Integrative tissue, cellular and molecular responsiveness of an innovative ex vivo model of the Staphylococcus aureus-mediated bone infection.

Osteomyelitis is a pathological condition of the bone, frequently associated with the presence of infectious agents - namely Staphylococcus aureus - that induce inflammation and tissue destruction. Recent advances in the understanding of its pathophysiology and the identification of innovative therapeutic approaches were gathered from experimental in vitro and in vivo systems. However, cell culture models offer limited representativeness of the cellular functionality and the cell-cell and cell-matrix interactions, further failing to mimic the three-dimensional tissue organization; and animal models allow for limited mechanistic assessment given the complex nature of systemic and paracrine regulatory systems and are endorsed with ethical constraints. Accordingly, this study aims at the establishment and assessment of a new ex vivo bone infection model, upon the organotypic culture of embryonic chicken femurs colonized with S. aureus, highlighting the model responsiveness at the molecular, cellular, and tissue levels. Upon infection with distinct bacterial inoculums, data reported an initial exponential bacterial growth, followed by diminished metabolic activity. At the tissue level, evidence of S. aureus-mediated tissue destruction was attained and demonstrated through distinct methodologies, conjoined with decreased osteoblastic/osteogenic and increased osteoclastic/osteoclastogenic functionalities-representative of the osteomyelitis clinical course. Overall, the establishment and characterization of an innovative bone tissue infection model that is simple, reproducible, easily manipulated, cost-effective, and simulates many features of human osteomyelitis, further allowing the maintenance of the bone tissue's three-dimensional morphology and cellular arrangement, was achieved. Model responsiveness was further demonstrated, showcasing the capability to improve the research pipeline in bone tissue infection-related research.

Blended intensive programme's implementation in dental education: post-pandemic evolution of learning.

Blended Intensive Programmes (BIP's) represent a valuable tool for gathering knowledge and summarising the latest trends in medicine and dentistry. Blended education has been found, even before the COVID-19 pandemic, to increase the level of education and stimulate effective learning for postgraduate healthcare professionals. Interprofessional education is critical for preparing students to enter the health workforce, where teamwork and collaboration are important competencies. This article outlines the key points of the Blended Intensive Programme's implementation in dental education organised by Wroclaw Medical University in Poland. BIP involved professors from 12 universities or research institutions from Europe and South America and 28 participants from 8 countries. The course was taught remotely and in person. In addition, it included a visit to the university and practical classes with artificial simulation and practice in dentistry. A structured questionnaire enabled measuring the evaluation of students' perception of the COVID-19 education before and after the pandemic. The European Region Action Scheme for the Mobility of University Students (ERASMUS) was fundamental to carrying out the BIP with the participation of several countries, allowing the exchange of knowledge, assessing the impact of the pandemic on dental universities, and strengthening international collaborations and the future project of research, education and clinical assistance. We conclude that hybrid teaching programmes broaden the learning spectrum in dental studies by allowing transnational and interdisciplinary approaches that make students aware of the importance of their work within the framework of the general health approach, as this differs from country to country.

Alendronate induces skeletal alterations in the chicken embryonic development model.

Alendronate, a nitrogen-containing bisphosphonate, has reported long-term clinical success in the management of distinct bone-related conditions, particularly in the modulation of post-menopausal osteoporosis. Nonetheless, whether the inhibitory activity over osteoclastic cells' functionality is widely acknowledged, contradictory evidence arises from the assessment of alendronate activity over osteoblastic populations. This may be of particular relevance in situations in which bone formation exceeds bone resorption, with further emphasis on embryonic development, since alendronate can cross the placental barrier and alendronate-based therapies are being extended into women of reproductive age. Accordingly, the present study aims to assess the effects of alendronate, at distinct concentrations (1.5E-10M to 1.5E-7M) on bone tissue development, within a translational animal model - the embryonic chicken development model. Embryos, at the beginning of osteogenesis (day 7) were exposed to different alendronate concentrations for 4 days. Embryos were following characterized for skeletal development by histomorphometric analysis upon histochemical staining, microtomographic analysis, and gene expression assessment of genes related to osteoclastogenic/osteoclastic and osteoblastogenic/osteogenic differentiation, as well as to the immuno-inflammatory activation. The findings revealed that exposure to alendronate had a dose-dependent impact on skeletal growth and mineralization. This effect was evidenced by diminished bone volume and reduced bone surface parameters, with the 1.5E-7M concentration leading to a remarkable reduction of over 50%. Additionally, a decreased osteoclastogenic/osteoclastic gene expression was verified, associated with a diminished osteoblastogenic/osteogenic program - within the 30-50% range for 1.5E-7 M, supporting the diminished bone formation process. An increased inflammatory activation may contribute, at least in part, to the attained outcomes. Overall present findings suggest a negative influence of alendronate on the embryonic bone development process in a dose-dependent manner, highlighting the potential risk of alendronate use during embryonic development.

