New Avenues for Capturing Mineralization Events at Biomaterial Interfaces with Liquid-Transmission Electron Microscopy.

Liquid-transmission electron microscopy (liquid-TEM) provides exciting potential for capturing mineralization events at biomaterial interfaces, though it is largely unexplored. To address this, we established a unique approach to visualize calcium phosphate (CaP)-titanium (Ti) interfacial mineralization events by combining the nanofabrication of Ti lamellae by focused ion beam with in situ liquid-TEM. Multiphasic CaP particles were observed to nucleate, adhere, and form different assemblies onto and adjacent to Ti lamellae. Here, we discuss new approaches for exploring the interaction between biomaterials and liquids at the nanoscale. Driving this technology is crucial for understanding and controlling biomineralization to improve implant osseointegration and direct new pathways for mineralized tissue disease treatment in the future.

Tooth acellular extrinsic fibre cementum incremental lines in humans are formed by parallel branched Sharpey's fibres and not by its mineral phase.

In humans, the growth pattern of the acellular extrinsic fibre cementum (AEFC) has been useful to estimate the age-at-death. However, the structural organization behind such a pattern remains poorly understood. In this study tooth cementum from seven individuals from a Mexican modern skeletal series were analyzed with the aim of unveiling the AEFC collagenous and mineral structure using multimodal imaging approaches. The organization of collagen fibres was first determined using: light microscopy, transmission electron microscopy (TEM), electron tomography, and plasma FIB scanning electron microscopy (PFIB-SEM) tomography. The mineral properties were then investigated using: synchrotron small-angle X-ray scattering (SAXS) for T-parameter (correlation length between mineral particles); synchrotron X-ray diffraction (XRD) for L-parameter (mineral crystalline domain size estimation), alignment parameter (crystals preferred orientation) and lattice parameters a and c; as well as synchrotron X-ray fluorescence for spatial distribution of calcium, phosphorus and zinc. Results show that Sharpey's fibres branched out fibres that cover and uncover other collagen bundles forming aligned arched structures that are joined by these same fibres but in a parallel fashion. The parallel fibres are not set as a continuum on the same plane and when they are superimposed project the AEFC incremental lines due to the collagen birefringence. The orientation of the apatite crystallites is subject to the arrangement of the collagen fibres, and the obtained parameter values along with the elemental distribution maps, revealed this mineral tissue as relatively homogeneous. Therefore, no intrinsic characteristics of the mineral phase could be associated with the alternating AEFC incremental pattern.

Unraveling the Supramolecular Structure and Nanoscale Dislocations of Bacterial Cellulose Ribbons Using Correlative Super-Resolution Light and Electron Microscopy.

Cellulose is a structural linear polysaccharide that is naturally produced by plants and bacteria, making it the most abundant biopolymer on Earth. The hierarchical structure of cellulose from the nano- to microscale is intimately linked to its biosynthesis and the ability to process this sustainable resource for materials applications. Despite this, the morphology of bacterial cellulose microfibrils and their assembly into higher order structures, as well as the structural origins of the alternating crystalline and disordered supramolecular structure of cellulose, have remained elusive. In this work, we employed high-resolution transmission electron and atomic force microscopies to study the morphology of bacterial cellulose ribbons at different levels of its structural hierarchy and provide direct visualization of nanometer-wide microfibrils. The non-persistent twisting of cellulose ribbons was characterized in detail, and we found that twists are associated with nanostructural defects at the bundle and microfibril levels. To investigate the structural origins of the persistent disordered regions that are present along cellulose ribbons, we employed a correlative super-resolution light and electron microscopy workflow and observed that the disordered regions that can be seen in super-resolution fluorescence microscopy largely correlated with the ribbon twisting observed in electron microscopy. Unraveling the hierarchical assembly of bacterial cellulose and the ultrastructural basis of its disordered regions provides insights into its biosynthesis and susceptibility to hydrolysis. These findings are important to understand the cell-directed assembly of cellulose, develop new cellulose-based nanomaterials, and develop more efficient biomass conversion strategies.

Simultaneous Visualization of Wet Cells and Nanostructured Biomaterials in SEM using Ionic Liquids.

