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Objective: This study aimed to compare the salivary protein profile in individuals with Type 2 Diabetes Mellitus (DM2) and periodontitis and their respective controls., Methods: Eighty participants were included in the study. The four groups were formed by individuals with DM2 and periodontitis (DM2 + P, n = 20), DM2 without periodontitis (DM2, n = 20), periodontitis without DM2 (P, n = 20) and individuals without periodontitis and without DM2 (H, n = 20). Periodontal clinical examinations were performed and unstimulated saliva was collected. Proteomic analysis was performed by shotgun mass spectrometry. The results were obtained by searching the Homo sapiens database of the UniProt catalog., Results: A total of 220 proteins were identified in saliva samples. In the comparison between DM2 + P and DM2 groups, 27 proteins were up-regulated [e.g. S100-A8 was 6 times up-regulated (humoral immune response pathway)]. The DM2 + P and P groups had 26 up-regulated proteins [e.g. Immunoglobulin lambda constant 7 more than 2 times up-regulated (complement activation pathway)]. The non-DM2 groups (P and H) presented 22 up-regulated proteins [e.g. Glyceraldehyde-3-phosphate dehydrogenase more than 2 times up-regulated (Peptidyl-cysteine S-nitrosylation pathway)]. The groups without periodontitis (DM2 and H) showed 23 were up-regulated proteins [e.g. Hemoglobin subunit alpha that was more than 10 times up-regulated (cellular oxidant detoxification pathway)]., Conclusion: The presence of DM2 and periodontitis significantly impacts the salivary proteome. Our proteomic analysis demonstrated that changes in the S100 family proteins (S100A8 and S100 A9) are highly related to the presence of DM2 and periodontitis., Clinical Relevance: Diabetes Mellitus (DM) and periodontitis are highly prevalent chronic diseases that present a wide variety of signs and symptoms. They present a bidirectional relationship, where patients with DM have a higher prevalence and severity of periodontitis, and patients with periodontitis have a higher prevalence of DM, worse glycemic control, and more diabetic complications. Diagnosing periodontitis requires specific clinical examinations, which require a highly trained operator. In this study, we used high throughput proteomics in order to evaluate non-invasive biomarkers for periodontitis in type 2 DM subjects. The results can contribute to earlier, more accurate, and less costly diagnosis of periodontitis in diabetic subjects, enabling better diabetes control., Competing Interests: Declarations. Ethics approval and consent to participate: This observational clinical study was approved by the University of Taubaté ethics committee (CAAE: 19319219.8.0000.5501). Eligible patients willing to participate signed an informed consent form after receiving oral and written information. Competing interests: The authors declare no competing interests., (© 2025. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Paternal exposure to environmental challenges is critical for the offspring's future health, and the transmission of acquired traits through generations increases the risk of offspring developing diseases. Ivermectin (IVM) is widely used in veterinary and human medicine to treat parasitosis. Our previous studies showed that IVM acute administration induced disorganization of the germinal epithelium and could cause damage to sperm production. Thus, this study investigated the effects of paternal exposure to repeated high ivermectin doses on paternal testis histology. After mating, their pups' development and sexual behavior in adult rats were examined. Method: Two groups of male rats were treated with IVM or its vehicle once a week for three weeks. We observed these males' body weight, organs and testis histology, and testosterone levels. These rats were mated with females without any treatment: the reproductive performance, the offspring development, and the male and female sexual behavior observed in adulthood. Relative to controls, the IVM paternal testis histology showed hypertrophy and hyperplasia of Leydig cells and increased diameter of the seminiferous tubules-no impairment in reproductive performance. In males and females, the physical and reflexes were modified. In adult age, female rats of the IVM group showed reduced sexual behavior and sexual preferences for the same sex, while male sexual behavior was not altered. Thus, it is possible that paternal exposure to IVM interfered with pups' hormonal and growth factors during development and in adult age. Further studies are needed to explore IVM transgenerational effects identifying possible mechanisms underpinning behavioral effects., Competing Interests: Declaration of Competing Interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Maria Martha Bernardi reports financial support was provided by Paulista University. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper, (Copyright © 2024 Elsevier Inc. All rights reserved.)

The action of Corepressor for Element Silencing Transcription Factor 1 (CoREST) is primarily related to neural fate decisions. However, the molecular mechanisms linking neuroinflammation to the histone modifying complex remain unclear. CoREST is a hub for several cofactors that play important roles in epigenetic remodeling and transcriptional regulation. It allows us to question their functions during the inflammatory response in the Central Nervous System. The impact of LPS-induced neuroinflammation on the transcriptional epigenetic control of members with CoRest repressive complex in the hippocampus was investigated. Characterizing the basal transcriptional profile in the hippocampus of members with CoREST repressive complex showed that the Rcor3 is the most expressed gene, and the Rcor2 is the least expressed one. It was also demonstrated that the levels of Lsd1, CoREST1 (Rcor1), and CoREST2 (Rcor2) transcripts increased in the hippocampus after LPS i.p. administration, while for CoREST3 (Rcor3), no significant difference was observed. A significant increase was noticed in the percentages of the 5-meC mark (Hypermethylation) for the Rcor1 and Lsd1 genes with a positive Pearson correlation between methylation and expression. However, the correlation was directly proportional, ruling out DNA methylation as the main mechanism in transcriptional control., Competing Interests: Declarations Competing interests The authors declare no competing interests. Ethical statements All experiments were conducted in accordance with the guidelines of the Animal Research Ethics Committee (PP00760/CEUA) of the Federal University of Santa Catarina, Florianópolis, SC, Brazil., (© 2024. The Author(s), under exclusive licence to Springer Nature B.V.)

