Single-cell epigenetic analysis reveals principles of chromatin states in H3.3-K27M gliomas.

Cancer cells are highly heterogeneous at the transcriptional level and epigenetic state. Methods to study epigenetic heterogeneity are limited in throughput and information obtained per cell. Here, we adapted cytometry by time-of-flight (CyTOF) to analyze a wide panel of histone modifications in primary tumor-derived lines of diffused intrinsic pontine glioma (DIPG). DIPG is a lethal glioma, driven by a histone H3 lysine 27 mutation (H3-K27M). We identified two epigenetically distinct subpopulations in DIPG, reflecting inherent heterogeneity in expression of the mutant histone. These two subpopulations are robust across tumor lines derived from different patients and show differential proliferation capacity and expression of stem cell and differentiation markers. Moreover, we demonstrate the use of these high-dimensional data to elucidate potential interactions between histone modifications and epigenetic alterations during the cell cycle. Our work establishes new concepts for the analysis of epigenetic heterogeneity in cancer that could be applied to diverse biological systems.

Nuclear chromosome locations dictate segregation error frequencies.

Chromosome segregation errors during cell divisions generate aneuploidies and micronuclei, which can undergo extensive chromosomal rearrangements such as chromothripsis1-5. Selective pressures then shape distinct aneuploidy and rearrangement patterns-for example, in cancer6,7-but it is unknown whether initial biases in segregation errors and micronucleation exist for particular chromosomes. Using single-cell DNA sequencing8 after an error-prone mitosis in untransformed, diploid cell lines and organoids, we show that chromosomes have different segregation error frequencies that result in non-random aneuploidy landscapes. Isolation and sequencing of single micronuclei from these cells showed that mis-segregating chromosomes frequently also preferentially become entrapped in micronuclei. A similar bias was found in naturally occurring micronuclei of two cancer cell lines. We find that segregation error frequencies of individual chromosomes correlate with their location in the interphase nucleus, and show that this is highest for peripheral chromosomes behind spindle poles. Randomization of chromosome positions, Cas9-mediated live tracking and forced repositioning of individual chromosomes showed that a greater distance from the nuclear centre directly increases the propensity to mis-segregate. Accordingly, chromothripsis in cancer genomes9 and aneuploidies in early development10 occur more frequently for larger chromosomes, which are preferentially located near the nuclear periphery. Our findings reveal a direct link between nuclear chromosome positions, segregation error frequencies and micronucleus content, with implications for our understanding of tumour genome evolution and the origins of specific aneuploidies during development.

Gastric Cancer: The Microbiome Beyond Helicobacter pylori.

Gastric cancer remains an important global health burden. Helicobacter pylori is the major etiological factor in gastric cancer, infecting the stomach of almost half of the population worldwide. Recent progress in microbiome research offered a new perspective on the complexity of the microbial communities of the stomach. Still, the role of the microbiome of the stomach beyond H. pylori in gastric carcinogenesis is not well understood and requires deeper investigation. The gastric bacterial communities of gastric cancer patients are distinct from those of patients without cancer, but the microbial alterations that occur along the process of gastric carcinogenesis, and the mechanisms through which microorganisms influence cancer progression still need to be clarified. Except for Epstein-Barr virus, the potential significance of the virome and of the mycobiome in gastric cancer have received less attention. This chapter updates the current knowledge regarding the gastric microbiome, including bacteria, viruses, and fungi, within the context of H. pylori-mediated carcinogenesis. It also reviews the possible roles of the local gastric microbiota, as well as the microbial communities of the oral and gut ecosystems, as biomarkers for gastric cancer detection. Finally, it discusses future perspectives and acknowledges limitations in the area of microbiome research in the gastric cancer setting, to which further research efforts should be directed. These will be fundamental not only to increase our current understanding of host-microbial interactions but also to facilitate translation of the findings into innovative preventive, diagnostic, and therapeutic strategies to decrease the global burden of gastric cancer.

Permission to pass: on the role of p53 as a gatekeeper for aneuploidy.

Aneuploidy-the karyotype state in which the number of chromosomes deviates from a multiple of the haploid chromosome set-is common in cancer, where it is thought to facilitate tumor initiation and progression. However, it is poorly tolerated in healthy cells: during development and tissue homeostasis, aneuploid cells are efficiently cleared from the population. It is still largely unknown how cancer cells become, and adapt to being, aneuploid. P53, the gatekeeper of the genome, has been proposed to guard against aneuploidy. Aneuploidy in cancer genomes strongly correlates with mutations in TP53, and p53 is thought to prevent the propagation of aneuploid cells. Whether p53 also participates in preventing the mistakes in cell division that lead to aneuploidy is still under debate. In this review, we summarize the current understanding of the role of p53 in protecting cells from aneuploidy, and we explore the consequences of functional p53 loss for the propagation of aneuploidy in cancer.

