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Leishmania presents a complex life cycle that involves both invertebrate and vertebrate hosts. By regulating gene expression, protein synthesis, and metabolism, the parasite can adapt to various environmental conditions. This regulation occurs mainly at the post-transcriptional level and may involve epitranscriptomic modifications of RNAs. Recent studies have shown that mRNAs in humans undergo a modification known as N4-acetylcytidine (ac4C) catalyzed by the enzyme N-acetyltransferase (NAT10), impacting mRNAs stability and translation. Here, we characterized the NAT10 homologue of L. mexicana, finding that the enzyme exhibits all the conserved acetyltransferase domains although failed to functionally complement the Kre33 mutant in Saccharomyces cerevisiae. We also discovered that LmexNAT10 is nuclear, and seems essential, as evidenced by unsuccessful attempts to obtain null mutant parasites. Phenotypic characterization of single-knockout parasites revealed that LmexNAT10 affects the multiplication of procyclic forms and the promastigote-amastigote differentiation. Additionally, in vivo infection studies using the invertebrate vector Lutzomyia longipalpis showed a delay in the parasite differentiation into metacyclics. Finally, we observed changes in the cell cycle progression and protein synthesis in the mutant parasites. Together, these results suggest that LmexNAT10 might be important for parasite differentiation, potentially by regulating ac4C levels., (© 2025 John Wiley & Sons Ltd.)

The development of effective biomaterials for tissue regeneration has led to the exploration of blood derivatives such as leucocyte- and platelet-rich fibrin (L-PRF). A novel variant, Albumin-Enriched Platelet-Rich Fibrin (Alb-PRF), has been introduced to improve structural stability and bioactivity, making it a promising candidate for bone regeneration. This study aimed to evaluate Alb-PRF's capacity for cytokine and growth factor release, along with its effects on the proliferation, differentiation, and mineralization of human osteoblasts in vitro. Alb-PRF membranes were analyzed using histological, scanning electron microscopy, and fluorescence microscopy techniques. Cytokine and growth factor release was quantified over seven days, and osteoinductive potential was evaluated with MG-63 osteoblast-like cells. Structural analysis showed Alb-PRF as a biphasic, highly cellularized material that releases lower levels of inflammatory cytokines and higher concentrations of platelet-derived growth factor (PDGF) and vascular endothelial growth factor (VEGF) compared to L-PRF. Alb-PRF exhibited higher early alkaline phosphatase activity and in vitro mineralization ( p < 0.05) and significantly increased the OPG/RANKL mRNA ratio ( p < 0.05). These results indicate that Alb-PRF has promising potential as a scaffold for bone repair, warranting further in vivo and clinical assessments to confirm its suitability for clinical applications.

The increasing demand for effective bone regeneration materials drives the exploration of biomaterials with enhanced bioactivity and biocompatibility, such as zinc-substituted compounds. This study investigates the in vitro cellular interactions with nanostructured spheres composed of alginate/carbonated hydroxyapatite (CHA), compared to zinc-substituted CHA (ZnCHA). This work aimed to compare the physicochemical properties and biological effects of ZnCHA and CHA on osteoblasts. ZnCHA was synthesized using a wet chemical method, followed by characterization through X-ray diffraction, Fourier transform infrared spectroscopy, total organic carbon analysis, Wavelength-dispersive X-ray spectroscopy, and BET surface area analysis to assess ion release and structural changes. Biological evaluation was conducted using cell viability, proliferation, and biomineralization assays on osteoblasts. Results showed successful incorporation of zinc and carbonate, leading to reduced crystallinity and increased surface area. Cell viability and proliferation assays indicated ZnCHA's cytocompatibility and enhanced osteoblastic activity, with increased mineralization nodules compared to CHA samples. The study concludes that ZnCHA composites are promising candidates for bone tissue engineering, demonstrating improved cytocompatibility and potential for further preclinical evaluations.

Zinc is known for its role in enhancing bone metabolism, cell proliferation, and tissue regeneration. Several studies proposed the incorporation of zinc into hydroxyapatite (HA) to produce biomaterials (ZnHA) that stimulate and accelerate bone healing. This systematic review aimed to understand the physicochemical characteristics of zinc-doped HA-based biomaterials and the evidence of their biological effects on osteoblastic cells. A comprehensive literature search was conducted from 2022 to 2024, covering all years of publications, in three databases (Web of Science, PUBMED, Scopus), retrieving 609 entries, with 36 articles included in the analysis according to the selection criteria. The selected studies provided data on the material's physicochemical properties, the methods of zinc incorporation, and the biological effects of ZnHA on bone cells. The production of ZnHA typically involves the wet chemical synthesis of HA and ZnHA precursors, followed by deposition on substrates using processes such as liquid precursor plasma spraying (LPPS). Characterization techniques confirmed the successful incorporation of zinc into the HA lattice. The findings indicated that zinc incorporation into HA at low concentrations is non-cytotoxic and beneficial for bone cells. ZnHA was found to stimulate cell proliferation, adhesion, and the production of osteogenic factors, thereby promoting in vitro mineralization. However, the optimal zinc concentration for the desired effects varied across studies, making it challenging to establish a standardized concentration. ZnHA materials are biocompatible and enhance osteoblast proliferation and differentiation. However, the mechanisms of zinc release and the ideal concentrations for optimal tissue regeneration require further investigation. Standardizing these parameters is essential for the effective clinical application of ZnHA.