In light of the complex origins of ectopic vascular calcification and its significant health implications, this study offers a comprehensive exploration of the molecular dynamics governing vascular smooth muscle cells (VSMCs). Focusing on epigenetic modulation, we investigate the transition from a contractile to a calcifying phenotype in VSMCs, with an emphasis on understanding the role of SIRT1. For this purpose, a single batch of human aortic SMCs, used at a specified passage number to maintain consistency, was subjected to calcium and phosphate overload for up to 72 h. Our findings, validated through RT q-PCR, Western blot, immunofluorescence, and DNA methylation analyses, reveal a complex interplay between acetyltransferases and deacetylases during this phenotypic transition. We highlight HAT1A's critical role in histone acetylation regulation and the involvement of HDACs, as evidenced by subcellular localization studies. Moreover, we demonstrate the modulation of SIRT1 expression, a class III deacetylase, during VSMC calcification, underscoring the influence of DNA methylation in this process. Importantly, the study addresses previously unexplored aspects of the dynamic protein expression patterns observed, providing insight into the counterintuitive expressions of key proteins such as Runx2 and osterix. This research underscores the significant impact of epigenetic mechanisms, particularly the modulation of SIRT1, in the transition from a contractile to a calcifying phenotype in VSMCs, offering potential avenues for further exploration in the context of vascular calcification., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Blood vessel growth and osteogenesis in the skeletal system are coupled; however, fundamental aspects of vascular function in osteoblast-to-osteocyte transition remain unclear. Our study demonstrates that vascular smooth muscle cells (VSMCs), but not endothelial cells, are sufficient to drive bone marrow mesenchymal stromal cell-derived osteoblast-to-osteocyte transition via β-catenin signaling and exosome-mediated communication. We found that VSMC-derived exosomes are loaded with transcripts encoding proteins associated with the osteocyte phenotype and members of the WNT/β-catenin signaling pathway. In contrast, endothelial cell-derived exosomes facilitated mature osteoblast differentiation by reprogramming the TGFB1 gene family and osteogenic transcription factors osterix (SP7) and RUNX2. Notably, VSMCs express significant levels of tetraspanins (CD9, CD63, and CD81) and drive the intracellular trafficking of exosomes with a lower membrane zeta potential than those from other cells. Additionally, the high ATP content within these exosomes supports mineralization mechanisms, as ATP is a substrate for alkaline phosphatase. Osteocyte function was further validated by RNA sequencing, revealing activity in genes related to intermittent mineralization and sonic hedgehog signaling, alongside a significant increase in TNFSF11 levels. Our findings unveil a novel role of VSMCs in promoting osteoblast-to-osteocyte transition, thus offering new insights into bone biology and homeostasis, as well as in bone-related diseases. Clinically, these insights could pave the way for innovative therapeutic strategies targeting VSMC-derived exosome pathways to treat bone-related disorders such as osteoporosis. By manipulating these signaling pathways, it may be possible to enhance bone regeneration and improve skeletal health in patients with compromised bone structure and function., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Caffeine has been extensively studied in the context of CNS pathologies as many researchers have shown that consuming it reduces pro-inflammatory biomarkers, potentially delaying the progression of neurodegenerative pathologies. Several lines of evidence suggest that adenosine receptors, especially A 1 and A 2 A receptors, are the main targets of its neuroprotective action. We found that caffeine pretreatment 15 min before LPS administration reduced the expression of Il1b in the hippocampus and striatum. The harmful modulation of caffeine-induced inflammatory response involved the downregulation of the expression of A 2 A receptors, especially in the hippocampus. Caffeine treatment alone promoted the downregulation of the adenosinergic receptor Adora2A; however, this promotion effect was reversed by LPS. Although administering caffeine increased the expression of the enzymes DNA methyltransferases 1 and 3A and decreased the expression of the demethylase enzyme Tet1, this effect was reversed by LPS in the hippocampus of mice that were administered Caffeine + LPS, relative to the basal condition; no significant differences were observed in the methylation status of the promoter regions of adenosine receptors. Finally, the bioinformatics analysis of the expanded network demonstrated the following results: the Adora2B gene connects the extended networks of the adenosine receptors Adora1 and Adora2A; the Mapk3 and Esr1 genes connect the extended Adora1 network; the Mapk4 and Arrb2 genes connect the extended Adora2A network with the extended network of the proinflammatory cytokine Il1β. These results indicated that the anti-inflammatory effects of acute caffeine administration in the hippocampus may be mediated by a complex network of interdependencies between the Adora2B and Adora2A genes., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Published by Elsevier B.V.)

Epigenetic changes, particularly histone compaction modifications, have emerged as critical regulators in the epigenetic pathway driving endothelial cell phenotype under constant exposure to laminar forces induced by blood flow. However, the underlying epigenetic mechanisms governing endothelial cell behavior in this context remain poorly understood. To address this knowledge gap, we conducted in vitro experiments using human umbilical vein endothelial cells subjected to various tensional forces simulating pathophysiological blood flow shear stress conditions, ranging from normotensive to hypertensive forces. Our study uncovers a noteworthy observation wherein endothelial cells exposed to high shear stress demonstrate a decrease in the epigenetic marks H3K4ac and H3K27ac, accompanied by significant alterations in the levels of HDAC (histone deacetylase) proteins. Moreover, we demonstrate a negative regulatory effect of increased shear stress on HOXA13 gene expression and a concomitant increase in the expression of the long noncoding RNA, HOTTIP, suggesting a direct association with the suppression of HOXA13. Collectively, these findings represent the first evidence of the role of histone-related epigenetic modifications in modulating chromatin compaction during mechanosignaling of endothelial cells in response to elevated shear stress forces. Additionally, our results highlight the importance of understanding the physiological role of HOXA13 in vascular biology and hypertensive patients, emphasizing the potential for developing small molecules to modulate its activity. These findings warrant further preclinical investigations and open new avenues for therapeutic interventions targeting epigenetic mechanisms in hypertensive conditions., (© 2024 International Federation for Cell Biology.)

During the morphological changes occurring in osteoblast differentiation, Sonic hedgehog (Shh) plays a crucial role. While some progress has been made in understanding this process, the epigenetic mechanisms governing the expression of Hh signaling members in response to bone morphogenetic protein 7 (BMP7) signaling in osteoblasts remain poorly understood. To delve deeper into this issue, we treated pre-osteoblasts (pObs) with 100 ng/mL of BMP7 for up to 21 days. Initially, we validated the osteogenic phenotype by confirming elevated expression of well-defined gene biomarkers, including Runx2, Osterix, Alkaline Phosphatase (Alp), and bone sialoprotein (Bsp). Simultaneously, Hh signaling-related members Sonic (Shh), Indian (Ihh), and Desert (Dhh) Hedgehog (Hh) exhibited nuanced modulation over the 21 days in vitro period. Subsequently, we evaluated epigenetic markers, and our data revealed a notable change in the CpG methylation profile, considering the methylation/hydroxymethylation ratio. CpG methylation is a reversible process regulated by DNA methyltransferases and demethylases, including Ten-eleven translocation (Tets), which also exhibited changes during the acquisition of the osteogenic phenotype. Specifically, we measured the methylation pattern of Shh-related genes and demonstrated a positive Pearson correlation for GLI Family Zinc Finger 1 (Gli1) and Patched (Ptch1). This data underscores the significance of the epigenetic machinery in modulating the BMP7-induced osteogenic phenotype by influencing the activity of Shh-related genes. In conclusion, this study highlights the positive impact of epigenetic control on the expression of genes related to hedgehog signaling during the morphogenetic changes induced by BMP7 signaling in osteoblasts., (© 2024 International Federation for Cell Biology.)

