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Abstract Dental enamel is subjected to a lifetime of de- and re-mineralization cycles in the oral environment, the cumulative effects of which cause embrittlement with age. However, the understanding of atomic scale mechanisms of dental enamel aging is still at its infancy, particularly regarding where compositional differences occur in the hydroxyapatite nanocrystals and what underlying mechanisms might be responsible. Here, we use atom probe tomography to compare enamel from a young (22 years old) and a senior (56 years old) adult donor tooth. Findings reveal that the concentration of fluorine is elevated in the shells of senior nanocrystals relative to young, with less significant differences between the cores or intergranular phases. It is proposed that the embrittlement of enamel is driven, at least in part, by the infusion of fluorine into the nanocrystals and that the principal mechanism is de- and re-mineralization cycles that preferentially erode and rebuild the nanocrystals shells.

Invasive plant species have substantial negative ecological and economic impacts. Geographic information on the potential and actual distributions of invasive plants is critical for their effective management. For many regions, numerous sources of predictive geographic information exist for invasive plants, often in the form of outputs from species distribution models (SDMs). The creation of a repository of consistently produced SDMs of regional- or national-scale information predicting the potential distribution of invasive plant species could provide information to managers in the prioritisation of invasive species management. Here, we present a novel set of not only habitat suitability models for occurrence for 259 manager requested invasive plant species in the contiguous United States (USA), but also habitat suitability models for abundance (≥ 5% cover) and high abundance (≥ 25% cover). These data provide an update to the Invasive Species Habitat Tool (INHABIT; gis.usgs.gov/inhabit). This tool contains information on the majority of invasive plant species in the contiguous USA with sufficient location data for model building. INHABIT provides a canonical set of predicted geographic distributions for invasive plants in the contiguous USA that can aid in the search for new populations of invasive plant species and help create watch lists for emerging invaders. As this tool contains information on nearly all of the most problematic invasive plants in the contiguous USA, it helps in prioritising management strategies by showing which plants are already present or abundant in a land management area and which may become present or abundant in the future.

IntroductionHeterotopic ossification (HO) occurs following orthopedic trauma, spinal cord injuries, brain trauma and limb amputations. Once symptomatic, HO causes pain, limited mobility and decreased quality of life. Current treatments are limited and have significant complications with high recurrence rates, underscoring the need for improved therapeutic interventions. Osteoclasts (OCs) are physiological bone resorptive cells that secrete enzymes and protons to degrade bone.MethodsIn this study, we describe the use of genetically engineered OCs as a novel cell therapy approach to treat HO. Inducible, engineered myeloid precursors (iRANK cells) treated with a chemical inducer of dimerization (CID) differentiated into TRAP+ multinucleated OCs and resorbed mineralized tissues in vitro.ResultsIn vivo, BMP-2-induced murine HO lesions were significantly regressed following treatment using iRANK cells with concomitant systemic administration of CID. Moreover, many OCs were TRAP+, MMP9+, and GFP+, indicating that they differentiated from delivered iRANK cells.DiscussionIn summary, these data con rm the ability of engineered myeloid precursors to differentiate into OCs and resorb HO in vivo paving the way for OC delivery as a promising approach for HO treatment.

Student-run free clinics serve three primary functions: 1) providing basic medical services for the uninsured and underserved in our communities, 2) supporting the training of medical providers, and 3) advocating for medically underserved communities. Despite the multiple benefits that student-run free clinics can provide, the process of founding a clinic is a significant challenge, even for highly motivated students. Here, we present 12 tips generated by students and residents at the University of Cincinnati College of Medicine based on their combined experience in founding the Bearcat Eye Service (BES), a clinic that provides basic ophthalmic screenings. These tips were developed to provide a framework for those interested in founding a student-run free clinic or expanding ophthalmic care in their communities.

Background Components of diet known as fallback foods are argued to be critical in shaping primate dental anatomy. Such foods of low(er) nutritional quality are often non-preferred, mechanically challenging resources that species resort to during ecological crunch periods. An oft-cited example of the importance of dietary fallbacks in shaping primate anatomy is the grey-cheeked mangabey Lophocebus albigena. This species relies upon hard seeds only when softer, preferred resources are not available, a fact which has been linked to its thick dental enamel. Another mangabey species with thick enamel, the sooty mangabey Cercocebus atys, processes a mechanically challenging food year-round. That the two mangabey species are both thickly-enameled suggests that both fallback and routine consumption of hard foods are associated with the same anatomical feature, complicating interpretations of thick enamel in the fossil record. We anticipated that aspects of enamel other than its thickness might differ between Cercocebus atys and Lophocebus albigena. We hypothesized that to function adequately under a dietary regime of routine hard-object feeding, the molars of Cercocebus atys would be more fracture and wear resistant than those of Lophocebus albigena. Methods Here we investigated critical fracture loads, nanomechanical properties of enamel, and enamel decussation in Cercocebus atys and Lophocebus albigena. Molars of Cercopithecus, a genus not associated with hard-object feeding, were included for comparison. Critical loads were estimated using measurements from 2D µCT slices of upper and lower molars. Nanomechanical properties (by nanoindentation) and decussation of enamel prisms (by SEM-imaging) in trigon basins of one upper second molar per taxon were compared. Results Protocone and protoconid critical fracture loads were significantly greater in Cercocebus atys than Lophocebus albigena and greater in both than in Cercopithecus. Elastic modulus, hardness, and elasticity index in most regions of the crown were greater in Cercocebus atys than in the other two taxa, with the greatest difference in the outer enamel. All taxa had decussated enamel, but that of Cercocebus atys uniquely exhibited a bundle of transversely oriented prisms cervical to the radial enamel. Quantitative comparison of in-plane and out-of-plane prism angles suggests that decussation in trigon basin enamel is more complex in Cercocebus atys than it is in either Lophocebus albigena or Cercopithecus cephus. These findings suggest that Cercocebus atys molars are more fracture and wear resistant than those of Lophocebus albigena and Cercopithecus. Recognition of these differences between Cercocebus atys and Lophocebus albigena molars sharpens our understanding of associations between hard-object feeding and dental anatomy under conditions of routine vs. fallback hard-object feeding and provides a basis for dietary inference in fossil primates, including hominins.