Pediatric asthma is a common condition, and its exacerbations can be associated with significant morbidity and mortality. The role of nebulised magnesium as adjunct therapy for children with asthma exacerbations is still unclear. To compare clinical and functional outcomes for children with asthma exacerbation taking either nebulised magnesium sulfate added to standard medical therapy (SMT) versus SMT alone. PubMed, Embase, and Cochrane Library were systematically searched for randomised clinical trials (RCT) comparing the use of SMT with vs. without nebulised magnesium. The outcomes were respiratory rate, heart rate, % predicted peak expiratory flow rate (PEFR), % predicted forced expiratory volume (FEV1), peripheral O 2 saturation, asthma severity scores, and need for intravenous (IV) bronchodilator use. Twelve RCTs and 2484 children were included. Mean age was 5.6 (range 2-17) years old, mean baseline % predicted FEV1 was 69.6%, and 28.66% patients were male. Children treated with magnesium had a significantly higher % predicted PEFR (mean difference [MD] 5.33%; 95% confidence interval [CI] 4.75 to 5.90%; p < 0.01). Respiratory rate was significantly lower in the magnesium group (MD -0.70 respirations per minute; 95% CI -1.24 to -0.15; p < 0.01). Need for IV bronchodilators, % predicted FEV1, heart rate, asthma severity scores, and O 2 saturation were not significantly different between groups., Conclusion: In children with asthma exacerbation, treatment with nebulised magnesium and SMT was associated with a statistically significant, but small improvement in predicted PEFR and respiratory rate, as compared with SMT alone., What Is Known: • Magnesium sulfate has bronchodilating properties and aids in the treatment of asthma exacerbation when administered intravenously. • There is no significant evidence of benefit of nebulised magnesium as an adjunct therapy to the standard medical treatment for children with asthma exacerbations., What Is New: • Our study suggests nebulised magnesium sulfate may have a statistically significant, but small benefit in respiratory rate and peak expiratory flow rate. The addition of nebulised magnesium does not seem to increase adverse events., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Objectives: The aim of the present study was to assess the cytocompatibility of epoxy resin-based AH Plus Jet (Dentsply De Trey, Konstanz, Germany), Sealer Plus (MK Life, Porto Alegre, Brazil), calcium silicate-based Bio-C Sealer (Angelus, Londrina, PR, Brazil), Sealer Plus BC (MK Life) and AH Plus BC (Dentsply) through a tridimensional (3D) culture model of human osteoblast-like cells., Methods: Spheroids of MG-63 cells were produced and exposed to fresh root canal sealers extracts by 24 h, and the cytotoxicity was assessed by the Lactate Dehydrogenase assay (LDH). The distribution of dead cells within the microtissue was assessed by fluorescence microscopy, and morphological effects were investigated by histological analysis. The secreted inflammatory mediators were detected in cell supernatants through flow luminometry (XMap Luminex)., Results: Cells incubated with AH Plus Jet, AH Plus BC, Sealer Plus BC and Bio-C Sealer extracts showed high rates of cell viability, while the Sealer Plus induced a significant reduction of cell viability, causing reduction on the spheroid structure. Sealer Plus and Seaker Plus BC caused alterations on 3D microtissue morphology. The AH Plus BC extract was associated with the downregulation of secretion of pro-inflammatory cytokines IL-5, IL-7, IP-10 and RANTES., Conclusions: The new AH Plus BC calcium silicate-based endodontic sealer did not reduce cell viability in vitro, while led to the downregulation of pro-inflammatory cytokines., Clinical Significance: Choosing the appropriate endodontic sealer is a crucial step. AH Plus BC demonstrated high cell viability and downregulation of pro-inflammatory cytokines, appearing reliable for clinical use, while Sealer Plus presented lower cytocompatibility., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Objectives: This study aimed to produce and purify Clostridium perfringens type C beta-toxin, sheep anti-beta toxin immunoglobulin G (IgG) and chicken immunoglobulin Y (IgY)., Methods: Two methods were used for beta-toxin purification: single-step metal affinity chromatography (MAC) using zinc as a chelator and ion exchange chromatography (IEX). The purified and inactivated beta-toxoids were then administered to sheep and chickens in order to produce IgG and IgY., Results: All assays using the IEX failed. In contrast, MAC purified more than 21 mg of toxin per run in a single-step protocol. The purified and inactivated beta-toxoids were then administered to sheep and chickens, and IgG and IgY were purified with a high yield, medium antibody titer of 50 IU/mL, and high avidity (73.2 %)., Conclusions: C. perfringens type C beta-toxin and sheep or chicken anti-beta toxin IgG and IgY antibodies were successfully produced and purified using a simple protocol. This protocol can be used for the production of components used in the diagnosis and research of necrotic enteritis caused by C. perfringens type C, as well as for the evaluation of existing vaccines and the development of new preventive methods against this disease., Competing Interests: Declaration of competing interest The authors declare that they have no conflicts of interest., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Introduction: Significant evidence suggests that plasma-rich in growth factors (PRGF) favor the repair of chronic wounds, enabling a rapid return to functionality. However, components of PRGF and their effects on persistent ulcers and epithelial tissues are not well characterized. The goals of this research were to analyze the biological properties of platelet-derived factors, to examine their effectiveness on healing of venous ulcers, and to establish a correlation with clinical and sociodemographic data., Methods: For the preparation of PRGF, the centrifugation technique was used, obtaining a 100 % autologous and biocompatible blood sample that was treated with sodium citrate and calcium chloride. The patients were attended weekly at the outpatient clinic for nursing consultation and wound dressing changes, with PRGF application every 15 days. The treatment protocols are described, and follow-up results are reported., Results: Initially, the patients' ulcers ranged in sizes from 4 to 84 cm 2 . After 12 weeks of treatment, there was a significant mean reduction of 46.2 % in ulcer area. At baseline, epithelial tissue was absent in all venous ulcers, but its presence grew significantly by the treatment period. However, the reduction of the area of the ulcers did not show significant correlation with the concentrations of the patient's growth factors., Conclusions: Using the established protocol for PRGF isolating, it was possible to obtain a product with the presence of the six growth factors related to tissue regeneration and observed a positive response on wound healing following treatment of venous ulcers, with capacity to accelerate re-epithelialization and restore the skin functional integrity., Competing Interests: We declare that this material is original research, has not been previously published and has not been submitted for publication elsewhere while under consideration and there is no conflict of interest., (© 2024 The Japanese Society for Regenerative Medicine. Production and hosting by Elsevier B.V.)

Platelet-rich Fibrin (PRF), a second-generation blood concentrate, offers a versatile structure for bone regeneration due to its composition of fibrin, growth factors, and cytokines, with adaptations like denatured albumin-enriched with liquid PRF (Alb-PRF), showing potential for enhanced stability and growth factor dynamics. Researchers have also explored the combination of PRF with other biomaterials, aiming to create a three-dimensional framework for enhanced cell recruitment, proliferation, and differentiation in bone repair studies. This study aimed to evaluate a combination of Alb-PRF with nanostructured carbonated hydroxyapatite microspheres (Alb-ncHA-PRF), and how this association affects the release capacity of growth factors and immunomodulatory molecules, and its impact on the behavior of MG63 human osteoblast-like cells. Alb-PRF membranes were prepared and associated with nanocarboapatite (ncHA) microspheres during polymerization. MG63 cells were exposed to eluates of both membranes to assess cell viability, proliferation, mineralization, and alkaline phosphatase (ALP) activity. The ultrastructural analysis has shown that the spheres were shattered, and fragments were incorporated into both the fibrin mesh and the albumin gel of Alb-PRF. Alb-ncHA-PRF presented a reduced release of growth factors and cytokines when compared to Alb-PRF ( p < 0.05). Alb-ncHA-PRF was able to stimulate osteoblast proliferation and ALP activity at lower levels than those observed by Alb-PRF and was unable to positively affect in vitro mineralization by MG63 cells. These findings indicate that the addition of ncHA spheres reduces the biological activity of Alb-PRF, impairing its initial effects on osteoblast behavior.

Bone critical-size defects and non-union fractures have no intrinsic capacity for self-healing. In this context, the emergence of bone engineering has allowed the development of functional alternatives. The aim of this study was to evaluate the capacity of ASC spheroids in bone regeneration using a synergic strategy with 3D-printed scaffolds made from poly (lactic acid) (PLA) and nanostructured hydroxyapatite doped with carbonate ions (CHA) in a rat model of cranial critical-size defect. In summary, a set of results suggests that ASC spheroidal constructs promoted bone regeneration. In vitro results showed that ASC spheroids were able to spread and interact with the 3D-printed scaffold, synthesizing crucial growth factors and cytokines for bone regeneration, such as VEGF. Histological results after 3 and 6 months of implantation showed the formation of new bone tissue in the PLA/CHA scaffolds that were seeded with ASC spheroids. In conclusion, the presence of ASC spheroids in the PLA/CHA 3D-printed scaffolds seems to successfully promote bone formation, which can be crucial for a significant clinical improvement in critical bone defect regeneration.

Bone tissue engineering seeks biomaterials that enable cell migration, angiogenesis, matrix deposition, and tissue regeneration. Blood concentrates like platelet-rich fibrin (L-PRF) offer a cost-effective source of cells and growth factors to enhance healing. The present study aimed to evaluate heated serum albumin with liquid PRF (Alb-PRF) and L-PRF clinically and biochemically after placement in dental sockets following mandibular third molar extraction. In a controlled, split-mouth study involving 10 volunteers, 20 extracted molars were treated with either Alb-PRF or L-PRF. Post-extraction, pain, trismus, infection presence, and swelling were measured. The concentrations of different analytes in the surgical sites were also examined. The data were statistically analyzed, with significance defined at p < 0.05 ( t -test). No significant difference was noted between the groups for pain and trismus, but Alb-PRF showed a significant reduction in swelling on day seven. The Alb-PRF group showed lower levels of pro-inflammatory cytokines (GM-CSF, IL-1b, IL-6, IFNy, IL-8, IL-15, RANTES, and MIP-1a) after seven days, with only higher expressions of MIP-1b, IL-1b, and MCP-1 found in the L-PRF group. Differences were observed in the release of analytes between L-PRF and Alb-PRF, with Alb-PRF significantly reducing edema after seven days. Alb-PRF reduced edema, while L-PRF increased inflammatory cytokines. When compared to L-PRF, Alb-PRF reduced edema and the release of inflammatory cytokines, suggesting promising effects in socket healing while underscoring the role of growth factors and cytokines in potential applications of blood concentrates.