Particle abrasion as a pre-bonding dentin surface treatment: A scoping review.

OBJECTIVE: This scoping review aims to assess the influence of air abrasion with aluminum oxide and bioactive glass on dentin bond strength. MATERIALS AND METHODS: An electronic search was conducted in three databases (PubMed, Cochrane Library, and Embase), on March 3rd, 2023, with previously identified MeSH Terms. A total of 1023 records were screened. Exclusion criteria include primary teeth, air abrasion of a substrate other than sound dentin, use of particles apart from aluminum oxide or bioactive glass, and studies in which bond strength was not assessed. RESULTS: Out of the 1023 records, title and abstract screening resulted in the exclusion of 895 and 67 studies, respectively, while full-text analysis excluded another 25 articles. In addition, 5 records were not included, as full texts could not be obtained after requesting the authors. Two cross-references were added. Thus, 33 studies were included in this review. It is important to emphasize the absence of standardization of air abrasion parameters. According to 63.6% of the studies, air abrasion does not influence dentin bond strength. Moreover, 30.3% suggest improving bonding performance, and 6.1% advocate a decrease. CONCLUSIONS: Air abrasion with aluminum oxide does not enhance or impair dentin bond strength. The available data on bioactive glass are limited, which hinders conclusive insights. CLINICAL SIGNIFICANCE: Dentin air abrasion is a widely applied technique nowadays, with numerous clinical applications. Despite the widespread adoption of this procedure, its potential impact on bonding performance requires a thorough analysis of the existing literature.

Effects of endodontic irrigation solutions on structural, chemical, and mechanical properties of coronal dentin: A scoping review.

OBJECTIVE: This review aims to assess structural, chemical, and mechanical properties of coronal dentin after endodontic irrigation. MATERIALS AND METHODS: Reporting followed the PRISMA extension for scoping reviews. An electronic search was carried out in PubMed, Embase, and Cochrane Library. Records filtered by language and published up to November 4, 2022 were independently screened by two researchers. Studies evaluating structural, chemical, or mechanical properties of human permanent coronal dentin after irrigation within the scope of nonsurgical root canal treatment were included. Data were extracted regarding study type, sample description and size, experimental groups, outcome, evaluation method, and main findings. RESULTS: From the initial 1916 studies, and by adding 2 cross-references, 11 in vitro studies were included. Seven studies provide ultrastructural and/or chemical characterization, and six assessed microhardness and/or flexural strength. One percent to 8% sodium hypochlorite (NaOCl) and 1%-17% ethylenediaminetetraacetic acid (EDTA) were the most commonly tested solutions, with contact times of 2-240 min (NaOCl) and 1-1440 min (EDTA) being evaluated. CONCLUSIONS: Overall, the literature is consensual regarding the inevitable impact of NaOCl and chelating agents on coronal dentin, with both deproteinizing and decalcifying effects being concentration- and time-dependent. The alteration of mechanical parameters further confirmed the surface and subsurface ultrastructural and chemical changes. CLINICAL SIGNIFICANCE: Endodontic treatment success highly depends on restorative sealing. Understanding the result of exposing coronal dentin, the main substrate for bonding, to irrigants' action is crucial. The deproteinizing and decalcifying effects of NaOCl and chelating agents are both concentration- and time-dependent, causing surface and subsurface ultrastructural, chemical, and mechanical alterations.

Chagas Disease: A Silent Threat for Dogs and Humans.

Chagas disease (CD) is a vector-borne Neglected Zoonotic Disease (NZD) caused by a flagellate protozoan, Trypanosoma cruzi, that affects various mammalian species across America, including humans and domestic animals. However, due to an increase in population movements and new routes of transmission, T. cruzi infection is presently considered a worldwide health concern, no longer restricted to endemic countries. Dogs play a major role in the domestic cycle by acting very efficiently as reservoirs and allowing the perpetuation of parasite transmission in endemic areas. Despite the significant progress made in recent years, still there is no vaccine against human and animal disease, there are few drugs available for the treatment of human CD, and there is no standard protocol for the treatment of canine CD. In this review, we highlight human and canine Chagas Disease in its different dimensions and interconnections. Dogs, which are considered to be the most important peridomestic reservoir and sentinel for the transmission of T. cruzi infection in a community, develop CD that is clinically similar to human CD. Therefore, an integrative approach, based on the One Health concept, bringing together the advances in genomics, immunology, and epidemiology can lead to the effective development of vaccines, new treatments, and innovative control strategies to tackle CD.

