SARS-CoV-2 ferritin nanoparticle vaccines produce hyperimmune equine sera with broad sarbecovirus activity.

The rapid emergence of SARS-CoV-2 variants of concern (VoC) and the threat of future zoonotic sarbecovirus spillover emphasizes the need for broadly protective next-generation vaccines and therapeutics. We utilized SARS-CoV-2 spike ferritin nanoparticle (SpFN), and SARS-CoV-2 receptor binding domain ferritin nanoparticle (RFN) immunogens, in an equine model to elicit hyperimmune sera and evaluated its sarbecovirus neutralization and protection capacity. Immunized animals rapidly elicited sera with the potent neutralization of SARS-CoV-2 VoC, and SARS-CoV-1 pseudoviruses, and potent binding against receptor binding domains from sarbecovirus clades 1b, 1a, 2, 3, and 4. Purified equine polyclonal IgG provided protection against Omicron XBB.1.5 virus in the K18-hACE2 transgenic mouse model. These results suggest that SARS-CoV-2-based nanoparticle vaccines can rapidly produce a broad and protective sarbecovirus response in the equine model and that equine serum has therapeutic potential against emerging SARS-CoV-2 VoC and diverse sarbecoviruses, presenting a possible alternative or supplement to monoclonal antibody immunotherapies.

Use of autologous micrografts associated with xenogeneic anorganic bone in vertical bone augmentation procedures with Barbell Technique®.

INTRODUCTION: Bidirectional vertical ridge augmentation in the posterior maxilla is very challenging. PURPOSE: To evaluate the regenerative potential of micrografts, derived from periosteum or bone tissue, added to an anorganic xenograft in vertical reconstruction of the posterior maxilla, by a prospective, controlled study. MATERIALS AND METHODS: After clinical selection and the analysis of CBCT scans, 24 posterior maxillary sites, in 19 patients, were treated by using Barbell Technique®. Sites requiring both inlay and onlay reconstruction were enrolled in the study. In the Control Group (CG, n = 8), a xenograft was used in the inlay site and for the onlay site, a 1:1 mix of xenograft and an autograft was used. In Test Group 1 (TG1, n = 8), both inlay and onlay sites were grafted with the xenograft associated with the micrografts derived from periosteum. In Test Group 2 (TG2, n = 8), both inlay and onlay sites were grafted with the xenograft associated with the micrografts derived from bone. Six months after the procedures, CBCT scans were obtained, and bone biopsy samples were harvested during implant placement surgery. The bone specimens were analyzed histomorphometrically, by measuring the percentages of vital mineralized tissue (VMT), non vital mineralized tissue (NVMT) and non mineralized tissue (NMT). Immunohistochemically, the levels of VEGF were categorized by a score approach. RESULTS: Histomorphometric analysis revealed, for the inlay grafts, no significant difference among the groups for VMT, NVMT and NMT. However, for onlay grafts, CG achieved a higher amount of VMT in comparison with TG2, and the opposite occurred for NMT values. In this regard, no statistical difference was observed between CG and TG1. Concerning immunohistochemistry, the VEGF values for CG and TG1 were slightly higher than those obtained by TG2 for both inlay and onlay grafts, but without statistical significance. CBCT analysis showed a similar level of gain for all groups, for both inlay and onlay bone augmentation sites. Clinically, one implant (in CG) within a total of 50 implants installed, had early failure and was replaced after 3 months. All patients received implant supported prosthesis. CONCLUSION: This study indicated that the clinical use of micrograft derived from periosteum may have some potential to increase bone formation in onlay reconstructions, unlike the micrograft derived from bone tissue.

Age- and Sex-Associated Wnt Signaling Dysregulation is Exacerbated from the Early Stages of Neuropathology in an Alzheimer's Disease Model.

