Three-dimensional cellular distribution in polymeric scaffolds for bone regeneration: a microCT analysis compared to SEM, CLSM and DNA content.

In orthopaedic surgery the tissues damaged by injury or disease could be replaced using constructs based on biocompatible materials, cells and growth factors. Scaffold design, porosity and early colonization are key components for the implant success. From biological point of view, attention may be also given to the number, type and size of seeded cells, as well as the seeding technique and cell morphological and volumetric alterations. This paper describes the use of the microCT approach (to date used principally for mineralized matrix quantification) to observe construct colonization in terms of cell localization, and make a direct comparison of the microtomographic sections with scanning electron microscopy images and confocal laser scanning microscope analysis. Briefly, polycaprolactone scaffolds were seeded at different cell densities with MG63 osteoblastic-like cells. Two different endpoints, 1 and 2 weeks, were selected for the three-dimensional colonization and proliferation analysis of the cells. By observing all images obtained, in addition to a more extensive distribution of cells on scaffolds surfaces than in the deeper layers, cell volume increased at 2 weeks compared to 1 week after seeding. Combining the cell number quantification by deoxyribonucleic acid analysis and the single cell volume changes by confocal laser scanning microscope, we validated the microCT segmentation method by finding no statistical differences in the evaluation of the cell volume fraction of the scaffold. Furthermore, the morphological results of this study suggest that an effective scaffold colonization requires a precise balance between different factors, such as number, type and size of seeded cells in addition to scaffold porosity.

Inflammatory tissue response in human soft tissue is caused by a higher particle load near carbon fiber-reinforced PEEK compared to titanium plates.

Titanium as the leading implant material in locked plating is challenged by polymers such as carbon fiber-reinforced polyetheretherketone (CFR-PEEK), which became the focus of interest of researchers and manufacturers in recent years. However, data on human tissue response to these new implant materials are rare. Osteosynthesis plates and peri­implant soft tissue samples of 16 healed proximal humerus fractures were examined (n = 8 CFR-PEEK, n = 8 titanium). Soft tissue was analyzed by immunohistochemistry and µCT. The entrapped foreign bodies were further examined for their material composition by FTIR. To gain insight into their origin and formation mechanism, explanted and new plates were evaluated by SEM, EDX, profilometry and HR-CT. In the peri­implant soft tissue of the CFR-PEEK plates, an inflammatory tissue reaction was detected. Tissues contained foreign bodies, which could be identified as tantalum wires, carbon fiber fragments and PEEK particles. Titanium particles were also found in the peri­implant soft tissue of the titanium plates but showed a less intense surrounding tissue inflammation in immunohistochemistry. The surface of explanted CFR-PEEK plates was rougher and showed exposed and broken carbon fibers as well as protruding and deformed tantalum wires, especially in used screw holes, whereas scratches were identified on the titanium plate surfaces. Particles were present in the peri­implant soft tissue neighboring both implant materials and could be clearly assigned to the plate material. Particles from both plate materials caused detectable tissue inflammation, with more inflammatory cells found in soft tissue over CFR-PEEK plates than over titanium plates. STATEMENT OF SIGNIFICANCE: Osteosynthesis plates are ubiquitously used in various medical specialties for the reconstruction of bone fractures and defects and are therefore indispensable for trauma surgeons, ENT specialists and many others. The leading implant material are metals such as titanium, but recently implants made of polymers such as carbon fiber-reinforced polyetheretherketone (CFR-PEEK) have become increasingly popular. However, little is known about human tissue reaction and particle generation related to these new implant types. To clarify this question, 16 osteosynthesis plates (n = 8 titanium and n = 8 CFR-PEEK) and the overlying soft tissue were analyzed regarding particle occurrence and tissue inflammation. Tissue inflammation is clinically relevant for the development of scar tissue, which is discussed to cause movement restrictions and thus contributes significantly to patient outcome.

3D micro-CT analysis of the interface voids associated with Thermafil root fillings used with AH Plus or a flowable MTA sealer.

