Beating the heart failure odds: long-term survival after myocardial ischemia in juvenile rainbow trout.

Salmonid fish include some of the most valued cultured fish species worldwide. Unlike most other fish, the hearts of salmonids, including Atlantic salmon and rainbow trout, have a well-developed coronary circulation. Consequently, their hearts' reliance on oxygenation through coronary arteries leaves them prone to coronary lesions, believed to precipitate myocardial ischemia. Here, we mimicked such coronary lesions by subjecting groups of juvenile rainbow trout to coronary ligation, assessing histomorphological myocardial changes associated with ischemia and scarring in the context of cardiac arrhythmias using electrocardiography (ECG). Notable ECG changes resembling myocardial ischemia-like ECG in humans, such as atrioventricular blocks and abnormal ventricular depolarization (prolonged and fragmented QRS complex), as well as repolarization (long QT interval) patterns, were observed during the acute phase of myocardial ischemia. A remarkable 100% survival rate was observed among juvenile trout subjected to coronary ligation after 24 wk. Recovery from coronary ligation occurred through adaptive ventricular remodeling, coupled with a fast cardiac revascularization response. These findings carry significant implications for understanding the mechanisms governing cardiac health in salmonid fish, a family particularly susceptible to cardiac diseases. Furthermore, our results provide valuable insights into comparative studies on the evolution, pathophysiology, and ontogeny of vertebrate cardiac repair and restoration.NEW & NOTEWORTHY Juvenile rainbow trout exhibit a remarkable capacity to recover from cardiac injury caused by myocardial ischemia. Recovery from cardiac damage occurs through adaptive ventricular remodeling, coupled with a rapid cardiac revascularization response. These findings carry significant implications for understanding the mechanisms governing cardiac health within salmonid fishes, which are particularly susceptible to cardiac diseases.

Ultrastructure and Nanoporosity of Human Bone Shown with Correlative On-Axis Electron and Spectroscopic Tomographies.

Mineralized collagen fibrils are the building block units of bone at the nanoscale. While it is known that collagen fibrils are mineralized both inside their gap zones (intra-fibrillar mineralization) and on their outer surfaces (extra-fibrillar mineralization), a clear visualization of this architecture in three dimensions (3D), combining structural and compositional information over large volumes, but without compromising the resolution, remains challenging. In this study, we demonstrate the use of on-axis Z-contrast electron tomography (ET) with correlative energy-dispersive X-ray spectroscopy (EDX) tomography to examine rod-shaped samples with diameters up to 700 nm prepared from individual osteonal lamellae in the human femur. Our work mainly focuses on two aspects: (i) low-contrast nanosized circular spaces ("holes") observed in sections of bone oriented perpendicular to the long axis of a long bone, and (ii) extra-fibrillar mineral, especially in terms of morphology and spatial relationship with respect to intra-fibrillar mineral and collagen fibrils. From our analyses, it emerges quite clearly that most "holes" are cross-sectional views of collagen fibrils. While this had been postulated before, our 3D reconstructions and reslicing along meaningful two-dimensional (2D) cross-sections provide a direct visual confirmation. Extra-fibrillar mineral appears to be composed of thin plates that are interconnected and span over several collagen fibrils, confirming that mineralization is cross-fibrillar, at least for the extra-fibrillar phase. EDX tomography shows mineral signatures (Ca and P) within the gap zones, but the signal appears weaker than that associated with the extra-fibrillar mineral, pointing toward the existence of dissimilarities between the two types of mineralization.

Leptin receptor gene deficiency minimally affects osseointegration in rats.

Metabolic syndrome represents a cluster of conditions such as obesity, hyperglycaemia, dyslipidaemia, and hypertension that can lead to type 2 diabetes mellitus and/or cardiovascular disease. Here, we investigated the influence of obesity and hyperglycaemia on osseointegration using a novel, leptin receptor-deficient animal model, the Lund MetS rat. Machined titanium implants were installed in the tibias of animals with normal leptin receptor (LepR+/+) and those harbouring congenic leptin receptor deficiency (LepR-/-) and were left to heal for 28 days. Extensive evaluation of osseointegration was performed using removal torque measurements, X-ray micro-computed tomography, quantitative backscattered electron imaging, Raman spectroscopy, gene expression analysis, qualitative histology, and histomorphometry. Here, we found comparable osseointegration potential at 28 days following implant placement in LepR-/- and LepR+/+ rats. However, the low bone volume within the implant threads, higher bone-to-implant contact, and comparable biomechanical stability of the implants point towards changed bone formation and/or remodelling in LepR-/- rats. These findings are corroborated by differences in the carbonate-to-phosphate ratio of native bone measured using Raman spectroscopy. Observations of hypermineralised cartilage islands and increased mineralisation heterogeneity in native bone confirm the delayed skeletal development of LepR-/- rats. Gene expression analyses reveal comparable patterns between LepR-/- and LepR+/+ animals, suggesting that peri-implant bone has reached equilibrium in healing and/or remodelling between the animal groups.