Background: Pain remains a prevalent and burdensome complaint for people living with human immunodeficiency virus and/or aquired immunodeficiency syndrome (LWHA). Positive Living (PL), a multimodal pain intervention, reduced pain in female South Africans LWHA. We investigated the efficacy of the PL programme in South African males living with human immunodeficiency virus and/or acquired immunodeficiency syndrome (MLWA) in a rural community. Aim: To determine the effects of a multimodal pain intervention in MLWHA. Setting: Various primary care clinics in Manguzi, Kwa-Zulu Natal, South Africa. Methodology: Therapeutic relationship (TR) intervention alone or in combination with the PL programme were allocated to HIV-positive men between the ages of 18–40. Pain intensity and interference were the primary outcome measures. Secondary outcome measures included physical function, health-related quality of life, depressive symptoms and self-efficacy. Results: Forty-seven men (mean age 35 ± 3 years) were recruited with baseline mean pain severity of 5.02 (± 3.01) and pain interference of 4.6 (± 3.18). Nineteen men were allocated to the TR intervention alone, 28 were allocated to the TR intervention and PL programme. Attendance at the intervention sessions varied from 10% to 36%. No changes in any outcomes were recorded. Conclusion: Poor attendance at the intervention and follow-up sessions make these results an unreliable reflection of the intervention. Contextual factors including internal migration and issues around employment were identified. These may influence healthcare utilisation for MLWHA living in rural settings. Contribution: Unmet healthcare needs of MLWHA in a rural community have been identified. If we are to ‘leave no one behind’, healthcare interventions should account for context and be ‘rural-proofed’.

Abstract Background The Patient Education Empowerment Programme (PEEP) is an interdisciplinary group intervention for people living with chronic pain. As a result of the COVID-19 pandemic, lockdown and restrictions on in-person group-based health care delivery in South Africa, PEEP was modified to a telehealth electronic format (ePEEP) and offered to patients on a waiting list at two interdisciplinary chronic pain clinics in Cape Town, South Africa. The purpose of this study was to explore the feasibility and acceptability of ePEEP through the perspectives of individuals with chronic pain who participated in ePEEP. Methods A qualitative, exploratory descriptive study was conducted. One month after completion of the 6-week ePEEP programme, individuals who participated, were recruited for the study. Data were collected through semi-structured interviews. Data analysis followed an iterative process of inductive content analysis. Results Six individuals, all women, consented and participated in the study. Three main themes emerged from the data. Theme one focussed on how ePEEP initiated a journey of personal development and discovery within the participants. In theme two, participants reflected on the importance and value of building peer and therapeutic relationships as part of ePEEP. In theme 3, participants shared that the online learning environment had features which influenced learning about pain in different ways. Conclusion ePEEP was found to be acceptable, feasible and valuable for the participants. ePEEP facilitated self-discovery, empowerment, relationship building and transformation in the participants, through experiential learning. Although barriers and facilitators to learning were present, both enhanced the learning experience. The positive reception of this telehealth initiative indicates potential for enhanced access to chronic pain management services in the South African setting.

Abstract Background Distributed training has been cited as an opportunity that offers transformative learning experiences in preparing a future workforce to address local needs. For this reason, rural and longitudinal placements are increasingly being adopted by medical schools across the world. Place, participation and person are considered integral in the process of transformation of medical students into responsive graduates on the distributed platform. This article aims to explore the experiences and perceptions of student learning on a rural training platform in South Africa while considering the interrelation between person, place and participation as a process of transformation to becoming a health care professional. The research forms part of a 5-year longitudinal case study, initiated in 2019 to explore a university-rural hospital collaboration on students, staff and the local health care system. Methods Data was collected using interviews and surveys from 63 purposively selected and consenting participants between January and November of 2019. All qualitative data were inductively analysed using an interpretivist approach to thematic analysis for the purposes of this article. All quantitative data was analysed descriptively using Microsoft Excel. Ethics and permission for this research was granted by the Stellenbosch University Human Research Ethics Committee, the Undergraduate Students Programme Committee and the Northern Cape Department of Health, South Africa. Findings Four themes, namely: authenticity of context; participation in a community of practice and social activities; supervision and reflection; and distance support were extracted from the data. These findings contribute to the theory of transformative learning on the distributed platform by expanding on the interrelationship of person, place and participation, specifically as it relates to participation within various communities and practices. The value of active participation in reflection and supervision, distance academic support and social support systems are explored. Conclusions The three dimensions and interrelationship of person, place and participation in the process of transformative learning on the rural training platform can be further unpacked by exploring the types of participation that have facilitated student learning in this research context. Participation in interprofessional teams; supervision, reflection and distance support appear to be the most crucial elements during this transformative learning process.