Microgap and microleakage of a hybrid connection platform-switched implant system in the absence or presence of a silicone-based sealing agent.

The present study aims to characterize, for the first time, the microgap and bacterial microleakage of a platform-switched implant system with hybrid connection, screwed at distinct torque values (manufacturer recommended torque-25 N cm-and a reduced torque-5 N cm-mimicking the long-term functional use), in the absence or presence of a silicon-based sealing agent. Microgap was determined through scanning electron microscopy and bacterial microleakage was evaluated in vitro, upon Enterococcus faecalis colonization of the system. The sealing efficacy was evaluated in the absence or presence of a commercially available silicon-based sealer. The cytotoxicity of the sealer was further addressed in vitro, with a fibroblastic cell line, in accordance with reference standards. A low microgap of the implant system was verified, regardless of the applied torque load-maximal values ranged around 0.25 and 1.25 µm, for 25 and 5 N cm torques, respectively. No bacterial microleakage was reported at 25 N cm, while at 5 N cm, leakage was verified on 38% of the samples. The application of a silicon-based sealer-with an adequate cytocompatible profile-was effective on preventing the bacterial microleakage on the assayed experimental setting. The assayed platform-switched implant system with hybrid connection presented a low interfacial misfit and an effective sealing capability at manufacturer recommended torque. Despite the increased microleakage at low torque conditions, the application of a cytocompatible silicon-based sealing agent restored the sealing effectiveness of the system. The use of a silicon-based sealing agent can assist on the maintenance of the sealing effectiveness even at low torque conditions.

Vascular biosafety of commercial hydroxyapatite particles: discrepancy between blood compatibility assays and endothelial cell behavior.

BACKGROUND: Vascular homeostasis is ensured by a dynamic interplay involving the endothelium, the platelets and the coagulation system. Thus, the vascular safety of particulate materials must address this integrated system, an approach that has been largely neglected. This work analysed the effects of commercial hydroxyapatite (HA) particles in blood compatibility and in endothelial cell behavior, due to their clinical relevance and scarcity of data on their vascular biosafety. RESULTS: Particles with similar chemical composition and distinct size and morphology were tested, i.e. rod-like, nano dimensions and low aspect ratio (HAp1) and needle-shape with wider size and aspect ratio (HAp2). HAp1 and HAp2, at 1 to 10 mg/mL, did not affect haemolysis, platelet adhesion, aggregation and activation, or the coagulation system (intrinsic and extrinsic pathways), although HAp2 exhibited a slight thrombogenic potential at 10 mg/mL. Notwithstanding, significantly lower levels presented dose-dependent toxicity on endothelial cells' behavior. HAp1 and HAp2 decreased cell viability at levels ≥ 250 and ≥ 50 µg/mL, respectively. At 10 and 50 µg/mL, HAp1 did not interfere with the F-actin cytoskeleton, apoptotic index, cell cycle progression, expression of vWF, VECad and CD31, and the ability to form a network of tubular-like structures. Comparatively, HAp2 caused dose-dependent toxic effects in these parameters in the same concentration range. CONCLUSION: The most relevant observation is the great discrepancy of HA particles' levels that interfere with the routine blood compatibility assays and the endothelial cell behavior. Further, this difference was also found to be dependent on the particles' size, morphology and aspect ratio, emphasizing the need of a complementary biological characterization, taking into consideration the endothelial cells' functionality, to establish the vascular safety of particulate HA.

Reviewing particulate delivery systems loaded with repurposed tetracyclines - From micro to nanoparticles.

Tetracyclines (TCs) are a class of broad-spectrum antibacterial agents recognized for their multifaceted properties, including anti-inflammatory, angiogenic and osteogenic effects. This versatility positions them as suitable candidates for drug repurposing, benefitting from well-characterized safety and pharmacological profiles. In the attempt to explore both their antibacterial and pleiotropic effects locally, innovative therapeutic strategies were set on engineering tetracycline-loaded micro and nanoparticles to tackle a vast number of clinical applications. Moreover, the conjoined drug carrier can function as an active component of the therapeutic approach, reducing off-target effects and accumulation, synergizing to an improvement of the therapeutic efficacy. In this comprehensive review we will critically evaluate recent advances involving the use of tetracyclines loaded onto micro- or nanoparticles, intended for biomedical applications, and discuss emerging approaches and current limitations associated with these drug carriers. Owing to their distinctive physical, chemical, and biological properties, these novel carriers have the potential to become a platform technology in personalized regenerative medicine and other therapeutic applications.