Targeting Pseudomonas aeruginosa quorum sensing with sodium salicylate modulates immune responses in vitro and in vivo.

Introduction: Chronic infections are a major clinical challenge in hard-to-heal wounds and implanted devices. Pseudomonas aeruginosa is a common causative pathogen that produces numerous virulence factors. Due to the increasing problem of antibiotic resistance, new alternative treatment strategies are needed. Quorum sensing (QS) is a bacterial communication system that regulates virulence and dampens inflammation, promoting bacterial survival. QS inhibition is a potent strategy to reduce bacterial virulence and alleviate the negative impact on host immune response. Aim: This study investigates how secreted factors from P. aeruginosa PAO1, cultured in the presence or absence of the QS inhibitor sodium salicylate (NaSa), influence host immune response. Material and methods: In vitro, THP-1 macrophages and neutrophil-like HL-60 cells were used. In vivo, discs of titanium were implanted in a subcutaneous rat model with local administration of P. aeruginosa culture supernatants. The host immune response to virulence factors contained in culture supernatants (+/-NaSa) was characterized through cell viability, migration, phagocytosis, gene expression, cytokine secretion, and histology. Results: In vitro, P. aeruginosa supernatants from NaSa-containing cultures significantly increased THP-1 phagocytosis and HL-60 cell migration compared with untreated supernatants (-NaSa). Stimulation with NaSa-treated supernatants in vivo resulted in: (i) significantly increased immune cell infiltration and cell attachment to titanium discs; (ii) increased gene expression of IL-8, IL-10, ARG1, and iNOS, and (iii) increased GRO-α protein secretion and decreased IL-1ß, IL-6, and IL-1α secretion, as compared with untreated supernatants. Conclusion: In conclusion, treating P. aeruginosa with NaSa reduces the production of virulence factors and modulates major immune events, such as promoting phagocytosis and cell migration, and decreasing the secretion of several pro-inflammatory cytokines.

Bone structure and composition in a hyperglycemic, obese, and leptin receptor-deficient rat: Microscale characterization of femur and calvarium.

Metabolic abnormalities, such as diabetes mellitus and obesity, can impact bone quantity and/or bone quality. In this work, we characterize bone material properties, in terms of structure and composition, in a novel rat model with congenic leptin receptor (LepR) deficiency, severe obesity, and hyperglycemia (type 2 diabetes-like condition). Femurs and calvaria (parietal region) from 20-week-old male rats are examined to probe bones formed both by endochondral and intramembranous ossification. Compared to the healthy controls, the LepR-deficient animals display significant alterations in femur microarchitecture and in calvarium morphology when analyzed by micro-computed X-ray tomography (micro-CT). In particular, shorter femurs with reduced bone volume, combined with thinner parietal bones and shorter sagittal suture, point towards a delay in the skeletal development of the LepR-deficient rodents. On the other hand, LepR-deficient animals and healthy controls display analogous bone matrix composition, which is assessed in terms of tissue mineral density by micro-CT, degree of mineralization by quantitative backscattered electron imaging, and various metrics extrapolated from Raman hyperspectral images. Some specific microstructural features, i.e., mineralized cartilage islands in the femurs and hyper-mineralized areas in the parietal bones, also show comparable distribution and characteristics in both groups. Overall, the altered bone microarchitecture in the LepR-deficient animals indicates compromised bone quality, despite the normal bone matrix composition. The delayed development is also consistent with observations in humans with congenic Lep/LepR deficiency, making this animal model a suitable candidate for translational research.

Fe and C additions decrease the dissolution rate of silicon nitride coatings and are compatible with microglial viability in 3D collagen hydrogels.

