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Pathogenic variants resulting in protein phosphatase 2A (PP2A) dysfunction result in mild to severe neurodevelopmental delay. PP2A is a trimer of a catalytic (C) subunit, scaffolding (A) subunit, and substrate binding/regulatory (B) subunit, encoded by 19 different genes. De novo missense variants in PPP2R5D (B56δ) or PPP2R1A (Aα) and de novo missense and loss-of-function variants in PPP2CA (Cα) lead to syndromes with overlapping phenotypic features, known as Houge-Janssens syndrome (HJS) types 1, 2, and 3, respectively. Here, we describe an additional condition in the HJS spectrum in 26 individuals with variants in PPP2R5C, encoding the regulatory B56γ subunit. Most changes were de novo and of the missense type. The clinical features were well within the HJS spectrum with strongest resemblance to HJS type 1, caused by B56δ variants. Common features were neurodevelopmental delay and hypotonia, with a high risk of epilepsy, behavioral problems, and mildly dysmorphic facial features. Head circumferences were above average or macrocephalic. The degree of intellectual disability was, on average, milder than in other HJS types. All variants affected either substrate binding (2/19), C-subunit binding (2/19), or both (15/19). Five variants were recurrent. Catalytic activity of the phosphatase was variably affected by the variants. Of note, PPP2R5C total loss-of-function variants could be inherited from a non-symptomatic parent. This implies that a dominant-negative mechanism on substrate dephosphorylation or general PP2A function is the most likely pathogenic mechanism., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2025 American Society of Human Genetics. Published by Elsevier Inc. All rights reserved.)

Purpose: This study details a novel syndromic form of autosomal recessive intellectual disability resulting from recessive variants in GTF3C3, encoding a key component of the DNA-binding transcription factor IIIC, which has a conserved role in RNA polymerase III-mediated transcription., Methods: Exome sequencing, minigene analysis, molecular modeling, RNA polymerase III reporter gene assays, and Drosophila knockdown models were utilized to characterize GTF3C3 variants., Results: Twelve affected individuals from 7 unrelated families were identified with homozygous or compound heterozygous missense variants in GTF3C3 including c.503C>T p.(Ala168Val), c.1268T>C p.(Leu423Pro), c.1436A>G p.(Tyr479Cys), c.2419C>T p.(Arg807Cys), and c.2420G>A p.(Arg807His). The cohort presented with intellectual disability, variable nonfamilial facial features, motor impairments, seizures, and cerebellar/corpus callosum malformations. Consistent with disruptions in intra- and intermolecular interactions observed in molecular modeling, RNA polymerase III reporter assays confirmed that the majority of missense variants resulted in a loss of function. Minigene analysis of the recurrent c.503C>T p.(Ala168Val) variant confirmed the introduction of a cryptic donor site into exon 4, resulting in mRNA missplicing. Consistent with the clinical features of this cohort, neuronal loss of Gtf3c3 in Drosophila induced seizure-like behavior, motor impairment, and learning deficits., Conclusion: These findings confirm that GTF3C3 variants result in an autosomal recessive form of syndromic intellectual disability., Competing Interests: Conflict of Interest The authors declare no conflicts of interest., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Around 60% of individuals with neurodevelopmental disorders (NDD) remain undiagnosed after comprehensive genetic testing, primarily of protein-coding genes 1 . Large genome-sequenced cohorts are improving our ability to discover new diagnoses in the non-coding genome. Here we identify the non-coding RNA RNU4-2 as a syndromic NDD gene. RNU4-2 encodes the U4 small nuclear RNA (snRNA), which is a critical component of the U4/U6.U5 tri-snRNP complex of the major spliceosome 2 . We identify an 18 base pair region of RNU4-2 mapping to two structural elements in the U4/U6 snRNA duplex (the T-loop and stem III) that is severely depleted of variation in the general population, but in which we identify heterozygous variants in 115 individuals with NDD. Most individuals (77.4%) have the same highly recurrent single base insertion (n.64_65insT). In 54 individuals in whom it could be determined, the de novo variants were all on the maternal allele. We demonstrate that RNU4-2 is highly expressed in the developing human brain, in contrast to RNU4-1 and other U4 homologues. Using RNA sequencing, we show how 5' splice-site use is systematically disrupted in individuals with RNU4-2 variants, consistent with the known role of this region during spliceosome activation. Finally, we estimate that variants in this 18 base pair region explain 0.4% of individuals with NDD. This work underscores the importance of non-coding genes in rare disorders and will provide a diagnosis to thousands of individuals with NDD worldwide., (© 2024. The Author(s).)

Purpose: Genome sequencing (GS)-specific diagnostic rates in prospective tightly ascertained exome sequencing (ES)-negative intellectual disability (ID) cohorts have not been reported extensively., Methods: ES, GS, epigenetic signatures, and long-read sequencing diagnoses were assessed in 74 trios with at least moderate ID., Results: The ES diagnostic yield was 42 of 74 (57%). GS diagnoses were made in 9 of 32 (28%) ES-unresolved families. Repeated ES with a contemporary pipeline on the GS-diagnosed families identified 8 of 9 single-nucleotide variations/copy-number variations undetected in older ES, confirming a GS-unique diagnostic rate of 1 in 32 (3%). Episignatures contributed diagnostic information in 9% with GS corroboration in 1 of 32 (3%) and diagnostic clues in 2 of 32 (6%). A genetic etiology for ID was detected in 51 of 74 (69%) families. Twelve candidate disease genes were identified. Contemporary ES followed by GS cost US$4976 (95% CI: $3704; $6969) per diagnosis and first-line GS at a cost of $7062 (95% CI: $6210; $8475) per diagnosis., Conclusion: Performing GS only in ID trios would be cost equivalent to ES if GS were available at $2435, about a 60% reduction from current prices. This study demonstrates that first-line GS achieves higher diagnostic rate than contemporary ES but at a higher cost., Competing Interests: Conflict of Interest The authors declare no conflicts of interest., (Copyright © 2024 American College of Medical Genetics and Genomics. Published by Elsevier Inc. All rights reserved.)