Emerging studies suggest that Wnt signaling is dysregulated in the brains of AD patients, suggesting that this pathway may also contribute to disease progression. However, it remains to be determined whether alterations in the Wnt pathway are the cause or consequence of this disease and which elements of Wnt signaling mainly contribute to the appearance of AD histopathological markers early in disease compared to what occurs during normal aging. The present study aimed to describe the status of several canonical Wnt pathway components and the expression of the AD marker p-tau in the hippocampi of female and male 3xTg-AD mice during disease progression compared to those during normal aging. We analyzed the levels of the canonical Wnt components Wnt7a, Dkk-1, LRP6 and GSK3ß as well as the levels of p-tau and BDNF at 3, 6, 9-12 and 18 months of age. We found a gradual increase in Dkk-1 levels during aging prior to Wnt7a and LRP5/6 depletion, which was strongly exacerbated in 3xTg-AD mice even at young ages and correlated with GSK3ß activation and p-tau-S202/Thr205 expression. Dkk-1 upregulation, as well as the level of p-tau, was significantly greater in females than in males. Our results suggest that Dkk-1 upregulation is involved in the expression of several features of AD at early stages, which supports the possibility of positively modulating the canonical Wnt pathway as a therapeutic tool to delay this disease at early stages.

Effects of photobiomodulation on different phases of in vitro osteogenesis.

The present study aimed to evaluate the effect of photobiomodulation therapy (PBM) on different stages of osteogenesis in vitro. For this, osteoblastic-like cells (Saos-2 cell lineage) were irradiated in two different periods: during the Proliferation phase (PP; from the second to the fourth day) and during the Differentiation phase (DP; from the seventh to the ninth day). The energy density used in the study was 1.5 J/ cm2. The following parameters were evaluated: 1) quantification of collagen type 1 (COL 1), osteopontin (OPN), and bone morphogenetic protein 2 (BMP-2); 2) quantification of alkaline phosphatase (ALP) activity; and 3) quantification of  extracellular matrix (ECM) mineralization. Non-irradiated cultures were used as controls. The data were analyzed using the Student's t-test or one-way ANOVA, considering a significance level of 5%. The results indicated that COL 1 and BMP-2 quantification was higher in Saos-2 irradiated during the DP in relation to the control group at day 10 (p < 0.05). No differences were observed for other comparisons at this time point (p > 0.05). OPN expression was greater in PP compared with the other experimental groups at day 10 (p < 0.05). Irradiation did not affect ALP activity in Saos-2 regardless of the exposure phase and the time point evaluated (p > 0.05). At day 14, ECM mineralization was higher in Saos-2 cultures irradiated during the DP in relation to the PP (p < 0.05). In conclusion, the results suggested that the effects of PBM on osteoblastic cells may be influenced by the stage of cell differentiation.

Differences in neutralizing antibody sensitivities and envelope characteristics indicate distinct antigenic properties of Nigerian HIV-1 subtype G and CRF02_AG.

The magnitude of the HIV-1 epidemic in Nigeria is second only to the subtype C epidemic in South Africa, yet the subtypes prevalent in Nigeria require further characterization. A panel of 50 subtype G and 18 CRF02_AG Nigerian HIV-1 pseudoviruses (PSV) was developed and envelope coreceptor usage, neutralization sensitivity and cross-clade reactivity were characterized. These PSV were neutralized by some antibodies targeting major neutralizing determinants, but potentially important differences were observed in specific sensitivities (eg. to sCD4, MPER and V2/V3 monoclonal antibodies), as well as in properties such as variable loop lengths, number of potential N-linked glycans and charge, demonstrating distinct antigenic characteristics of CRF02_AG and subtype G. There was preferential neutralization of the matched CRF/subtype when PSV from subtype G or CRF02_AG were tested using pooled plasma. These novel Nigerian PSV will be useful to study HIV-1 CRF- or subtype-specific humoral immune responses for subtype G and CRF02_AG.

A Minimally Invasive Surgical Procedure to Harvest Palate Periosteum as a Source of Mesenchymal Stromal/Stem Cells for Bone Tissue Engineering.