Silicon nitride (SiN) coatings may reduce unwanted release of metal ions from metallic implants. However, as SiN slowly dissolves in aqueous solutions, additives that reduce this dissolution rate would likely increase the lifetime and functionality of implants. Adding iron (Fe) and carbon (C) permits tuning of the SiN coatings' mechanical properties, but their effect on SiN dissolution rates, and their capacity to reduce metal ion release from metallic implant substrates, have yet to be investigated. Such coatings have recently been proposed for use in spinal implants; therefore, it is relevant to assess their impact on the viability of cells expected at the implant site, such as microglia, the resident macrophages of the central nervous system (CNS). To study the effects of Fe and C on the dissolution rate of SiN coatings, compositional gradients of Si, Fe and C in combination with N were generated by physical vapor deposition onto CoCrMo discs. Differences in composition did not affect the surface roughness or the release of Si, Fe or Co ions (the latter from the CoCrMo substrate). Adding Fe and C reduced ion release compared to a SiN reference coating, which was attributed to altered reactivity due to an increase in the fraction of stabilizing Si-C or Fe-C bonds. Extracts from the SiN coatings containing Fe and C were compatible with microglial viability in 2D cultures and 3D collagen hydrogels, to a similar degree as CoCrMo and SiN coated CoCrMo reference extracts. As Fe and C reduced the dissolution rate of SiN-coatings and did not compromise microglial viability, the capacity of these additives to extend the lifetime and functionality of SiN-coated metallic implants warrants further investigation.

Magnesium implant degradation provides immunomodulatory and proangiogenic effects and attenuates peri-implant fibrosis in soft tissues.

Implants made of magnesium (Mg) are increasingly employed in patients to achieve osteosynthesis while degrading in situ. Since Mg implants and Mg2+ have been suggested to possess anti-inflammatory properties, the clinically observed soft tissue inflammation around Mg implants is enigmatic. Here, using a rat soft tissue model and a 1-28 d observation period, we determined the temporo-spatial cell distribution and behavior in relation to sequential changes of pure Mg implant surface properties and Mg2+ release. Compared to nondegradable titanium (Ti) implants, Mg degradation exacerbated initial inflammation. Release of Mg degradation products at the tissue-implant interface, culminating at 3 d, actively initiated chemotaxis and upregulated mRNA and protein immunomodulatory markers, particularly inducible nitric oxide synthase and toll-like receptor-4 up to 6 d, yet without a cytotoxic effect. Increased vascularization was demonstrated morphologically, preceded by high expression of vascular endothelial growth factor. The transition to appropriate tissue repair coincided with implant surface enrichment of Ca and P and reduced peri-implant Mg2+ concentration. Mg implants revealed a thinner fibrous encapsulation compared with Ti. The detailed understanding of the relationship between Mg material properties and the spatial and time-resolved cellular processes provides a basis for the interpretation of clinical observations and future tailoring of Mg implants.

Complex geometry and integrated macro-porosity: Clinical applications of electron beam melting to fabricate bespoke bone-anchored implants.

The last decade has witnessed rapid advancements in manufacturing technologies for biomedical implants. Additive manufacturing (or 3D printing) has broken down major barriers in the way of producing complex 3D geometries. Electron beam melting (EBM) is one such 3D printing process applicable to metals and alloys. EBM offers build rates up to two orders of magnitude greater than comparable laser-based technologies and a high vacuum environment to prevent accumulation of trace elements. These features make EBM particularly advantageous for materials susceptible to spontaneous oxidation and nitrogen pick-up when exposed to air (e.g., titanium and titanium-based alloys). For skeletal reconstruction(s), anatomical mimickry and integrated macro-porous architecture to facilitate bone ingrowth are undoubtedly the key features of EBM manufactured implants. Using finite element modelling of physiological loading conditions, the design of a prosthesis may be further personalised. This review looks at the many unique clinical applications of EBM in skeletal repair and the ground-breaking innovations in prosthetic rehabilitation. From a simple acetabular cup to the fifth toe, from the hand-wrist complex to the shoulder, and from vertebral replacement to cranio-maxillofacial reconstruction, EBM has experienced it all. While sternocostal reconstructions might be rare, the repair of long bones using EBM manufactured implants is becoming exceedingly frequent. Despite the various merits, several challenges remain yet untackled. Nevertheless, with the capability to produce osseointegrating implants of any conceivable shape/size, and permissive of bone ingrowth and functional loading, EBM can pave the way for numerous fascinating and novel applications in skeletal repair, regeneration, and rehabilitation. STATEMENT OF SIGNIFICANCE: Electron beam melting (EBM) offers unparalleled possibilities in producing contaminant-free, complex and intricate geometries from alloys of biomedical interest, including Ti6Al4V and CoCr. We review the diverse range of clinical applications of EBM in skeletal repair, both as mass produced off-the-shelf implants and personalised, patient-specific prostheses. From replacing large volumes of disease-affected bone to complex, multi-material reconstructions, almost every part of the human skeleton has been replaced with an EBM manufactured analog to achieve macroscopic anatomical-mimickry. However, various questions regarding long-term performance of patient-specific implants remain unaddressed. Directions for further development include designing personalised implants and prostheses based on simulated loading conditions and accounting for trabecular bone microstructure with respect to physiological factors such as patient's age and disease status.