Significance: FT-IR is an important and emerging tool, providing information related to the biochemical composition of biofluids. It is important to demonstrate that there is an efficacy in separating healthy and diseased groups, helping to establish FT-IR uses as fast screening tool., Aim: Via saliva diagnosis evaluate the accuracy of FT-IR associate with machine learning model for classification among healthy (control group), diabetic (D) and periodontitis (P) patients and the association of both diseases (DP)., Approach: Eighty patients diagnosed with diabetes and periodontitis through conventional methods were recruited and allocated in one of the four groups. Saliva samples were collected from participants of each group (n = 20) and were processed using Bruker Alpha II spectrometer in a FT-IR spectral fingerprint region between 600 and-1800 cm -1 , followed by data preprocessing and analysis using machine learning tools., Results: Various FTI-R peaks were detectable and attributed to specific vibrational modes, which were classified based on confusion matrices showed in paired groups. The highest true positive rates (TPR) appeared between groups C vs D (93.5 % ± 2.7 %), groups C vs. DP (89.2 % ± 4.1 %), and groups D and P (90.4 % ± 3.2 %). However, P vs DP presented higher TPR for DP (84.1 % ±3.1 %) while D vs. DP the highest rate for DP was 81.7 % ± 4.3 %. Analyzing all groups together, the TPR decreased., Conclusion: The system used is portable and robust and can be widely used in clinical environments and hospitals as a new diagnostic technique. Studies in our groups are being conducted to solidify and expand data analysis methods with friendly language for healthcare professionals. It was possible to classify healthy patients in a range of 78-93 % of accuracy. Range over 80 % of accuracy between periodontitis and diabetes were observed. A general classification model with lower TPR instead of a pairwise classification would only have advantages in scenarios where no prior patient information is available regarding diabetes and periodontitis status., Competing Interests: Declaration of competing interest The authors reported no conflict of interest related to this study., (Copyright © 2024. Published by Elsevier B.V.)

Microplastics (MPs) composed of different polymers with various shapes, within a vast granulometric distribution (1 μm - 5 mm) and with a wide variety of physicochemical surface and bulk characteristics spiral around the globe, with different atmospheric, oceanic, cryospheric, and terrestrial residence times, while interacting with other pollutants and biota. The challenges of microplastic pollution are related to the complex relationships between the microplastic generation mechanisms (physical, chemical, and biological), their physicochemical properties, their interactions with other pollutants and microorganisms, the changes in their properties with aging, and their small sizes that facilitate their diffusion and transportation between the air, water, land, and biota, thereby promoting their ubiquity. Early career researchers (ERCs) constitute an essential part of the scientific community committed to overcoming the challenges of microplastic pollution with their new ideas and innovative scientific perspectives for the development of remediation technologies. However, because of the enormous amount of scientific information available, it may be difficult for ERCs to determine the complexity of this environmental issue. This mini-review aims to provide a quick and updated overview of the essential insights of microplastic pollution to ERCs to help them acquire the background needed to develop highly innovative physical, chemical, and biological remediation technologies, as well as valorization proposals and environmental education and awareness campaigns. Moreover, the recommendations for the development of holistic microplastic pollution remediation strategies presented here can help ERCs propose technologies considering the environmental, social, and practical dimensions of microplastic pollution while fulfilling the current government policies to manage this plastic waste., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Nicoly Milhardo Lourenco Nohara reports financial support was provided by Sao Paulo Research Foundation (FAPESP). Erika Iveth Cedillo-Gonzalez reports financial support was provided by University of Modena and Reggio Emilia. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Objectives: This study aimed to evaluate the effect of microwave sintering temperature and cooling rate (MS) on 3Y-TZP ceramics and its influence on the ceramic microstructure and mechanical properties. Specifically, to optimize the sintering process, reducing the total sintering time compared to conventional sintering., Materials and Methods: Eighty-four pre-sintered Y-TZP discs (Vipi block Zirconn, VIPI) (ISO 6872) were divided into seven groups (n = 12) according to the sintering conditions: conventional sintering (CS) at 1530 °C for 120 min and microwave sintering at 1400 °C (MS1400) and 1450 °C (MS1450) for 15 min followed by different cooling conditions: rapid cooling (RC), cooling at 400 °C (C400) and 25 °C (C25). The specimens were submitted to apparent density measurements, X-ray diffraction analysis (XRD), scanning electron microscopy, and biaxial flexural strength test. Data was statistically analyzed through two-way ANOVA, Tukey, Sidak, Dunnett and Weibull (α = 0.05)., Results: All MS1400 groups presented lower density values than the CS and MS1450 groups. Two-way ANOVA revealed that the MS temperature and cooling rate affected the biaxial flexural strength of the Y-TZP (p < 0.01). Group MS1400RC presented lower biaxial flexural strength values (681.9 MPa) than MS1450RC (824.7 MPa). The cooling rate did not statistically decrease the biaxial strength among the groups submitted to microwave sintering at 1450 °C. XRD analysis showed that the sintering and cooling temperature did not induce tetragonal to monoclinic phase transformation., Conclusions: Microwave sintering at 1450 °C for 15 min followed by rapid cooling can be a viable fast alternative protocol for Y-TZP sintering, compared with the conventional sintering, reducing the total sintering time by 75% and reducing the energy used for the sintering process without affecting the Y-TZP biaxial flexural strength and relative density compared to the conventional sintering. Moreover, the microwave technique promoted smaller grains and did not induce monoclinic phase formation., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Climate science has long explored whether higher resolution regional climate models (RCMs) provide improved simulation of regional climates over global climate models (GCMs). The advent of convective-permitting RCMs (CPRCMs), where sufficiently fine-scale grids allow explicitly resolving rather than parametrising convection, has created a clear distinction between RCM and GCM formulations. This study investigates the simulation of tropical-extratropical (TE) cloud bands in a suite of pan-South America convective-permitting Met Office Unified Model (UM) and Weather Research and Forecasting (WRF) climate simulations. All simulations produce annual cycles in TE cloud band frequency within 10-30% of observed climatology. However, too few cloud band days are simulated during the early summer (Nov-Dec) and too many during the core summer (Jan-Feb). Compared with their parent forcing, CPRCMs simulate more dry days but systematically higher daily rainfall rates, keeping the total rain biases low. During cloud band systems, the CPRCMs correctly reproduced the observed changes in tropical rain rates and their importance to climatology. Circulation analysis suggests that simulated lower subtropical rain rates during cloud bands systems, in contrast to the higher rates in the tropics, are associated with weaker northwesterly moisture flux from the Amazon towards southeast South America, more evident in the CPRCMs. Taken together, the results suggest that CPRCMs tend to be more effective at producing heavy daily rainfall rates than parametrised simulations for a given level of near-surface moist energy. The extent to which this improves or degrades biases present in the parent simulations is strongly region-dependent., Competing Interests: Conflict of interestThe authors have no relevant financial or non-financial interests to disclose., (© The Author(s) 2024.)