The aim of this study is to validate a minimally invasive surgical procedure to harvest palate periosteum as a source of tissue for mesenchymal stromal/stem cells. We performed a standardized procedure to harvest the palate periosteum in ten subjects, which consisted of a 3 mm disposable punch and a Molt periosteal elevator to harvest a small full-thickness fragment of soft tissue at the hard palate area, between the upper bicuspids, 3 to 4 mm apical to the cement enamel junction. The one-third inner portion was fragmented, and following standard cell culture procedures, the adherent cells were cultured for three passages, after obtaining 70-90% confluence. Cell morphology analysis, flow cytometry analysis, and viability and osteogenic differentiation assays were performed. In all 10 cases, uneventful healing was observed, with no need for analgesic intake. The evaluation of cell morphology showed elongated spindle-shaped cells distributed in woven patterns. A high viability range was verified as well as an immunophenotype compatible with mesenchymal stem cell lineage. The differentiation assay showed the potential of the cells to differentiate into the osteogenic lineage. These results demonstrate that the minimally invasive proposed surgical technique is capable of supplying enough periosteum source tissue for stem cell culture and bone tissue engineering.

The cardiovascular effects of large hiatal hernias: a narrative review of cases and studies.

Hiatal hernia (HH) is a common disease in the general population. It is often asymptomatic, but if it does present clinical manifestations, these are usually gastrointestinal. Gastroesophageal reflux is the main symptom that accompanies it. Depending on the severity of the hernia, it is classified into several subtypes from I-IV. Especially, IV type (giant HH) can lead to various cardiopulmonary symptoms with several degrees of severity. It is necessary to keep this possibility in mind among the various differential diagnoses that may occur in this clinical setting. The current paper aims to review the literature on classic and novel information on the HH - cardiovascular system relationship. Epidemiological data, physiological aspects of the heart compressed by HH, cardiovascular symptoms, electrocardiographic changes, echocardiographic alterations and clinical implications are discussed.

Normally, the stomach and the heart are not in direct contact because they are in different cavities, the thorax and the abdomen, respectively. When part of the stomach moves toward the chest through the diaphragm, we say there is a hiatal hernia (HH). Most of the time the HH symptoms are mild and clearly digestive. In severe cases, surgical repair of the HH is required. Even in these circumstances, digestive symptoms continue to be the most frequent. However, some patients present cardiovascular symptoms and few or no digestive symptoms. This easily creates diagnostic confusion, which leads to incorrect treatments and unnecessary expenses. In extreme cases, as seen in giant HH, the degree of cardiovascular involvement is very serious. There are documented cases that have suffered cardiac arrest, arrhythmias of different types and symptoms like classic acute myocardial infarction. It is required that clinical doctors and surgeons are aware that this complication exists. Only with this in mind can a timely diagnosis be achieved. Some emergency measures have been saving, gastric decompression with a tube being the most important. The main mechanism that explains the serious cardiovascular consequences of giant HH is cardiac compression. The dissemination of this knowledge can help save lives.

[Associated factors with the occurrence of in-hospital cardiac arrest in patients admitted to internal medicine wards for non-cardiovascular causes]. / Factores asociados a la ocurrencia de paro cardiaco intrahospitalario en pacientes ingresados en salas de medicina interna por causas no cardiovasculares.

BACKGROUND AND OBJECTIVE: In-hospital cardiac arrest (IHCA) has a low survival rate, so it is essential to recognize the cases with the highest probability of developing it. The aim of this study is to identify factors associated with the occurrence of IHCA. MATERIAL AND METHODS: A single-center case-control study was conducted including 65 patients admitted to internal medicine wards for non-cardiovascular causes who experienced IHCA, matched with 210 admitted controls who did not present with IHCA. RESULTS: The main reason for admission was pneumonia. The most prevalent comorbidity was arterial hypertension. Four characteristics were strongly and independently associated with IHCA presentation, these are electrical left ventricular hypertrophy (LVH) (OR: 13.8; 95% IC: 4.7-40.7), atrial fibrillation (OR: 9.4: 95% CI: 4.3-20.6), the use of drugs with known risk of torsades de pointes (OR: 2.7; 95% CI: 1.3-5.5) and the combination of the categories known risk plus conditional risk (OR: 17.1; 95% CI: 6.7-50.1). The first two detected in the electrocardiogram taken at the time of admission. CONCLUSION: In admitted patients for non-cardiovascular causes, the use of drugs with a known risk of torsades de pointes, as well as the detection of electrical LVH and atrial fibrillation in the initial electrocardiogram, is independently associated with a higher probability of suffering a IHCA.