Background: In 2018, Stellenbosch University's Ukwanda Centre for Rural Health led a faculty initiative to expand undergraduate health professions training to a new site, 9 hours drive from the health sciences campus in the sparsely populated Northern Cape Province of South Africa in the town of Upington. This is part of a faculty strategy to extend undergraduate health sciences training into an under-resourced part of the country, where there is no medical school. During 2019, the first year of implementation, four final year medical students undertook a longitudinal integrated clerkship at this site, while final year students from other programmes undertook short 5-week rotations, with plans for extending rotations and including more disciplines in 2020. The aim of this study was to understand stakeholder perceptions regarding the development of Upington as a rural clinical training site and how this influenced existing services, workforce sustainability and health professions education.Methods: An iterative thematic analysis of qualitative data collected from 55 participants between January and November 2019 was conducted as part of the case study. A constructivist approach to data collection was utilized to explore participants' perceptions, experiences and understanding of the new training site. Triangulation of data collection and reflexive thematic analysis contributed to the trustworthiness of the data and credibility of the findings.Findings: The perceptions of three key groups of stakeholders are reported: (1) Dr. Harry Surtie Hospital and Academic Programme Managers; (2) Supervising and non-supervising clinical staff and (3) Students from three undergraduate programs of the Faculty. Five themes emerged regarding the development of the site. The themes include the process of development; the influence on the health service; workforce sustainability; a change in perspective and equipping a future workforce.Discussion: This case study provides data to support the value of establishing a rural clinical training platform in a resource constrained environment. The influence of the expansion initiative on the current workforce speaks to the potential for improved capacity and competence in patient management with an impact on encouraging a rural oriented workforce. Using this case study to explore how the establishment of a new rural clinical training site is perceived to influence rural workforce sustainability and pathways, may have relevance to other institutions in similar settings. The degree of sustainability of the clinical training initiative is explored.

Abstract. Pain affects over half of the people living with HIV/AIDS (LWHA), and pharmacological treatment has limited efficacy. Preliminary evidence supports nonpharmacological interventions. We previously piloted a multimodal intervention in amaXhosa women LWHA and chronic pain in South Africa with improvements seen in all outcomes, in both intervention and control groups. A multicentre, single-blind randomised controlled trial with 160 participants recruited was conducted to determine whether the multimodal peer-led intervention reduced pain in different populations of both male and female South Africans LWHA. Participants were followed up at weeks 4, 8, 12, 24, and 48 to evaluate effects on the primary outcome of pain, and on depression, self-efficacy, and health-related quality of life. We were unable to assess the efficacy of the intervention due to a 58% loss to follow-up (LTFU). Secondary analysis of the LTFU found that sociocultural factors were not predictive of LTFU. Depression, however, did associate with LTFU, with greater severity of depressive symptoms predicting LTFU at week 8 (P = 0.01). We were unable to evaluate the effectiveness of the intervention due to the high LTFU and the risk of retention bias. The different sociocultural context in South Africa may warrant a different approach to interventions for pain in HIV compared with resource-rich countries, including a concurrent strategy to address barriers to health care service delivery. We suggest that assessment of pain and depression need to occur simultaneously in those with pain in HIV. We suggest investigation of the effect of social inclusion on pain and depression.

We investigate the magnetic stability and endurance of MgO-based magnetic tunnel junctions (MTJs) with an exchange-biased synthetic antiferromagnetic (SAF) pinned layer. When a uniaxially cycling switching field is applied along the easy axis of the free magnetic layer, the magnetoresistance varies only by 1.7% logarithmically with the number of cycles, while no such change appears in the case of a rotating field. This observation is consistent with the effect of the formation and motion of domain walls in the free layer, which create significant stray fields within the pinned hard layer. Unlike in previous studies, the decay we observed only occurs during the first few starting cycles (

Synthetic hydroxyapatite (HA) is a widely studied bioceramic for bone tissue engineering (BTE) due to its similarity to the mineral component of bone. As bone mineral contains various ionic substitutions that play a crucial role in bone metabolism, the bioactivity of HA can be improved by adding small amounts of physiologically relevant ions into its crystal structure, with silicate-substituted HA (Si-HA) showing particularly promising results. Nevertheless, it remains unclear how distinct material characteristics influence the bioactivity due to the intertwined nature of surface properties. A coculture methodology was optimized and applied for in vitro quantification of the biological response. Initially, HA and Si-HA samples were produced and characterized. To compare the bioactivity of the samples, a method was developed to measure interactions in an increasingly complex environment, first including fibronectin (FN) adsorption and subsequently cell adhesion in mono and coculture using primary human osteoblasts (hOBs) and human dermal microvascular endothelial cells (HDMECs), with and without FN precoating. An experimental set-up was designed to assess to what extent different surface features of the samples contribute to the induced biological response. An 8-nm gold sputter coating was applied to eradicate the electrochemical differences and polishing and abrading was used to reduce the differences in surface topographies. Overall, 1.25 wt% Si-HA exhibited most nanoscale variations in surface potential. In terms of bioactivity, 1.25 wt% Si-HA samples induced the highest osteoblast attachment and vessel formation. Additionally, in vitro vessel formation was established on Si-HA surfaces using a hOB:HDMEC cell ratio of 70:30 and a methodology was established that enabled the assessment of the relative effect of topographical and electrochemical features induced by silicon substitution in the HA lattice on their bioactivity. It was found that the difference in the amount of protein attached to HA and 1.25 wt% Si-HA after 2 h was affected by topographical differences. Conversely, electrochemical differences induced different vessel-like structure formation in coculture with a FN precoating. Without an FN precoating, both topographical and electrochemical differences dictated the differences in angiogenic response. Overall, 1.25 wt% Si-HA surface features appear to induce the most favorable protein adsorption and cell adhesion in mono and coculture with and without FN precoating.