Bone without borders - Monetite-based calcium phosphate guides bone formation beyond the skeletal envelope.

Calcium phosphates (CaP) represent an important class of osteoconductive and osteoinductive biomaterials. As proof-of-concept, we show how a multi-component CaP formulation (monetite, beta-tricalcium phosphate, and calcium pyrophosphate) guides osteogenesis beyond the physiological envelope. In a sheep model, hollow dome-shaped constructs were placed directly over the occipital bone. At 12 months, large amounts of bone (∼75%) occupy the hollow space with strong evidence of ongoing remodelling. Features of both compact bone (osteonal/osteon-like arrangements) and spongy bone (trabeculae separated by marrow cavities) reveal insights into function/need-driven microstructural adaptation. Pores within the CaP also contain both woven bone and vascularised lamellar bone. Osteoclasts actively contribute to CaP degradation/removal. Of the constituent phases, only calcium pyrophosphate persists within osseous (cutting cones) and non-osseous (macrophages) sites. From a translational perspective, this multi-component CaP opens up exciting new avenues for osteotomy-free and minimally-invasive repair of large bone defects and augmentation of the dental alveolar ridge.

Multimodal and Multiscale Characterization of the Bone-Bacteria Interface in a Case of Medication-Related Osteonecrosis of the Jaw.

Medication-related osteonecrosis of the jaw (MRONJ) is a known side effect of bisphosphonates (BPs). Although bacterial infection is usually present, the etiology of MRONJ remains unknown. Here we apply a multimodal and multiscale (micro-to-nano) characterization approach to investigate the interface between necrotic bone and bacteria in MRONJ. A non-necrotic bone sample was used as control. Both necrotic and non-necrotic bone samples were collected from the jaw of a female individual affected by MRONJ after using BPs for 23 years. For the first time, resin cast etching was used to expose bacteria at the necrotic site. The bone-bacteria interface was also resolved at the nanoscale by scanning transmission electron microscopy (STEM). Nanosized particulates, likely corresponding to degraded bone mineral, were often noted in close proximity to or enclosed by the bacteria. STEM also revealed that the bone-bacteria interface is composed of a hypermineralized front fading into a highly disordered region, with decreasing content of calcium and phosphorus, as assessed by electron energy loss spectroscopy (EELS). This, combined with the variation in calcium, phosphorus, and carbon across the necrotic bone-bacteria interface evaluated by scanning electron microscopy (SEM)-energy dispersive X-ray spectroscopy (EDX) and the lower mineral-to-matrix ratio measured by micro-Raman spectroscopy in necrotic bone, indicates the absence of a mineralization front in MRONJ. It appears that the bone-bacteria interface originates not only from uncontrolled mineralization but also from the direct action of bacteria degrading the bone matrix. © 2022 The Authors. JBMR Plus published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research.

The impact of medication on osseointegration and implant anchorage in bone determined using removal torque-A review.

Permanently anchored metal implants are frequently used in dental, craniomaxillofacial, and orthopaedic rehabilitation. The success of such therapies is owed to the phenomenon of osseointegration-the direct connection between the living bone and the implant. The extent of biomechanical anchorage (i.e., physical interlocking between the implant and bone) can be assessed with removal torque (RTQ) measurement. Implant anchorage is strongly influenced by underlying bone quality, involving physicochemical and biological properties such as composition and structural organisation of extracellular matrix, extent of micro-damage, and bone turnover. In this review, we evaluated the impact of various pharmacological agents on osseointegration, from animal experiments conducting RTQ measurements. In addition to substances whose antiresorptive and/or anti-catabolic effects on bone are well-documented (e.g., alendronate, zoledronate, ibandronate, raloxifene, human parathyroid hormone, odanacatib, and the sclerostin monoclonal antibody), positive effects on RTQ have been reported for substances that do not primarily target bone (e.g., aminoguanidine, insulin, losartan, simvastatin, bone morphogenetic protein, alpha-tocopherol, and the combination of silk fibroin powder and platelet-rich fibrin). On the contrary, several substances (e.g., prednisolone, cyclosporin A, cisplatin, and enamel matrix derivative) tend to adversely impact RTQ. While morphometric parameters such as bone-implant contact appear to influence the biomechanical anchorage, increased or decreased RTQ is not always accompanied by corresponding fluctuations in bone-implant contact. This further confirms that factors such as bone quality underpin biomechanical anchorage of metal implants. Several fundamental questions on drug metabolism and bioavailability, drug dosage, animal-to-human translation, and the consequences of treatment interruption remain yet unanswered.