Background: Caffeine is commonly used as therapy for apnea of prematurity and has shown potential in preventing other conditions in preterm neonates. However, the optimal timing for caffeine therapy remains uncertain., Objective: This study aimed to compare the outcomes of early versus late administration of caffeine in preterm neonates., Methods: PubMed, Embase, and Cochrane Library were searched for studies comparing 0-2 days to ≥3 days caffeine introduction in preterm neonates. Outcomes included were mortality, bronchopulmonary dysplasia (BPD), intraventricular hemorrhage (IVH), necrotizing enterocolitis (NEC), retinopathy of prematurity (ROP), patent ductus arteriosus (PDA), late-onset sepsis, length of hospital stay, and the composite of BPD or death. RevMan 5.4.1 was used for statistical analysis., Results: A total of 122,579 patients from 11 studies were included, 2 were randomized controlled trials (RCTs), and 63.9% of the neonates received early caffeine administration. The rates of BPD (OR: 0.70; 95% CI: [0.60-0.81]; p &lt; 0.0001), IVH (OR: 0.86; 95% CI: [0.82-0.90]; p &lt; 0.0001), ROP (OR: 0.80; 95% CI: [0.74-0.86]; p &lt; 0.0001), late-onset sepsis (OR: 0.84; 95% CI: [0.79-0.89]; p &lt; 0.00001), and PDA (OR: 0.60; 95% CI: [0.47-0.78]; p &lt; 0.0001) were significantly reduced in the early caffeine group. The composite outcome of BPD or death was also lower in the early caffeine group (OR: 0.76; 95% CI: [0.66-0.88]; p &lt; 0.0003). Mortality rate was higher in the early caffeine group (OR: 1.20; 95% CI: 1.12-1.29; p &lt; 0.001)., Conclusion: As compared with late caffeine administration, early caffeine is associated with a reduction in BPD, IVH, ROP, late-onset sepsis, and PDA in preterm neonates, albeit increased mortality. Additional RCTs are warranted to confirm these findings and evaluate whether the effect on mortality may be related to survival bias in observational studies favoring the late treatment group., (© 2023 S. Karger AG, Basel.)

The increase in greenhouse gasses (GHG) anthropogenic emissions and deforestation over the last decades have led to many chemical and physical changes in the climate system, affecting the atmosphere's energy and water balance. A process that could be affected is the Amazonian moisture transport in the South American continent (including La Plata basin), which is crucial to the southeast Brazilian water regime. The focus of our research is on evaluating how local (i.e. Amazon deforestation) and global forcings (increase of atmospheric GHG concentration) may modify this moisture transport under climate change scenarios. We used two coupled land-atmosphere models forced by CMIP6 sea surface temperatures to simulate these processes for two scenarios: i) increase in carbon dioxide (CO 2 ) - RCP8.5 atmospheric levels (00DEF), and ii) total Amazon deforestation simultaneous with atmospheric CO 2 levels increased (100DEF). These scenarios were compared with a control simulation, set with a constant CO 2 of 388 ppm and present-day Amazon Forest cover. The 30-year Specific Warming Level 2 (SWL2) index evaluated from the simulations is set to be reached 2 years earlier due to Amazon deforestation. A reduction in precipitation was observed in the Amazon basin (-3.1 mm·day -1 ) as well as in La Plata Basin (-0.5 mm·day -1 ) due to reductions in the Amazon evapotranspiration (-0.9 mm·day -1 ) through a stomatal conductance decrease (00DEF) and land cover change (100DEF). In addition, the income moisture transport decreased (22 %) in the northern La Plata basin in both scenarios and model experiments. Our results indicated a worse scenario than previously found in the region. Both Amazon and La Plata hydrological regimes are connected (moisture and energy transport), indicating that a large-scale Amazon deforestation will have additional climate, economic and social implications for South America., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Crown Copyright © 2023. Published by Elsevier B.V. All rights reserved.)

The literature describes multiple ways to stimulate wound healing to reduce the patient's perception of pain. This systematic review aimed to evaluate if methods that enhance wound healing can reduce the patient's perception of pain after free gingival graft removal from the palate region compared to natural healing. A systematic review protocol was written following the PRISMA checklist. Electronic searches of five databases were performed to identify randomized clinical trials (RCTs) that assessed the patient's perception of pain after the removal of a free gingival graft from the palate. The primary outcome was the visual analog scale (VAS) score assessing the patient's perception of pain 7 days after the free gingival graft removal from the palate region. Of the 1,622 potentially relevant articles retrieved from the electronic databases, 16 RCTs were selected for qualitative analysis, and of these, 6 RCTs were included in the meta-analysis. RCTs showed a significant VAS reduction associated with the use of methods to enhance wound healing. The pooled estimates revealed a significant overall VAS reduction of 2.20 (95% CI 2.32, 2.07) 7 days after surgery. The methods that presented the greatest reduction in the perception of pain were platelet-rich fibrin, hyaluronic acid, and autologous fibrin glue. Methods that enhance wound healing, including platelet-rich fibrin, hyaluronic acid, and autologous fibrin glue, can reduce pain perception after free gingival graft removal in the palate region. However, only 1 RCT investigated each approach, which hinders the conclusion regarding the best procedure to reduce the perception of pain.