This work presents a successful methodology to image mammalian cells adhered to nanostructured titanium by using scanning electron microscopy (SEM) operating in low-vacuum mode following ionic liquid treatment. Human osteoblast-like Saos-2 cells were treated with a room-temperature ionic liquid, 1-ethyl-3-methylimidazolium tetrafluoroborate, and subsequently imaged on titanium by SEM. Titanium substrates were modified to create laser-induced periodic surface structures (LIPSS) for visualization at the submicron scale. By using a combination of fluorescence-based cell metabolism along with light microscopy and SEM image analysis, the shape and location of irradiated cells were confirmed to be unchanged after multiple irradiation sessions; the viability of minimally irradiated cells was also unaltered. The wet imaging conditions combined with a rapid facile protocol using ionic liquid allows this technique to fulfill a niche in examining cellular behavior on biomaterials with submicron surface features. The demonstrated method to track observed cell adhesion to submicron surface features by SEM has great implications for understanding cell migration on nanostructured surfaces as well as the exploration of simpler SEM preparation methods for cellular imaging.

Ti-5Al-5Mo-5V-3Cr bone implants with dual-scale topography: a promising alternative to Ti-6Al-4V.

Modifications to the compositional, topographical and morphological aspects of bone implants can lead to improved osseointegration, thus increasing the success of bone implant procedures. This study investigates the creation of dual-scale topography on Ti-5Al-5Mo-5V-3Cr (Ti5553), an alloy not presently used in the biomedical field, and compares it to Ti-6Al-4V (Ti64), the most used Ti alloy for bone implants. Dual-scale surface topography was obtained by combining selective laser melting (SLM) and electrochemical anodization, which resulted in micro- and nanoscale surface features, respectively. Ti5553 and Ti64 samples were manufactured by SLM and showed comparable surface topography. Subsequent electrochemical anodization succeeded in forming titania nanotubes (TNTs) on both alloys, with larger nanotubes obtained with Ti5553 at all investigated anodization voltages. At an anodization voltage of 40 V, a minimum time of 20 min was necessary to have nanotube formation on the surface of either alloy, while only nanopores were evident for shorter times. Seeded Saos-2 cells showed ideal interactions with surface-modified structures, with filopodia extending to both surface microparticles characteristic of SLM and to the interior of TNTs. Attractiveness of Ti5553 lies in its lower elastic modulus (E = 72 GPa) compared to Ti64, which should mitigate stress-shielding phenomena in vivo. This, combined with the analogous results obtained in terms of dual-scale surface topography and cell-substrate interaction, could indicate Ti5553 as a promising alternative to the widely-employed Ti64 for bone implant device manufacturing.

Micrometer-Sized Magnesium Whitlockite Crystals in Micropetrosis of Bisphosphonate-Exposed Human Alveolar Bone.

Osteocytes are contained within spaces called lacunae and play a central role in bone remodelling. Administered frequently to prevent osteoporotic fractures, antiresorptive agents such as bisphosphonates suppress osteocyte apoptosis and may be localized within osteocyte lacunae. Bisphosphonates also reduce osteoclast viability and thereby hinder the repair of damaged tissue. Osteocyte lacunae contribute to toughening mechanisms. Following osteocyte apoptosis, the lacunar space undergoes mineralization, termed "micropetrosis". Hypermineralized lacunae are believed to increase bone fragility. Using nanoanalytical electron microscopy with complementary spectroscopic and crystallographic experiments, postapoptotic mineralization of osteocyte lacunae in bisphosphonate-exposed human bone was investigated. We report an unprecedented presence of ∼80 nm to ∼3 µm wide, distinctly faceted, magnesium whitlockite [Ca18Mg2(HPO4)2(PO4)12] crystals and consequently altered local nanomechanical properties. These findings have broad implications on the role of therapeutic agents in driving biomineralization and shed new insights into a possible relationship between bisphosphonate exposure, availability of intracellular magnesium, and pathological calcification inside lacunae.

Mesoscale characterization of osseointegration around an additively manufactured genistein-coated implant.