Long-term osseointegration of laser-ablated hearing implants in sheep cranial bone.

Osseointegration, the ability for an implant to be anchored in bone tissue with direct bone-implant contact and allowing for continuous adaptive remodelling, is clinically used in different reconstructive fields, such as dentistry, orthopedics and otology. The latter uses a bone conducting sound processor connected to a skin-penetrating abutment that is mounted on a titanium implant placed in the temporal bone, thereby acting as a path for transmission of the vibrations generated by the sound processor. The success of the treatment relies on bone healing and osseointegration, which could be improved by surface modifications. The aim of this study was to evaluate the long-term osseointegration in a sheep skull model and compare a laser-ablated implant surface with a machined implant. Commercially available 4 mm titanium implants, either with a machined (Wide Ponto) or a laser-ablated surface (Ponto BHX, Oticon Medical, Sweden), were used in the current study. The surfaces were evaluated by scanning electron microscopy. The implantation was performed with a full soft tissue flap and the osteotomy was prepared using the MIPS drill kit (Oticon Medical, Sweden) prior to installation of the implants in the frontal bone of eight female sheep. After five months, biopsies including the implant and surrounding bone tissue obtained, processed and analysed using histology, histomorphometry, scanning electron microscopy and Raman spectroscopy. The animals healed well, without signs of adverse events. Histomorphometry showed a large amount of bone tissue around both implant types, with 75% of the threaded area occupied by bone for both implant types. A large amount of bone-implant contact was observed for both implant types, with 67%-71% of the surface covered by bone. Both implant types were surrounded by mature remodelled lamellar bone with high mineral content, corroborating the histological observations. The current results show that the laser-ablated surface induces healing similar to the well-known clinically used machined surface in ovine cranial bone. In conclusion, the present long-term experimental results indicate that a laser-ablated implant performs equally well as a clinically used implant with a machined surface. This, together with previously reported, improved early biomechanical anchorage, suggests future, safe and efficient clinical potential.

Bone mineral organization at the mesoscale: A review of mineral ellipsoids in bone and at bone interfaces.

Much debate still revolves around bone architecture, especially at the nano- and microscale. Bone is a remarkable material where high strength and toughness coexist thanks to an optimized composition of mineral and protein and their hierarchical organization across several distinct length scales. At the nanoscale, mineralized collagen fibrils act as building block units. Despite their key role in biological and mechanical functions, the mechanisms of collagen mineralization and the precise arrangement of the organic and inorganic constituents in the fibrils remains not fully elucidated. Advances in three-dimensional (3D) characterization of mineralized bone tissue by focused ion beam-scanning electron microscopy (FIB-SEM) revealed mineral-rich regions geometrically approximated as prolate ellipsoids, much larger than single collagen fibrils. These structures have yet to become prominently recognized, studied, or adopted into biomechanical models of bone. However, they closely resemble the circular to elliptical features previously identified by scanning transmission electron microscopy (STEM) in two-dimensions (2D). Herein, we review the presence of mineral ellipsoids in bone as observed with electron-based imaging techniques in both 2D and 3D with particular focus on different species, anatomical locations, and in proximity to natural and synthetic biomaterial interfaces. This review reveals that mineral ellipsoids are a ubiquitous structure in all the bones and bone-implant interfaces analyzed. This largely overlooked hierarchical level is expected to bring different perspectives to our understanding of bone mineralization and mechanical properties, in turn shedding light on structure-function relationships in bone. STATEMENT OF SIGNIFICANCE: In bone, the hierarchical organization of organic (mainly collagen type I) and inorganic (calcium-phosphate mineral) components across several length scales contributes to a unique combination of strength and toughness. However, aspects related to the collagen-mineral organization and to mineralization mechanisms remain unclear. Here, we review the presence of mineral prolate ellipsoids across a variety of species, anatomical locations, and interfaces, both natural and with synthetic biomaterials. These mineral ellipsoids represent a largely unstudied feature in the organization of bone at the mesoscale, i.e., at a level connecting nano- and microscale. Thorough understanding of their origin, development, and structure can provide valuable insights into bone architecture and mineralization, assisting the treatment of bone diseases and the design of bio-inspired materials.