Spanish cohort of VEXAS syndrome: clinical manifestations, outcome of treatments and novel evidences about UBA1 mosaicism.

BACKGROUND: The vacuoles, E1-enzyme, X linked, autoinflammatory and somatic (VEXAS) syndrome is an adult-onset autoinflammatory disease (AID) due to postzygotic UBA1 variants. OBJECTIVES: To investigate the presence of VEXAS syndrome among patients with adult-onset undiagnosed AID. Additional studies evaluated the mosaicism distribution and the circulating cytokines. METHODS: Gene analyses were performed by both Sanger and amplicon-based deep sequencing. Patients' data were collected from their medical charts. Cytokines were quantified by Luminex. RESULTS: Genetic analyses of enrolled patients (n=42) identified 30 patients carrying UBA1 pathogenic variants, with frequencies compatible for postzygotic variants. All patients were male individuals who presented with a late-onset disease (mean 67.5 years; median 67.0 years) characterised by cutaneous lesions (90%), fever (66.7%), pulmonary manifestations (66.7%) and arthritis (53.3%). Macrocytic anaemia and increased erythrocyte sedimentation rate and ferritin were the most relevant analytical abnormalities. Glucocorticoids ameliorated the inflammatory manifestations, but most patients became glucocorticoid-dependent. Positive responses were obtained when targeting the haematopoietic component of the disease with either decitabine or allogeneic haematopoietic stem cell transplantation. Additional analyses detected the UBA1 variants in both haematopoietic and non-haematopoietic tissues. Finally, analysis of circulating cytokines did not identify inflammatory mediators of the disease. CONCLUSION: Thirty patients with adult-onset AID were definitively diagnosed with VEXAS syndrome through genetic analyses. Despite minor interindividual differences, their main characteristics were in concordance with previous reports. We detected for the first time the UBA1 mosaicism in non-haematopoietic tissue, which questions the previous concept of myeloid-restricted mosaicism and may have conceptual consequences for the disease mechanisms.

Ketamine-dexmedetomidine combination for sedation in pediatric major surgery in a low-income country.

Anorectal malformations are one of the most frequent congenital malformations treated by pediatric surgeons. In low-income countries, the surgical and anesthetic management of children in need of these procedures can be challenging. Limited oxygen supply, lack of equipment, especially pediatric, and intensive care units make the use of regional anesthesia appealing. We present a series of four cases of anorectal malformations corrections in Guinea Bissau, in children up to 13 months of age, under regional anesthesia and sedation with ketodex, a mixture of ketamine and dexmedetomidine (in a proportion of 1 mg to 1 µg). No child developed respiratory depression requiring airway intervention or supplemental oxygen, or had hemodynamic instability.

Motivation and Knowledge of Portuguese Community Pharmacists Towards the Reporting of Suspected Adverse Reactions to Medicines: A Cross-Sectional Survey.

The close contact between patients and community pharmacists, along with the extensive geographical distribution of pharmacies in Portugal, offer exceptional conditions to detect and report adverse drug reactions (ADR). This study aimed to evaluate the motivation and knowledge of spontaneous reporting of ADR by community pharmacists of Porto, Portugal. Secondly, we aimed to generate real-world evidence on the main factors determining ADR report and at raising potential alternatives to the current reporting procedure in community pharmacy. We performed a descriptive, cross-sectional, observational, anonymous web survey-based study. Between April and July 2021, a web survey was implemented, targeting community pharmacists in the Porto district, Portugal. We validated 217 surveys from pharmacists. Regular notifiers seem to be more familiarised than non-regular notifiers with the Portuguese Pharmacovigilance System (PPS), with the Portal RAM for reporting suspected ADR, and with the update of the concept of ADR. Moreover, regular notifiers seem to be more proactive with their care in questioning patients about ADR and have more self-knowledge to identify suspected ADR. Conversely, non-regular notifiers, seem to be more reluctant to be judged by their ADR reporting activities. Respondents suggested to simplify and optimise the reporting process (31% of the suggestions), or to integrate a reporting platform into the pharmacy's software (27%). This study identified opportunities to promote the ADR reporting process by community pharmacists, namely receiving feedback from the PPS on the reported case and its regulatory implications, implementing training programs in pharmacovigilance, and creating solutions to simplify the reporting process.