Given the hierarchical nature of bone and bone interfaces, osseointegration, namely the formation of a direct bone-implant contact, is best evaluated using a multiscale approach. However, a trade-off exists between field of view and spatial resolution, making it challenging to image large volumes with high resolution. In this study, we combine established electron microscopy techniques to probe bone-implant interfaces at the microscale and nanoscale with plasma focused ion beam-scanning electron microscopy (PFIB-SEM) tomography to evaluate osseointegration at the mesoscale. This characterization workflow is demonstrated for bone response to an additively manufactured Ti-6Al-4V implant which combines engineered porosity to facilitate bone ingrowth and surface functionalization via genistein, a phytoestrogen, to counteract bone loss in osteoporosis. SEM demonstrated new bone formation at the implant site, including in the internal implant pores. At the nanoscale, scanning transmission electron microscopy and energy-dispersive X-ray spectroscopy confirmed the gradual nature of the bone-implant interface. By leveraging mesoscale analysis with PFIB-SEM tomography that captures large volumes of bone-implant interface with nearly nanoscale resolution, the presence of mineral ellipsoids varying in size and orientation was revealed. In addition, a well-developed lacuno-canalicular network and mineralization fronts directed both towards the implant and away from it were highlighted.

Nanoscale implant anchorage aided by cement line deposition into titanium dioxide nanotubes.

The success of titanium dental implants relies on osseointegration, the load-bearing connection between bone tissue and the device that, in contact osteogenesis, comprises the deposition of bony cement line matrix onto the implant surface. Titanium dioxide nanotubes (NTs) are considered a promising surface for improved osseointegration, yet the mechanisms of cement line integration with such features remains elusive. Herein, we illustrate cement line deposition into NTs on the surface of titanium implants with two underlaying microstructures: a machined surface or a blasted/acid etched surface placed in the tibiae of Wistar rats. After retrieval, scanning electron microscopy of tissue reflected from the implant surface indicated minimal incursion of the cement line matrix into the NTs. To investigate this further, focused ion beam was utilized to prepare cross-sectional samples that could be characterized using scanning transmission electron microscopy. The cement line matrix covered NTs regardless of underlying microstructure, which was further confirmed by elemental analysis. In some instances, cement line infiltration into the NTs was noted, which reveals a mechanism of nanoscale anchorage. This study is the first to demonstrate cement line deposition into titanium NTs, suggesting nano-anchorage as a mechanism for the success of the NT modified surfaces in vivo.

Atom probe tomography for biomaterials and biomineralization.

Biominerals and biomaterials are part of our daily lives, from our skeleton and teeth to coral reefs and carbon-capturing single-cell organisms in the oceans, to engineered ceramics comprising our toothpaste and bone replacements. Many biominerals are hierarchically structured with remarkable material properties that arise from their unique combination of organic and inorganic components. Such structural hierarchy is often formed through a process of biomineralization. However, many fundamental questions remain regarding mineralization events in bones or teeth, and near biomaterials, partly due to the challenges in characterizing three-dimensional (3D) structure and chemical composition simultaneously at the nanometer scale. Atom probe tomography (APT) is a 3D characterization technique that combines both sub-nanometer spatial resolution and compositional sensitivity down to tens of parts per million. While APT is well-established in application to conventional engineering materials, recent years have seen its expansion into biomineralization research. Here, we focus our review on APT applications to biominerals, biomaterials and biointerfaces, providing a high-level summary of findings, as well as a primer on theory and best practices specific to the biomineralization community. We show that APT is a promising characterization tool, where its unique ability to quantify 3D chemical composition is not only complementary to other microscopy techniques but could become an integral part of biomaterial research. With the emerging trends of correlative and cryogenic workflow, notwithstanding the challenges outlined herein, APT has the potential to improve understanding of a broader range of biomaterials, while deriving innovative perspectives on clinical applications and strategies for biomaterial design. STATEMENT OF SIGNIFICANCE: Atom probe tomography (APT) is a three-dimensional characterization technique that can provide quantitative elemental and isotopic analysis with sub-nanometer resolution and compositional sensitivity down to tens of parts per million. These capabilities make it uniquely positioned for the analysis of biomineralized materials, both natural and synthetic. Here, we review the various applications of APT to the field of biomineralization, including applications in biominerals, biomaterials, biointerfaces and other biological materials, such as cells or proteins. A brief but comprehensive summary of the relevant technical concepts, limitations, and future perspectives to enable growth in this field are also included. Although APT is relatively new to the field of biomineralization, it has shown the potential to transform our basic understanding of biomineralization mechanisms and better inform biomaterials design.

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.

PTH 1-34-functionalized bioactive glass improves peri-implant bone repair in orchiectomized rats: Microscale and ultrastructural evaluation.