Applied to the epicardium in-vivo, regenerative cardiac patches support the ventricular wall, reduce wall stresses, encourage ventricular wall thickening, and improve ventricular function. Scaffold engraftment, however, remains a challenge. After implantation, scaffolds are subject to the complex, time-varying, biomechanical environment of the myocardium. The mechanical capacity of engineered tissue to biomimetically deform and simultaneously support the damaged native tissue is crucial for its efficacy. To date, however, the biomechanical response of engineered tissue applied directly to live myocardium has not been characterized. In this paper, we utilize optical imaging of a Langendorff ex-vivo cardiac model to characterize the native deformation of the epicardium as well as that of attached engineered scaffolds. We utilize digital image correlation, linear strain, and 2D principal strain analysis to assess the mechanical compliance of acellular ice templated collagen scaffolds. Scaffolds had either aligned or isotropic porous architecture and were adhered directly to the live epicardial surface with either sutures or cyanoacrylate glue. We demonstrate that the biomechanical characteristics of native myocardial deformation on the epicardial surface can be reproduced by an ex-vivo cardiac model. Furthermore, we identified that scaffolds with unidirectionally aligned pores adhered with suture fixation most accurately recapitulated the deformation of the native epicardium. Our study contributes a translational characterization methodology to assess the physio-mechanical performance of engineered cardiac tissue and adds to the growing body of evidence showing that anisotropic scaffold architecture improves the functional biomimetic capacity of engineered cardiac tissue. STATEMENT OF SIGNIFICANCE: Engineered cardiac tissue offers potential for myocardial repair, but engraftment remains a challenge. In-vivo, engineered scaffolds are subject to complex biomechanical stresses and the mechanical capacity of scaffolds to biomimetically deform is critical. To date, the biomechanical response of engineered scaffolds applied to live myocardium has not been characterized. In this paper, we utilize optical imaging of an ex-vivo cardiac model to characterize the deformation of the native epicardium and scaffolds attached directly to the heart. Comparing scaffold architecture and fixation method, we demonstrate that sutured scaffolds with anisotropic pores aligned with the native alignment of the superficial myocardium best recapitulate native deformation. Our study contributes a physio-mechanical characterization methodology for cardiac tissue engineering scaffolds., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Additive manufacturing and electrospinning are widely used to create degradable biomedical components. This work presents important new data showing that the temperature used in accelerated tests has a significant impact on the degradation process in amorphous 3D printed poly-l-lactic acid (PLLA) fibres. Samples (c. 100 μ m diameter) were degraded in a fluid environment at 37 ° C, 50 ° C and 80   ° C over a period of 6 months. Our findings suggest that across all three fluid temperatures, the fibres underwent bulk homogeneous degradation. A three-stage degradation process was identified by measuring changes in fluid pH, PLLA fibre mass, molecular weight and polydispersity index. At 37   ° C, the fibres remained amorphous but, at elevated temperatures, the PLLA crystallised. A short-term hydration study revealed a reduction in glass transition (Tg), allowing the fibres to crystallise, even at temperatures below the dry Tg. The findings suggest that degradation testing of amorphous PLLA fibres at elevated temperatures changes the degradation pathway which, in turn, affects the sample crystallinity and microstructure. The implication is that, although higher temperatures might be suitable for testing bulk material, predictive testing of the degradation of amorphous PLLA fibres (such as those produced via 3D printing or electrospinning) should be conducted at 37   ° C., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Malone, Best and Cameron.)

Collagen is a naturally occurring polymer that can be freeze-dried to create 3D porous scaffold architectures for potential application in tissue engineering. The process comprises the freezing of water in an aqueous slurry followed by sublimation of the ice via a pre-determined temperature-pressure regime and these parameters determine the arrangement, shape and size of the ice crystals. However, ice nucleation is a stochastic process, and this has significant and inherent limitations on the ability to control scaffold structures both within and between the fabrication batches. In this paper, we demonstrate that it is possible to overcome the disadvantages of the stochastic process via the use of low-frequency ultrasound (40 kHz) to trigger nucleation, on-demand, in type I insoluble bovine collagen slurries. The application of ultrasound was found to define the nucleation temperature of collagen slurries, precisely tailoring the pore architecture and providing important new structural and mechanistic insights. The parameter space includes reduction in average pore size and narrowing of pore size distributions while maintaining the percolation diameter. A set of core principles are identified that highlight the huge potential of ultrasound to finely tune the scaffold architecture and revolutionise the reproducibility of the scaffold fabrication protocol.

Regenerative cardiac tissue is a promising field of study with translational potential as a therapeutic option for myocardial repair after injury, however, poor electrical and contractile function has limited translational utility. Emerging research suggests scaffolds that recapitulate the structure of the native myocardium improve physiological function. Engineered cardiac constructs with anisotropic extracellular architecture demonstrate improved tissue contractility, signaling synchronicity, and cellular organization when compared to constructs with reduced architectural order. The complexity of scaffold fabrication, however, limits isolated variation of individual structural and mechanical characteristics. Thus, the isolated impact of scaffold macroarchitecture on tissue function is poorly understood. Here, we produce isotropic and aligned collagen scaffolds seeded with embryonic stem cell derived cardiomyocytes (hESC-CM) while conserving all confounding physio-mechanical features to independently assess the effects of macroarchitecture on tissue function. We quantified spatiotemporal tissue function through calcium signaling and contractile strain. We further examined intercellular organization and intracellular development. Aligned tissue constructs facilitated improved signaling synchronicity and directional contractility as well as dictated uniform cellular alignment. Cells on aligned constructs also displayed phenotypic and genetic markers of increased maturity. Our results isolate the influence of scaffold macrostructure on tissue function and inform the design of optimized cardiac tissue for regenerative and model medical systems., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Sanjay Sinha reports financial support was provided by British Heart Foundation Oxbridge Centre for Regenerative Medicine. Sanjay Sinha reports financial support was provided by British Heart Foundation Senior Fellowship. Sanjay Sinha reports financial support was provided by British Heart Foundation Centre for Cardiovascular Research Excellence. Maria Colzani reports financial support was provided by British Heart Foundation Oxbridge Centre for Regenerative Medicine. Maria Colzani reports financial support was provided by British Heart Foundation. Jamie A. Cyr reports financial support was provided by the Gates Cambridge Fellowship. Ruth E. Cameron reports financial support was provided by the Engineering and Physical Sciences Research Council Established Career Fellowship. Serena M. Best reports financial support was provided by the Engineering and Physical Sciences Research Council Established Career Fellowship. Vera Graup reports financial support was provided by the British Heart Foundation Cambridge Centre of Excellence. Sanjay Sinha reports financial support was provided by the Wellcome Trust Medical Research Council Cambridge Stem Cell Institute., (Copyright © 2023 The Authors. Published by Elsevier B.V. All rights reserved.)