Clinical in-situ evaluation of the effect of rubber dam isolation on bond strength to enamel.

OBJECTIVE: To evaluate the effect of rubber dam isolation on shear bond strength of two different adhesive systems to enamel. MATERIALS AND METHODS: The mesial, distal, lingual, and vestibular enamel surfaces of thirty human third molars were prepared (total n = 120). A custom splint was made to fit a volunteer's maxilla, holding the specimens in place in the oral cavity. Four composite resin cylinders were bonded to each tooth with one of two bonding agents (OptiBond FL and Prime&Bond active) with or without rubber dam isolation. Shear bond strength was tested in a universal testing machine and failure modes were assessed. Significance level for statistical analyses was set at 5%. RESULTS: All pairwise comparisons revealed statistical differences (p < 0.05). The highest mean shear bond strength values were obtained in rubber dam experimental groups, regardless of the adhesive system. Group OptiBond FL with rubber dam presented the highest mean bond strength values. Fracture modes for specimens bonded without rubber dam isolation were adhesive and cohesive within enamel, while rubber dam experimental groups revealed only cohesive fractures. CONCLUSIONS: Absolute isolation with rubber dam increases bond strength to enamel, independent of the adhesive system. The three-step total-etch system OptiBond FL provided significantly higher bond strength values than Prime&Bond active under both experimental conditions. CLINICAL SIGNIFICANCE: Rubber dam isolation has a significant effect on bond strengths to enamel, independent of the adhesive system. Its application is, therefore, advised whenever adhesive procedures are performed. A filled three-step etch-and-rinse adhesive performed superiorly, with or without rubber dam isolation, when bonding to enamel compared to an isopropanol-based universal adhesive.

Evaluation of the Cytotoxic Effect of Pd2Spm against Prostate Cancer through Vibrational Microspectroscopies.

Regarding the development of new antineoplastic agents, with a view to assess the selective antitumoral potential which aims at causing irreversible damage to cancer cells while preserving the integrity of their healthy counterparts, it is essential to evaluate the cytotoxic effects in both healthy and malignant human cell lines. In this study, a complex with two Pd(II) centers linked by the biogenic polyamine spermine (Pd2Spm) was tested on healthy (PNT-2) and cancer (LNCaP and PC-3) prostate human cell lines, using cisplatin as a reference. To understand the mechanisms of action of both cisplatin and Pd2Spm at a molecular level, Fourier Transform Infrared (FTIR) and Raman microspectroscopies were used. Principal component analysis was applied to the vibrational data, revealing the major metabolic changes caused by each drug, which were found to rely on DNA, lipids, and proteins, acting as biomarkers of drug impact. The main changes were observed between the B-DNA native conformation and either Z-DNA or A-DNA, with a higher effect on lipids having been detected in the presence of cisplatin as compared to Pd2Spm. In turn, the Pd-agent showed a more significant impact on proteins.

A Non-Conventional Platinum Drug against a Non-Small Cell Lung Cancer Line.

A dinuclear Pt(II) complex with putrescine as bridging polyamine ligand ([Pt2Put2(NH3)4]Cl4) was synthesized and assessed as to its potential anticancer activity against a human non-small cell lung cancer line (A549), as well as towards non-cancer cells (BEAS-2B). This effect was evaluated through in vitro cytotoxicity assays (MTT and SRB) coupled to microFTIR and microRaman spectroscopies, the former delivering information on growth-inhibiting and cytotoxic abilities while the latter provided very specific information on the metabolic impact of the metal agent (at the sub-cellular level). Regarding cancer cells, a major impact of [Pt2Put2(NH3)4]Cl4 was evidenced on cellular proteins and lipids, as compared to DNA, particularly via the Amide I and Amide II signals. The effect of the chelate on non-malignant cells was lower than on malignant ones, evidencing a promising low toxicity towards healthy cells.

Adenosine A2A Receptors Contribute to the Radial Migration of Cortical Projection Neurons through the Regulation of Neuronal Polarization and Axon Formation.