Around 60% of individuals with neurodevelopmental disorders (NDD) remain undiagnosed after comprehensive genetic testing, primarily of protein-coding genes 1 . Increasingly, large genome-sequenced cohorts are improving our ability to discover new diagnoses in the non-coding genome. Here, we identify the non-coding RNA RNU4-2 as a novel syndromic NDD gene. RNU4-2 encodes the U4 small nuclear RNA (snRNA), which is a critical component of the U4/U6.U5 tri-snRNP complex of the major spliceosome 2 . We identify an 18 bp region of RNU4-2 mapping to two structural elements in the U4/U6 snRNA duplex (the T-loop and Stem III) that is severely depleted of variation in the general population, but in which we identify heterozygous variants in 119 individuals with NDD. The vast majority of individuals (77.3%) have the same highly recurrent single base-pair insertion (n.64_65insT). We estimate that variants in this region explain 0.41% of individuals with NDD. We demonstrate that RNU4-2 is highly expressed in the developing human brain, in contrast to its contiguous counterpart RNU4-1 and other U4 homologs, supporting RNU4-2 's role as the primary U4 transcript in the brain. Overall, this work underscores the importance of non-coding genes in rare disorders. It will provide a diagnosis to thousands of individuals with NDD worldwide and pave the way for the development of effective treatments for these individuals., Competing Interests: Competing interests NW receives research funding from Novo Nordisk and has consulted for ArgoBio studio. SJS receives research funding from BioMarin Pharmaceutical. AODL is on the scientific advisory board for Congenica, was a paid consultant for Tome Biosciences and Ono Pharma USA Inc., and received reagents from PacBio to support rare disease research. HLR has received support from Illumina and Microsoft to support rare disease gene discovery and diagnosis. MHW has consulted for Illumina and Sanofi and received speaking honoraria from Illumina and GeneDx. SBM is an advisor for BioMarin, Myome and Tenaya Therapeutics. SMS has received honoraria for educational events or advisory boards from Angelini Pharma, Biocodex, Eisai, Zogenix/UCB and institutional contributions for advisory boards, educational events or consultancy work from Eisai, Jazz/GW Pharma, Stoke Therapeutics, Takeda, UCB and Zogenix. The Department of Molecular and Human Genetics at Baylor College of Medicine receives revenue from clinical genetic testing completed at Baylor Genetics Laboratories. JMMH is a full-time employee of Novo Nordisk and holds shares in Novo Nordisk A/S. DGM is a paid consultant for GlaxoSmithKline, Insitro, and Overtone Therapeutics and receives research support from Microsoft.

Heterozygous ARID1B variants result in Coffin-Siris syndrome. Features may include hypoplastic nails, slow growth, characteristic facial features, hypotonia, hypertrichosis, and sparse scalp hair. Most reported cases are due to ARID1B loss of function variants. We report a boy with developmental delay, feeding difficulties, aspiration, recurrent respiratory infections, slow growth, and hypotonia without a clinical diagnosis, where a previously unreported ARID1B missense variant was classified as a variant of uncertain significance. The pathogenicity of this variant was refined through combined methodologies including genome-wide methylation signature analysis (EpiSign), Machine Learning (ML) facial phenotyping, and LIRICAL. Trio exome sequencing and EpiSign were performed. ML facial phenotyping compared facial images using FaceMatch and GestaltMatcher to syndrome-specific libraries to prioritize the trio exome bioinformatic pipeline gene list output. Phenotype-driven variant prioritization was performed with LIRICAL. A de novo heterozygous missense variant, ARID1B p.(Tyr1268His), was reported as a variant of uncertain significance. The ACMG classification was refined to likely pathogenic by a supportive methylation signature, ML facial phenotyping, and prioritization through LIRICAL. The ARID1B genotype-phenotype has been expanded through an extended analysis of missense variation through genome-wide methylation signatures, ML facial phenotyping, and likelihood-ratio gene prioritization., (© 2023 The Authors. American Journal of Medical Genetics Part C: Seminars in Medical Genetics published by Wiley Periodicals LLC.)

Lettuce produces natural rubber (NR) with an average M w of > 1 million Da in laticifers, similar to NR from rubber trees. As lettuce is an annual, self-pollinating, and easily transformable plant, it is an excellent model for molecular genetic studies of NR biosynthesis. CRISPR/Cas9 mutagenesis was optimized using lettuce hairy roots, and NR-deficient lettuce was generated via bi-allelic mutations in cis-prenyltransferase (CPT). This is the first null mutant of NR deficiency in plants. In the CPT mutant, orthologous CPT counterparts from guayule (Parthenium argentatum) and goldenrod (Solidago canadensis) were expressed under a laticifer-specific promoter to examine how the average M w of NR is affected. No developmental defects were observed in the NR-deficient mutants. The lettuce mutants expressing guayule and goldenrod CPT produced 1.8 and 14.5 times longer NR, respectively, than the plants of their origin. This suggests that, although goldenrod cannot synthesize a sufficiently lengthy NR, goldenrod CPT has the catalytic competence to produce high-quality NR in the cellular context of lettuce laticifers. Thus, CPT alone does not determine the length of NR. Other factors, such as substrate concentration, additional proteins, and/or the nature of protein complexes including CPT-binding proteins, influence CPT activity in determining NR length., (© 2023 The Authors. New Phytologist © 2023 New Phytologist Foundation.)