Objectives: Twin pregnancy is currently an exclusion criterion for prenatal repair of open spina bifida (OSB). The main objective of this study was to report on our experience of treating twin pregnancies with OSB using the skin-over-biocellulose for antenatal fetoscopic repair (SAFER) technique. We also discuss reconsideration of the current exclusion criteria for fetal OSB repair., Methods: Eight fetuses with OSB from seven twin pregnancies underwent successful prenatal repair. Six pregnancies were dichorionic diamniotic with only one twin affected, and one was monochorionic diamniotic with both twins affected. Percutaneous fetoscopy was performed under CO 2 insufflation of the sac of the affected twin. Neurosurgical repair was performed using a biocellulose patch to protect the placode, with the skin sutured to hold the patch in place, with or without a myofascial flap. Neurodevelopment was assessed using the pediatric evaluation of disability inventory scale in babies older than 6 months of adjusted age, whereas the Alberta scale was used for babies younger than 6 months of adjusted age., Results: All 14 fetuses were liveborn and none required additional repair. Gestational age at surgery ranged from 27.3 to 31.1 weeks, and gestational age at birth ranged from 31.6 to 36.0 weeks. Four out of eight affected twins developed sepsis, but had a good recovery. No sequela of prematurity was found in any of the unaffected twins. Short-term neurodevelopment was normal in all evaluated unaffected twins (5/5) and in all but one affected twins (7/8). In the affected group, only one baby required ventriculoperitoneal shunt placement., Conclusions: Prematurity is frequent after fetal surgery, and the risk is increased in twin pregnancy. Nevertheless, prenatal surgery using the SAFER technique is feasible, with low risk to both twins and their mother when performed by a highly experienced team. Long-term cognitive assessment of the unaffected twin is needed. © 2023 The Authors. Ultrasound in Obstetrics & Gynecology published by John Wiley & Sons Ltd on behalf of International Society of Ultrasound in Obstetrics and Gynecology., (© 2023 The Authors. Ultrasound in Obstetrics & Gynecology published by John Wiley & Sons Ltd on behalf of International Society of Ultrasound in Obstetrics and Gynecology.)

Background: Nanoparticles (NPs) have gained significant attention in various fields due to their unique properties and potential applications. Polymethyl methacrylate (PMMA) is an acrylic resin widely used in dentistry and medicine. However, the effect of different types of NP fillers on the physical properties of PMMA-based resins has not been thoroughly explored in the literature., Objectives: The present study aimed to evaluate the effects of 3 different types of NP fillers on the physical properties of an experimental PMMA-based resin as a function of the NP content and concentration., Material and Methods: Ten groups (n = 10) were designed. The specimens were composed of an acrylic resin, silicon dioxide (SiO2), cerium dioxide (CeO2), and titanium dioxide (TiO2) at the following ratios (wt%): group 1 (G1) - control; group 2 (G2) - 0.5% SiO2; group 3 (G3) - 1% SiO2; group 4 (G4) - 3% SiO2; group 5 (G5) - 0.5% CeO2; group 6 (G6) - 1% CeO2; group 7 (G7) - 3% CeO2; group 8 (G8) - 0.5% TiO2; group 9 (G9) - 1% TiO2; and group 10 (G10) - 3% TiO2. Transmission electron microscopy (TEM) was used to assess the quality of NP dispersion. Thermal stability was assessed with thermogravimetric analysis (TGA) and differential scanning calorimetry (DSC). The effects of the abovementioned NPs on the properties of the resin were evaluated using the Archimedes principle for density, the Vickers hardness (VH) test and the impact strength (IS) test. Data analysis employed the oneand two-way analysis of variance (ANOVA), followed by Duncan's post hoc test at a significance level of 0.05., Results: Transmission electron microscopy showed partial NP dispersion. All types of NPs enhanced the mechanical properties of the acrylic resin except for IS, which was similar to that of the control group. Among the types of NPs, irrespective of the weight percentage, CeO2 showed higher thermal stability and higher IS for 0.5 wt% and 1 wt% as compared to other groups, as well as the highest values of density at 0.5 wt%, 1 wt% and 3 wt%. Titanium oxide at 1 wt% presented a higher VH as compared to other groups. The fracture pattern was the same for all groups., Conclusions: Incorporating the tested NPs into the acrylic resin resulted in enhanced physical properties, primarily attributed to a lower NP content.

Objective: The aims of this study were to compare the bond strength between metallic brackets and two different glass ceramics and to evaluate the ceramic surface roughness after different finishing protocols., Methods: The surface roughness of lithium disilicate and resin matrix ceramic samples was measured (initial). All samples were treated with hydrofluoric acid and silane and bonded to metallic brackets with orthodontic cement adhesive. Shear bond strength tests were performed using a universal testing machine (n=12). The surface roughness was measured again (intermediate, n=6) after removing the remaining cement adhesive from the ceramic surfaces with a diamond or 24-blade bur after polishing the ceramic surfaces (final, n=6)., Results: The resin matrix ceramic had the highest bond strength. The rotatory instrument used for the removal of cement adhesive did not affect the surface roughness of the resin matrix ceramic or lithium disilicate (p=0.985 and p=0.504, respectively), but did affect the evaluation time (p<0.001) for both restorative materials. The intermediate roughness was the highest. For the resin matrix ceramic, polishing promoted a final surface roughness similar to the initial condition; however, changes in the surface shape of this ceramic could be visibly observed when using a 24-blade bur., Conclusion: The bond strength of metallic brackets bonded on resin-matrix ceramics is higher than bonding on lithium disilicate. The use of diamond burs for the removal of the remaining adhesive from the resin matrix ceramics is highly recommended.