The objective of this work was to investigate the use of Biogran® functionalized with parathyroid hormone (PTH) 1-34 by sonochemistry for the local delivery of this anabolic agent to the implant site. The effects of Biogran® and topical administration of PTH 1-34 on peri-implant bone regeneration were evaluated from the microscale to ultrastructural levels in healthy (SHAM) and orchiectomized (ORQ). While some animals only received a titanium implant in their tibial metaphyses (CLOT group), in others the peri-implant defect was first filled with Biogran® either without or with PTH 1-34 functionalization (BG and BGPTH groups, respectively) prior to implant installation. Osseointegration was characterized from a biomechanical perspective by measuring the removal torque with the counter-torque technique. Micro-CT was used to evaluate the percentage of bone volume, trabecular thickness, number and separation, and bone-implant contact (BIC). Dynamics of new bone formation were assessed by measuring fluorochrome area, daily mineral apposition rate, and neoformed bone area using confocal laser microscopy. RT-PCR was performed to evaluate ALP and osteocalcin expression. The interface between newly formed bone and Biogran® was examined using scanning electron microscopy (SEM) and scanning transmission electron microscopy (STEM) at the micro-and nanoscale, respectively, while elemental analyses were completed in SEM with energy-dispersive X-ray spectroscopy (EDS). STEM imaging demonstrated the intimate attachment of bone to Biogran® (nanoscale level). Overall, the results suggest that the effectiveness of the topical administration of PTH 1-34 at the implant site seems enhanced in osteoporotic bone, promoting peri-implant bone regeneration to comparable levels in healthy conditions.

Synthesis and release of trace elements from hollow and porous hydroxyapatite spheres.

It is known that organic species regulate fabrication of hierarchical biological forms via solution methods. However, in this study, we observed that the presence of inorganic ions plays an important role in the formation and regulation of biological spherical hydroxyapatite formation. We present a mineralization method to prepare ion-doped hydroxyapatite spheres with a hierarchical structure that is free of organic surfactants and biological additives. Porous and hollow strontium-doped hydroxyapatite spheres were synthesized via controlling the concentration of strontium ions in a calcium and phosphate buffer solution. Similarly, fluoride and silicon-doped hydroxyapatite spheres were synthesized. While spherical particle formation was attainable at low and high temperature for Sr-doped hydroxyapatite, it was only possible at high temperature in the F/Si-doped system. The presence of inorganic ions not only plays an important role in the formation and regulation of biological spherical hydroxyapatite, but also could introduce pharmaceutical effects as a result of trace element release. Such ion release results showed a sustained release with pH responsive behavior, and significantly influenced the hydroxyapatite re-precipitation. These ion-doped hydroxyapatite spheres with hollow and porous structure could have promising applications as bone/tooth materials, drug delivery systems, and chromatography supports.

Free form fabricated features on CoCr implants with and without hydroxyapatite coating in vivo: a comparative study of bone contact and bone growth induction.

The current study evaluates the in vivo response to free form fabricated cobalt chromium (CoCr) implants with and without hydroxyapatite (HA) plasma sprayed coatings. The free form fabrication method allowed for integration of complicated pyramidal surface structures on the cylindrical implant. Implants were press fit into the tibial metaphysis of nine New Zealand white rabbits. Animals were sacrificed and implants were removed and embedded. Histological analysis, histomorphometry and electron microscopy studies were performed. Focused ion beam was used to prepare thin sections for high-resolution transmission electron microscopy examination. The fabricated features allowed for effective bone in-growth and firm fixation after 6 weeks. Transmission electron microscopy investigations revealed intimate bone-implant integration at the nanometre scale for the HA coated samples. In addition, histomorphometry revealed a significantly higher bone contact on HA coated implants compared to native CoCr implants. It is concluded that free form fabrication in combination with HA coating improves the early fixation in bone under experimental conditions.

Visualization of Collagen-Mineral Arrangement Using Atom Probe Tomography.