Rosehip Extract-Functionalized Magnesium Hydroxide Nanoparticles and Its Effect on Osteoblastic and Osteoclastic Cells.

Considering the role of magnesium in bone metabolism and the increasing relevance of plant-mediated green-synthesis, this work compares the bone cytocompatibility of magnesium hydroxide nanoparticles (NPs) produced by using pure water, Mg(OH)2, or a rosehip (RH) aqueous extract, Mg(OH)2RH. The NPs were evaluated for dose- and time-dependent effects on human osteoblastic and osteoclastic response, due to the direct involvement of the two cell types in bone metabolism. Mg(OH)2 NPs presented nanoplatelet-like morphology (mean diameter ~90 nm) and a crystalline structure (XRD analysis); the RH-mediated synthesis yielded smaller rounded particles (mean diameter <10 nm) with decreased crystallinity. On the ATR-FTIR spectra, both NPs presented the characteristic Mg-OH peaks; Mg(OH)2RH exhibited additional vibration bands associated with the presence of phytochemicals. On osteoblastic cells, NPs did not affect cell growth and morphology but significantly increased alkaline phosphatase (ALP) activity; on osteoclastic cells, particles had little effect in protein content, tartrate-resistant acid phosphatase (TRAP) activity, percentage of multinucleated cells, and cell area. However, compared with Mg(OH)2, Mg(OH)2RH increased osteoblastic differentiation by inducing ALP activity and promoting the expression of Runx2, SP7, Col1a1, and ALP, and had a negative effect on the expression of the osteoclastic genes NFATC1, CA2, and CTSK. These observations suggest the potential usefulness of Mg(OH)2RH NPs in bone regeneration.

Molecular and Cellular Aspects of Socket Healing in the Absence and Presence of Graft Materials and Autologous Platelet Concentrates: a Focused Review.

OBJECTIVES: The present manuscript aims to critically detail the physiologic process of socket healing, in the absence or presence of grafting materials or platelet concentrates, addressing the associated molecular and cellular events that culminate in the restoration of the lost tissue architecture and functionality. MATERIAL AND METHODS: An electronic search in the National Library of Medicine database MEDLINE through its online site PubMed and Web of Science from inception until May 2019 was conducted to identify articles concerning physiologic process of socket healing, in the absence or presence of grafting materials or platelet concentrates. The search was restricted to English language articles without time restriction. Additionally, a hand search was carried out in oral surgery, periodontology and dental implants related journals. RESULTS: In total, 122 literature sources were obtained and reviewed. The detailed biological events, at the molecular and cellular level, that occur in the alveolus after tooth extraction and socket healing process modulated by grafting materials or autologous platelet concentrates were presented as two entities. CONCLUSIONS: Tooth extraction initiates a convoluted set of orderly biological events in the alveolus, aiming wound closure and socket healing. The healing process comprises a wide range of events, regulated by the interplay of cytokines, chemokines and growth factors that determine cellular recruitment, proliferation and differentiation in the healing milieu, in a space- and time-dependent choreographic interplay. Additionally, the healing process may further be modulated by the implantation of grafting materials or autologous platelet concentrates within the tooth socket, aiming to enhance the regenerative outcome.

The 2nd Baltic Osseointegration Academy and Lithuanian University of Health Sciences Consensus Conference 2019. Summary and Consensus Statements: Group I - Biological Aspects of Tooth Extraction, Socket Healing and Indications for Socket Preservation.

INTRODUCTION: The task of Group I was to review and update the existing data concerning the physiologic process of socket healing, in the absence or presence of grafting materials or platelet concentrates, addressing the associated molecular and cellular events that culminate in the restoration of the lost tissue architecture and functionality. The second task was to review current literature concerning extraction socket classification immediately following tooth extraction and the rationales for socket preservation/augmentation procedures and with reference to it suggest novel clinical decision tree for extraction socket preservation/augmentation in aesthetic and non-aesthetic area. MATERIAL AND METHODS: The main areas indicated by this group were as follows: socket healing process, including haemostasis and coagulation, inflammatory phase, proliferative phase, bone tissue modelling and remodelling; socket healing with graft materials and autologous platelet concentrates; extraction socket classifications; indications and reasons for extraction socket preservation/augmentation. The systematic reviews and/or meta-analyses were registered in PROSPERO, an international prospective register of systematic reviews: http://www.crd.york.ac.uk/PROSPERO/. The literature in the corresponding areas of interest was screened and reported following the PRISMA (Preferred Reporting Item for Systematic Review and Meta-Analysis) Statement: http://www.prisma-statement.org/. Method of preparation of the systematic reviews, based on comprehensive search strategies, was discussed and standardized. The summary of the materials and methods employed by the authors in preparing the systematic reviews and/or meta-analyses is presented in Preface chapter. RESULTS: The results and conclusions of the review process are presented in the respective papers. One theoretical review-analysis and one systematic review were performed. The group's general commentaries, consensus statements, clinical recommendations and implications for research are presented in this article.