Brain Long non-coding RNA (lncRNA) and microRNAs (miRs) play essential roles in the regulation of several important biological processes, including neuronal activity, cognitive processes, neurogenesis, angiogenesis, and neuroinflammation. In this context, the transcriptional repressor, RE1 silencing transcription factor (Rest), acts regulating the expression of neuronal genes as well as of lncRNAs and multiple miRNAs in the central nervous system. Nevertheless, its role in neuroinflammation was less explored. Here, we demonstrate, using an in vivo model of neuroinflammation induced by i.p. injection of LPS (0.33 mg/kg), that neuroinflammation increases gene expression of pro-inflammatory cytokines concomitant with the native and truncated forms of Rest and of non-coding RNAs. Additionally, the increased expression of enzymes Drosha ribonuclease III) (Drosha), Exportin 5 (Xpo5) and Endoribonuclease dicer (Dicer), associated with high expression of neuroprotective miRs 22 and 132 are indicative that the activation of biogenesis of miRs in the hippocampal region is a Central Nervous System (CNS) protective mechanism for the deleterious effects of neuroinflammation. Our results indicate that positive regulation of Rest gene expression in the hippocampal region by neuroinflammation correlates directly with the expression of miRs 22 and 132 and inversely with miR 335. In parallel, the confirmation of the possible alignment between the lncRNAs with miR 335 by bioinformatics corroborates with the sponge effect of Hottip and Hotair hybridizing and inhibiting the pro-inflammatory action of miR 335. This suggests the existence of a possible correlation between the activation of miR biogenesis machinery with increased expression of the transcription factor Rest, contributing to neuroprotection., Competing Interests: Declaration of Competing Interest None., (Copyright © 2023. Published by Elsevier B.V.)

Background: Prospective studies have reported conflicting results regarding the adjunctive effect of antimicrobial photodynamic therapy (aPDT) on clinical and microbiological parameters in individuals under periodontal maintenance therapy (PMT). This study aimed to evaluate the clinical and microbiological adjunctive effects of aPDT using indocyanine green (ICG) in residual sites with probing depth (PD) ≥5 mm during PMT in comparison with scaling and root planing (SRP) alone., Methods: A split-mouth randomized controlled clinical trial was conducted with 24 individuals in a PMT program. Contralateral quadrants with eligible residual sites were randomly assigned to either SRP + aPDT (test group) or SRP alone (control). aPDT included ICG dye and diode laser (909 nm) performed together with SRP and repeated 15 days after. Periodontal clinical parameters, periodontal inflamed surface area (PISA) index, and subgingival biofilm samples were collected at baseline (T1), 3 (T2), and 6 months later (T3). Microbiological analyses were performed by quantitative real-time polymerase chain reaction., Results: Significant improvements were observed in all clinical and microbiological parameters in both groups from T1 to T3. However, no significant differences were observed regarding plaque index, PD, and clinical attachment level. Test group showed significantly greater reductions in bleeding on probing (BOP), PISA index, and Porphyromonas gingivalis and Aggregatibacter actinomycetemcomitans levels, when compared with controls., Conclusions: Both treatments resulted in significant clinical periodontal improvements, but with no significant differences between groups except from inflammation parameters. aPDT using ICG resulted in significant reductions in BOP and PISA index, as well as in P. gingivalis and A. actinomycetemcomitans levels., (© 2023 American Academy of Periodontology.)

Objective: To evaluate the mechanical properties of different CAD/CAM ceramic systems and the post-fatigue fracture and stress distribution when used as cemented crowns., Materials and Methods: Sixty (60) CAD/CAM monolithic crowns were milled using three different ceramic materials (FD - Feldspathic [Vita Mark II]), LE - Leucite-based ceramic [IPS Empress CAD] and LD - Lithium Disilicate [IPS e.max CAD]) and adhesively cemented on resin composite dyes. Specimens were stored in distillated water (37 °C) for 7 days. After, half of the crowns were submitted to immediate fracture load test while the other half was submitted to fatigue cycling. The average cement layer of approximately 80 μm was assessed using scanning electron microscopy (SEM). The average thickness was used in the three-dimensional (3D) Finite Element Analysis (FEA). For each ceramic material, the density, Poisson ratio, shear modulus, Young modulus, fracture toughness, and true hardness were assessed (n = 3). The data was used to assess the Maximum Principal Stress throughout 3D-FEA according to each material during load to fail and post-fatigue. Data were submitted to two-way ANOVA and Tukey test (α = 0.05)., Results: LD showed the highest compression load, density, shear modulus, Young modulus, fracture toughness and true hardness values. While LE presented the lowest mechanical properties values. There is no difference in the Poisson ratio between the evaluated ceramics., Conclusion: LD was susceptible to aging process but presented stronger physicomechanical properties, showing the highest post-fatigue fracture load and highest stress magnitude., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2023 The Authors.)

Statement of Problem: Investigations on the effectiveness of new methods for optimizing the fabrication of oral devices are lacking., Purpose: The purpose of this in vitro study was to evaluate stress distribution with photoelastic analysis in the periradicular area of teeth supporting occlusal devices fabricated by 5 different processes., Material and Methods: The occlusal devices were fabricated by vacuum thermoforming, heat-polymerized acrylic resin, chemical polymerized acrylic resin, 3-dimensional printing, and milling (computer-aided manufacturing). The devices were evaluated regarding initial fit, number of adjustments for passive fit, and stress distribution under 100-N and 400-N loads in the periradicular locations of posterior teeth., Results: The 3-dimensional printing device did not require any adjustment for initial adaptation to the photoelastic model and presented a little friction with the model. The heat-polymerized acrylic resin device did not seat initially, requiring more sites of adjustment until passive adaptation. At 100-N and 400-N loads, the use of the computer-aided manufacturing occlusal device resulted in the lowest stresses in periradicular areas (0.744 and 1.583, respectively), and the 3-dimensional printing occlusal device produced the highest stresses with a 400-N load application (2.427). The lowest mean of fringe pattern was observed for the computer-aided manufacturing device, and the highest mean of fringe pattern was observed for the vacuum thermoforming device., Conclusions: The computer-aided design and computer-aided manufacturing milled occlusal device presented the best initial adaptation and transferred lower stresses to the periradicular areas than the other evaluated devices., (Copyright © 2021 Editorial Council for the Journal of Prosthetic Dentistry. Published by Elsevier Inc. All rights reserved.)

