 9, 13 and 14 (MMP2, MMP9, MMP13 and MMP14)), could be achieved in co-culture systems, yielding an accumulation of ECM (specifically for 2:1 and 4:1 VSMC:monocyte culture systems). This study has significant implications for the tissue engineering field (including vascular tissue engineering), not only because it identified important cytokines and proteases that control ECM accumulation/degradation within synthetic tissue engineering scaffolds, but also because the established culture systems could be applied to improve the development of different types of tissue constructs. Statement of Significance Sufficient extracellular matrix accumulation within cardiovascular and connective tissue engineered constructs is a prerequisite for their appropriate function in vivo . This study established co-culture systems with tissue specific cells (vascular smooth muscle cells (VSMCs)) and defined ratios of immune cells (monocytes) to investigate extracellular matrix (ECM) generation and degradation processes, revealing important mechanisms underlying ECM turnover during vascular tissue regeneration/remodelling. A specific growth factor (IGF-1), as well as hydrolytic proteases (e.g. MMP2, MMP9, MMP13 and MMP14), were identified as playing important roles in these processes. ECM accumulation was found to be dependent on achieving a desired release profile of these ECM-promoting and ECM-degrading factors within the multi-cellular microenvironment. The findings enhance our understanding of ECM deposition and degradation during in vitro tissue engineering and would be applicable to the repair or regeneration of a variety of tissues. [ABSTRACT FROM AUTHOR]

Biomaterial blood compatibility is a complex process that involves four key pathways, including the coagulation cascade, the complement system, platelets, and leukocytes. While many studies have addressed the initial contact of blood with homopolymeric (e.g. Teflon) or simple copolymeric (e.g. Dacron) biomaterials, relatively less attention has been given to investigating blood coagulation with respect to complex copolymeric systems containing well defined and diverse function. The current study sought to assess the hemocompatibility of a complex polyurethane (PU) containing a unique combination of polar, hydrophobic, and ionic domains (D-PHI). This included a whole blood (WB) study, followed by tests on the intrinsic and extrinsic coagulation pathways, complement activation, platelet activation, and an assessment of the effect of leukocytes on platelet-biomaterial interactions. A small increase in blood clot formation was observed on D-PHI in WB; however, there was no significant increase in clotting via the intrinsic coagulation cascade. No significant increase in platelet adhesion and only a very slight increase in platelet activation were observed in comparison to albumin-coated substrates (negative control). D-PHI showed mild complement activation and increased initiation of the extrinsic pathway of coagulation, along with the observation that leukocytes were important in mediating platelet-biomaterial interactions. It is proposed that complement is responsible for activating coagulation by inciting leukocytes to generate tissue factor (TF), which causes extrinsic pathway activation. This low level of blood clotting on D-PHI’s surface may be necessary for the beneficial wound healing of vascular constructs that has been previously reported for this material. Statement of Significance Understanding the hemocompatibility of devices intended for blood-contacting applications is important for predicting device failure. Hemocompatibility is a complex parameter (affected by at least four different mechanisms) that measures the level of thrombus generation and immune system activation resulting from blood-biomaterial contact. The complexity of hemocompatibility implies that homopolymers are unlikely to solve the clotting challenges that face most biomaterials. Diversity in surface chemistry (containing hydrophobic, ionic, and polar domains) obtained from engineered polyurethanes can lead to favourable interactions with blood. The current research considered the effect of a highly functionalized polyurethane biomaterial on all four mechanisms in order to provide a comprehensive in vitro measure of the hemocompatibility of this unique material and the important mechanisms at play. [ABSTRACT FROM AUTHOR]