The effect of root canal preparation tapers on planktonic bacteria and biofilm reduction in the apical third: A correlative microtomography and microbiological laboratory study.

AIM: To evaluate the influence of different preparation tapers on the reduction in planktonic bacteria and biofilms of Enterococcus faecalis and Candida albicans in the apical third (4 mm) of the mesial roots of mandibular molars, correlating decontamination with canal shape. METHODOLOGY: After microtomography analysis for morphological standardization of the canals, 48 mandibular molar roots, each containing two canals (96 canals), were contaminated with E. faecalis and C. albicans and divided into four groups (n = 11) for canal instrumentation using ProDesign Logic 2 files with different tapers G (.03): # 25.03; G (.04): # 25.04; G (.05): # 25.05; and G (.06): # 25.06 and irrigation with 2.5% sodium hypochlorite. Four roots were examined under a scanning electron microscope (SEM) to qualitatively assess biofilm formation. Eight roots were used as the negative control group (samples were not contaminated). Bacteriological samples were taken exclusively from the apical third of the roots before and after chemical-mechanical preparation and bacterial counts were determined (CFU/mL). The final micro-CT scan was used to quantify the volume variation and unprepared canal area in the apical third. Statistical analysis was performed using the Kruskal-Wallis, Student-Newman-Keuls and Wilcoxon tests for analysis of microbiological data. anova and the Tukey or Games-Howell test were used for analysis of micro-CT data and Spearman's test for correlations (α = 5%). RESULTS: All groups showed a significant reduction in bacteria (p < .05), with no statistically significant difference between groups. There was no significant difference in per cent volume increase between groups. The unprepared area (Δ%) was affected by the file used (p = .026) and was significantly lower for G (.06) compared to G (.03). There was no statistically significant correlation among bacterial reduction, volume and unprepared area (p > .05). CONCLUSION: The different preparation tapers influenced root canal shaping in the apical third but did not improve decontamination in this region.

Diverse array of neutralizing antibodies elicited upon Spike Ferritin Nanoparticle vaccination in rhesus macaques.

The repeat emergence of SARS-CoV-2 variants of concern (VoC) with decreased susceptibility to vaccine-elicited antibodies highlights the need to develop next-generation vaccine candidates that confer broad protection. Here we describe the antibody response induced by the SARS-CoV-2 Spike Ferritin Nanoparticle (SpFN) vaccine candidate adjuvanted with the Army Liposomal Formulation including QS21 (ALFQ) in non-human primates. By isolating and characterizing several monoclonal antibodies directed against the Spike Receptor Binding Domain (RBD), N-Terminal Domain (NTD), or the S2 Domain, we define the molecular recognition of vaccine-elicited cross-reactive monoclonal antibodies (mAbs) elicited by SpFN. We identify six neutralizing antibodies with broad sarbecovirus cross-reactivity that recapitulate serum polyclonal antibody responses. In particular, RBD mAb WRAIR-5001 binds to the conserved cryptic region with high affinity to sarbecovirus clades 1 and 2, including Omicron variants, while mAb WRAIR-5021 offers complete protection from B.1.617.2 (Delta) in a murine challenge study. Our data further highlight the ability of SpFN vaccination to stimulate cross-reactive B cells targeting conserved regions of the Spike with activity against SARS CoV-1 and SARS-CoV-2 variants.

Antibody targeting of conserved sites of vulnerability on the SARS-CoV-2 spike receptor-binding domain.

Given the continuous emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants of concern (VoCs), immunotherapeutics that target conserved epitopes on the spike (S) glycoprotein have therapeutic advantages. Here, we report the crystal structure of the SARS-CoV-2 S receptor-binding domain (RBD) at 1.95 Å and describe flexibility and distinct conformations of the angiotensin-converting enzyme 2 (ACE2)-binding site. We identify a set of SARS-CoV-2-reactive monoclonal antibodies (mAbs) with broad RBD cross-reactivity including SARS-CoV-2 Omicron subvariants, SARS-CoV-1, and other sarbecoviruses and determine the crystal structures of mAb-RBD complexes with Ab246 and CR3022 mAbs targeting the class IV site, WRAIR-2134, which binds the recently designated class V epitope, and WRAIR-2123, the class I ACE2-binding site. The broad reactivity of class IV and V mAbs to conserved regions of SARS-CoV-2 VoCs and other sarbecovirus provides a framework for long-term immunotherapeutic development strategies.