The minichromosome maintenance (MCM) complex acts as a DNA helicase during DNA replication, and thereby regulates cell cycle progression and proliferation. In addition, MCM-complex components localize to centrosomes and play an independent role in ciliogenesis. Pathogenic variants in genes coding for MCM components and other DNA replication factors have been linked to growth and developmental disorders as Meier-Gorlin syndrome and Seckel syndrome. Trio exome/genome sequencing identified the same de novo MCM6 missense variant p.(Cys158Tyr) in two unrelated individuals that presented with overlapping phenotypes consisting of intra-uterine growth retardation, short stature, congenital microcephaly, endocrine features, developmental delay and urogenital anomalies. The identified variant affects a zinc binding cysteine in the MCM6 zinc finger signature. This domain, and specifically cysteine residues, are essential for MCM-complex dimerization and the induction of helicase activity, suggesting a deleterious effect of this variant on DNA replication. Fibroblasts derived from the two affected individuals showed defects both in ciliogenesis and cell proliferation. We additionally traced three unrelated individuals with de novo MCM6 variants in the oligonucleotide binding (OB)-fold domain, presenting with variable (neuro)developmental features including autism spectrum disorder, developmental delay, and epilepsy. Taken together, our findings implicate de novo MCM6 variants in neurodevelopmental disorders. The clinical features and functional defects related to the zinc binding residue resemble those observed in syndromes related to other MCM components and DNA replication factors, while de novo OB-fold domain missense variants may be associated with more variable neurodevelopmental phenotypes. These data encourage consideration of MCM6 variants in the diagnostic arsenal of NDD., (© 2023. The Author(s).)

Purpose: This study aimed to establish variants in CBX1, encoding heterochromatin protein 1β (HP1β), as a cause of a novel syndromic neurodevelopmental disorder., Methods: Patients with CBX1 variants were identified, and clinician researchers were connected using GeneMatcher and physician referrals. Clinical histories were collected from each patient. To investigate the pathogenicity of identified variants, we performed in vitro cellular assays and neurobehavioral and cytological analyses of neuronal cells obtained from newly generated Cbx1 mutant mouse lines., Results: In 3 unrelated individuals with developmental delay, hypotonia, and autistic features, we identified heterozygous de novo variants in CBX1. The identified variants were in the chromodomain, the functional domain of HP1β, which mediates interactions with chromatin. Cbx1 chromodomain mutant mice displayed increased latency-to-peak response, suggesting the possibility of synaptic delay or myelination deficits. Cytological and chromatin immunoprecipitation experiments confirmed the reduction of mutant HP1β binding to heterochromatin, whereas HP1β interactome analysis demonstrated that the majority of HP1β-interacting proteins remained unchanged between the wild-type and mutant HP1β., Conclusion: These collective findings confirm the role of CBX1 in developmental disabilities through the disruption of HP1β chromatin binding during neurocognitive development. Because HP1β forms homodimers and heterodimers, mutant HP1β likely sequesters wild-type HP1β and other HP1 proteins, exerting dominant-negative effects., Competing Interests: Conflict of Interest The authors declare no conflicts of interest., (Copyright © 2023 American College of Medical Genetics and Genomics. Published by Elsevier Inc. All rights reserved.)

Purpose: ZMYND8 encodes a multidomain protein that serves as a central interactive hub for coordinating critical roles in transcription regulation, chromatin remodeling, regulation of super-enhancers, DNA damage response and tumor suppression. We delineate a novel neurocognitive disorder caused by variants in the ZMYND8 gene., Methods: An international collaboration, exome sequencing, molecular modeling, yeast two-hybrid assays, analysis of available transcriptomic data and a knockdown Drosophila model were used to characterize the ZMYND8 variants., Results: ZMYND8 variants were identified in 11 unrelated individuals; 10 occurred de novo and one suspected de novo; 2 were truncating, 9 were missense, of which one was recurrent. The disorder is characterized by intellectual disability with variable cardiovascular, ophthalmologic and minor skeletal anomalies. Missense variants in the PWWP domain of ZMYND8 abolish the interaction with Drebrin and missense variants in the MYND domain disrupt the interaction with GATAD2A. ZMYND8 is broadly expressed across cell types in all brain regions and shows highest expression in the early stages of brain development. Neuronal knockdown of the DrosophilaZMYND8 ortholog results in decreased habituation learning, consistent with a role in cognitive function., Conclusion: We present genomic and functional evidence for disruption of ZMYND8 as a novel etiology of syndromic intellectual disability., Competing Interests: Conflict of Interest S.V.M., T.B.P., and M.J.G.S. are employees of GeneDx, Inc. All other authors declare no conflicts of interest., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

Neurodevelopmental disorders are highly heterogenous conditions resulting from abnormalities of brain architecture and/or function. FBXW7 (F-box and WD-repeat-domain-containing 7), a recognized developmental regulator and tumor suppressor, has been shown to regulate cell-cycle progression and cell growth and survival by targeting substrates including CYCLIN E1/2 and NOTCH for degradation via the ubiquitin proteasome system. We used a genotype-first approach and global data-sharing platforms to identify 35 individuals harboring de novo and inherited FBXW7 germline monoallelic chromosomal deletions and nonsense, frameshift, splice-site, and missense variants associated with a neurodevelopmental syndrome. The FBXW7 neurodevelopmental syndrome is distinguished by global developmental delay, borderline to severe intellectual disability, hypotonia, and gastrointestinal issues. Brain imaging detailed variable underlying structural abnormalities affecting the cerebellum, corpus collosum, and white matter. A crystal-structure model of FBXW7 predicted that missense variants were clustered at the substrate-binding surface of the WD40 domain and that these might reduce FBXW7 substrate binding affinity. Expression of recombinant FBXW7 missense variants in cultured cells demonstrated impaired CYCLIN E1 and CYCLIN E2 turnover. Pan-neuronal knockdown of the Drosophila ortholog, archipelago, impaired learning and neuronal function. Collectively, the data presented herein provide compelling evidence of an F-Box protein-related, phenotypically variable neurodevelopmental disorder associated with monoallelic variants in FBXW7., Competing Interests: Declaration of interests I.E.S. has served on scientific advisory boards for UCB, Eisai, GlaxoSmithKline, BioMarin, Nutricia, Rogcon, Chiesi, Encoded Therapeutics, Xenon Pharmaceuticals, and Knopp Biosciences; has received speaker honoraria from GlaxoSmithKline, UCB, BioMarin, Biocodex, and Eisai; has received funding for travel from UCB, Biocodex, GlaxoSmithKline, Biomarin and Eisai; has served as an investigator for Zogenix, Zynerba, Ultragenyx, GW Pharma, UCB, Eisai, Anavex Life Sciences, Ovid Therapeutics, Epygenyx, Encoded Therapeutics and Marinus; and has consulted for Zynerba Pharmaceuticals, Atheneum Partners, Ovid Therapeutics, Care Beyond Diagnosis, Epilepsy Consortium and UCB. She may accrue future revenue on pending patent WO2009/086591; her patent for SCN1A testing is held by Bionomics and is licensed to various diagnostic companies; and she has a patent for a molecular diagnostic/therapeutic target for benign familial infantile epilepsy (BFIE) (PRRT2), WO/2013/059884. She receives and/or has received research support from the National Health and Medical Research Council of Australia, Medical Research Future Fund, Health Research Council of New Zealand, CURE, Australian Epilepsy Research Fund, and the National Institute of Neurological Disorders and Stroke of the National Institutes of Health. J.P. is co-chief scientific officer for Global Gene Corp. All other authors declare no competing interests., (Copyright © 2022 American Society of Human Genetics. Published by Elsevier Inc. All rights reserved.)