Platelet-rich fibrin (PRF) is a second-generation blood concentrate that serves as an autologous approach for both soft and hard tissue regeneration. It provides a scaffold for cell interaction and promotes the local release of growth factors. PRF has been investigated as an alternative to bone tissue therapy, with the potential to expedite wound healing and bone regeneration, though the mechanisms involved are not yet fully understood. This review aims to explore the in vitro evidence of PRF's effects on the behavior of mineralizing cells related to bone tissue regeneration. A systematic electronic search was conducted up to August 2023, utilizing three databases: PubMed, Web of Science, and Scopus. A total of 76 studies were selected, which presented in vitro evidence of PRF's usefulness, either alone or in conjunction with other biomaterials, for bone tissue treatment. PRF membranes' influence on the proliferation, differentiation, and mineralization of bone cells is linked to the constant release of growth factors, resulting in changes in crucial markers of bone cell metabolism and behavior. This further reinforces their therapeutic potential in wound healing and bone regeneration. While there are some notable differences among the studies, the overall results suggest a positive effect of PRF on cell proliferation, differentiation, mineralization, and a reduction in inflammation. This points to its therapeutic potential in the field of regenerative medicine. Collectively, these findings may help enhance our understanding of how PRF impacts basic physiological processes in bone and mineralized tissue.

Experimental research is critical for advancing medical knowledge and enhancing patient outcomes, including in vitro and in vivo preclinical assessments. Platelet-rich fibrin (PRF) is a blood by-product that has garnered attention in the medical and dental fields due to its potential for tissue regeneration and wound healing. Animal models, such as rabbits and rats, have been used to produce PRF and examine its properties and applications. PRF has demonstrated potential in the dental and medical fields for reducing inflammation, promoting tissue repair, and accelerating wound healing. This narrative review aims to compare existing evidence and provide guidelines for PRF animal research, emphasizing the importance of standardizing animal models, following ethical considerations, and maintaining transparency and accountability. The authors highlight the necessity to use the correct relative centrifugal force (RCF), standardize centrifugal calibration, and report detailed information about blood collection and centrifuge parameters for reproducible results. Standardizing animal models and techniques is crucial for narrowing the gap between laboratory research and clinical applications, ultimately enhancing the translation of findings from bench to bedside.

Sticky bone, a growth factor-enriched bone graft matrix, is a promising autologous material for bone tissue regeneration. However, its production is strongly dependent on manual handling steps. In this sense, a new device was developed to simplify the confection of the sticky bone, named Sticky Bone Preparation Device (SBPD ® ). The purpose of this pilot study was to investigate the suitability of the SBPD ® to prepare biomaterials for bone regeneration with autologous platelet concentrates. The SBPD ® allows the blending of particulate samples from synthetic, xenograft, or autogenous bone with autologous platelet concentrates, making it easy to use and avoiding the need of further manipulations for the combination of the materials. The protocol for the preparation of sticky bone samples using the SBPD ® is described, and the resulting product is compared with hand-mixed SB preparations regarding in vitro parameters such as cell content and the ability to release growth factors and cytokines relevant to tissue regeneration. The entrapped cell content was estimated, and the ability to release biological mediators was assessed after 7 days of incubation in culture medium. Both preparations increased the leukocyte and platelet concentrations compared to whole-blood samples ( p < 0.05), without significant differences between SB and SBPD ® . SBPD ® samples released several growth factors, including VEGF, FGFb, and PDGF, at concentrations physiologically equivalent to those released by SB preparations. Therefore, the use of SBPD ® results in a similar product to the standard protocol, but with more straightforward and shorter preparation times and less manipulation. These preliminary results suggest this device as a suitable alternative for combining bone substitute materials with platelet concentrates for bone tissue regeneration.

Herd vaccination is an important preventive measure against enterotoxemia in ruminants. Vaccination in goats should be performed every four months, and recent studies have shown that immunity in cattle lasts for less than one year. One of the mechanisms for increasing the duration of the immune response is to use purified toxoids as immunogens. The aim of the present study was to evaluate the humoral response in cattle and goats after vaccination with purified and semi-purified Clostridium perfringens type D epsilon toxoid. The following three different vaccines were used: vaccine 1 (V1), a semi-purified toxoid adsorbed to aluminum hydroxide; vaccine 2 (V2), a purified toxoid adsorbed to aluminum hydroxide; and vaccine (V3), a purified toxoid adsorbed on chitosan microparticles. Groups of cattle (n = 6-7) and goats (n = 6-7) were vaccinated on days 0 and 30, and serum samples for antitoxin titration were collected every 30 days for one-year post-vaccination. Goats were revaccinated on day 360, and their serum was evaluated on days 367 and 374. The antibody peaks ranged between 6.90 and 11.47 IU/mL in cattle and from 1.11 to 4.40 IU/mL in goats. In cattle administered with the V1 and V2 vaccines, we observed that the antibody titers were maintained above 0.2 IU/mL until the end of the experiment. In goats, V2 elicited long-lasting antibodies, and all animals maintained the protective titers for 210 days after the first dose. In conclusion, the purified toxoid vaccine with aluminum hydroxide adjuvant was able to induce strong and long-lasting humoral responses in both species and could be an alternative for improving the immunization schedule against enterotoxemia in goats and cattle., Competing Interests: Declaration of competing interest The authors declare that there are no conflicts of interest., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Access to genetic resources (GR) and/or traditional knowledge associated with genetic resources (ATK) has been regulated in Brazil since 2001. The law 13,123 / 2015 determined a significant change in the theme, mainly on the rules of distribution of benefits obtained for conservation and sustainable use of biodiversity, the access to technology and technology transfer, the exploitation of products or reproductive material from the GR or ATK and consignment to the outside of part or all the living or dead organism shipped for GR. The implementation of international treaties on GR and ATK for research, biotechnological development and bioprospecting have been causing difficulties for Brazilian researchers, mainly due to the lack of information and dissemination available for compliance with the legislation. In this work, the members of the Committee for Access to Genetic Resources and Associated Traditional Knowledge of the Federal Fluminense University (UFFGEN) - Brazil, and collaborators performed a critical reflection on the new law, helping Brazilian researchers with information necessary to understand the changes made by the new legislation, especially in the field of Biotechnology associated with Brazilian Biodiversity.

Successful biomaterials for bone tissue therapy must present different biocompatible properties, such as the ability to stimulate the migration and proliferation of osteogenic cells on the implantable surface, to increase attachment and avoid the risks of implant movement after surgery. The present work investigates the applicability of a three-dimensional (3D) model of bone cells (osteospheres) in the evaluation of osteoconductive properties of different implant surfaces. Three different titanium surface treatments were tested: machined (MA), sandblasting and acid etching (BE), and Hydroxyapatite coating by plasma spray (PSHA). The surfaces were characterized by Scanning Electron Microscopy (SEM) and atomic force microscopy (AFM), confirming that they present very distinct roughness. After seeding the osteospheres, cell-surface interactions were studied in relation to cell proliferation, migration, and spreading. The results show that BE surfaces present higher densities of cells, leaving the aggregates towards than titanium surfaces, providing more evidence of migration. The PSHA surface presented the lowest performance in all analyses. The results indicate that the 3D model allows the focal analysis of an in vitro cell/surfaces interaction of cells and surfaces. Moreover, by demonstrating the agreement with the clinical data observed in the literature, they suggest a potential use as a predictive preclinical tool for investigating osteoconductive properties of novel biomaterials for bone therapy.