Reducing physical restraints in pediatrics: A quality improvement mixed-methods analysis of implementing a clinical debriefing process after behavioural health emergencies in a Children's Hospital.

INTRODUCTION: An increasing number of pediatric patients with mental and behavioral health (MBH) conditions present to Emergency Department (ED) and inpatient settings with behavioral events that require physical restraint (PR). PR usage is associated with adverse outcomes. Clinical debriefing (CD) programs have been associated with improved performance but have not been studied in this population. After implementing an MBH-CD program in our Children's Hospital, we aimed to decrease the baseline (7/2018-3/2021) rate of a second PR episode (2PR) by 50 % in the ED and inpatient settings over two years. METHODS: A multidisciplinary team implemented an MBH-CD process in April 2021 for hospital teams to use immediately after behavioral events. We included patients ≤18 years old, with an ED or inpatient discharge MBH diagnosis, between July 2018 and June 2023. Pre- and post-implementation secondary outcomes included the ED median duration of PR and the ED PR time per 1000 h of ED care. ED and inpatient mean length of stay (LOS) and mean monthly visits (MMV) in pre- and post-implementation were also compared. Qualitative analysis identified major themes. RESULTS: Post-implementation, the ED significantly decreased 2PR rate by 67 %; in inpatients, no significant change was demonstrated. Median duration of ED PR decreased from 112 to 71 min (p = 0.006) and ED PR time significantly decreased by 82 % (14.8 to 2.7 h per 1000 h). In the post-implementation period, mean LOS (ED and inpatient) and MMV (ED only) were significantly higher. Fifty-one percent of 494 behavioral alerts were debriefed. Median debriefing duration was 6 min (IQR 4,10). Common themes included cooperation and coordination (23 %) and clinical standards (14 %). DISCUSSION: Clinical debriefing implementation was associated with significant improvement in ED patient outcomes. Inpatient outcomes were unchanged, but debriefings in both settings should enable frontline teams to continuously identify opportunities to improve future outcomes.

In vitro cytotoxicity of resin cement and its influence on the expression of antioxidant genes.

AIM: This study evaluated cytotoxicity and antioxidant gene expression of resin cements on human gingival fibroblasts (hGF). MATERIALS AND METHOD: RelyX Ultimate™(RXU), Variolink™II(VLII), and RelyXU200™(RXU200) resin cements were incubated with culture medium for 24 h to obtain eluates. Then, the eluates were applied over hGF to assess cell viability at 24 h, 48 h, and 72 h and antioxidant gene expression at 24 h. hGF cultures non-exposed to the eluates were used as Control. Data were submitted to ANOVA and Bonferroni tests (α≤0.05). RESULTS: RXU and RXU200 reduced the number of viable cells in 24 h. Longer exposure to cement extracts caused cell death. Gene expression showed peroxiredoxin 1 (PRDX1) induction by all resin cement types, and superoxide dismutase 1 (SOD1) induction by RXU200 and VLII. Moreover, RXU200 induced not only PRDX1 and SOD1, but also glutathione peroxidase 1 (GPX1), catalase (CAT), and glutathione synthetase (GSS). CONCLUSIONS: All resin cements showed toxicity, and induced antioxidant genes in hGF. Antioxidant gene induction is at least partly associated with cytotoxicity of tested cements to oxidative stress experience.