Abstract: Periodontal diseases cause the breakdown of the tooth-supporting gingival tissue. In treatments aimed at gingival tissue regeneration, tissue engineering is preferred over the common treatments such as scaling. Perfused (dynamic) culture has been shown to increase cell growth in tissue-engineered scaffolds. Since gingival tissues are highly vascularized, it was desired to investigate the influence of perfusion on the function of human gingival fibroblasts (HGF) when cultured in a degradable/polar/hydrophobic/ionic polyurethane scaffold during the early culture phase (4weeks) of engineering gingival tissues. It was observed that the growth of HGF was continuous over 28days in dynamic culture (3-fold increase, p <0.05), while it was reduced after 14days in static culture (i.e. no flow condition). Cell metabolic activity, as measured by a WST-1 assay, and total protein production show that HGF were in different metabolic states in the dynamic vs. static cultures. Observations from scanning electron microscopy and type I collagen (Col I) production measured by Western blotting suggest that medium perfusion significantly promoted collagen production in HGF after the first 4weeks of culture (p <0.05). The different proliferative and metabolic states for HGF in the perfused scaffolds suggest a different cell phenotype which may favour tissue regeneration. [Copyright &y& Elsevier]

Abstract: In vascular tissue, elastin is an essential extracellular matrix protein that plays an important biomechanical and biological signalling role. Native elastin is insoluble and is difficult to extract from tissues, which results in its relatively rare use for the fabrication of vascular tissue engineering scaffolds. Recombinant elastin-like polypeptide-4 (ELP4), which mimics the structure and function of native tropoelastin, represents a practical alternative to the native elastic fibre for vascular applications. In this study, electrospinning was utilized to fabricate fibrous scaffolds which were subsequently surface modified with ELP4 and used as substrates for smooth muscle cell culture. ELP4 surface modified materials demonstrated enhanced smooth muscle cell (SMC) adhesion and maintenance of cell numbers over a 1-week period relative to controls. SMCs seeded on the ELP4 surface modified materials were also shown to exhibit the cell morphology and biological markers of a contractile phenotype including a spindle-like morphology, actin filament organization and smooth muscle myosin heavy chain expression. Competitive inhibition experiments demonstrated that the elastin–laminin cell surface receptor and its affinity for the VGVAPG peptide sequence on ELP4 molecules are likely involved in the initial SMC contact with the ELP4 modified materials. Elastin-like polypeptides show promise as surface modifiers for candidate scaffolds for engineering contractile vascular tissues. [Copyright &y& Elsevier]

Abstract: Activated monocytes can promote inflammation or wound repair, depending on the nature of the implant environment. Recent work showed that a degradable, polar-hydrophobic-ionic polyurethane (D-PHI) induced an anti-inflammatory monocyte phenotype. In the current study it is hypothesized that wound-healing phenotype monocytes (activated by D-PHI material chemistry) will promote human vascular smooth muscle cells (hVSMC) to attach and migrate into porous D-PHI scaffolds. hVSMC migration is necessary for hVSMC population of the scaffold and tissue formation to occur, and then, once tissue formation is complete, the monocyte should promote contractile phenotype markers in the hVSMC. hVSMC were cultured for up to 28days with or without monocytes and analyzed for cell viability, attachment (DNA) and migration. Lysates were analyzed for the hVSMC contractile phenotype markers calponin and α-smooth muscle actin (α-SMA) as well as urokinase plasminogen activator (uPA; pro-migration marker) using immunoblotting analysis. Histological staining showed that hVSMC alone remained around the perimeter of the scaffold, whereas co-culture samples had co-localization of monocytes with hVSMC in the pores, a more even cell distribution throughout the scaffold and increased total cell attachment (P <0.05). Co-culture samples had higher cell numbers and more DNA than the addition of both single cell cultures. The water-soluble tetrazolium-1 data suggested that cells were not dying over the 28day culture period. Calponin, also linked to cell motility, was maintained up to 28days in the co-culture and hVSMC alone, whereas α-SMA disappeared after 7days. Co-cultures on D-PHI showed that monocytes were activated to a wound-healing phenotype (low TNF-α, elevated IL-10), while promoting uPA expression. In summary, this study showed that, by co-culturing monocytes with hVSMC, the latter showed increased total cell attachment and infiltration into the D-PHI scaffold compared with hVSMC alone, suggesting that monocytes may promote hVSMC migration, a condition necessary for ultimately achieving uniform tissue formation in porous scaffolds. [Copyright &y& Elsevier]