The helical arrangement of cardiac muscle fibres underpins the contractile properties of the heart chamber. Across the heart wall, the helical angle of the aligned fibres changes gradually across the range of 90-180°. It is essential to recreate this structural hierarchy in vitro for developing functional artificial tissue. Ice templating can achieve single-oriented pore alignment via unidirectional ice solidification with a flat base mould design. We hypothesise that the orientation of aligned pores can be controlled simply via base topography, and we propose a scalable base design to recapitulate the transmural fibre orientation. We have utilised finite element simulations for rapid testing of base designs, followed by experimental confirmation of the Bouligand-like orientation. X-ray microtomography of experimental samples showed a gradual shift of 106 ± 10°, with the flexibility to tailor pore size and spatial helical angle distribution for personalised medicine.

This paper investigates drug release from a novel series of mPEG-functionalised PLLA polymers whose individual components (PEG and PLLA) have regulatory FDA approval. Two processing methods were explored to understand their effect on the morphology and drug release profiles of the polymers, with and without mPEG functionalisation. In the first method the polymer and Propranolol.HCl drug powders were mixed together before injection moulding. In the second method, supercritical CO 2 was used to mix the polymer and drug before injection moulding. When non-functionalised PLLA was processed through injection moulding alone, there were no signs of polymer-drug interaction, and the drug was confined to crystals on the surface. This resulted in up to 85 wt% burst release of propranolol.HCl after one day of incubation. By contrast, injection moulding of mPEG-functionalised polymers resulted in the partial dissolution of drug in the polymer matrix and a smaller burst (50 wt% drug) followed by sustained release. This initial burst release was completely eliminated from the profile of mPEG-functionalised polymers processed via supercritical CO 2 . The addition of mPEG facilitated the distribution of the drug into the bulk matrix of the polymer. Paired with supercritical CO 2 processing, the drug release profile showed a slow, sustained release throughout the 4 months of the study., (© 2022. The Author(s).)

Bone tissue engineering (BTE) aims to improve the healing of bone fractures using scaffolds that mimic the native extracellular matrix. For successful bone regeneration, scaffolds should promote simultaneous bone tissue formation and blood vessel growth for nutrient and waste exchange. However, a significant challenge in regenerative medicine remains the development of grafts that can be vascularized successfully. Amongst other things, optimization of physicochemical conditions of scaffolds is key to achieving appropriate angiogenesis in the period immediately following implantation. Calcium phosphates and collagen scaffolds are two of the most widely studied biomaterials for BTE, due to their close resemblance to inorganic and organic components of bone, respectively, and their bioactivity, tunable biodegradability and the ability to produce tailored architectures. While various strategies exist to enhance vascularization of these scaffolds in vivo , further in vitro assessment is crucial to understand the relation between physicochemical properties of a biomaterial and its ability to induce angiogenesis. While mono-culture studies can provide evidence regarding cell-material interaction of a single cell type, a co-culture procedure is crucial for assessing the complex mechanisms involved in angiogenesis. A co-culture more closely resembles the natural tissue both physically and biologically by stimulating natural intercellular interactions and mimicking the organization of the in vivo environment. Nevertheless, a co-culture is a complex system requiring optimization of various parameters including cell types, cell ratio, culture medium and seeding logistics. Gaining fundamental knowledge of the mechanism behind the bioactivity of biomaterials and understanding the contribution of surface and architectural features to the vascularization of scaffolds, and the biological response in general, can provide an invaluable basis for future optimization studies. This review gives an overview of the available literature on scaffolds for BTE, and trends are extracted on the relationship between architectural features, biochemical properties, co-culture parameters and angiogenesis., (© The Author(s) 2023. Published by Oxford University Press.)

Collagen-based biomaterials are used widely as tissue engineering scaffolds because of their excellent bioactivity and their similarity to the natural ECM. The regeneration of healthy bone tissue requires simultaneous support for both osteoblasts and, where angiogenesis is intended, endothelial cells. Hence it is important to tailor carefully the biochemical and structural characteristics of the scaffold to suit the needs of each cell type. This work describes for the first time a systematic study to gain insight into the cell type-specific response of primary human osteoblast (hOBs) and human dermal microvascular endothelial cells (HDMECs) to insoluble collagen-based biomaterials. The behaviour was evaluated on both 2D films and 3D scaffolds, produced using freeze-drying. The collagen was cross-linked at various EDC/NHS concentrations and mono-cultured with hOBs and HDMECs to assess the effect of architectural features and scaffold stabilization on cell behaviour. It was observed that 3D scaffolds cross-linked at 30% of the standard conditions in literature offered an optimal combination of mechanical stiffness and cellular response for both cell types, although endothelial cells were more sensitive to the degree of cross-linking than hOBs. Architectural features have a time-dependent impact on the cell migration profile, with alignment being the most influential parameter overall., (© The Author(s) 2023. Published by Oxford University Press.)