This review evaluated the knowledge of health professionals (HP) about breastfeeding and factors that leading the weaning. A search was performed in four electronics databases and the grey literature. The search strategy included Mesh terms and synonyms. No language or date restrictions were adopted. Studies that evaluated the knowledge of HP about breastfeeding and weaning were considered eligible. The studies retrieved by the searches were evaluated by two independently examiners. From 1,417 studies retrieved, 35 were included. Many countries and professionals from different health areas were analyzed. No studies evaluated the dentists' knowledge. Although the included HP know the benefits of breastfeeding for health, the length of breastfeeding recommended by the World Health Organization, exclusive or not, was not aligned with all professionals' endorsement. Information about weaning is scarce; however, HP are mindful of the main potential causes of early weaning. The knowledge of HP is conflicting about breastfeeding and unusual about weaning. Furthermore, no studies were found that presented data on the knowledge of dentists on the subject. Thus, assessments of dentists' knowledge and education measures for HP are necessary since they are frequently questioned about these issues.

Massively parallel sequencing has markedly improved mendelian diagnostic rates. This study assessed the effects of custom alterations to a diagnostic genomic bioinformatic pipeline in response to clinical need and derived practice recommendations relative to diagnostic rates and efficiency. The Genomic Annotation and Interpretation Application (GAIA) bioinformatics pipeline was designed to detect panel, exome, and genome sample integrity and prioritize gene variants in mendelian disorders. Reanalysis of selected negative cases was performed after improvements to the pipeline. GAIA improvements and their effect on sensitivity are described, including addition of a PubMed search for gene-disease associations not in the Online Mendelian Inheritance of Man database, inclusion of a process for calling low-quality variants (known as QPatch), and gene symbol nomenclature consistency checking. The new pipeline increased the diagnostic rate and reduced staff costs, resulting in a saving of US$844.34 per additional diagnosis. Recommendations for genomic analysis pipeline requirements are summarized. Clinically responsive bioinformatics pipeline improvements increase diagnostic sensitivity and increase cost-effectiveness., (Crown Copyright © 2021. Published by Elsevier Inc. All rights reserved.)

ARGONAUTE-2 and associated miRNAs form the RNA-induced silencing complex (RISC), which targets mRNAs for translational silencing and degradation as part of the RNA interference pathway. Despite the essential nature of this process for cellular function, there is little information on the role of RISC components in human development and organ function. We identify 13 heterozygous mutations in AGO2 in 21 patients affected by disturbances in neurological development. Each of the identified single amino acid mutations result in impaired shRNA-mediated silencing. We observe either impaired RISC formation or increased binding of AGO2 to mRNA targets as mutation specific functional consequences. The latter is supported by decreased phosphorylation of a C-terminal serine cluster involved in mRNA target release, increased formation of dendritic P-bodies in neurons and global transcriptome alterations in patient-derived primary fibroblasts. Our data emphasize the importance of gene expression regulation through the dynamic AGO2-RNA association for human neuronal development.

Objectives: This randomized controlled clinical trial evaluated the efficacy of sealing carious dentin in controlling the progression of lesions in primary molars for 2-year follow-up., Materials and Methods: Children (6.79 ± 1.81 years, n = 28) presenting primary molars with occlusal caries in the outer half of dentine were randomized and allocated into 2 groups: test (sealing caries with a flowable resin - SC) and control (partial removal of caries followed by restoration - PRC). The primary outcomes were: the clinical success of restorations evaluated by USPHS criteria and the radiographic analysis of caries progression. The children anxiety was evaluated by a Facial Image Scale; and the time required to perform the treatments was registered., Results: In 21 patients evaluated after 2 years, 48 primary molars were analyzed. Clinically, there was no difference between the groups. There was no difference between treatments (p = 0.848) considering lesion progression. The anxiety level did not change after the two interventions (p = 0.650). The treatment time of SC (9.03 ± 1.91 min) was lower (p = 0.002) than the PRC time (17.13 ± 5.26 min)., Conclusion: Sealing carious dentin may be used in dentistry since it did not alter the children anxiety, reduced the chair time and demonstrated clinical success rate and no radiographic difference in relation to the partial caries removal followed by restoration., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Background: Epileptic encephalopathies are a devastating group of neurological conditions in which etiological diagnosis can alter management and clinical outcome. Exome sequencing and gene panel testing can improve diagnostic yield but there is no cost-effectiveness analysis of their use or consensus on how to best integrate these tests into clinical diagnostic pathways., Methods: We conducted a retrospective cost-effectiveness study comparing trio exome sequencing with a standard diagnostic approach, for a well-phenotyped cohort of 32 patients with epileptic encephalopathy, who remained undiagnosed after "first-tier" testing. Sensitivity analysis was included with a range of commercial exome and multigene panels., Results: The diagnostic yield was higher for the exome sequencing (16/32; 50%) than the standard arm (2/32; 6.2%). The trio exome sequencing pathway was cost-effective compared to the standard diagnostic pathway with a cost saving of AU$5,236 (95% confidence intervals $2,482; $9,784) per additional diagnosis; the standard pathway cost approximately 10 times more per diagnosis. Sensitivity analysis demonstrated that the majority of commercial exome sequencing and multigene panels studied were also cost-effective. The clinical utility of all diagnoses was reported., Conclusion: Our study supports the integration of exome sequencing and gene panel testing into the diagnostic pathway for epileptic encephalopathy, both in terms of cost effectiveness and clinical utility. We propose a diagnostic pathway that integrates initial rapid screening for treatable causes and comprehensive genomic screening. This study has important implications for health policy and public funding for epileptic encephalopathy and other neurological conditions., (© 2017 The Authors. Molecular Genetics & Genomic Medicine published by Wiley Periodicals, Inc.)