This study aimed to analyze Fibroblast Growth Factor-2 (FGF-2) levels in the peri-implant crevicular fluid throughout supportive mucositis therapy. Twenty-six participants with Branemark protocol prosthesis were divided into two groups: the control group, characterized by healthy peri-implants, and the mucositis group, presenting a diagnosis of peri-implant mucositis. All participants underwent clinical examination, radiographic analysis, prosthesis removal, and non-invasive peri-implant therapy (mechanical debridement associated with chlorhexidine 0.12%) during a period of 36 days divided into three intervals. Peri-implant crevicular fluid samples were collected at each interval in order to analyze FGF-2 levels by immuno-enzymatic assay. The control and mucositis groups showed difference in keratinized mucosa. The smaller the range of keratinized mucosa the higher susceptibility of peri-implant mucositis. Throughout the treatment intervals, participants were diagnosed in different groups indicating whether or not the non-invasive therapy was able to treat peri-implant mucositis. There was a significant difference of FGF-2 levels between groups, with the higher FGF-2 levels in the control group (p=0.01). After supportive therapy, the mucositis group showed significantly increased FGF-2 levels (p<0.01) compared to initial levels. After 36 days of supportive therapy, there was a reduction of peri-implant mucositis from 70% to 23%. Clinical and laboratory outcomes showed a clear correlation since FGF-2 levels increased after 36 days. It was concluded that the therapy protocol was effective and promoted a regenerative reaction and FGF-2 can be considered a future target for peri-implant mucositis understanding.

The Bacterial Endotoxin Test (BET) is a method for exclusion of endotoxin-related pyrogen contamination in pharmaceutical products, as an alternative to the Rabbit Pyrogen Test (RPT). However, BET does not detect a broad range of biologically relevant pyrogens, and interferences can limit its practical use for different medical products. This work aimed to scope the evidence in the scientific literature for case-by-case validity assessments of BET in different uses for medical products. A search strategy was conducted in PubMed, Scopus, and Web of Science in April 2020, according to the PRISMA-ScR statement. Twenty-two references were included, evaluating medical products for endotoxin contamination through both BET and RPT according to standardized protocols. A critical appraisal was performed through ToxRTool, followed by data extraction and qualitative synthesis of outcomes and methodological issues. Four classes of products assessed by BET were identified, including nanoparticles, drugs, blood and biological products. A considerable variation was observed on the BET methods used. Collectively, the evidence indicates different factors influencing the outcome of BET, including the chemical nature of samples that may cause interference depending on the selected method. While some applications to medical products appear adequate, others, such as nanoparticles, may require the use of different in vitro pyrogen testing methods, reinforcing the need for case-by-case validation for each BET method and type of medical product., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

The aim of this study was to purify Clostridium perfringens type D epsilon toxin and produce and purify anti-epsilon chicken immunoglobulin Y (IgY). A single-step ion exchange chromatography resulted in a high-yield and high-purity toxin, while ion exchange chromatography followed by gel filtration resulted in the highest purity of the toxin, but at a lower yield. Purified and inactivated epsilon toxin were then administered in chickens via four inoculations and IgY was obtained at a high purity and yield, with an antibody titer of 50 IU/mL and high levels of avidity (73.2%). In summary, C. perfringens type D epsilon toxin and chicken anti-epsilon IgY were successfully produced and purified, and may be used for the diagnosis of enterotoxemia caused by the epsilon toxin, as well as in potency tests of existing and future vaccines against enterotoxemia., Competing Interests: Declaration of competing interest The authors declare that there are no conflicts of interest., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Guided bone regeneration (GBR) is a process that involves the regeneration of bone defects through the application of occlusive membranes that mechanically exclude the population of non-osteogenic cells from the surrounding soft tissue. Interestingly, platelet-rich fibrin (PRF) has previously been proposed as an autologous GBR membrane despite its short-term resorption period of 2-3 weeks. Recent clinical observations have demonstrated that, by heating a liquid platelet-poor plasma (PPP) layer and mixing the cell-rich buffy coat zone, the resorption properties of heated albumin gel with liquid-PRF (Alb-PRF) can be significantly improved. The aim of this study was to evaluate the inflammatory reaction, biocompatibility, and extended degradation properties of a new autologous Alb-PRF membrane in comparison to commonly utilized standard PRF after nude mice implantation, according to ISO 10993-6/2016. Two standard preparations of PRF (L-PRF and H-PRF) were compared to novel Alb-PRF following subcutaneous implantation at 7, 14, and 21 days. All groups demonstrated excellent biocompatibility owing to their autologous sources. However, it is worth noting that, while both L-PRF and H-PRF membranes demonstrated significant or complete resorption by 21 days, the Alb-PRF membrane remained volume-stable throughout the duration of the study. This study demonstrates-for the first time, to the best of our knowledge-a marked improvement in the membrane stability of Alb-PRF. This indicates its future potential for use as a biological barrier membrane for GBR procedures with a long-lasting half-life, or as a biological filler material in esthetic medicine applications. Thus, further studies are warranted to explore future clinical applications in various fields of medicine.

Evaluate the cytocompatibility of Calen ® /ZO, Calcicur ® , Vitapex ® , Endoflas ® , and zinc oxide/eugenol-based (ZOE) root canal pastes (RCP) to human primary osteoblasts (HPO) through a simplified model for primary teeth. The model employed pipette tips filled with 0.037 g of paste, exposed to 185 µL of culture medium for 24 h (n = 6). Release of components was analysed by Proton Nuclear Magnetic Resonance Spectroscopy ( 1 H-NMR). HPO were exposed to conditioned media for 24 h. Cell viability was assessed by cell density and metabolic activity, and release of interleukin 6 (IL-6), vascular endothelial growth factor (VEGF) and fibroblast growth factor (bFGF) by immunological assay. Physicochemical properties and antimicrobial efficacy were also evaluated. 1 H-NMR spectra analysis showed similarity between ZOE, Endoflas ® , Calcicur ® , and Vitapex ® compared to Calen ® /ZO and positive control, which showed distinct released components. Calen ® /ZO and Calcicur ® exhibited high alkaline pH in all periods and showed similar solubility. Calen ® /ZO, ZOE, and Vitapex ® showed similar flow rate. Calen ® /ZO, Calcicur ® , and Vitapex ® did not exhibit antimicrobial efficacy. Calen ® /ZO presented cytotoxicity (p < 0.05). Pastes did not increase IL-6 release compared to control. Apart from Vitapex ® , all pastes significantly induced VEGF/bFGF release. Interactive effects among released products may affect biological response to filling pastes. Calcicur ® , ZOE, Endoflas ® and Calen ® /ZO presented good to moderate cytocompatibility, with low impact on pro-inflammatory cytokine release and induction of growth factors of interest to tissue repair. This simplified model, specific for the evaluation of the cytocompatibility of RCPs on primary teeth, suggests how these pastes might contribute to bone repair in clinical situations of apical periodontitis in children.