AIM: To investigate nondestructively the percentage of 3D voids and marginal gaps in a pre-defined interface volume of interest (VOI) within root fillings produced by Thermafil Obturators with either a hydrophobic epoxy-resin-based sealer (AH Plus) or a hydrophilic flowable calcium-silicate sealer [mineral trioxide aggregate (MTA) Flow]. METHODOLOGY: Sixteen single root canals from extracted premolar teeth were prepared with ProTaper rotary instruments, randomly allocated into two groups (n = 8) and filled with size 30 Thermafil Obturators in association with AH Plus or MTA Flow sealers. The filled roots were stored at 37 °C in 5 mL of Hank's balanced salt solution (HBSS) used to represent body fluids and scanned after 7 days and 6 months using a high-resolution micro-CT. From each root, images of 3000 sections were analysed in 3D and binarized using a high-resolution micro-CT (4-µ resolution). The 3D distribution of voids (porosity and marginal gaps) at the gutta-percha-sealer-dentine interface was detected through a threshold grey level and expressed as percentage of the 40-µ-thick pre-defined interface VOI (20 µ of interface dentine and 20 µ of gutta-percha/sealer). A method of analysis based on the root canal segmentation was used, and coronal, middle and apical thirds considered separately. The percentage of 3D void volume was compared statistically using one-way anova (significance for P < 0.05). Environmental Scanning Electron Microscope with Energy Dispersive X-ray (ESEM-EDX) analysis was performed on the surface of both sealers after soaking in HBSS. RESULTS: Micro-CT detected gaps at the dentine-sealer interface in both groups. Void volumes wider than 10.21 µm(3) with 1.35 µm diameter were detected. Cul-de-sac-type voids (blind pores) and through-and-through voids (continuous pores) were discriminated. The apical thirds had a significantly lower 3D void volume (P < 0.05) than the middle and coronal thirds. The 3D void volume reduced significantly (P < 0.05) over time. ESEM-EDX analysis revealed that MTA Flow sealer created a dense apatite layer after 7 days of immersion in HBSS, whilst only sparse calcium phosphate deposits were detected on AH Plus even after 28 days. CONCLUSIONS: Micro-CT proved to be a powerful nondestructive 3D analysis tool for visualizing the porous internal microstructure of dental/endodontic materials at the interface with dentine. The proportion of voids was least in the apical third of root canals. Voids reduced over time in the presence of simulated body fluid.

Pulsed electromagnetic fields and platelet rich plasma alone and combined for the treatment of wear-mediated periprosthetic osteolysis: An in vivo study.

Wear-mediated osteolysis is a common complication occurring around implanted prosthesis, which ultimately leads to bone loss with mechanical instability and the need for surgical revision. At the moment, revision surgery is the only effective treatment. The aim of this study was to assess the efficacy of pulsed electromagnetic fields (PEMFs) and platelet rich plasma (PRP), alone and in association, in a clinically relevant in vivo model of periprosthetic osteolysis. Titanium alloy pins were implanted intramedullary in distal femurs of male inbred rats and, after osseointegration, polyethylene particles were injected intra-articularly to induce osteolysis. Animals were divided in four groups of treatment: PEMFs, PRP, PEMFs + PRP and no treatment. Microtomography was performed during the course of experiments to monitor bone stock and microarchitecture. Histology, histomorphometry, immunohistochemistry and biomechanics were evaluated after treatments. Biophysical and biological stimulations significantly enhanced bone to implant contact, bone volume and bone microhardness and reduced fibrous capsule formation and the number of osteoclasts around implants. Among treatments, PEMFs alone and in association with PRP exerted better results than PRP alone. Present data suggest that biophysical stimulation, with or without the enrichment with platelet derived growth factors, might be a safe, mini-invasive and conservative therapy for counteracting osteolysis and prompting bone formation around implants. STATEMENT OF SIGNIFICANCE: Pulsed electromagnetic fields (PEMFs) and platelet rich plasma (PRP) show anabolic and anti-inflammatory effects and they are already been used in clinical practice, but separately. To date, there are no preclinical in vivo studies evaluating their combined efficacy in periprosthetic osteolysis, in bone tissue microarchitecture and in biomechanics. The aim of the present study was to evaluate the effects of PEMFs and PRP in vivo, when administered individually and in combination in the treatment of periprosthetic wear mediated ostelysis, and in restoring the osteogenetic properties of perimplant bone tissue and its biomechanical competence. The combination of PEMFs and PRP could be employed for counteracting the ostelysis process in a conservative and non surgical manner.