Certain mutations can cause proteins to accumulate in neurons, leading to neurodegeneration. We recently showed, however, that upregulation of a wild-type protein, Ataxin1, caused by haploinsufficiency of its repressor, the RNA-binding protein Pumilio1 (PUM1), also causes neurodegeneration in mice. We therefore searched for human patients with PUM1 mutations. We identified eleven individuals with either PUM1 deletions or de novo missense variants who suffer a developmental syndrome (Pumilio1-associated developmental disability, ataxia, and seizure; PADDAS). We also identified a milder missense mutation in a family with adult-onset ataxia with incomplete penetrance (Pumilio1-related cerebellar ataxia, PRCA). Studies in patient-derived cells revealed that the missense mutations reduced PUM1 protein levels by ∼25% in the adult-onset cases and by ∼50% in the infantile-onset cases; levels of known PUM1 targets increased accordingly. Changes in protein levels thus track with phenotypic severity, and identifying posttranscriptional modulators of protein expression should identify new candidate disease genes., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Early infantile epileptic encephalopathies (EOEE) are a debilitating spectrum of disorders associated with cognitive impairments. We present a clinical report of a KCNT2 mutation in an EOEE patient. The de novo heterozygous variant Phe240Leu SLICK was identified by exome sequencing and confirmed by Sanger sequencing. Phe240Leu rSlick and hSLICK channels were electrophysiologically, heterologously characterized to reveal three significant alterations to channel function. First, [Cl - ] i sensitivity was reversed in Phe240Leu channels. Second, predominantly K + -selective WT channels were made to favor Na + over K + by Phe240Leu. Third, and consequent to altered ion selectivity, Phe240Leu channels had larger inward conductance. Further, rSlick channels induced membrane hyperexcitability when expressed in primary neurons, resembling the cellular seizure phenotype. Taken together, our results confirm that Phe240Leu is a "change-of-function" KCNT2 mutation, demonstrating unusual altered selectivity in K Na channels. These findings establish pathogenicity of the Phe240Leu KCNT2 mutation in the reported EOEE patient., (Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.)

Objective: To compare the risk for caries in children as determined by Cariogram ® software (CS; Stockholm, Sweden) with and without its microbiological component and by a form based on Cariogram ® (FBC)., Methods: Children (n = 28) aged 3-9 years were included. Data were collected clinically and from anamnesis. The salivary levels of Streptococcus mutans (SM) were evaluated. A linear regression model was used to determine which variables were predictive for each type of risk analysis. Caries risk was the dependent variable and the independent variables were caries experience, related disease, plaque amount, diet frequency, salivary levels of SM, fluoride sources and clinical judgment. A paired Student t-test was used for the following comparisons: (a) CS with and without SM; (b) CS without SM and FBC; (c) CS with SM and FBC., Results: The mean dmft/DMFT was 5.56 ± 2.51. There was no difference between the methods (p < .05). Regardless of caries risk, the children presented the same levels of SM (p = .889). Caries experience, plaque amount, diet frequency and fluoride sources were predictors of caries risk in all assessment methods. Clinical judgment was a significant predictor in CS., Conclusions: Caries experience, plaque amount, diet frequency and fluoride sources are valuable predictors of caries risk; microbiological tests are not necessary for evaluating caries risk in children, which can be assessed similarly by CS without SM and FBC.

Studies have been showing a decrease of bond strength in dentin treated with sodium hypochlorite (NaOCl). The aim of this study was to evaluate the effect of non-thermal argon plasma on the bond strength of a self-etch adhesive system to dentin exposed to NaOCl. Thirty-two flat dentin surfaces of bovine incisors were immersed in 2.5% NaOCl for 30 min to simulate the irrigation step during endodontic treatment. The specimens were divided into four groups (n=8), according to the surface treatment: Control (without plasma treatment), AR15 (argon plasma for 15 s), AR30 (argon plasma for 30 s) and AR45 (argon plasma for 45 s). For microtensile bond strength test, 5 specimens were used per group. In each group, the specimens were hybridized with a self-etch adhesive system (Clearfil SE Bond) and resin composite buildups were constructed. After 48 h of water storage, specimens were sectioned into sticks (5 per tooth, 25 per group) and subjected to microtensile bond strength test (μTBS) until failure, evaluating failure mode. Three specimens per group were analyzed under FTIR spectroscopy to verify the chemical modifications produced in dentin. μTBS data were analyzed using ANOVA and Tamhane tests (p<0.05). AR30 showed the highest μTBS (20.86±9.0). AR15 (13.81±6.4) and AR45 (11.51±6.8) were statistically similar to control (13.67±8.1). FTIR spectroscopy showed that argon plasma treatment produced chemical modifications in dentin. In conclusion, non-thermal argon plasma treatment for 30 s produced chemical changes in dentin and improved the μTBs of Clearfil SE Bond to NaOCl-treated dentin.