This study evaluated the impact of rotor angle and time of storage after centrifugation on the in vitro biological properties of platelet-rich fibrin (PRF) membranes. Blood samples (n = 9) were processed with a vertical fixed-angle (V) or a swing-out horizontal (H) centrifuge, with 20-60 min of sample storage after centrifugation. Leukocytes, platelets, and red blood cells were counted, and fibrin architecture was observed by scanning electron microscopy (SEM). The release of FGF2, PDGFbb, VEGF, IL-6, and IL-1β was measured after incubation on culture media for 7-21 days. Cell content was equivalent in all experimental groups (p > .05). The fibrin matrix was similar for fixed-angle and horizontal centrifugation. Horizontal centrifugation induced a twofold increase in PDGF and 1.7× increase on FGF release as compared to V samples, while IL-1β was significantly reduced (p < .05). No significant difference was observed on the release of growth factors and cytokines at different times after centrifugation (p < .05). These data suggest that both angles of centrifugation produce PRF membranes with similar structure and cellularity, but horizontal centrifugation induces a higher release of growth factors. Higher times of storage after centrifugation did not impact on cell content and the release of growth factors., (© 2020 Wiley Periodicals LLC.)

This study aimed to assess the response of 3D printed polylactic acid (PLA) scaffolds biomimetically coated with apatite on human primary osteoblast (HOb) spheroids and evaluate the biological response to its association with Bone Morphogenetic Protein 2 (rhBMP-2) in rat calvaria. PLA scaffolds were produced via 3D printing, soaked in simulated body fluid (SBF) solution to promote apatite deposition, and characterized by physical-chemical, morphological, and mechanical properties. PLA-CaP scaffolds with interconnected porous and mechanical properties suitable for bone repairing were produced with reproducibility. The in vitro biological response was assessed with human primary osteoblast spheroids. Increased cell adhesion and the rise of in vitro release of growth factors (Platelet-Derived Growth Factor (PDGF), Basic Fibroblast Growth Factor (bFGF), Vascular Endothelial Growth Factor (VEGF) was observed for PLA-CaP scaffolds, when pre-treated with fetal bovine serum (FBS). This pre-treatment with FBS was done in a way to enhance the adsorption of serum proteins, increasing the number of bioactive sites on the surface of scaffolds, and to partially mimic in vivo interactions. The in vivo analysis was conducted through the implantation of 3D printed PLA scaffolds either alone, coated with apatite (PLA-CaP) or PLA-CaP loaded with rhBMP-2 on critical-sized defects (8 mm) of rat calvaria. PLA-CaP+rhBMP2 presented higher values of newly formed bone (NFB) than other groups at all in vivo experimental periods ( p < 0.05), attaining 44.85% of NFB after six months. These findings indicated two new potential candidates as alternatives to autogenous bone grafts for long-term treatment: (i) 3D-printed PLA-CaP scaffold associated with spheroids, since it can reduce the time of repair in situ by expression of biomolecules and growth factors; and (ii) 3D-printed PLA-CaP functionalized rhBMP2 scaffold, a biocompatible, bioactive biomaterial, with osteoconductivity and osteoinductivity.

This study determined the effect of thiourethane-functionalized fillers (TU) on the antimicrobial properties, cytotoxicity, degree of conversion (DC), water sorption (Wsp) and solubility (Wsl) of experimental composites. TU-modified fillers were added at different ratios in experimental composites: 0 (Control-TU0), 25% (TU25), 50% (TU50), 75% (TU75) and 100wt% (TU100). The antimicrobial properties were detected through the exhaustion test and counting of Streptococus mutans colonies for biofilm formation. Cytotoxicity to human gingival fibroblasts was evaluated in three different parameters: XTT (2,3-Bis-(2-Methoxy-4-Nitro-5-Sulfophenyl)-2H-Tetrazolium-5-Carboxanilide), NRU (Neutral Red Uptake assay) and CVDE (Crystal Violet Dye Exclusion test)) at the same cells. ELISA was used to measure the IL-6 and b-FGF biomarkers. DC was determined by Fourier-transformed infrared spectroscopy, while Wsp and Wsl by mass variations. Inhibitory capacity of biofilm formation was not observed for any material. All groups presented at least 70% of cell survival within the observed periods (24h and 7 days). Positive control (toxic) had high IL-6 values and low b-FGF values. No significant variations in DC, Wsp, and Wsl were observed among the experimental groups. The use of thiourethane did not present antimicrobial and cytotoxic activity and the tested materials presented equivalent properties to those conventionally used in dentistry.

Objectives: This systematic review aimed to compare the cytotoxicity of root canal filling materials (RCFMs) assessed using tetrazolium salt-based tests (TSBT), including the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay, with those obtained using other cell viability assays., Methods: A search was performed on PubMed, Scopus, Web of Science and OpenGrey up to March 2019, followed by a manual search. According to the Participants, Exposure, Comparator and Outcomes (PECO) criteria, in vitro studies that evaluated the cytotoxic effect of RCFMss on animal and/or human cells through TSBT and at least one other viability assay were compared. The methodological quality of selected papers was assessed using ToxRTool ® and SciRAP ® . Data were analysed using Wilcoxon's signed-rank test for paired samples and linear weighting kappa., Results: A total of 230 non-duplicated records were identified. After applying the eligibility criteria, 55 studies were selected for methodological evaluation, seven were selected by manual searching, 22 were excluded for methodological reasons, and 40 were included. A total of 410 comparisons were performed between TSBT and distinct cell viability tests (DCVT). MTT had moderate concordance with DCVT using human cells (n = 138 samples) (P = 0.507; k = 0.4225) and animal cells (n = 122 samples) (P = 0.124; k = 0.5775). XTT had good concordance using human (n = 110 samples) (P = 0.507; k = 0.6336) and animal cells (n = 12 samples) (P = 0.564; k = 0.6604). MTT, XTT, WST and MTS assays showed moderate concordance with DCVT (n = 410 samples) (P = 0.375; k = 0.5138) and complete agreement in 226 samples., Discussion: The included studies had methodological heterogeneity that was minimized by the systematic review methodology., Conclusions: MTT and XTT do not cause over- or underestimation of cell viability during cytotoxicity screening of root canal filling materials, implying that these assays can be considered reliable for this purpose. Nonetheless, the development of protocols for the cytotoxic screening of these materials on 3D tissue-like cultures aiming to improve their predictability in the clinical scenario is suggested., (© 2020 International Endodontic Journal. Published by John Wiley & Sons Ltd.)

Objective: Different studies in the last decade have proposed that gene expression alterations that are independent of the DNA sequence may also play an important role in periapical disease. The present study aimed to assess the available evidence supporting a relationship between these alterations and apical periodontitis through a scoping review., Design: Specific strategies were developed for different databases (MEDLINE via PubMed, Cochrane Library, Scopus, Web of Science, and Virtual Health Library) and a search performed by March 1st, 2019. The evidence sources were selected according to the eligibility criteria and underwent a critical appraisal of methodological quality., Results: The initial search retrieved 212 references, with eight eligible articles after the removal of replicates and application of exclusion criteria. Five studies identified altered DNA methylation on inflammatory response genes (FOXP3, CXCL3, FADD, MMP2, MMP9, IFNG, IL4, IL12) on AP patients. Three others identified the alterations on the expression of several microRNAs (miR-29b, 106b, 125b, 143, 146a, 155, 198) during AP. No evidence was identified regarding mechanisms of histone methylation, or of epigenetic heritability or stability., Conclusions: There is available evidence for the involvement of different genetic regulatory mechanisms independent of changes in DNA sequence in the development or severity of apical periodontitis. However, due to methodological limitations, further research must be performed before novel therapies and diagnostic tools for AP may arise from these data., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

This paper investigates factors associated with motherhood among adolescents from 14 to 16 years of age in Porto Alegre, Brazil. This is a case-control study with 431 adolescent mothers (cases) and 862 adolescents who had never given birth (controls). D. ata were obtained through home visits by an interviewer-applied questionnaire. Sociodemographic characteristics, quality of social and family relationships, lifestyle and history of abuse were studied as potential determinants to early adolescent motherhood. Conditional logistic regression was used for data analysis according to a two-stage hierarchical model. Results showed that lower economic class, schooling failure, tobacco consumption, alcoholic drunkenness at least once in life and having a mother who gave birth before 20 years of age were positively associated with early adolescent motherhood. Later menarche and having relatives or having friends in whom to trust remained as protective factors. Schooling failure, which obtained the highest risk, points to the important role of the school in this population's development and its potential to stimulate healthy life habits.