Subchondral bone response to injected adipose-derived stromal cells for treating osteoarthritis using an experimental rabbit model.

Although articular cartilage is the target of osteoarthritis (OA), its deterioration is not always clearly associated with patient symptoms. Because a functional interaction between cartilage and bone is crucial, the pathophysiology of OA and its treatment strategy must focus also on subchondral bone. We investigated whether adipose-derived stromal cells (ASCs) injected into a joint at two different concentrations could prevent subchondral bone damage after the onset of mild OA in a rabbit model. We measured both volumetric and densitometric aspects of bone remodeling. Although OA can stimulate bone remodeling either catabolically or anabolically over time, the accelerated turnover does not allow complete mineralization of new bone and therefore gradually reduces its density. We measured changes in morphometric and densitometric bone parameters using micro-CT analysis and correlated them with the corresponding parameters in cartilage and meniscus. We found that ASCs promoted cartilage repair and helped counteract the accelerated bone turnover that occurs with OA.

Customized hybrid biomimetic hydroxyapatite scaffold for bone tissue regeneration.

Three-dimension (3D) scaffolds for bone tissue regeneration were produced combining three different phases: nanometric hydroxyapatite (HA) was synthesized by precipitation method and the crystals nucleation took place directly within collagen fibrils following a biologically inspired mineralization process; polycaprolactone was employed to give the material a 3D structure. The chemico-physical analysis carried out to test the material's properties and composition revealed a high similarity in composition and morphology with biologically mineralized collagen fibrils and a scaffold degradation pattern suitable for physiological processes. The micro- computerized tomography (micro-CT) showed 53.53% porosity and a 97.86% mean interconnected pores. Computer-aided design and computer-aided manufacturing (CAD-CAM) technology was used for molding the scaffold's volume (design/shape) and for guiding the surgical procedure (cutting guides). The custom made scaffolds were implanted in sheep mandible using prototyped surgical guides and customized bone plates. After three months healing, scanning electron microscopy (SEM) analysis of the explanted scaffold revealed a massive cell seeding of the scaffold, with cell infiltration within the scaffold's interconnected pores. The micro-CT of the explanted construct showed a good match between the scaffold and the adjacent host's bone, to shield the implant primary stability. Histology confirmed cell penetration and widely documented neoangiogenesis within the entire scaffold's volume. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 105B: 723-734, 2017.

[Althesin or CT 1341 as a drug to induce balanced anesthesia with ethrane or fluothane associated with type II NLA in dental, maxillofacial and plastic surgery of the head area]. / L'Althesin o CT 1341 come farmaco di induzione alla narcosi bilanciata con ethrane o fluothane associati alla N.L.A. tipo II in chirurgia stomatologica, maxillo-facciale e plastica dell'estremo cranio-cefalico.

The Authors have tried Althesin as a medicine for the induction and maintainement of the narcosis in the stomatological, maxillo-facial and extra and intra-oral surgery. They have deduced that CT 1341 does not constitute to the anaesthetics Ethrane and Fluothane because of the anaesthetic-level instability (restlessness of the patient and clonic movements of the limbs during the most painful operations in particular). The Authors, supporters of the balanced anaesthesia, suggest using Althesin in the narcosis induction, the maintaining of which is carried on by the N.L.A. type II medicine and by the Ethrane or Fluothane. The latest ones are to be given in very small, not dangerous quantities. The Authors have so obtained a remarkable oxygenation of the patient, a ready, calm and lucid awakening without any collateral and dangerous consequences.