The identification of recurrent driver mutations by whole-exome sequencing (WES) of fresh-frozen human cancers and the subsequent development of novel targeted therapies have recently transformed the treatment of many cancers including melanoma. In routine clinical practice, fresh-frozen tissue is rarely available and mutation testing usually needs to be carried out on archival formalin fixed, paraffin embedded (FFPE) tissue, from which DNA is typically fragmented, cross-linked and of lower quality. In this study we aimed to determine whether WES data generated from genomic DNA (gDNA) extracted from FFPE tissues can be produced reliably and of clinically-actionable standard. In this study of ten melanoma patients, we compared WES data produced from analysis of gDNA isolated from FFPE tumour tissue with that isolated from fresh-frozen tumour tissue from the same specimen. FFPE samples were sequenced using both Illumina's Nextera and NimbleGen SeqCap exome capture kits. To examine mutations between the two tissue sources and platforms, somatic mutations in the FFPE exomes were called using the matched fresh tissue sequence as a reference. Of the 10 FFPE DNA samples, seven Nextera and four SeqCap samples passed library preparation. On average, there were 5341 and 2246 variants lost in FFPE compared to matched fresh tissue utilising Nextera and SeqCap kits, respectively. In order to explore the feasibility of future clinical implementation of WES, FFPE variants in 27 genes of important clinical relevance in melanoma were assessed. The average concordance rate was 43.2% over a total of 1299 calls for the chosen genes in the FFPE DNA. For the current clinically most important melanoma mutations, 0/3 BRAF and 6/8 (75%) NRAS FFPE calls were concordant with the fresh tissue result, which was confirmed using a Sequenom OncoCarta Panel. The poor performance of FFPE WES indicates that specialised library construction to account for low quality DNA and further refinements will be necessary before this approach could be used for routine clinical decision making over currently preferred techniques., (Copyright © 2016 Royal College of Pathologists of Australasia. Published by Elsevier B.V. All rights reserved.)

Asparagine Synthetase Deficiency is a recently described cause of profound intellectual disability, marked progressive cerebral atrophy and variable seizure disorder. To date there has been limited functional data explaining the underlying pathophysiology. We report a new case with compound heterozygous mutations in the ASNS gene (NM&#95;183356.3:c. [866G>C]; [1010C>T]). Both variants alter evolutionarily conserved amino acids and were predicted to be pathogenic based on in silico protein modelling that suggests disruption of the critical ATP binding site of the ASNS enzyme. In patient fibroblasts, ASNS expression as well as protein and mRNA stability are not affected by these variants. However, there is markedly reduced proliferation of patient fibroblasts when cultured in asparagine-limited growth medium, compared to parental and wild type fibroblasts. Restricting asparagine replicates the physiology within the blood-brain-barrier, with limited transfer of dietary derived asparagine, resulting in reliance of neuronal cells on intracellular asparagine synthesis by the ASNS enzyme. These functional studies offer insight into the underlying pathophysiology of the dramatic progressive cerebral atrophy associated with Asparagine Synthetase Deficiency., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Purpose. This study tested the null hypothesis that different classes of direct restorative dental materials: silorane-based resin, low-shrinkage and conventional (non-flowable and flowable) resin-based composite (RBC) do not differ from each other with regard to polymerization shrinkage, depth of cure or microhardness. Methods. 140 RBC samples were fabricated and tested by one calibrated operator. Polymerization shrinkage was measured using a gas pycnometer both before and immediately after curing with 36 J/cm(2) light energy density. Depth of cure was determined, using a penetrometer and the Knoop microhardness was tested from the top surface to a depth of 5 mm. Results. Considering polymerization shrinkage, the authors found significant differences (p < 0.05) between different materials: non-flowable RBCs showed lower values compared to flowable RBCs, with the silorane-based resin presenting the smallest shrinkage. The low shrinkage flowable composite performed similarly to non-flowable with significant statistical differences compared to the two other flowable RBCs. Regarding to depth of cure, low-shrinkage flowable RBC, were most effective compared to other groups. Microhardness was generally higher for the non-flowable vs. flowable RBCs (p < 0.05). However, the values for low-shrinkage flowable did not differ significantly from those of non-flowable, but were significantly higher than those of the other flowable RBCs. Clinical Significance. RBCs have undergone many modifications as they have evolved and represent the most relevant restorative materials in today's dental practice. This study of low-shrinkage RBCs, conventional RBCs (non-flowable and flowable) and silorane-based composite-by in vitro evaluation of volumetric shrinkage, depth of cure and microhardness-reveals that although filler content is an important determinant of polymerization shrinkage, it is not the only variable that affects properties of materials that were tested in this study.

This study used atomic force microscopy to evaluate the effects of an in-office dental bleaching protocol on the surface roughness of 3 resins: microfilled, microhybrid, and nanofilled. Two disks of each resin were prepared and evaluated. Twenty-four areas (5 x 5 µm) were scanned using an atomic force microscope. Then, each disk was treated with 35% hydrogen peroxide activated by a light emitting diode. After bleaching, the disks were scanned again. Data from the 24 areas before and after bleaching were evaluated qualitatively using topographical and 3-dimensional images, as well as profile lines. Quantitatively, roughness data (mean roughness and root mean square) were statistically evaluated using standard ANOVA and Student-Newman-Keuls tests (P < 0.05). The bleaching procedure increased roughness for each resin analyzed. Based on the results, it was concluded that in-office bleaching with 35% hydrogen peroxide-based gel significantly increased the surface roughness of microfilled, microhybrid, and nanofilled resins.

The aim of this study was to evaluate, using atomic force microscopy, the effect of two different bleaching agents on the modification of dental composites materials. This modification will be judged by analyzing the variation of surface roughness and surface morphology of two different composites: one containing nanoparticles and other consisting of microhybrid resin. The bleaching was performed by using two different concentrations of hydrogen peroxide: HP Blue (20% hydrogen peroxide) and Whiteness HP Maxx (35% hydrogen peroxide). Disks of Esthet X and Filtek™ Z350 composites were used. Atomic force microscopy was used for analyses of the same place of the sample before and after treatment. A total of 12 analyses were performed per group (n = 12). The samples were analyzed qualitatively by evaluating morphological changes in the images and quantitatively by using roughness parameters (Ra). Data were analyzed statistically using Kruskal-Wallis, Mann-Whitney, and Friedman tests (P < 0.05). Changes were observed both qualitatively and quantitatively only in the groups where Esthet X resin was used. The use of hydrogen peroxide bleaching agents caused changes only in the surface of microhybrid composites, with no changes being observed in the composite containing nanoparticles. Despite being even significant, these alterations are clinically slight and can be eliminated by polishing them., (Copyright © 2013 Wiley Periodicals, Inc.)