OBJETIVO: O objetivo deste estudo foi avaliar a toxicidade dos cimentos resinosos Rely X Ultimate 2, Rely X U200 e Variolink II, bem como sua influência na expressão de genes antioxidantes em fibroblastos gengivais humanos. Materiais e Método: Corpos de prova de cada cimento foram colocados em meio de cultura por 24 h e os extratos correspondentes foram aplicados aos fibroblastos. A viabilidade celular foi avaliada após 24, 48 e 72 h de exposição pelo ensaio de exclusão do azul de tripano e MTT. A expressão gênica foi avaliada por PCR quantitativo após 24 h de exposição aos extratos. Estes parâmetros foram comparados aos das células não expostas aos cimentos. Os dados foram submetidos ao teste ANOVA, seguido pelo pós-teste de Bonferroni (a≤0.05). RESULTADOS: Os resultados demonstraram que todos os cimentos promoveram redução do número de células viáveis e da atividade mitocondrial nos períodos de 48 e de 72 h (p< 0,01), sendo que o Variolink II apresentou o menor efeito e os cimentos Rely X Ultimate e Rely X U200 promoveram similarmente os maiores efeitos. A análise de expressão gênica evidenciou influência significativa em todos os cimentos avaliados sobre os níveis de transcritos de PRDX1, SOD1, GPX1 e GSS (p> 0,05), com um aumento considerável no Rely X U200. Conclusão: A indução de genes antioxidantes está, pelo menos em parte, associada à citotoxicidade dos cimentos testados para a experiência de estresse oxidativo.

Surface topography of resorbable porcine collagen membranes, and their effect on early osteogenesis: An in vitro study.

OBJECTIVE: Guided tissue regeneration (GTR) is based on the use of different membranes that function as sealants and barriers in specific clinical situations. Among the several tissue production methods and origins, resorbable porcine-derived membranes are the most commonly used. Because these membranes are so diverse, and have several different clinical applications, doubts linger as to their effect in stimulating osteogenesis. The objective of this study was to make an in vitro evaluation of the viability and differentiation of osteoblastic cells cultured on the surface of the following collagen membranes: Jason® (Botiss Biomaterials), Collprotect® (Botiss Biomaterials), and Bio-Gide® (Geistlich). MATERIAL AND METHODS: Fragments of the 3 resorbable collagen membranes (5 × 5 mm) were used, and pre-osteoblastic SAOS-2 cells (ATCC, USA) were plated on their porous surfaces. Evaluation of the membranes was performed at 3, 5 and 7 days, considering the following parameters: (1) topographic analysis of the different surfaces by scanning electron microscope; (2) cellular viability by MTT, (3) quantification of type I collagen and osteopontin by Elisa. The quantitative analyses were carried out using a significance level of 5%. RESULTS: Collprotect® and Jason® membranes presented a rough surface with an irregular aspect on both sides, while double-layered Bio-Gide® had one layer with a smooth surface and the other with a rough surface along each respective length. The viability assays revealed that the cells cultured the cells grown on Collprotect® showed higher viability than those grown in Bio-Gide® or Jason®, especially after 5 and 7 days. After 3 and 5 days, evaluation of type I collagen showed that the cells plated on the Jason® and Collprotect® surfaces had greater collagen secretion than those plated on BioGide®. After 7 days, an increase in osteopontin levels was observed when the cells were plated on all the experimental membranes, compared with the control group. CONCLUSION: All the tested membranes were suitable for use in GTR clinical procedures. Their indication in specific regenerative cases depends on the mechanical and biological properties of their originating tissues, thus enabling better results and assertive choices by dental professionals.

Conditioned Media from Human Pulp Stem Cell Cultures Improve Bone Regeneration in Rat Calvarial Critical-Size Defects.

The aim of this study was to test whether lyophilized conditioned media from human dental pulp mesenchymal stem cell cultures promote the healing of critical-size defects created in the calvaria of rats. Prior to the surgical procedure, the medium in which dental pulp stem cells were cultured was frozen and lyophilized. After general anesthesia, an 8 mm diameter bone defect was created in the calvaria of twenty-four rats. The defects were filled with the following materials: xenograft alone (G1) or xenograft associated with lyophilized conditioned medium (G2). After 14 or 42 days, the animals were euthanized, and the specimens processed for histologic and immunohistochemical analysis. Bone formation at the center of the defect was observed only in the G2 at 42 days. At both timepoints, increased staining for VEGF, a marker for angiogenesis, was observed in G2. Consistent with this, at 14 days, G2 also had a higher number of blood vessels detected by immunostaining with an anti-CD34 antibody. In conclusion, conditioned media from human dental pulp mesenchymal stem cell cultures had a positive effect on the regenerative process in rat critical-size bone defects. Both the formation of bone and enhancement of vascularization were stimulated by the conditioned media.