Cortical interneurons born in the subpallium reach the cortex through tangential migration, whereas pyramidal cells reach their final position by radial migration. Purinergic signaling via P2Y1 receptors controls the migration of intermediate precursor cells from the ventricular zone to the subventricular zone. It was also reported that the blockade of A2A receptors (A2AR) controls the tangential migration of somatostatin+ interneurons. Here we found that A2AR control radial migration of cortical projection neurons. In A2AR-knockout (KO) mouse embryos or naïve mouse embryos exposed to an A2AR antagonist, we observed an accumulation of early-born migrating neurons in the lower intermediate zone at late embryogenesis. In utero knockdown of A2AR also caused an accumulation of neurons at the lower intermediate zone before birth. This entails the presently identified ability of A2AR to promote multipolar-bipolar transition and axon formation, critical for the transition of migrating neurons from the intermediate zone to the cortical plate. This effect seems to require extracellular ATP-derived adenosine since a similar accumulation of neurons at the lower intermediate zone was observed in mice lacking ecto-5'-nucleotidase (CD73-KO). These findings frame adenosine as a fine-tune regulator of the wiring of cortical inhibitory and excitatory networks.

Luting indirect restorations with resin cements versus composite resins: Effects of preheating and ultrasound energy on film thickness.

OBJECTIVE: This study aims to evaluate and compare the film thickness obtained with a resin cement and two composite resins, preheated and/or ultrasonically vibrated, as luting agents. MATERIALS AND METHODS: One hundred and twenty-six (126) pairs of resin discs were randomly assigned to six experimental groups (n = 21) according to luting agent (Variolink Esthetic LC, IPS Empress Direct or Estelite Omega) and cementation technique (preheating at 68°C and/or ultrasonic vibration). Specimens were luted by applying a controlled force. Following sectioning and film thickness measurement through field emission gun scanning electron microscopy, statistical analysis was carried out considering a 5% significance level. RESULTS: Statistically significant lower film thickness was observed in Variolink Esthetic LC group when compared to all composite resin groups (p < 0.001), except IPS Empress Direct preheated and ultrasonically vibrated group (p = 0.073). IPS Empress Direct with ultrasonic vibration yielded statistically lower film thickness values than Estelite Omega groups, regardless of luting technique (p < 0.05). Ultrasonically vibrated Estelite Omega groups showed statistically lower film thickness values than solely preheated groups (p < 0.05). CONCLUSIONS: Both Variolink Esthetic LC and IPS Empress Direct preheated and ultrasonically vibrated provided the lowest film thickness. The addition of ultrasonic vibration during cementation proved to be effective in reducing film thickness of both tested composite resins. CLINICAL SIGNIFICANCE: The cementation technique will have variable results depending on the luting material. Adhesive cementation protocols with composite resins should mainly consider ultrasonic vibration, but also preheating, as strategies for reducing film thickness. The tested resin cement, alongside with IPS Empress Direct composite resin preheated and ultrasonically vibrated, provided the lowest film thickness among the tested materials and techniques.

Metallodrug-protein interaction probed by synchrotron terahertz and neutron scattering spectroscopy.

This experimental work applied coherent synchrotron-radiation terahertz spectroscopy and inelastic neutron scattering to address two processes directly associated with the mode of action of metal-based anticancer agents that can severely undermine chemotherapeutic treatment: drug binding to human serum albumin, occurring during intravenous drug transport, and intracellular coordination to thiol-containing biomolecules (such as metallothioneins) associated with acquired drug resistance. Cisplatin and two dinuclear platinum (Pt)- and palladium (Pd)-polyamine agents developed by this research group, which have yielded promising results toward some types of human cancers, were investigated. Complementary synchrotron-radiation-terahertz and inelastic neutron scattering data revealed protein metalation, through S- and N-donor ligands from cysteine, methionine, and histidine residues. A clear impact of the Pt and Pd agents was evidenced, drug binding to albumin and metallothionein having been responsible for significant changes in the overall protein conformation, as well as for an increased flexibility and possible aggregation.

Purinergic signalling and brain development.

ATP and adenosine are released from cells as a function of their metabolic activity, being important cell-to-cell communication signals. Both purines are also released from neurons in an activity-dependent manner, with several established roles to fine tune brain function in adults, as best heralded by the effects of caffeine, an antagonist of adenosine receptors. Purines are also dynamically released from early neurogenesis and different purine receptors are dynamically expressed throughout development. Accordingly, emerging evidence supports multiple roles for purinergic signalling in the control of different processes of brain development, such as embryonic neurogenesis, migration of principal neurons and interneurons, guidance for neuronal connectivity, synaptogenesis and synaptic stability/elimination. Although major efforts are still required to unravel the time and space-related engagement of the different components of the purinergic system, the relevance of purines in brain development is heralded by their association with neurodevelopmental disorders, positing novel opportunities to understand and correct brain wiring.