Background: Deep sequencing of single cell-derived cDNAs offers novel insights into oncogenesis and embryogenesis. However, traditional library preparation for RNA-seq analysis requires multiple steps with consequent sample loss and stochastic variation at each step significantly affecting output. Thus, a simpler and better protocol is desirable. The recently developed hyperactive Tn5-mediated library preparation, which brings high quality libraries, is likely one of the solutions., Results and Conclusions: Here, we tested the applicability of hyperactive Tn5-mediated library preparation to deep sequencing of single cell cDNA, optimized the protocol, and compared it with the conventional method based on sonication. This new technique does not require any expensive or special equipment, which secures wider availability. A library was constructed from only 100 ng of cDNA, which enables the saving of precious specimens. Only a few steps of robust enzymatic reaction resulted in saved time, enabling more specimens to be prepared at once, and with a more reproducible size distribution among the different specimens. The obtained RNA-seq results were comparable to the conventional method. Thus, this Tn5-mediated preparation is applicable for anyone who aims to carry out deep sequencing for single cell cDNAs., (Copyright © 2012 Wiley Periodicals, Inc.)

The aim of this study was to evaluate the effects of different pretreatments on the bond strength of a dual-cure resin core to 3 types of fiber posts. Bond strength was measured using a push-out design. One-sided t-Test of Hypothesis with unknown variance was performed (p-values < 5%). Sandblasting abrasion with 50 micro alumina particles at a specific distance, pressure and time was the only surface treatment in DT Light Post and Transluma Post that increased the bond strength to dual cure resin composite cores. FRC Postec Plus post did not shown an increase in bond strength in any group.

The objective of this study was to evaluate the effect of two bleaching agents (10% and 35% hydrogen peroxide) on the color stability and surface roughness of two composites, one nanohybrid and one nanoparticle. Specimens were polished, aged, stained, bleached and polished again. The action of the bleaching agents on the composites was analyzed using a profilometer (surface roughness) and a spectrophotometer (color stability). The effect of polishing the composites on the surface roughness and the resumption of the composite color was also evaluated. The results were analyzed statistically by ANOVA and Tukey's test at 5% significance level. The analysis indicated that the nanohybrid composite was more affected by staining. The bleaching agents were not able to promote bleaching of either composite over the evaluation period. Surface polishing returned nanohybrid composite to its original color condition, which did not occur for the nanoparticle composite. Additionally, polishing did not return the surface roughness of either composite to its original value. It may be concluded that polishing surface after bleaching should not be the treatment of choice, as it was not possible to reverse the roughness of the composites to their original values, suggesting that a more extensive and irreversible degradation might have occurred.

Using variants from the 1000 Genomes Project pilot European CEU dataset and data from additional resequencing studies, we densely genotyped 183 non-HLA risk loci previously associated with immune-mediated diseases in 12,041 individuals with celiac disease (cases) and 12,228 controls. We identified 13 new celiac disease risk loci reaching genome-wide significance, bringing the number of known loci (including the HLA locus) to 40. We found multiple independent association signals at over one-third of these loci, a finding that is attributable to a combination of common, low-frequency and rare genetic variants. Compared to previously available data such as those from HapMap3, our dense genotyping in a large sample collection provided a higher resolution of the pattern of linkage disequilibrium and suggested localization of many signals to finer scale regions. In particular, 29 of the 54 fine-mapped signals seemed to be localized to single genes and, in some instances, to gene regulatory elements. Altogether, we define the complex genetic architecture of the risk regions of and refine the risk signals for celiac disease, providing the next step toward uncovering the causal mechanisms of the disease.

The aim of this study was to describe a technique and report the success of a clinical case in which a modified technique of in-office bleaching with hydrogen peroxide at 35% was performed for two sessions in a premolar pigmented by amalgam followed by placement of a posterior direct composite resin restoration.

Dentinal surfaces prepared with air abrasion have considerably different characteristics from those prepared with conventional instruments. Different hybrid layer morphology and thickness occur, which can result in differences in the quality of restorations placed on dentinal surfaces prepared with a diamond bur compared to surfaces prepared using air abrasion. The objective of this study was to compare the hybrid layer thickness and morphology formed utilizing Scotchbond Multi-Purpose Plus (SBMP) on dentin prepared with a diamond bur in a high-speed handpiece and on dentin prepared using air abrasion. Flat dentin surfaces obtained from five human teeth were prepared using each method, then treated with the dentin adhesive system according to manufacturer's instructions. After a layer of composite was applied, specimens were sectioned, flattened, polished, and prepared for scanning electron microscopy. Ten different measurements of hybrid layer thickness were obtained along the bonded surface in each specimen. SBMP produced a 3.43 ± 0.75 µm hybrid layer in dentin prepared with diamond bur. This hybrid layer was regular and found consistently. In the air abrasion group, SBMP produced a 4.94 ± 1.28 µm hybrid layer, which was regular and found consistently. Statistical ANOVA (P = 0.05) indicated that there was a statistically significant difference between the groups. These data indicate that the air abrasion, within the parameters used in this study, provides a thick hybrid layer formation.

Monozygotic (MZ) twin pair discordance for childhood-onset Type 1 Diabetes (T1D) is ∼50%, implicating roles for genetic and non-genetic factors in the aetiology of this complex autoimmune disease. Although significant progress has been made in elucidating the genetics of T1D in recent years, the non-genetic component has remained poorly defined. We hypothesized that epigenetic variation could underlie some of the non-genetic component of T1D aetiology and, thus, performed an epigenome-wide association study (EWAS) for this disease. We generated genome-wide DNA methylation profiles of purified CD14+ monocytes (an immune effector cell type relevant to T1D pathogenesis) from 15 T1D-discordant MZ twin pairs. This identified 132 different CpG sites at which the direction of the intra-MZ pair DNA methylation difference significantly correlated with the diabetic state, i.e. T1D-associated methylation variable positions (T1D-MVPs). We confirmed these T1D-MVPs display statistically significant intra-MZ pair DNA methylation differences in the expected direction in an independent set of T1D-discordant MZ pairs (P = 0.035). Then, to establish the temporal origins of the T1D-MVPs, we generated two further genome-wide datasets and established that, when compared with controls, T1D-MVPs are enriched in singletons both before (P = 0.001) and at (P = 0.015) disease diagnosis, and also in singletons positive for diabetes-associated autoantibodies but disease-free even after 12 years follow-up (P = 0.0023). Combined, these results suggest that T1D-MVPs arise very early in the etiological process that leads to overt T1D. Our EWAS of T1D represents an important contribution toward understanding the etiological role of epigenetic variation in type 1 diabetes, and it is also the first systematic analysis of the temporal origins of disease-associated epigenetic variation for any human complex disease., Competing Interests: The authors have declared that no competing interests exist.