The architectural and physiomechanical properties of regenerative scaffolds have been shown to improve engineered tissue function at both a cellular and tissue level. The fabrication of regenerative three-dimensional scaffolds that precisely replicate the complex hierarchical structure of native tissue, however, remains a challenge. The aim of this work is therefore two-fold: i) demonstrate an innovative multidirectional freeze-casting system to afford precise architectural control of ice-templated collagen scaffolds; and ii) present a predictive simulation as an experimental design tool for bespoke scaffold architecture. We used embedded heat sources within the freeze-casting mold to manipulate the local thermal environment during solidification of ice-templated collagen scaffolds. The resultant scaffolds comprised complex and spatially varied lamellar orientations that correlated with the imposed thermal environment and could be readily controlled by varying the geometry and power of the heat sources. The complex macro-architecture did not interrupt the hierarchical features characteristic of ice-templated scaffolds, but pore orientation had a significant impact on the stiffness of resultant structures under compression. Furthermore, our finite element model (FEM) accurately predicted the thermal environment and illustrated the freezing front topography within the mold during solidification. The lamellar orientation of freeze-cast scaffolds was also predicted using thermal gradient vector direction immediately prior to phase change. In combination our FEM and bespoke freeze-casting system present an exciting opportunity for tailored architectural design of ice-templated regenerative scaffolds that mimic the complex hierarchical environment of the native extracellular matrix. STATEMENT OF SIGNIFICANCE: Biomimetic scaffold structure improves engineered tissue function, but the fabrication of three-dimensional scaffolds that precisely replicate the complex hierarchical structure of native tissue remains a challenge. Here, we leverage the robust relationship between thermal gradients and lamellar orientation of ice-templated collagen scaffolds to develop a multidirectional freeze-casting system with precise control of the thermal environment and consequently the complex lamellar structure of resultant scaffolds. Demonstrating the diversity of our approach, we identify heat source geometry and power as control parameters for complex lamellar orientations. We simultaneously present a finite element model (FEM) that describes the three-dimensional thermal environment during solidification and accurately predicts lamellar structure of resultant scaffolds. The model serves as a design tool for bespoke regenerative scaffolds., (Copyright © 2022 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Recreating the cell niche of virtually all tissues requires composite materials fabricated from multiple extracellular matrix (ECM) macromolecules. Due to their wide tissue distribution, physical attributes and purity, collagen, and more recently, tropoelastin, represent two appealing ECM components for biomaterials development. Here we blend tropoelastin and collagen, harnessing the cell-modulatory properties of each biomolecule. Tropoelastin was stably co-blended into collagen biomaterials and was retained after EDC-crosslinking. We found that human dermal fibroblasts (HDF), rat glial cells (Rugli) and HT1080 fibrosarcoma cells ligate to tropoelastin via EDTA-sensitive and EDTA-insensitive receptors or do not ligate with tropoelastin, respectively. These differing elastin-binding properties allowed us to probe the cellular response to the tropoelastin-collagen composites assigning specific bioactivity to the collagen and tropoelastin component of the composite material. Tropoelastin addition to collagen increased total Rugli cell adhesion, spreading and proliferation. This persisted with EDC-crosslinking of the tropoelastin-collagen composite. Tropoelastin addition did not affect total HDF and HT1080 cell adhesion; however, it increased the contribution of cation-independent adhesion, without affecting the cell morphology or, for HT1080 cells, proliferation. Instead, EDC-crosslinking dictated the HDF and HT1080 cellular response. These data show that a tropoelastin component dominates the response of cells that possess non-integrin based tropoelastin receptors. EDC modification of the collagen component directs cell function when non-integrin tropoelastin receptors are not crucial for cell activity. Using this approach, we have assigned the biological contribution of each component of tropoelastin-collagen composites, allowing informed biomaterial design for directed cell function via more physiologically relevant mechanisms. STATEMENT OF SIGNIFICANCE: Biomaterials fabricated from multiple extracellular matrix (ECM) macromolecules are required to fully recreate the native tissue niche where each ECM macromolecule engages with a specific repertoire of cell-surface receptors. Here we investigate combining tropoelastin with collagen as they interact with cells via different receptors. We identified specific cell lines, which associate with tropoelastin via distinct classes of cell-surface receptor. These showed that tropoelastin, when combined with collagen, altered the cell behaviour in a receptor-usage dependent manner. Integrin-mediated tropoelastin interactions influenced cell proliferation and non-integrin receptors influenced cell spreading and proliferation. These data shed light on the interplay between biomaterial macromolecular composition, cell surface receptors and cell behaviour, advancing bespoke materials design and providing functionality to specific cell populations., Competing Interests: Declaration of Competing Interest Anthony S Weiss is the Scientific Founder of Elastagen Pty Ltd. now sold to Allergan and AbbVie. All other authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2021. Published by Elsevier Ltd.)