Nanomedicine is an emerging research area which has brought new possibilities and promising applications in image, diagnosis, and treatment. Nanoparticles (NPs) for medicinal purposes can be made of several material types such as silica, carbon, different polymers, and metals as silver, copper, titanium, and gold. Gold NPs (AuNPs) are the most studied and used, mostly due to their characteristics including simple preparation, controllable size and distribution, biocompatibility, good acceptance of surface modifications, and specific surface plasmon resonance (SPR). This study reviews the scientific literature regarding the potential applications of AuNPs in the development of new diagnostic and therapeutic strategies for nanomedicine, including their biomedical use as a drug carrier, as an agent in radio and phototherapy, and bioimaging for image diagnosis. While it becomes clear that much research remains to be done to improve the use of these nanoparticles, with particular concern for safety issues, the evidence from the literature already points to the great potential of AuNPs in nanomedicine., (© 2019 Wiley Periodicals, Inc.)

The aim of this study was to evaluate the titers of neutralizing antibodies in cattle inoculated with multivalent commercial clostridial vaccines containing C. botulinum type C (BoNTC), C. botulinum type D (BoNTD), and C. perfringens epsilon (ETX) toxoids for a period of one year. Cattle (Bos taurus), aged 4-6 months and not previously immunized, were vaccinated under four different protocols at days 0 and 30 and followed over one year. Individual serum titration was performed by a serum neutralization test in mice or in MDCK cells. The number of animals with detectable neutralizing antibodies ranged from 40.6% to 78.1%, but only 12.5% of animals showed neutralizing antibodies against all tested antigens. Neutralizing antibodies were found only until 60 days for ETX, 120 days for BoNTC, and 180 days for BoNTD. The absence of detectable neutralizing antibodies against the three antigens before 360 days, suggests that cattle remained unprotected for a long period before the recommended booster vaccination., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Nanostructured carbonated hydroxyapatite (nCHA) is a promising biomaterial for bone tissue engineering due to its chemical properties, similar to those of the bone mineral phase and its enhanced in vivo bioresorption. However, the biological effects of nCHA nanoparticles on cells and tissues are not sufficiently known. This study assessed the impact of exposing pre-osteoblasts to suspensions with high doses of nCHA nanoparticles with high or low crystallinity. MC3T3-E1 pre-osteoblasts were cultured for 1 or 7 days in a culture medium previously exposed to CHA nanoparticles for 1 day. Control groups were produced by centrifugation for removal of bigger nCHA aggregates before exposure. Interaction of nanoparticles with the culture medium drastically changed medium composition, promoting Ca, P, and protein adsorption. Transmission Electron microscopy revealed that exposed cells were able to internalize both materials, which seemed concentrated inside endosomes. No cytotoxicity was observed for both materials, regardless of centrifugation, and the exposure did not induce alterations in the release of pro-and anti-inflammatory cytokines. Morphological analysis revealed strong interactions of nCHA aggregates with cell surfaces, however without marked alterations in morphological features and cytoskeleton ultrastructure. The overall in vitro biocompatibility of nCHA materials, regardless of physicochemical characteristics such as crystallinity, encourages further studies on their clinical applications., (© 2019 Wiley Periodicals, Inc.)

Purpose: To evaluate the clinical outcomes of platelet-rich fibrin (PRF) application for hemostasis after dental extraction in patients receiving anticoagulant therapy with factor Xa inhibitors., Methods: In total, 25 patients receiving anticoagulant therapy with rivaroxaban or apixaban who required routine dental extraction were evaluated. In all patients, PRF was used for hemostasis in addition to adapting sutures. Bleeding was subjectively assessed using a sterilize gauze pad at 24, 48, and 72 h after the procedure., Results: All invited participants (n = 25) consented to participate. The PRF clots successfully arrested bleeding after extraction in all patients, with no complications at any time point after the procedure. Favorable soft tissue healing was observed during suture removal at 10 days after the procedure in all patients, with no signs of infection or late healing., Conclusions: The results of this case series indicate that PRF is a promising natural hemostatic agent for the management of bleeding after dental extraction in patients receiving factor Xa inhibitor therapy. Further controlled clinical studies with larger patient samples are necessary to clarify the findings of this case series.

Objectives: The increased demand for esthetics and minimally invasive tooth restorations resulted in a rapid development of adhesive dentistry. However, much controversy remains about the safe use and cytotoxic effects of different groups of dental adhesives. The present study performed a systematic review to identify the answer to the following question: are self-etch adhesives more cytotoxic than those employing the etch-and-rinse system?, Methods: This systematic review was performed in accordance with the PRISMA statement; a quality assessment for in vitro studies was conducted using the ToxRTool. Specific search strategies were developed and performed in the electronic databases MEDLINE via PubMed, Cochrane Library, Scopus, Web of Science, and LILACS/BBO. After removal of duplicated studies and application of the exclusion criteria, ten eligible articles were selected and submitted to a qualitative descriptive analysis comparing both groups of dental adhesives. Most in vitro test systems employed pulp cells or gingival fibroblasts., Results: The methodologies presented great variability regarding the exposure to the test materials. Only four studies assessed the role of the degree of conversion of the materials in their toxicity, with conflicting results., Significance: While the lack of methodological standardization among the studies still hinders the establishment of a relationship between type of dental adhesive and toxicity, studies employing dentin barrier systems indicate greater cytotoxicity for etch-and-rinse adhesives., (Copyright © 2018 The Academy of Dental Materials. Published by Elsevier Inc. All rights reserved.)

The oral administration of non-toxigenic strains of Clostridioides difficile (NTCD) is currently showing promising results for the prevention of Clostridioides difficile infection (CDI) in humans and animals, and is being considered as a possible commercial product to be used in the near future. The aim of this work was to evaluate five culture media for the growth and sporulation of one NTCD (Z31) and evaluate the viability of a lyophilized spore solution of NTCD Z31 stored at 4 °C or at 25 °C for 2 years. Reinforced clostridial medium (RCM) and brain heart infusion broth (BHI) provided the highest production of NTCD Z31 spores. In the first 6 months of the storage of the lyophilized solution, a reduction in spore count of approximately 0.3 Log 10  CFU/mL was observed; however, no further significant reduction in spore count was observed up to 24 months. No difference in spore concentration was found between the two storage temperatures from 6 to 24 months of storage. The present work showed BHI and RCM to be the best choices for the growth and sporulation of NTCD Z31 and suggested that the spores of NTCD Z31 are stable for up to 2 years under both temperature conditions.