The effects of controlled nanotopography, machined topography and their combination on molecular activities, bone formation and biomechanical stability during osseointegration.

The initial cellular and molecular activities at the bone interface of implants with controlled nanoscale topography and microscale roughness have previously been reported. However, the effects of such surface modifications on the development of osseointegration have not yet been determined. This study investigated the molecular events and the histological and biomechanical development of the bone interface in implants with nanoscale topography, microscale roughness or a combination of both. Polished and machined titanium implants with and without controlled nanopatterning (75 nm protrusions) were produced using colloidal lithography and coated with a thin titanium layer to unify the chemistry. The implants were inserted in rat tibiae and subjected to removal torque (RTQ) measurements, molecular analyses and histological analyses after 6, 21 and 28 days. The results showed that nanotopography superimposed on microrough, machined, surfaces promoted an early increase in RTQ and hence produced greater implant stability at 6 and 21 days. Two-way MANOVA revealed that the increased RTQ was influenced by microscale roughness and the combination of nanoscale and microscale topographies. Furthermore, increased bone-implant contact (BIC) was observed with the combined nanopatterned machined surface, although MANOVA results implied that the increased BIC was mainly dependent on microscale roughness. At the molecular level, the nanotopography, per se, and in synergy with microscale roughness, downregulated the expression of the proinflammatory cytokine tumor necrosis factor alpha (TNF-α). In conclusion, controlled nanotopography superimposed on microrough machined implants promoted implant stability during osseointegration. Nanoscale-driven mechanisms may involve attenuation of the inflammatory response at the titanium implant site. STATEMENT OF SIGNIFICANCE: The role of combined implant microscale and nanotopography features for osseointegration is incompletely understood. Using colloidal lithography technique, we created an ordered nanotopography pattern superimposed on screwshaped implants with microscale topography. The midterm and late molecular, bone-implant contact and removal torque responses were analysed in vivo. Nanotopography superimposed on microrough, machined, surfaces promoted the implant stability, influenced by microscale topography and the combination of nanoscale and microscale topographies. Increased bone-implant contact was mainly dependent on microscale roughness whereas the nanotopography, per se, and in synergy with microscale roughness, attenuated the proinflammatory tumor necrosis factor alpha (TNF-α) expression. It is concluded that microscale and nanopatterns provide individual as well as synergistic effects on molecular, morphological and biomechanical implant-tissue processes in vivo.

A stochastic micro to macro mechanical model for the evolution of bone-implant interface stiffness.

Upon placement of an implant into living bone, an interface is formed through which various biochemical, biological, physical, and mechanical interactions take place. This interface evolves over time as the mechanical properties of peri-implant bone increase. Owing to the multifactorial nature of interfacial processes, it is challenging to devise a comprehensive model for predicting the mechanical behavior of the bone-implant interface. We propose a simple spatio-temporally evolving mechanical model - from an elementary unit cell comprising randomly oriented mineralized collagen fibrils having randomly assigned stiffness all the way up to a macroscopic bone-implant interface in a gap healing scenario. Each unit cell has an assigned Young's modulus value between 1.62 GPa and 25.73 GPa corresponding to minimum (i.e., 0) and maximum (i.e., 0.4) limits of mineral volume fraction, respectively, in the overlap region of the mineralized collagen fibril. Gap closure and subsequent stiffening are modeled to reflect the two main directions of peri-implant bone formation, i.e., contact osteogenesis and distance osteogenesis. The linear elastic stochastic finite element model reveals highly nonlinear temporal evolution of bone-implant interface stiffness, strongly dictated by the specific kinetics of contact osteogenesis and distance osteogenesis. The bone-implant interface possesses a small stiffness until gap closure, which subsequently evolves into a much higher stiffness, and this transition is reminiscent of a percolation transition whose threshold corresponds to gap closure. The model presented here, albeit preliminary, can be incorporated into future calculations of the bone-implant system where the interface is well-defined mechanically. STATEMENT OF SIGNIFICANCE: A simple, physically informed model for the mechanical characteristics of the bone-implant interface is still missing. Here, we start by extending the reported mechanical characteristics of a one cubic micrometre unit cell to a 250 µm long interface made of 1 µm thick layers. The stiffness of each cell (based on mineral content) is assigned randomly to mimic bone micro-heterogeneity. The numerical study of this interface representative structure allows for the simultaneous determination of the spatio-temporal evolution of the mechanical response at local (discrete element) and global (overall model) scales. The proposed model is the first of this kind that can easily be incorporated into realistic future models of bone-implant interaction with emphasis on implant stability and different loading conditions.