Bone is a functional material comprised of mainly two phases: an organic collagenous phase and an inorganic mineral phase. Collagen-mineral arrangement has implications for bone function, aging, and disease. However, theories on collagen-mineral arrangement have been confined to studies with low spatial and/or compositional resolution resulting in an extensive debate over the location of mineral with respect to collagen. Herein, a strategy is developed to extract a single mineralized collagen fibril from bone and analyze its composition and structure atom-by-atom with 3D sub-nanometer accuracy and compositional clarity using atom probe tomography (APT). It is shown for the first time a method to probe fibril-level mineralization and collagen-mineral arrangement from an in vivo system with both the spatial and compositional precision required to comment on nanoscale collagen-mineral arrangement. APT of leporine bone shows distinct and helical collagen fibrils with mineralized deposits both encapsulating and incorporated into the collagenous structures. This study demonstrates a novel fibril-level detection method that can be used to probe the composition of bone and contribute new insights to the structure and organization of mineralized materials such as bones and teeth.

Implant resonance and the mechanostat theory: Applications of therapeutic ultrasound for porous metallic scaffolds.

The development of treatment strategies for improving secondary stability at the bone-implant interface is a challenge. Porous implants are one solution for improving long-term implant stability, but the osteoconduction process of implants into the bone can be slow. Strain-driven osteogenesis from the mechanostat theory offers insight into pathways for post-operative treatment but mechanisms to deliver strain to the bone-implant interface need refinement. In this work, the use of therapeutic ultrasound is simulated to induce resonance into a porous implant structure. Local strains through the scaffold are measured by varying systemic variables such as damping ratio, applied vibrational force, primary bone-implant stability, and input frequency. At the natural frequency of the system with applied forces of 0.5 N and a damping ratio of 0.5%, roughly half of the nodes in the simulated environment exceed the microstrain threshold of 1000 µÎµ required for new bone formation. A high degree of sensitivity was noted upon changing input frequency, with minor sensitivities arising from damping ratio and applied vibrational force. These findings suggest that the application of therapeutic resonance to improve osseointegration of the bone-implant interface may be viable for applications including dental implants or segmental bone defects.

From tissue retrieval to electron tomography: nanoscale characterization of the interface between bone and bioactive glass.

The success of biomaterials for bone regeneration relies on many factors, among which osseointegration plays a key role. Biogran (BG) is a bioactive glass commonly employed as a bone graft in dental procedures. Despite its use in clinical practice, the capability of BG to promote osseointegration has never been resolved at the nanoscale. In this paper, we present the workflow for characterizing the interface between newly formed bone and BG in a preclinical rat model. Areas of bone-BG contact were first identified by backscattered electron imaging in a scanning electron microscope. A focused ion beam in situ lift-out protocol was employed to prepare ultrathin samples for transmission electron microscopy analysis. The bone-BG gradual interface, i.e. the biointerphase, was visualized at the nanoscale with unprecedented resolution thanks to scanning transmission electron microscopy. Finally, we present a method to view the bone-BG interface in three dimensions using electron tomography.

Flexible Hierarchical Wraps Repel Drug-Resistant Gram-Negative and Positive Bacteria.

Healthcare acquired infections are a major human health problem, and are becoming increasingly troublesome with the emergence of drug resistant bacteria. Engineered surfaces that reduce the adhesion, proliferation, and spread of bacteria have promise as a mean of preventing infections and reducing the use of antibiotics. To address this need, we created a flexible plastic wrap that combines a hierarchical wrinkled structure with chemical functionalization to reduce bacterial adhesion, biofilm formation, and the transfer of bacteria through an intermediate surface. These hierarchical wraps were effective for reducing biofilm formation of World Health Organization-designated priority pathogens Gram positive methicillin-resistant Staphylococcus aureus (MRSA) and Gram negative Pseudomonas aeruginosa by 87 and 84%, respectively. In addition, these surfaces remain free of bacteria after being touched by a contaminated surface with Gram negative E. coli. We showed that these properties are the result of broad liquid repellency of the engineered surfaces and the presence of reduced anchor points for bacterial adhesion on the hierarchical structure. Such wraps are fabricated using scalable bottom-up techniques and form an effective cover on a variety of complex objects, making them superior to top-down and substrate-specific surface modification methods.

Niobium pentoxide and hydroxyapatite particle loaded electrospun polycaprolactone/gelatin membranes for bone tissue engineering.