Abstract: Tissue culture polystyrene (TCPS) is a ubiquitous substrate used by many researchers in the biomedical and biological sciences. Different parameters involved in the production of TCPS, including the treatment time and the use of reactive gases and chemical agents, can have a significant influence on the ultimate surface properties achieved. The assumption that they will all yield a consistent and controlled product has not proven to be true. To provide a better insight into the bioactivity differences in TCPS supplied by different manufacturers, TCPS from three different companies (Sarstedt, Wisent Corp., and Becton Dickinson (BD)) were analyzed for their surface properties, protein adsorption characteristics, and interactions with human monocytes. Marked differences were observed in terms of surface wettability and surface chemistry. Furthermore, Wisent TCPS adsorbed more than twice the amount of serum proteins compared with BD and Sarstedt TCPS. Sarstedt showed significantly more cell retention (more DNA) compared with both BD and Wisent TCPS brands over a 7day culture period. Cytokine release from monocytes adherent on the three different TCPS also differed significantly, suggesting that the differences in the surface properties were sufficient to differentially mediate monocyte activation. These results have important implications for TCPS research use, in terms of appreciating the interpretation of the data when TCPS is used as a control substrate as well as when it is used where a pre-conditioned state would influence the outcome of the study. [Copyright &y& Elsevier]

Abstract: A degradable, polar, hydrophobic, ionic polyurethane (D-PHI), with physical properties comparable to those of peripheral arterial vascular tissue, was evaluated for monocyte interactions with two different physical forms: two-dimensional films and three-dimensional porous scaffolds. Monocytes, isolated from human whole blood, were seeded onto D-PHI films and scaffolds, and differentiated to monocyte-derived macrophages (MDM) for up to 28days. The effect of surface structure on the MDM phenotype was assessed by assaying: cell attachment (DNA), activation (intracellular protein expression, esterase and acid phosphatase (AP) activity) as well as pro- and anti-inflammatory cytokines (TNF-α and IL-10, respectively). The cells on scaffolds exhibited an initial peak in total protein synthesized per DNA at 3days; however, both substrates generated similar protein levels per DNA at all other time points. While scaffolds generated more esterase and AP per cell than for films, the cells on films expressed significantly more of these two proteins relative to their total protein produced. At day 7 (acute phase of monocyte activation), cells on films were significantly more activated than monocytes on the scaffolds as assessed by cell morphology and tumor necrosis factor-α and interleukin-10 levels. Histological analysis of scaffolds showed that cells were able to migrate throughout the three-dimensional matrix. By inducing a low inflammatory, high wound-healing phenotype monocyte, the negative effects of the foreign body reaction in vivo may be controlled in a manner possible to direct the vascular tissue cells into the appropriate functional phenotypes necessary for successful tissue engineering. [ABSTRACT FROM AUTHOR]

Abstract: High porosity and pore interconnectivity are important features of a successful tissue engineering scaffold. The objective of this work was to optimize the pore interconnectivity and to increase the porosity of an elastomeric degradable/polar/hydrophobic/ionic (D-PHI) polyurethane porous scaffold while maintaining its mechanical integrity in order to allow for the transfer of mechanical stimulus to vascular smooth muscle cells (VSMCs) seeded onto the scaffold. The effect of varying porogen (sodium bicarbonate (salt) and polyethylene glycol (PEG)) composition and concentration on the mechanical properties, degree of swelling and porosity of the scaffolds was investigated. It was found that the use of 10wt.% PEG and 65wt.% salt in scaffold fabrication (D-PHI-75T) resulted in micropore (1–5μm) formation, a high porosity (79±3%) and mechanical properties (elastic modulus=0.16±0.03MPa, elongation-at-yield=31±5% and tensile strength=0.04±0.01MPa) required to withstand the physiologically relevant mechanical strain experienced by VMSCs in vivo. This study also investigated the influence of cyclic mechanical strain (CMS) on select molecular markers of A10 VSMCs when seeded into the optimized D-PHI scaffold. To study the interaction of A10 cells with the optimized D-PHI-75T scaffold in the presence of uniaxial strain (10%, 1Hz), a CMS bioreactor was designed and constructed. Molecular marker studies showed a statistical increase in DNA mass and calponin expression after 3 and 7days of CMS when compared to static samples, indicating that the translation of mechanical loading from the novel polyurethane elastomeric scaffold onto VSMCs will be important to consider with regard to modulating cell phenotype. [ABSTRACT FROM AUTHOR]