Multimodal Analysis of the Tissue Response to a Bone-Anchored Hearing Implant: Presentation of a Two-Year Case Report of a Patient With Recurrent Pain, Inflammation, and Infection, Including a Systematic Literature Review.

Osseointegration is a well-established concept used in applications including the percutaneous Bone-Anchored Hearing System (BAHS) and auricular rehabilitation. To date, few retrieved implants have been described. A systematic review including cases where percutaneous bone-anchored implants inserted in the temporal bone were retrieved and analyzed was performed. We also present the case of a patient who received a BAHS for mixed hearing loss. After the initial surgery, several episodes of soft tissue inflammation accompanied by pain were observed, leading to elective abutment removal 14 months post-surgery. Two years post-implantation, the implant was removed due to pain and subjected to a multiscale and multimodal analysis: microbial DNA using molecular fingerprinting, gene expression using quantitative real-time polymerase chain reaction (qPCR), X-ray microcomputed tomography (micro-CT), histology, histomorphometry, backscattered scanning electron microscopy (BSE-SEM), Raman spectroscopy, and fluorescence in situ hybridization (FISH). Evidence of osseointegration was provided via micro-CT, histology, BSE-SEM, and Raman spectroscopy. Polymicrobial colonization in the periabutment area and on the implant, including that with Staphylococcus aureus and Staphylococcus epidermidis, was determined using a molecular analysis via a 16S-23S rDNA interspace [IS]-region-based profiling method (IS-Pro). The histology suggested bacterial colonization in the skin and in the peri-implant bone. FISH confirmed the localization of S. aureus and coagulase-negative staphylococci in the skin. Ten articles (54 implants, 47 patients) met the inclusion criteria for the literature search. The analyzed samples were either BAHS (35 implants) or bone-anchored aural epitheses (19 implants) in situ between 2 weeks and 8 years. The main reasons for elective removal were nonuse/changes in treatment, pain, or skin reactions. Most samples were evaluated using histology, demonstrating osseointegration, but with the absence of bone under the implants' proximal flange. Taken together, the literature and this case report show clear evidence of osseointegration, despite prominent complications. Nevertheless, despite implant osseointegration, chronic pain related to the BAHS may be associated with a chronic bacterial infection and raised inflammatory response in the absence of macroscopic signs of infection. It is suggested that a multimodal analysis of peri-implant health provides possibilities for device improvements and to guide diagnostic and therapeutic strategies to alleviate the impact of complications.

A Body Weight Sensor Regulates Prepubertal Growth via the Somatotropic Axis in Male Rats.

In healthy conditions, prepubertal growth follows an individual specific growth channel. Growth hormone (GH) is undoubtedly the major regulator of growth. However, the homeostatic regulation to maintain the individual specific growth channel during growth is unclear. We recently hypothesized a body weight sensing homeostatic regulation of body weight during adulthood, the gravitostat. We now investigated if sensing of body weight also contributes to the strict homeostatic regulation to maintain the individual specific growth channel during prepubertal growth. To evaluate the effect of increased artificial loading on prepubertal growth, we implanted heavy (20% of body weight) or light (2% of the body weight) capsules into the abdomen of 26-day-old male rats. The body growth, as determined by change in biological body weight and growth of the long bones and the axial skeleton, was reduced in rats bearing a heavy load compared with light load. Removal of the increased load resulted in a catch-up growth and a normalization of body weight. Loading decreased hypothalamic growth hormone releasing hormone mRNA, liver insulin-like growth factor (IGF)-1 mRNA, and serum IGF-1, suggesting that the reduced body growth was caused by a negative feedback regulation on the somatotropic axis and this notion was supported by the fact that increased loading did not reduce body growth in GH-treated rats. Based on these data, we propose the gravitostat hypothesis for the regulation of prepubertal growth. This states that there is a homeostatic regulation to maintain the individual specific growth channel via body weight sensing, regulating the somatotropic axis and explaining catch-up growth.

In situ bone regeneration of large cranial defects using synthetic ceramic implants with a tailored composition and design.