Ustilago maydis effector Jsi1 interacts with Topless corepressor, hijacking plant jasmonate/ethylene signaling.

Ustilago maydis is the causal agent of maize smut disease. During the colonization process, the fungus secretes effector proteins that suppress immune responses and redirect the host metabolism in favor of the pathogen. As effectors play a critical role during plant colonization, their identification and functional characterization are essential to understanding biotrophy and disease. Using biochemical, molecular, and transcriptomic techniques, we performed a functional characterization of the U. maydis effector Jasmonate/Ethylene signaling inducer 1 (Jsi1). Jsi1 interacts with several members of the plant corepressor family Topless/Topless related (TPL/TPR). Jsi1 expression in Zea mays and Arabidopsis thaliana leads to transcriptional induction of the ethylene response factor (ERF) branch of the jasmonate/ethylene (JA/ET) signaling pathway. In A. thaliana, activation of the ERF branch leads to biotrophic susceptibility. Jsi1 likely activates the ERF branch via an EAR (ET-responsive element binding-factor-associated amphiphilic repression) motif, which resembles EAR motifs from plant ERF transcription factors, that interacts with TPL/TPR proteins. EAR-motif-containing effector candidates were identified from different fungal species, including Magnaporthe oryzae, Sporisorium scitamineum, and Sporisorium reilianum. Interaction between plant TPL proteins and these effector candidates from biotrophic and hemibiotrophic fungi indicates the convergent evolution of effectors modulating the TPL/TPR corepressor hub.

HIV-1 impact on oral health-related quality of life: a cross-sectional study.

This cross-sectional study compared the Oral-Health-Related Quality of Life (OHRQoL) in HIV negative patients (Group 1, n=129, mean age: 39.9 ± 15.6, 75 females) and HIV+ patients (Group 2, n=670, mean age: 43.2 ± 9.8, 246 females) from the same socio-economic environment using the OHIP-49 questionnaire. OHIP total score were determined by simple summing. A multiple linear regression model was carried out to predict OHIP scores in which HIV+ patients experienced a significantly (p<0.001) worst OHRQoL for total and every dimension. A general linear model was used for estimating the means in the two groups adjusted for covariates included in the previous model. Adjusted means for subscale and total OHIP scores were significantly higher for Group 2 (61.6 ± 6.26 vs. 119.8 ± 3.31) with a large effect size (0.94). HIV+ infection, decayed teeth, prosthodontic and surgical needs, care index, drug use, employment and age presented an independent effect on questionnaire scores. This study shows that HIV+ infection has an independent and negative impact on the OHRQoL while care index presents a positive impact. Additional factors like high decayed teeth, prosthodontic treatment needs and drug use are independently associated with total OHIP scores, presenting a negative effect on OHRQoL.

Effect of restorative timing on shear bond strength of composite resin/calcium silicate-based cements adhesive interfaces.

OBJECTIVES: The aim of this study was to evaluate the shear bond strength inherent to immediate and delayed definitive composite resin restorations, over three different biomaterials. METHODS: Fifty-four aluminum blocks were randomly assigned to six experimental groups (n = 9) according to the biomaterial (Biodentine, TotalFill BC Putty, or PCM) and restoration timing (immediate or 7 days delayed). Regarding the restorative procedures, a two-step self-etch bonding system (Clearfil SE Bond) was applied and subsequently restored using a flowable composite resin (SDR). After shear bond strength testing, fracture pattern analysis was performed. The significance level was set at α = 0.05. RESULTS: Within the immediate groups, Biodentine showed the highest mean shear bond strength value, with no statistically significant differences (P > 0.05) being detected when compared with both TotalFill (P = 0.072) and PCM (P = 0.232) groups. Regarding the delayed restoration groups, Biodentine and TotalFill presented the highest mean shear bond strength value, with statistical differences when compared with PCM (P = 0.002 and P = 0.043, respectively). No statistically significant differences (P > 0.05) were verified between both immediate and delayed groups for each tested biomaterial. Also, a statistically significant association was found between the fracture type and the biomaterial (P < 0.001). CONCLUSIONS AND CLINICAL RELEVANCE: Our findings suggest that all tested biomaterials present suitable alternatives which allow to perform restorative procedures immediately after pulp capping biomaterial placement (3 or 12 min, depending on the bioactive cement), therefore requiring one single appointment to complete the procedure. Biodentine and TotalFill BC Putty have shown superior shear bond strength results when compared with PCM, within a delayed timeframe (7 days).