Abstract: The current study analyzes the biodegradation of a polycarbonate polyurethane scaffold intended for the growth of a tissue-engineered annulus fibrosus (AF) disc component. Electrospun scaffolds with random and aligned nanofiber configurations were fabricated using a biodegradable polycarbonate urethane with and without an anionic surface modifier (anionic dihydroxyl oligomer), and the mechanical behavior of the scaffolds was examined during a 4week biodegradation study. Both the tensile strength and initial modulus of aligned scaffolds (σ =14±1MPa, E =46±3MPa) were found to be higher than those of random fiber scaffolds (σ =1.9±0.4MPa, E =2.1±0.2MPa) prior to degradation. Following initial wetting of the scaffold, the initial modulus of the aligned samples showed a significant decrease (dry: 46±3MPa; pre-wetted: 9±1MPa, p <0.001). The modulus remained relatively constant during the remainder of the 4week incubation period (aligned at 4weeks: 8.0±0.3MPa). The tensile strength for aligned fiber scaffolds was affected in the same manner. Similar changes were not observed for the initial modulus of the random scaffold configuration. Biodegradation of the scaffold in the presence of cholesterol esterase (a monocyte derived enzyme) yielded a 0.5mgweek–1 weight loss. The soluble and non-soluble degradation products were found to be non-toxic to bovine AF cells grown in vitro. The consistent rate of material degradation along with stable mechanical properties comparable to those of native AF tissue and the absence of cytotoxic effects make this polymer a suitable biomaterial candidate for further investigation into its use for tissue-engineering annulus fibrosus. [ABSTRACT FROM AUTHOR]

Monocyte-derived macrophages (MDM) and multinucleated foreign body giant cells (FBGC) are the primary cell types that remain at the cell–material interface of polyurethane (PU)-based medical devices as a result of chronic inflammatory responses. In vitro studies have demonstrated that MDM possess degradative potential toward PU, which can result in device failure. Because most studies have followed the degradation potential, morphology, and function of these cells only once fully differentiated, the current study investigated the influence of a non-degradable control tissue culture-grade polystyrene (TCPS) surface relative to two degradable model polycarbonate-urethanes (PCNU), of different chemistry, on various parameters of MDM morphology and function during a 14-day differentiation time course. The differentiation of human monocytes isolated from whole blood on PCNU materials resulted in increased cell attachment, decreased multinucleation, and significant decreases in cell spreading when compared with cells differentiated on TCPS. Actin-stained podosome-like cell adhesion structures were increased in PCNU-adherent cells, accompanied by an alteration in β-actin and vinculin protein expression. The expression of the CD68 macrophage marker was reduced when cells were adherent to the PCNU materials and compared with TCPS, suggesting altered cell activation by the degradable relative to non-degradable materials. The degradative potential of these cells was altered by the material surface they were exposed to as measured by esterase activity and protein expression of monocyte-specific esterase. This was also supported by physical material breakdown evident in scanning electron microscopy images that illustrated holes in the PCNU films generated by the presence of differentiating MDM. It was concluded from these studies that PCNU materials significantly alter the function and morphology of differentiating MDM. This must be taken into consideration when studying cell–material interactions because these cells will receive cues from their immediate environment (including the biomaterial) upon differentiation, thereby affecting their resulting phenotype. [ABSTRACT FROM AUTHOR]