Objective: The combination of calcium phosphate with blood-derived growth factors (BDGF) has been widely used in bone regeneration procedures although its benefits are still unclear. The purpose of this study was to evaluate whether or not BDGF improves the efficacy of a modified carbonated calcium phosphate biomaterial in sinus floor augmentation., Material and Methods: Ten patients underwent 20 sinus floor augmentation procedures using nanostructured carbonated hydroxyapatite (cHA) microspheres alone or associated with BDGF in a randomized controlled clinical trial. The in vitro release of growth factors was assessed by an elution assay. Bone grafts were randomly implanted in the right and left maxillary sinuses of each participant, associated either with a 0.9% saline solution or BDGF. Bone gain was evaluated through cone beam tomography after 180 days., Results: Nine women and one man composed the sample. The blood-derived concentrates were able to release high levels of growth factors and cytokines. A significant clinical advantage was observed in the use of the BDGF after fibrin polymerization around the biomaterial microspheres, optimizing the surgical procedures, thereby reducing the time and displacement, and improving the adaptation of the biomaterial in the maxillary sinus. No synergistic effect was observed in bone formation when cHA was associated with BDGF (p > 0.05)., Conclusion: Equivalent new bone formation was observed for cHA in the presence or absence of the BDGF concentrate in bilateral sinus floor elevation after 6 months. Blood-derived growth factors did not improve bone repair when associated with calcium phosphate in sinus lift procedures.

During adolescence, body image is linked to subjective well-being. This study aimed to identify variables related to overall well-being (life satisfaction, health perception, happiness, and optimism) that discriminate between teenagers who are satisfied with body image and those who are dissatisfied. A cross-sectional, school- based study was conducted involving a sample of 1,460 ninth-grade students attending state schools in Canoas in the State of Rio Grande do Sul, Brazil. The following instruments were used: a sociodemographic questionnaire; the Economic Classification Criteria Brazil; the Body Shape Questionnaire; the Brief Multidimensional Students' Life Satisfaction Scale; the Happiness Measures; and health perception and perceived optimism. The data was analyzed using univariate analysis and multiple discriminant analysis. A total of 1,091 participants (74.7%) were satisfied with their body image. The variables that contributed most to the discrimination between groups were sex (0.680), self satisfaction (0.644), health perception (0.630), and level of happiness with health (0.601). Adopting an approach where having a positive body image is seen as a key element of mental and physical health can help make health care services more accessible to adolescents and promote more effective and less regulatory care.

Impact Statement: This study evaluated for the first time the composition and bioactivity of platelet-rich fibrin (PRF) produced from small animal blood by reducing the initial blood volume needed for the preparation of PRF from 10 to 3 mL. The results showed that different preparation protocols of PRF produced using 3 mL of animal blood exhibit the same composition, properties, and bioactivity as PRF prepared using 10 mL human blood.

The control of postoperative bleeding represents one of the main intercurrent events associated with soft tissue surgical procedures in the oral cavity. In this context, platelet-rich fibrin (PRF) membranes are materials with great potential for optimizing soft tissue healing and induction of hemostasis. This interventional case series describes the treatment of 10 patients with excisional biopsy of benign oral cavity lesions, following a screening sequence at the surgery clinic of a Brazilian dental school between the years of 2015 and 2017. After treatment with PRF, patients presented mean time for postoperative hemostasis of 10.3 ± 2.5 s, requiring the average use of three membranes to cover the surgical area. The results suggest that the use of platelet-rich fibrin membranes may represent a feasible alternative hemostatic material for the treatment of oral lesions.

This study aimed to compare the cytotoxicity of the Vita AC12, Lava Ultimate, Vita Enamic and InSync indirect restorative materials. Extracts of each material were prepared by incubation for 1, 7 and 40 days, with daily washing. Human gingival fibroblasts were exposed to the extracts, and cell viability was evaluated by sequential assessment of mitochondrial activity (XTT), membrane integrity (NRU) and cell density (CVDE). Extracts of polystyrene beads and latex fragments were used as negative and positive controls, respectively. Differences between groups and experimental times were evaluated by analysis of variance. At the 24 h extraction, significant differences between the control and both Vita AC-12 and InSync were observed in the XTT assay (p<0.05), and between the control and both Enamic and Lava Ultimate, in the CVDE assay (p<0.05). AC12, Lava Ultimate, and InSync presented significantly lower cell viability than Enamic and the control group, in the NRU assay (p<0.05). The Vita Enamic and Lava Ultimate hybrid ceramic-like materials presented better biocompatibility at the 24 h extraction time point than the AC12 and InSync ceramic materials. However, a simulation of the removal of toxic components by biological fluids, conducted by using longer extraction times and daily washing, led to the absence of cytotoxicity in all the tested restorative materials. These findings can be viewed as positive for the clinical indication of these restorative materials, considering their contact with adjacent soft tissues for extended periods of time.

Objective: This replication study aimed to evaluate an association between caries experience and polymorphisms in DEFB1 and miRNA202 in two different Brazilian groups., Design: The population consisted in 312 Brazilian children. Genomic DNA for was extracted from buccal cells isolated from saliva. The genotyping analysis of the polymorphisms in DEFB1 and miRNA202 was performed by real-time polymerase chain reactions. The associations between caries experience, genotype and allele distribution was performed, with an alpha of 0.05., Results: A statistical significant difference was observed between allele distribution and the polymorphism rs12355840 in the miRNA202 for permanent dentition in the Manaus group, in which individuals that carry the allele C had almost three times more chance to have caries (p = .021; OR = 2.7, 95% CI = 1.1-6.7). In the Ribeirão Preto group there was a statistical significant difference for the polymorphism rs11362 in the DEFB1 for both dentition in alleles (p = .043) and genotype (p = .020) distributions. The T allele increased in two times the chance to have caries (OR = 2.03; 95% CI = 1.05-4.07)., Conclusion: In conclusion, the allelic distribution of the polymorphism rs12355840 in miRNA202 was associated with caries experience in the Manaus group. In the Ribeirão Preto group, the allelic and genotypic distributions in the polymorphism rs11362 in DEFB1 were associated with caries experience., (Copyright © 2018. Published by Elsevier Ltd.)

Platelet-rich fibrin membranes are biomaterials widely used for therapeutic purposes, and canonically produced through the processing of peripheral blood with fixed-angle rotor centrifuges. In this work, we evaluate the in vitro stability and release of cytokines and growth factors when these biomaterials are produced with a horizontal swing-out clinical centrifuge. Membranes produced from the blood of 14 donors were morphologically evaluated by scanning electron microscopy and fluorescence microscopy, and their stability was assessed by photographic recording after incubation in culture medium for up to 28 days. The release of 27 cytokines and growth factors was monitored for three weeks through a multiparametric immunoassay. The fibrin membranes presented complex three-dimensional structure with a high density of nucleated cells. A large release of growth factors [platelet derived growth factor, fibroblastic growth factor (bFGF), and vascular endothelial growth factor] was detected in the first 24 h, followed by time-dependent decay, maintaining significant concentrations after three weeks. Both anti-inflammatory and pro-inflammatory cytokines presented different release peaks, maintaining high rates of elution for up to 21 days. Chemokines of relevance in tissue repair [RANTES, granulocyte colony-stimulating factor (G-CSF)] were also produced in large quantities throughout the experimental period. The present results demonstrate that blood-derived fibrin membranes with high structural stability and cell content can be generated by horizontal centrifugation, being able of a prolonged production/release of growth factors and pro- and anti-inflammatory cytokines. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 106A: 1373-1380, 2018., (© 2018 Wiley Periodicals, Inc.)