Particulate reinforcement of polymeric matrices is a powerful technique for tailoring the mechanical and degradation properties of bioresorbable implant materials. Dispersion of inorganic particles is critical to achieving optimal properties, however established techniques such as twin-screw extrusion or solvent casting can have significant drawbacks including excessive thermal degradation or particle agglomeration. We present a facile method for production of polymer-inorganic composites that reduces the time at elevated temperature and the time available for particle agglomeration. Glass slurry was added to a dissolved PLLA solution, and ethanol was added to precipitate polymer onto the glass particles. Characterisation of parts formed by subsequent micro-injection moulding of composite precipitate revealed a significant reduction in agglomeration, with d 0.9 reduced from 170 to 43 μm. This drastically improved the ductility (ɛ B ) from 7% to 120%, without loss of strength or stiffness. The method is versatile and applicable to a wide range of polymer and filler materials., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

X-ray micro-computed tomography (μ-CT) can be used to provide both qualitative and quantitative information on the structure of three-dimensional (3D) bioactive scaffolds. When performed in a dry state, μ-CT accurately reflects the structure of collagen-based scaffolds, but imaging in a wet state offers challenges with radiolucency. Here we have used phosphotungstic acid (PTA) as a contrast agent to visualise fully hydrated collagen scaffolds in a physiologically relevant environment. A systematic investigation was performed to understand the effects of PTA on the results of μ-CT imaging by varying sample processing variables such as crosslinking density, hydration medium and staining duration. Immersing samples in 0.3% PTA solution overnight completely stained the samples and the treatment provided a successful route for μ-CT analysis of crosslinked samples. However, significant structural artefacts were observed for samples which were either non-crosslinked or had low levels of crosslinking, which had a heterogeneous interior architecture with collapsed pores at the scaffold periphery. This work highlights the importance of optimising the choice of processing and staining conditions to ensure accurate visualisation for hydrated 3D collagen scaffolds in an aqueous medium., Competing Interests: None., (© 2021 The Authors.)

Biocompatible neural guidance conduits are alternatives to less abundant autologous tissue grafts for small nerve gap injuries. To address larger peripheral nerve injuries, it is necessary to design cell selective biomaterials that attract neuronal and/or glial cells to an injury site while preventing the intrusion of fibroblasts that cause inhibitory scarring. Here, we investigate a potential method for obtaining this selective cellular response by analysing the responses of rat Schwann cells and human dermal fibroblasts to isoleucine-lysine-valine-alanine-valine (IKVAV)-capped dendrimer-activated collagen films. A high quantity of nanoscale IKVAV-capped dendrimers incorporated onto pre-crosslinked collagen films promoted rat Schwann cell attachment and proliferation, and inhibited human dermal fibroblast proliferation. In addition, while pre-crosslinked dendrimer-activated films inhibited fibroblast proliferation, non-crosslinked dendrimer-activated films and films that were crosslinked after dendrimer-activation (post-crosslinked films) did not. The different cellular responses to pre-crosslinked and post-crosslinked films highlight the importance of having fully exposed, non-covalently bound biochemical motifs (pre-crosslinked films) directing certain cellular responses. These results also suggest that high concentrations of nanoscale IKVAV motifs can inhibit fibroblast attachment to biological substrates, such as collagen, which inherently attract fibroblasts. Therefore, this work points toward the potential of IKVAV-capped dendrimer-activated collagen biomaterials in limiting neuropathy caused by fibrotic scarring at peripheral nerve injury sites.

Due to its ubiquity and versatility in the human body, collagen is an ideal base material for tissue-engineering constructs. Chemical crosslinking treatments allow precise control of the biochemical and mechanical properties through macromolecular modifications to the structure of collagen. In this work, three key facets regarding the collagen crosslinking process are explored. Firstly, a comparison is drawn between the carbodiimide-succinimide (EDC-NHS) system and two emerging crosslinkers utilising alternate chemistries: genipin and tissue transglutaminase (TG2). By characterising the chemical changes upon treatment, the effect of EDC-NHS, genipin and TG2 crosslinking mechanisms on the chemical structure of collagen, and thus the mechanical properties conferred to the substrate is explored. Secondly, the relative importance of mechanical and biochemical cues on cellular phenomena are investigated, including cell viability, integrin-specific attachment, spreading and proliferation. Here, we observe that for human dermal fibroblasts, long-term, stable proliferation is preconditioned by the availability of suitable binding sites, irrespective of the substrate modulus post-crosslinking. Finally, as seen in the graphical abstract we show that by choosing the appropriate crosslinker chemistries, a materials selection map can be drawn for collagen films, encompassing both a range of tensile modulus and fibroblast proliferation which can be modified independently. Thus, in addition to a range of parameters that can be modified in collagen constructs, we demonstrate a route to obtaining tunable bioactivity and mechanics in collagen constructs is uncovered, that is exclusively driven by the crosslinking process., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Diffusion of nutrients to cells cultured within three-dimensional scaffolds is fundamental for cell survival during development of the tissue construct, when no vasculature is present to aid transport. Significant efforts have been made to characterize the effect of structure on solute diffusivity in nanoporous hydrogels, yet a similar thorough characterization has not been attempted for microporous scaffolds. Here, we make use of freeze-dried collagen scaffolds, possessing pore sizes in the range 150-250 μm and isotropic or aligned morphology, to study the diffusivity of fluorescent dextran molecules. Fluorescence recovery after photobleaching is used to measure the self diffusivity of the solutes within single pores, while Fickian diffusion over scales larger than the pore size is studied by assessing the solute concentration profile within the materials over time. We show that, not only do the morphological parameters of the scaffolds significantly affect the diffusivity of the solutes, but also that the assessment of such diffusivity depends on the length scale of diffusion of the molecules under investigation, with the resulting diffusion coefficients being differently affected by the scaffold structure. The results provided can guide the design of scaffolds with tailored diffusivity and nutrient concentration profiles.