Breast cancer is a heterogeneous disease, and its spread involves a succession of clinical and pathological stages. Screening is predominantly based on mammography, which has critical limitations related to the effectiveness and production of false-positive or false-negative results, generating discomfort and low adherence. In this context, infrared with attenuated total reflection Fourier transform infrared (ATR FT-IR) spectroscopy emerges as a non-destructive sample tool, which is non-invasive, label-free, has a low operating-cost, and requires only a small amount of sample, including liquid plasma samples. We sought to evaluate the clinical applicability of ATR FT-IR in breast cancer screening. ATR FT-IR spectroscopy through its highest potential spectral biomarker could distinguish, by liquid plasma biopsy, breast cancer patients and healthy controls, obtaining a sensitivity of 97%, specificity of 93%, a receiver operating characteristic ROC curve of 97%, and a prediction accuracy of 94%. The main variance between the groups was mainly in the band 1511 cm -1 of the control group, 1502 and 1515 cm -1 of the cancer group, which are the peaks of the bands referring to proteins and amide II. ATR FT-IR spectroscopy has demonstrated to be a promising tool for breast cancer screening, given its time efficiency, cost of approach, and its high ability to distinguish between the liquid plasma samples of breast cancer patients and healthy controls.

Diabetes mellitus (DM) is a chronic metabolic disease, mainly characterized by increased blood glucose and insulin dysfunction. In response to the persistent systemic hyperglycemic state, numerous metabolic and physiological complications have already been well characterized. However, its relationship to bone fragility, cognitive deficits and increased risk of dementia still needs to be better understood. The impact of chronic hyperglycemia on bone physiology and architecture was assessed in a model of chronic hyperglycemia induced by a single intraperitoneal administration of streptozotocin (STZ; 55 mg/kg) in Wistar rats. In addition, the bone-to-brain communication was investigated by analyzing the gene expression and methylation status of genes that encode the main osteokines released by the bone [Fgf23 (fibroblast growth factor 23), Bglap (bone gamma-carboxyglutamate protein) and Lcn2 (lipocalin 2) and their receptors in both, the bone and the brain [Fgfr1 (fibroblast growth factor receptor 1), Gpr6A (G-protein coupled receptor family C group 6 member A), Gpr158 (G protein-coupled receptor 158) and Slc22a17 (Solute carrier family 22 member 17)]. It was observed that chronic hyperglycemia negatively impacted on bone biology and compromised the balance of the bone-brain endocrine axis. Ultrastructural disorganization was accompanied by global DNA hypomethylation and changes in gene expression of DNA-modifying enzymes that were accompanied by changes in the methylation status of the osteokine promoter region Bglap and Lcn2 (lipocalin 2) in the femur. Additionally, the chronic hyperglycemic state was accompanied by modulation of gene expression of the osteokines Fgf23 (fibroblast growth factor 23), Bglap (bone gamma-carboxyglutamate protein) and Lcn2 (lipocalin 2) in the different brain regions. However, transcriptional regulation mediated by DNA methylation was observed only for the osteokine receptors, Fgfr1(fibroblast growth factor receptor 1) in the striatum and Gpr158 (G protein-coupled receptor 158) in the hippocampus. This is a pioneer study demonstrating that the chronic hyperglycemic state compromises the crosstalk between bone tissue and the brain, mainly affecting the hippocampus, through transcriptional silencing of the Bglap receptor by hypermethylation of Gpr158 gene., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Aims: This study investigated the possible relationships between the expression of the Kiss1 and Gpr54 gene expressions and the pituitary-gonadal hormones with the female onset of puberty and sexual behavior. The Kiss1 and Gpr54 gene expressions were examined because they are critical to controlling the hypothalamic activation of GnRH neurons and, in turn, the pituitary-gonadal hormones related to the early onset of puberty and sexual behavior. Further, it was evaluated that the pituitary and gonadal hormones involved in the vaginal opening and the expression of sexual behavior., Methods: Pregnant rats exposed to PRS from gestation days 17 to 20 were evaluated for maternal and open-field behaviors. The maternal behavior was analyzed because it may alter brain sexual organization affecting the pups development. It was observed in female pups the physical and development and, in adult age, the open-field behavior, the anxiety-like behavior, the estrous cycle, the sexual behavior, the serum FSH, LH, estrogen, progesterone, and testosterone levels, and the gene expression of kisspeptin protein (Kiss1) and Gpr54 in the hypothalamus., Results: the maternal and open-field behaviors were unaffected. In the F1 generation, PRS reduced weight at weaning, delayed the day of the vaginal opening and reduced the intensity of lordosis, the estrogen levels, and the Kiss1 and Gpr54 gene expression. These effects were attributed to hypothalamic kisspeptidergic system downregulation of transcripts genes and the reduced estrogen levels affected by the PRS., Competing Interests: Declaration of Competing Interest The authors declare no conflict of interest, (Copyright © 2022 Elsevier Inc. All rights reserved.)

It is important to understand whether endothelial cells are epigenetically affected by titanium-enriched media when angiogenesis is required during bone development and it is expected to be recapitulated during osseointegration of biomaterials. To better address this issue, titanium-enriched medium was obtained from incubation of titanium discs for up to 24 h as recommended by ISO 10993-5:2016, and further used to expose human umbilical vein endothelial cells (HUVECs) for up to 72 h, when the samples were properly harvested to allow molecular analysis and epigenetics. In general, our data show an important repertoire of epigenetic players in endothelial cells responding to titanium, reinforcing protein related to the metabolism of acetyl and methyl groups, as follows: Histone deacetylases (HDACs) and NAD-dependent deacetylase sirtuin-1 (Sirt1), DNA methyltransferases (DNMTs) and ten-eleven translocation (TET) methylcytosine dioxygenases, which in conjunction culminate in driving chromatin condensation and the methylation profile of DNA strands, respectively. Taking our data into consideration, HDAC6 emerges as important player of this environment-induced epigenetic mechanism in endothelial cells, while Sirt1 is required in response to stimulation of reactive oxygen species (ROS) production, as its modulation is relevant to vasculature surrounding implanted devices. Collectively, all these findings support the hypothesis that titanium keeps the surrounding microenvironment dynamically active and so affects the performance of endothelial cells by modulating epigenetics. Specifically, this study shows the relevance of HDAC6 as a player in this process, possibly correlated with the cytoskeleton rearrangement of those cells. Furthermore, as those enzymes are druggable, it opens new perspectives to consider the use of small molecules to modulate their activities as a biotechnological tool in order to improve angiogenesis and accelerate bone growth with benefits of a fast recovery time for patients.