This study evaluated comparatively two configurations (powder and putty) of a composite biomaterial based on PLGA (Poly(lactide-co-glycolide)/nanoescale hydroxyapatite (ReOss®, Intra-Lock International) through microscopic morphology, in vitro cytotoxicity, biocompatibility and in vivo response as a bone substitute. SEM and EDS characterized the biomaterials before/after grafting. Cytocompatibility was assessed with murine pre-osteoblasts. Osteoconductivity and biocompatibility were evaluated in White New Zealand rabbits. Both configurations were implanted in the calvaria of eighteen animals in non-critical size defects, with blood clot as the control group. After 30, 60 and 90 days, the animals were euthanized and the fragments containing the biomaterials and controls were harvested. Bone blocks were embedded in paraffin (n=15) aiming at histological and histomorphometric analysis, and in resin (n=3) aiming at SEM and EDS. Before implantation, the putty configuration showed both a porous and a fibrous morphological phase. Powder revealed porous particles with variable granulometry. EDS showed calcium, carbon, and oxygen in putty configuration, while powder also showed phosphorus. After implantation EDS revealed calcium, carbon, and oxygen in both configurations. The materials were considered cytotoxic by the XTT test. Histological analysis showed new bone formation and no inflammatory reaction at implant sites. However, the histomorphometric analysis indicated that the amount of newly formed bone was not statistically different between experimental groups. Although both materials presented in vitro cytotoxicity, they were biocompatible and osteoconductive. The configuration of ReOss® affected morphological characteristics and the in vitro cytocompatibility but did not impact on the in vivo biological response, as measured by the present model.

Although the discipline of cell biology (CB) is part of the curricula of predoctoral dental schools, students often fail to recognize its practical relevance. The aim of this study was to assess the effectiveness of a practical-theoretical project-based course in closing the gaps among CB, scientific research, and dentistry for dental students. A project-based learning course was developed with nine sequential lessons to evaluate 108 undergraduate dental students enrolled in CB classes of a Brazilian school of dentistry during 2013-16. To highlight the relevance of in vitro studies in the preclinical evaluation of dental materials at the cellular level, the students were challenged to complete the process of drafting a protocol and performing a cytocompatibility assay for a bone substitute used in dentistry. Class activities included small group discussions, scientific database search and article presentations, protocol development, lab experimentation, and writing of a final scientific report. A control group of 31 students attended only one laboratory class on the same theme, and the final reports were compared between the two groups. The results showed that the project-based learning students had superior outcomes in acknowledging the relevance of in vitro methods during biocompatibility testing. Moreover, they produced scientifically sound reports with more content on methodological issues, the relationship with dentistry, and the scientific literature than the control group (p<0.05). The project-based learning students also recognized a higher relevance of scientific research and CB to dental practice. These results suggest that a project-based approach can help contextualize scientific research in dental curricula.

Denture adhesives (DA) improve the retention and stability of ill-fitting dentures, especially for older adults. These materials should be biocompatible, i.e., they cannot cause undesired biological responses and be non-cytotoxic to oral tissues. However, in vitro testing of DA biocompatibility employing primary cell culture may possibly be affected by other factors, such as the donor age., Objective: To compare the cytotoxicity of three different denture adhesives when assessed in primary gingival fibroblasts from a young donor or from an older donor, as well as the release of the basic fibroblast growth factor (bFGF), and the inflammatory response marker interleukin-6 (IL-6)., Material and Methods: Gingival fibroblasts isolated from a 30- and a 62-year-old donor were assayed for proliferation (1-7 days) and sensitivity to latex (positive control). Fibroblasts were indirectly exposed to Corega Ultra (cream), Corega powder and Fixodent Original for a 24 h period and assayed by XTT and Crystal Violet tests. The release of IL-6 and bFGF by exposed cells was determined by ELISA., Results: While cells from the young donor presented higher cell growth after 7 days, the sensitivity to increasing concentrations of latex extracts was very similar between young and older cells. Both XTT and CVDE detected no difference between the DA and the control group. All materials induced higher levels of IL-6 and bFGF compared to control. Cells from the older donor exposed to Corega Ultra released lower levels of cytokine and growth factor., Conclusions: All materials were considered non-cytotoxic, but affected cytokine and growth factor release. The biological differences found between fibroblasts from both donors could be due to individual or age-related factors. The authors suggest the use of cells from older donors on studies of dental products aimed at older patients, to better simulate their physiological response.

The application of in silico methods is increasing on toxicological risk prediction for human and environmental health. This work aimed to evaluate the performance of three in silico freeware models (OSIRIS v.2.0, LAZAR, and Toxtree) on the prediction of carcinogenicity and mutagenicity of thirty-eight volatile organic compounds (VOC) related to chemical risk assessment for occupational exposure. Theoretical data were compared with assessments available in international databases. Confusion matrices and ROC curves were used to evaluate the sensitivity, specificity, and accuracy of each model. All three models (OSIRIS, LAZAR and Toxtree) were able to identify VOC with a potential carcinogenicity or mutagenicity risk for humans, however presenting differences concerning the specificity, sensitivity, and accuracy. The best predictive performances were found for OSIRIS and LAZAR for carcinogenicity and OSIRIS for mutagenicity, as these softwares presented a combination of negative predictive power and lower risk of false positives (high specificity) for those endpoints. The heterogeneity of results found with different softwares reinforce the importance of using a combination of in silico models to occupational toxicological risk assessment., (Copyright © 2017 Elsevier Inc. All rights reserved.)

The epsilon toxin, produced by Clostridium perfringens, is responsible for enterotoxemia in ruminants and is a potential bioterrorism agent. In the present study, 15 regions of the toxin were recognized by antibodies present in the serum, with different immunodominance scales, and may be antigen determinants that can be used to formulate subunit vaccines., (Copyright © 2017 Sociedade Brasileira de Microbiologia. Published by Elsevier Editora Ltda. All rights reserved.)

Dental caries is an oral pathology associated with both lifestyle and genetic factors. The caries process can be influenced by salivary composition, which includes ions and proteins. Studies have described associations between salivary protein polymorphisms and dental caries experience, while others have shown no association with salivary proteins genetic variability. The aim of this study is to assess the influence of salivary protein polymorphisms on the risk of dental caries by means of a systematic review of the current literature. An electronic search was performed in PubMed, Scopus, and Virtual Health Library. The following search terms were used: "dental caries susceptibility," "dental caries," "polymorphism, genetics," "saliva," "proteins," and "peptides." Related MeSH headings and free terms were included. The inclusion criteria comprised clinical investigations of subjects with and without caries. After application of these eligibility criteria, the selected articles were qualified by assessing their methodological quality. Initially, 338 articles were identified from the electronic databases after exclusion of duplicates. Exclusion criteria eliminated 322 articles, and 16 remained for evaluation. Eleven articles found a consistent association between salivary protein polymorphisms and risk of dental caries, for proteins related to antimicrobial activity (beta defensin 1 and lysozyme-like protein), pH control (carbonic anhydrase VI), and bacterial colonization/adhesion (lactotransferrin, mucin, and proline-rich protein Db). This systematic review demonstrated an association between genetic polymorphisms and risk of dental caries for most of the salivary proteins.

Context: A novel root-filling material based on the incorporation of ultrafine alkaline bioactive glass particles (bioactive gutta-percha, [BGP]) was developed to work without sealer., Aim: In the present study, the objective was to verify the in vitro biological response to this material by assessing its cytocompatibility., Materials and Methods: Prototypes of BGP were compared to conventional gutta-percha (GP), dense polystyrene beads as a negative control and fragments of latex as a positive control. Extracts of each material were prepared according to ISO 10993-5:2009, and human osteoblast-like cells in primary culture were exposed to all extracts for 24 h. Cell viability was assayed sequentially for three different parameters: mitochondrial activity, membrane integrity, and cell density., Statistical Analysis Used: Nonparametric analysis (using Kruskal-Wallis test combined with post hoc Dunn's test) was performed for comparison among groups, with significance established at 5%., Results: BGP reduced mitochondrial activity to 62% of control, but presented no toxicity on membrane integrity and proliferation assays. BGP effect on metabolism was dose-dependent and reduced to acceptable levels with dilution., Conclusion: The novel GP material presented slight dose-dependent effects on cell metabolism but did not affect cell survival., Competing Interests: There are no conflicts of interest.