Effective methods of accelerating the bone regeneration healing process are in demand for a number of bone-related diseases and trauma. This work developed scaffolds with improved properties for bone tissue engineering by electrospinning composite polycaprolactone-gelatin-hydroxyapatite-niobium pentoxide (PGHANb) membranes. Composite membranes, with average fiber diameters ranging from 123 to 156 nm, were produced by adding hydroxyapatite (HA) and varying concentrations of niobium pentoxide (Nb2O5) particles (0, 3, 7, and 10 wt%) to a polycaprolactone (PCL) and gelatin (GL) matrix prior to electrospinning. The morphology, mechanical, chemical and biological properties of resultant membranes were evaluated. Bioactivity was assessed using simulated body fluid (SBF) and it confirmed that the presence of particles induced the formation of hydroxyapatite crystals on the surface of the membranes. Samples were hydrophilic and cell metabolism results showed that the niobium-containing membranes were non-toxic while improving cell proliferation and differentiation compared to controls. This study demonstrated that electrospun membranes containing HA and Nb2O5 particles have potential to promote cell adhesion and proliferation while exhibiting bioactive properties. PGHANb membranes are promising candidates for bone tissue engineering applications.

Cross-linked cellulose nanocrystal aerogels as viable bone tissue scaffolds.

Chemically cross-linked cellulose nanocrystal (CNC) aerogels possess many properties beneficial for bone tissue scaffolding applications. CNCs were extracted using sulfuric acid or phosphoric acid, to produce CNCs with sulfate and phosphate half-ester surface groups, respectively. Hydrazone cross-linked aerogels fabricated from the two types of CNCs were investigated using scanning electron microscopy, X-ray micro-computed tomography, X-ray photoelectron spectroscopy, nitrogen sorption isotherms, and compression testing. CNC aerogels were evaluatedin vitrowith osteoblast-like Saos-2 cells and showed an increase in cell metabolism up to 7 days while alkaline phosphatase assays revealed that cells maintained their phenotype. All aerogels demonstrated hydroxyapatite growth over 14 days while submerged in simulated body fluid solution with a 0.1 M CaCl2 pre-treatment. Sulfated CNC aerogels slightly outperformed phosphated CNC aerogels in terms of compressive strength and long-term stability in liquid environments, and were implanted into the calvarian bone of adult male Long Evans rats. Compared to controls at 3 and 12 week time points, sulfated CNC aerogels showed increased bone volume fraction of 33% and 50%, respectively, compared to controls, and evidence of osteoconductivity. These results demonstrate that cross-linked CNC aerogels are flexible, porous and effectively facilitate bone growth after they are implanted in bone defects. STATEMENT OF SIGNIFICANCE: Due to the potential complications associated with autografts, there is a need for synthetic bone tissue scaffolds. Here, we report a new naturally-based aerogel material for bone regeneration made solely from chemically cross-linked cellulose nanocrystals (CNC). These highly porous CNC aerogels were shown to promote the proliferation of bone-like cells and support the growth of hydroxyapatite on their surface in vitro. The first in vivo study on these materials was conducted in rats and showed their osteconductive properties and an increase in bone volume up to 50% compared to sham sites. This study demonstrates the potential of using functionalized cellulose nanocrystals as the basis for aerogel scaffolds for bone tissue engineering.

Osseointegration of a 3D Printed Stemmed Titanium Dental Implant: A Pilot Study.

In this pilot study, a 3D printed Grade V titanium dental implant with a novel dual-stemmed design was investigated for its biocompatibility in vivo. Both dual-stemmed (n = 12) and conventional stainless steel conical (n = 4) implants were inserted into the tibial metaphysis of New Zealand white rabbits for 3 and 12 weeks and then retrieved with the surrounding bone, fixed, dehydrated, and embedded into epoxy resin. The implants were analyzed using correlative histology, microcomputed tomography, scanning electron microscopy (SEM), and transmission electron microscopy (TEM). The histological presence of multinucleated osteoclasts and cuboidal osteoblasts revealed active bone remodeling in the stemmed implant starting at 3 weeks and by 12 weeks in the conventional implant. Bone-implant contact values indicated that the stemmed implants supported bone growth along the implant from the coronal crest at both 3- and 12-week time periods and showed bone growth into microporosities of the 3D printed surface after 12 weeks. In some cases, new bone formation was noted in between the stems of the device. Conventional implants showed mechanical interlocking but did have indications of stress cracking and bone debris. This study demonstrates the comparable biocompatibility of these 3D printed stemmed implants in rabbits up to 12 weeks.