The repair of large cranial defects with bone is a major clinical challenge that necessitates novel materials and engineering solutions. Three-dimensionally (3D) printed bioceramic (BioCer) implants consisting of additively manufactured titanium frames enveloped with CaP BioCer or titanium control implants with similar designs were implanted in the ovine skull and at s.c. sites and retrieved after 12 and 3 mo, respectively. Samples were collected for morphological, ultrastructural, and compositional analyses using histology, electron microscopy, and Raman spectroscopy. Here, we show that BioCer implants provide osteoinductive and microarchitectural cues that promote in situ bone regeneration at locations distant from existing host bone, whereas bone regeneration with inert titanium implants was confined to ingrowth from the defect boundaries. The BioCer implant promoted bone regeneration at nonosseous sites, and bone bonding to the implant was demonstrated at the ultrastructural level. BioCer transformed to carbonated apatite in vivo, and the regenerated bone displayed a molecular composition indistinguishable from that of native bone. Proof-of-principle that this approach may represent a shift from mere reconstruction to in situ regeneration was provided by a retrieved human specimen, showing that the BioCer was transformed into well-vascularized osteonal bone, with a morphology, ultrastructure, and composition similar to those of native human skull bone.

The effect of two types of resorbable augmentation materials - a cement and an adhesive - on the screw pullout pullout resistance in human trabecular bone.

Augmentation materials, such as ceramic and polymeric bone cements, have been frequently used to improve the physical engagement of screws inserted into bone. While ceramic, degradable cements may ultimately improve fixation stability, reports regarding their effect on early fixation stability have been inconsistent. On the other hand, a newly developed degradable ceramic adhesive that can bond with tissues surrounding the screw, may improve the pullout performance, ensure early stability, and subsequent bony integration. The aim of this study was to investigate failure mechanisms of screw/trabecular bone constructs by comparing non-augmented screws with screws augmented with a calcium phosphate cement or an adhesive, i.e. a phosphoserine-modified calcium phosphate. Pullout tests were performed on screws inserted into trabecular cylinders extracted from human femoral bone. Continuous and stepwise pullout loading was applied with and without real-time imaging in a synchrotron radiation micro-computed tomograph, respectively. Statistical analysis that took the bone morphology into account confirmed that augmentation with the adhesive supported significantly higher pullout loads compared to cement-augmented, or non-augmented screws. However, the adhesive also allowed for a higher injection volume compared to the cement. In-situ imaging showed cracks in the vicinity of the screw threads in all groups, and detachment of the augmentation materials from the trabecular bone in the augmented specimens. Additional cracks at the periphery of the augmentation and the bone-material interfaces were only observed in the adhesive-augmented specimen, indicating a contribution of surface bonding to the pullout resistance. An adhesive that has potential for bonding with tissues, displayed superior pullout resistance, compared to a brushite cement, and may be a promising material for cementation or augmentation of implants.

Transformation of bone mineral morphology: From discrete marquise-shaped motifs to a continuous interwoven mesh.

Continual bone apposition at the cranial sutures provides the unique opportunity to understand how bone is built. Bone harvested from 16-week-old Sprague Dawley rat calvaria was either (i) deproteinised to isolate the inorganic phase (i.e., bone mineral) for secondary electron scanning electron microscopy or (ii) resin embedded for X-ray micro-computed tomography, backscattered electron scanning electron microscopy, and micro-Raman spectroscopy. Interdigitated finger-like projections form the interface between frontal and parietal bones. Viewed from the surface, bone mineral at the mineralisation front is comprised of nanoscale mineral platelets arranged into discrete, ~0.6-3.5 µm high and ~0.2-1.5 µm wide, marquise-shaped motifs that gradually evolve into a continuous interwoven mesh of mineralised bundles. Marquise-shaped motifs also contribute to the burial of osteoblastic-osteocytes by contributing to the roof over the lacunae. In cross-section, apices of the finger-like projections resemble islands of mineralised tissue, where new bone apposition at the surface is evident as low mineral density areas, while the marquise-shaped motifs appear as near-equiaxed assemblies of mineral platelets. Carbonated apatite content is higher towards the internal surface of the cranial vault. Up to 4 µm from the bone surface, strong Amide III, Pro, Hyp, and Phe signals, distinct PO4 3- bands, but negligible CO3 2- signal indicate recent bone formation and/or delayed maturation of the mineral. We show, for the first time, that the extracellular matrix of bone is assembled into micrometre-sized units, revealing a superstructure above the mineralised collagen fibril level, which has significant implications for function and mechanical competence of bone.