Sema4D is diminished in leukocyte platelet-rich fibrin and impairs pre-osteoblastic MC3T3-E1 cells' functionality.

OBJECTIVE: Semaphorin 4D (Sema4D) is a coupling factor expressed on osteoclasts that may hinder osteoblast differentiation. Since the leukocyte platelet-rich fibrin (L-PRF) membrane promotes growth factor concentration, this study aims to quantify the amount of Sema4D in L-PRF membranes, and analyze the impact of Sema4D on osteoblast cell function in vitro. DESIGN: Enzyme-linked immunosorbent assay (ELISA) was used to quantify the levels of Sema4D in both L-PRF and whole blood (serum). To analyze the impairment of Sema4D on osteoblasts, MC3T3-E1 cells were induced to osteogenic differentiation and exposed to Sema4D ranging from 10 to 500 ng/ml concentrations. The following parameters were assayed: 1) cell viability by MTT assay after 24, 48, and 72 h; 2) matrix mineralization by Alizarin Red staining after 14 days, 3) Runt-related transcription factor 2 (RUNX-2), osteocalcin (OCN), osteonectin (ONC), bone sialoprotein (BSP) and alkaline phosphatase (ALP) gene expression by qPCR. For all data, the significance level was set at 5%. RESULTS: The amount of Sema4D in the whole blood (serum) was higher than in L-PRF. Osteoblasts exposed to Sema4D at all tested concentrations exhibited a decrease in matrix mineralization formation as well in RUNX-2, OCN, ONC, BSP, and ALP gene expression (p < 0.05). CONCLUSION: The presence of Sema4D, a molecule known for suppressing osteoblast activity, diminishes within L-PRF, enhancing its ability to facilitate bone regeneration.

In vitro cytotoxicity of resin cement and its influence on the expression of antioxidant genes / Avaliação in vitro da toxicidade de cimentos resinosos e sua influência na expressão de genes antioxidantes em fibroblastos humanos

ABSTRACT Aim: This study evaluated cytotoxicity and antioxidant gene expression of resin cements on human gingival fibroblasts (hGF). Materials and Method: RelyX Ultimate™(RXU), Variolink™II(VLII), and RelyXU200™(RXU200) resin cements were incubated with culture medium for 24 h to obtain eluates. Then, the eluates were applied over hGF to assess cell viability at 24 h, 48 h, and 72 h and antioxidant gene expression at 24 h. hGF cultures non-exposed to the eluates were used as Control. Data were submitted to ANOVA and Bonferroni tests (α≤0.05). Results: RXU and RXU200 reduced the number of viable cells in 24 h. Longer exposure to cement extracts caused cell death. Gene expression showed peroxiredoxin 1 (PRDX1) induction by all resin cement types, and superoxide dismutase 1 (SOD1) induction by RXU200 and VLII. Moreover, RXU200 induced not only PRDX1 and SOD1, but also glutathione peroxidase 1 (GPX1), catalase (CAT), and glutathione synthetase (GSS). Conclusions: All resin cements showed toxicity, and induced antioxidant genes in hGF. Antioxidant gene induction is at least partly associated with cytotoxicity of tested cements to oxidative stress experience.