It has become increasingly evident that the mechanical and electrical environment of a cell is crucial in determining its function and the subsequent behavior of multicellular systems. Platforms through which cells can directly interface with mechanical and electrical stimuli are therefore of great interest. Piezoelectric materials are attractive in this context because of their ability to interconvert mechanical and electrical energy, and piezoelectric nanomaterials , in particular, are ideal candidates for tools within mechanobiology, given their ability to both detect and apply small forces on a length scale that is compatible with cellular dimensions. The choice of piezoelectric material is crucial to ensure compatibility with cells under investigation, both in terms of stiffness and biocompatibility. Here, we show that poly-l-lactic acid nanotubes, grown using a melt-press template wetting technique, can provide a "soft" piezoelectric interface onto which human dermal fibroblasts readily attach. Interestingly, by controlling the crystallinity of the nanotubes, the level of attachment can be regulated. In this work, we provide detailed nanoscale characterization of these nanotubes to show how differences in stiffness, surface potential, and piezoelectric activity of these nanotubes result in differences in cellular behavior., Competing Interests: The authors declare no competing financial interest., (Copyright © 2020 American Chemical Society.)

The endometrium is the secretory lining of the uterus that undergoes dynamic changes throughout the menstrual cycle in preparation for implantation and a pregnancy. Recently, endometrial organoids (EO) were established to study the glandular epithelium. We have built upon this advance and developed a multi-cellular model containing both endometrial stromal and epithelial cells. We use porous collagen scaffolds produced with controlled lyophilization to direct cellular organization, integrating organoids with primary isolates of stromal cells. The internal pore structure of the scaffold was optimized for stromal cell culture in a systematic study, finding an optimal average pore size of 101 µm. EO seeded organize to form a luminal-like epithelial layer, on the surface of the scaffold. The cells polarize with their apical surface carrying microvilli and cilia that face the pore cavities and their basal surface attaching to the scaffold with the formation of extracellular matrix proteins. Both cell types are hormone responsive on the scaffold, with hormone stimulation resulting in epithelial differentiation and stromal decidualization., Competing Interests: We declare we have no competing interests., (© 2020 The Authors.)

Micro-computed X-ray tomography (MicroCT) is one of the most powerful techniques available for the three-dimensional characterization of complex multi-phase or porous microarchitectures. The imaging and analysis of porous networks are of particular interest in tissue engineering due to the ability to predict various large-scale cellular phenomena through the micro-scale characterization of the structure. However, optimizing the parameters for MicroCT data capture and analyses requires a careful balance of feature resolution and computational constraints while ensuring that a structurally representative section is imaged and analysed. In this work, artificial datasets were used to evaluate the validity of current analytical methods by considering the effect of noise and pixel size arising from the data capture, and intrinsic structural anisotropy and heterogeneity. A novel 'segmented percolation method' was developed to exclude the effect of anomalous, non-representative features within the datasets, allowing for scale-invariant structural parameters to be obtained consistently and without manual intervention for the first time. Finally, an in-depth assessment of the imaging and analytical procedures are presented by considering percolation events such as micro-particle filtration and cell sieving within the context of tissue engineering. Along with the novel guidelines established for general pixel size selection for MicroCT, we also report our determination of 3 μm as the definitive pixel size for use in analysing connectivity for tissue engineering applications.

This study aimed to document the prevalence and factors associated with complete mental health (CMH) among Aboriginal peoples living in Canada. CMH is comprised of three parts: 1) the absence of major depressive episode, anxiety disorders, bipolar disorder, serious suicidal thoughts, and substance dependence in the past year as measured by the World Health Organization (WHO) versions of the Composite International Diagnostic Interview (WHO-CIDI), 2) happiness and/or satisfaction with life in the past month, and 3) psychological and social well-being. The method involved secondary analysis of Statistics Canada's 2012 Canadian Community Health Survey-Mental Health (CCHS-MH). Responses from Aboriginal peoples living in Canada off-reserve ( n  = 965) were examined to determine what percentage were in CMH and what characteristics are associated with being in CMH. Data analysis involved both bivariate and multivariate analytic techniques to examine factors associated with CMH among Aboriginal peoples. Overall, two-thirds of Aboriginal peoples (67.9%) living in Canada were in CMH. Those with a post-secondary degree, who had a confidant, and those who were free of disabling chronic pain were more likely to be in CMH. Additionally, the odds of CMH were higher among those without a history of suicidal ideation, major depression, alcohol dependence, drug dependence, anxiety disorder, or difficulty sleeping. Findings from this study provide indications of substantial resiliency among Aboriginal peoples in Canada.

To explore factors associated with recovery from suicidal ideation among Aboriginal peoples living off reserve in Canada. Recovery is defined as being free of serious suicidal thoughts for the past year. Data for this study came from the 2012 Aboriginal Peoples Survey, a nationally representative sample of Aboriginal peoples in Canada who are living off-reserve. The sample consisted of those who had seriously considered suicide at some point in their life (n = 2,680). Those who had been suicidal in the past year were compared to those who were no longer suicidal using Pearson chi-square and logistic regression analyses. Several factors were associated with recovery among Aboriginal peoples living off-reserve in Canada who had previously been suicidal. Recovery was higher among women, individuals who were older, and those who were food secure, spoke an Aboriginal language, had a high school degree, had a confidant, and had no previous diagnosis of mood disorders or learning disability. Several sociodemographic factors appear to influence recovery from suicidality among Aboriginal peoples. Intervention approaches to promote recovery from suicidal ideation would benefit from targeted outreach, a strength-based, culturally-specific approach using traditional practices, and encouraging involvement of various community members to foster resilience and formation of relationships.