Novel cellulose/hydroxyapatite scaffolds for bone tissue regeneration: In vitro and in vivo study.

Cellulose scaffolds containing nano- or micro-hydroxyapatite (nHA or µHA) were prepared by the regeneration of cellulose from its acetylated derivative and the mechanical immobilization of inorganic particles, followed by freeze-drying. Microtomographic (micro-computed tomography) evaluation revealed that both scaffolds presented a highly interconnected porous structure, with a mean pore diameter of 490 ± 94 and 540 ± 132 µm for cellulose/nHA and cellulose/µHA, respectively. In vitro and in vivo characterizations of the developed scaffolds were investigated. Commercially available bone allograft was used as a control material. For the in vitro characterization, osteoblastic cell cultures were used and characterized over time to evaluate cell adhesion, metabolic activity, and functional output (alkaline phosphatase activity and osteoblastic gene expression). The results revealed greater spreading cell distribution alongside an increased number of filopodia, higher MTT values, and significantly increased expression of osteoblastic genes (Runx-2, alkaline phosphatase, and BMP-2) for cellulose/nHA, compared with cellulose/µHA and the control. The in vivo biocompatibility was evaluated in a rabbit calvarial defect model. The investigated scaffolds were implanted in circular rabbit calvaria defects. Four- and 12-week bone biopsies were investigated using micro-computed tomography and histological analysis. Although both cellulose/HA scaffolds outperformed the assayed control, a significantly higher amount of newly formed mineralized tissue was found within the defects loaded with cellulose/nHA. Within the limitations of this study, the developed cellulose/HA scaffolds showed promising results for bone regeneration applications. The biological response to the scaffold seems to be greatly dependent on the HA particles' characteristics, with cellulose scaffolds loaded with nHA eliciting an enhanced bone response.

Osteogenic and Angiogenic Response to Calcium Silicate-based Endodontic Sealers.

INTRODUCTION: Calcium silicate-based endodontic sealers are reported to favor the regeneration of periradicular tissues, a process requiring concerted osteogenic and angiogenic events. This study compared 4 calcium silicate-based sealers for the effects of their extracts on osteogenic and angiogenic cell behavior. METHODS: Extracts from ProRoot MTA (Dentsply Tulsa Dental, Tulsa, OK), MTA Plus (Prevest Denpro Limited, Jammu City, India), MTA Fillapex (Angelus, Londrina, PR, Brazil), and Biodentine (Septodont, Saint-Maur-des-Fosses, France) were prepared from freshly mixed sealers (0.1 g/cm(2)/mL extraction medium) and diluted (1:2-1:20). The sealers were compared for the dose- and time-dependent effects on the proliferation and differentiation of human mesenchymal stem cells (hMSCs) and human umbilical vein endothelial cells (HUVECs). An ex vivo osteogenic assay (regeneration of neonatal mice parietal bone defects) and an in vivo angiogenesis assay (chorioallantoic membrane assay) were performed. RESULTS: Diluted extracts from MTA ProRoot and MTA Plus had evident stimulatory effects on the proliferation of hMSCs, alkaline phosphatase activity, and ex vivo regeneration of bone defects. They also increased HUVEC growth; allowed normal tubularlike network organization; and, in vivo, did not affect angiogenesis. Comparatively, Biodentine also elicited a favorable response on hMSCs and HUVECs, but the overall osteogenic and angiogenic outcome was slightly lower. MTA Fillapex exhibited the highest toxicity in hMSCs and HUVECs and, unlike the other sealers, only allowed a partial regeneration of bone defects. CONCLUSIONS: The sealers caused dose- and time-dependent effects on the osteoblastic and endothelial response, eliciting similar cytocompatibility profiles. Results suggest that the induction of both osteogenic and angiogenic events may contribute to the sealers' regenerative outcome.