RESUMO Objetivo: O objetivo deste estudo foi avaliar a toxicidade dos cimentos resinosos Rely X Ultimate 2, Rely X U200 e Variolink II, bem como sua influência na expressão de genes antioxidantes em fibroblastos gengivais humanos. Materiais e Método: Corpos de prova de cada cimento foram colocados em meio de cultura por 24 h e os extratos correspondentes foram aplicados aos fibroblastos. A viabilidade celular foi avaliada após 24, 48 e 72 h de exposição pelo ensaio de exclusão do azul de tripano e MTT. A expressão gênica foi avaliada por PCR quantitativo após 24 h de exposição aos extratos. Estes parâmetros foram comparados aos das células não expostas aos cimentos. Os dados foram submetidos ao teste ANOVA, seguido pelo pós-teste de Bonferroni (a≤0.05). Resultados: Os resultados demonstraram que todos os cimentos promoveram redução do número de células viáveis e da atividade mitocondrial nos períodos de 48 e de 72 h (p < 0,01), sendo que o Variolink II apresentou o menor efeito e os cimentos Rely X Ultimate e Rely X U200 promoveram similarmente os maiores efeitos. A análise de expressão gênica evidenciou influência significativa em todos os cimentos avaliados sobre os níveis de transcritos de PRDX1, SOD1, GPX1 e GSS (p> 0,05), com um aumento considerável no Rely X U200. Conclusão: A indução de genes antioxidantes está, pelo menos em parte, associada à citotoxicidade dos cimentos testados para a experiência de estresse oxidativo.

Targeting the Spike Receptor Binding Domain Class V Cryptic Epitope by an Antibody with Pan-Sarbecovirus Activity.

Novel therapeutic monoclonal antibodies (MAbs) must accommodate comprehensive breadth of activity against diverse sarbecoviruses and high neutralization potency to overcome emerging variants. Here, we report the crystal structure of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) receptor binding domain (RBD) in complex with MAb WRAIR-2063, a moderate-potency neutralizing antibody with exceptional sarbecovirus breadth, that targets the highly conserved cryptic class V epitope. This epitope overlaps substantially with the spike protein N-terminal domain (NTD) -interacting region and is exposed only when the spike is in the open conformation, with one or more RBDs accessible. WRAIR-2063 binds the RBD of SARS-CoV-2 WA-1, all variants of concern (VoCs), and clade 1 to 4 sarbecoviruses with high affinity, demonstrating the conservation of this epitope and potential resiliency against variation. We compare structural features of additional class V antibodies with their reported neutralization capacity to further explore the utility of the class V epitope as a pan-sarbecovirus vaccine and therapeutic target. IMPORTANCE Characterization of MAbs against SARS-CoV-2, elicited through vaccination or natural infection, has provided vital immunotherapeutic options for curbing the COVID-19 pandemic and has supplied critical insights into SARS-CoV-2 escape, transmissibility, and mechanisms of viral inactivation. Neutralizing MAbs that target the RBD but do not block ACE2 binding are of particular interest because the epitopes are well conserved within sarbecoviruses and MAbs targeting this area demonstrate cross-reactivity. The class V RBD-targeted MAbs localize to an invariant site of vulnerability, provide a range of neutralization potency, and exhibit considerable breadth against divergent sarbecoviruses, with implications for vaccine and therapeutic development.

Antibacterial nanocomposite of chitosan/silver nanocrystals/graphene oxide (ChAgG) development for its potential use in bioactive wound dressings.

An adequate wound dressing reduces time of healing, provides cost-effective care, thereby improving patients' quality life. An antimicrobial bioactivity is always desired, for that reason, the objective of this work is to design an antimicrobial nanocomposite of chitosan/silver nanocrystals/graphene oxide (ChAgG). ChAgG nanostructured composite material is composed of chitosan from corn (Ch), and silver nanocrystals from garlic (Allium sativum). The nanocomposite obtained is the result of a series of experiments combining the graphene oxide (GrOx) with two members of the Amaryllidaceae family; garlic and onion (Allium cebae), which contain different sulfur materials. The characterization arrays confirmed the successful production of silver crystal, graphene oxidation and the blending of both components. The role of the chitosan as a binder between graphene and silver nanocrystals is proved. Moreover, the study discusses garlic as an optimal source that permits the synthesis of silver nanocrystals (AgNCs) (⁓ 2 to 10 nm) with better thermal and crystallinity properties. It was also confirmed the successful production of the ChAgG nanocomposite. Escherichia coli and Staphylococcus aureus were used to demonstrate the antibacterial bioactivity and L-929 fibroblast cells were utilized to visualize their biocompatibility. The proposed ChAgG nanomaterial will be useful for functionalizing specific fiber network that represents current challenging research in the fabrication of bioactive wound dressings.