Cobalt-chromium (Co-Cr)-based alloys are emerging with important characteristics for use in dentistry, but the knowledge of epigenetic mechanisms in endothelial cells has barely been achieved. In order to address this issue, we have prepared a previously Co-Cr-enriched medium to further treat endothelial cells (HUVEC) for up to 72 h. Our data show there is important involvement with epigenetic machinery. Based on the data, it is believed that methylation balance in response to Co-Cr is finely modulated by DNMTs (DNA methyltransferases) and TETs (Tet methylcytosine dioxygenases), especially DNMT3B and both TET1 and TET2. Additionally, histone compaction HDAC6 (histone deacetylase 6) seems to develop a significant effect in endothelial cells. The requirement of SIRT1 seems to have a crucial role in this scenario. SIRT1 is associated with a capacity to modulate the expression of HIF-1α in response to hypoxia microenvironments, thus presenting a protective effect. As mentioned previously, cobalt is able to prevent HIF1A degradation and maintain hypoxia-related signaling in eukaryotic cells. Together, our results show, for the first time, a descriptive study reporting the relevance of epigenetic machinery in endothelial cells responding to cobalt-chromium, and it opens new perspectives to better understand their repercussions as prerequisites for driving cell adhesion, cell cycle progression, and angiogenesis surrounding this Co-Cr-based implantable device.

Evidence has shown that caffeine administration reduces pro-inflammatory biomarkers, delaying fatigue and improving endurance performance. This study examined the effects of caffeine administration on the expression of inflammatory-, adenosine receptor- (the targets of caffeine), epigenetic-, and oxidative metabolism-linked genes in the vastus lateralis muscle of mice submitted to lipopolysaccharide (LPS)-induced inflammation. We showed that caffeine pre-treatment before LPS administration reduced the expression of Il1b , Il6 , and Tnfa , and increased Il10 and Il13 . The negative modulation of the inflammatory response induced by caffeine involved the reduction of inflammasome components, Asc and Casp1 , promoting an anti-inflammatory scenario. Caffeine treatment per se promoted the upregulation of adenosinergic receptors, Adora1 and Adora2A , an effect that was counterbalanced by LPS. Moreover, there was observed a marked Adora2A promoter hypermethylation, which could represent a compensatory response towards the increased Adora2A expression. Though caffeine administration did not alter DNA methylation patterns, the expression of DNA demethylating enzymes, Tet1 and Tet2 , was increased in mice receiving Caffeine+LPS, when compared with the basal condition. Finally, caffeine administration attenuated the LPS-induced catabolic state, by rescuing basal levels of Ampk expression. Altogether, the anti-inflammatory effects of caffeine in the muscle can be mediated by modifications on the epigenetic landscape.

Sonic Hedgehog (Shh) signaling plays a critical role during central nervous system (CNS) development, and its dysregulation leads to neurological disorders. Nevertheless, little is known about Shh signaling regulation in the adult brain. Here, we investigated the contribution of DNA methylation on the transcriptional control of Shh signaling pathway members and its basal distribution impact on the brain, as well as its modulation by inflammation. The methylation status of the promoter regions of these members and the transcriptional profile of DNA-modifying enzymes (DNA Methyltransferases - DNMTs and Tet Methylcytosine Dioxygenase - TETs) were investigated in a murine model of neuroinflammation by qPCR. We showed that, in the adult brain, methylation in the CpG promoter regions of the Shh signaling pathway members was critical to determine the endogenous differential transcriptional pattern observed between distinct brain regions. We also found that neuroinflammation differentially modulates gene expression of DNA-modifying enzymes. This study reveals the basal transcriptional profile of DNMTs and TETs enzymes in the CNS and demonstrates the effect of neuroinflammation on the transcriptional control of members of the Shh Signaling pathway in the adult brain., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper, (Copyright © 2022 Elsevier B.V. All rights reserved.)

Background: In certain clinical situations, root canal treatment in teeth with apical periodontitis is performed in multiple visits, with the use of intracanal dressing between visits, aiming to reduce microorganisms and their by-products of the root canal system prior to filling. However, in recent years, discussions have been growing about the real need for the use of intracanal dressing in these cases. The use of ultrasonic activation of the auxiliary chemical substance has increased the potential for decontamination promoted during the chemomechanical preparation of the root canal. Thus, this study is designed to explore whether the use of intracanal dressing between visits during endodontic treatment favors periradicular repair in teeth with apical periodontitis., Methods: This is a randomized, prospective, double-blinded, controlled clinical trial designed to evaluate 3 distinct clinical approaches used during endodontic therapy: group 1-root canal treatment in a single visit (RCT-SV); group 2-root canal treatment in two visits with intracanal dressing (RCT-TVWD); and group 3-root canal treatment in two visits without intracanal dressing (RCT-TVWOD). A total of 150 adult patients aged 18 to 60, with at least one tooth diagnosed with asymptomatic apical periodontitis and periradicular lesion (confirmed with a cone beam computed tomography (CBCT)), will be randomized and will undergo one of the types of clinical approaches during endodontic therapy. Patients' postoperative pain levels will also be recorded in periods of 24, 48, and 72 h and 7 days. Subsequently, clinical findings and long-term follow-up evaluations, with periradicular repair, will be performed at 6 and 12 months by intraoral periapical radiograph (IOPAR) and CBCT at the 24-month follow-up., Discussion: This study will evaluate the periradicular repair of mandibular molar teeth with apical periodontitis, providing information about the efficacy, benefits, and safety of performing the endodontic treatment in a single and two visits, with and without the use of calcium hydroxide dressing. All endodontic therapy procedures will be performed under a dental operating microscope and using ultrasonic activation of auxiliary chemical substances. These results may contribute to changes in the clinical approaches adopted during endodontic therapy of teeth with apical periodontitis and reveal the potential of complementary approaches that aim to enhance the decontamination of the root canal system during the preparation stage., Trial Registration: ClinicalTrials.gov NCT05256667. Registered on 24 February 2022., (© 2023. The Author(s).)