Dentinal surfaces prepared with an Er:YAG laser have distinctly different characteristics compared to those prepared with conventional instruments. The objective of this study was to evaluate the shear bond strength of a total-etch adhesive system and a self-etch adhesive system to dentin, prepared conventionally or with an Er:YAG laser. Specimens that were prepared with a diamond bur and treated with the total-etch adhesive showed the highest mean bond strength, followed by specimens prepared with the laser and treated with the total-etch adhesive, specimens prepared with a diamond bur and treated with the self-etch adhesive, and specimens prepared with the laser and treated with the self-etch adhesive, but there were no differences among the groups (P > 0.05). These data indicate that the tested adhesive systems promote similar shear bond strengths, whether the dentin is prepared with a diamond bur or an Er:YAG laser, and that an Er:YAG laser is a suitable cavity preparation method that promotes an adequate surface for a posterior adhesive procedure.

This study investigated the effects of the cement type and the water storage time on the push-out bond strength of a glass fiber post. Glass fiber posts (Fibrekor, Jeneric Pentron) were luted to post spaces using a self-cured resin cement (C&B Cement [CB]), a glass ionomer cement (Ketac Cem [KC]) or a resin-modified glass ionomer cement (GC FujiCEM [FC]) according to the manufacturers' instructions. For each luting agent, the specimens were exposed to one of the following water storage times (n=5): 1 day (T1), 7 days (T7), 90 days (T90) and 180 days (T180). Push-out tests were performed after the storage times. Control specimens were not exposed to water storage, but subjected to the push-out test 10 min after post cementation. Data (in MPa) were analyzed by Kruskal-Wallis and Dunn`s test (α=0.05). Cement type and water storage time had a significant effect (p<0.05) on the push-out bond strength. CB showed significantly higher values of retention (p<0.05) than KC and FC, irrespective of the water storage time. Water storage increased significantly the push-out bond strength in T7 and T90, regardless of the cement type (p<0.05). The results showed that fiber posts luted to post spaces with the self-cured resin cement exhibited the best bonding performance throughout the 180-day water storage period. All cements exhibited a tendency to increase the bond strength after 7 and 90 days of water storage, decreasing thereafter.

A tridimensional surface roughness test evaluation is a nondestructive method that can be used to perform a topographic analysis of different surface treatments for glass and quartz fiber posts. This study divided 75 fiber posts into three groups according to their manufacturer. Each group was divided into five subgroups (n = 5), according to the surface treatment each received: immersion in hydrofluoric acid, sandblasting, immersion in hydrogen peroxide, sandblasting followed by immersion in hydrofluoric acid, or sandblasting followed by immersion in hydrogen peroxide. Surface roughness was measured using a tridimensional surface roughness test and analyzed with three-dimensional analysis software. Results were statistically analyzed using Student's t-test. The only surface treatment to modify the surface topography of glass and quartz fiber posts and provide a significant increase in roughness was sandblasting airborne-particle abrasion with 50 micro alumina at a distance of 30 mm, using 2.5 bars of pressure for five seconds.

To investigate the influence of the C-factor (Cf) and light-curing mode (LCM) on gap formation in resin composite (RC) restorations. Cylindrical Class I cavities with a 5.0 mm diameter andthree different depths (1.0, 2.0 and 3.0 mm) were prepared in the occlusal surfaces of 60 human molars and restored with P60 (P) and Supreme (Su). RCs were light-cured in accordance with two modes: Standard (S)--850mW/cm2/20 seconds and Ramp (R)-100 up to 1000mW/cm2/10 seconds +1000mW/cm2/10 seconds. After storage in distilled water, the restorations were cut into three slices and the gap widths were analyzed in a 3D-scanning system. The data were analyzed by ANOVA and Student-Newman-Keul's test (alpha = 0.05). ANOVA detected significant influence for the RC, Cf and LCM independent factors and for the double interactions RC vs Cf and LCM vs Cf. Smaller gap formation was found for cavities restored with Su. R was responsible for the smaller gap formation. The highest gap formation was found for cavities with Cf = 3.4, followed by Cf = 2.6 and 1.8 without statistical differences between them. These findings suggest that Cf played an essential role in gap formation. R LCM may allow RC relaxation during polymerization reaction. Finally, nanocomposites (Su) may lead to less gap formation at the resin-dentin interface.

The aim of this study was to evaluate the stress distribution in the cervical region of a sound upper central incisor in two clinical situations, standard and maximum masticatory forces, by means of a 3D model with the highest possible level of fidelity to the anatomic dimensions. Two models with 331,887 linear tetrahedral elements that represent a sound upper central incisor with periodontal ligament, cortical and trabecular bones were loaded at 45 masculine in relation to the tooth's long axis. All structures were considered to be homogeneous and isotropic, with the exception of the enamel (anisotropic). A standard masticatory force (100 N) was simulated on one of the models, while on the other one a maximum masticatory force was simulated (235.9 N). The software used were: PATRAN for pre- and post-processing and Nastran for processing. In the cementoenamel junction area, tensile forces reached 14.7 MPa in the 100 N model, and 40.2 MPa in the 235.9 N model, exceeding the enamel's tensile strength (16.7 MPa). The fact that the stress concentration in the amelodentinal junction exceeded the enamel's tensile strength under simulated conditions of maximum masticatory force suggests the possibility of the occurrence of non-carious cervical lesions such as abfractions.

This study compared the compressive strength of a composite resin and compomer photoactivated with a conventional quartz-tungsten halogen-light (XL 3000, 3M/SPE) and a blue light-emitting diode (LED) (SmartLite PS; Dentsply/De Trey). Forty disc-shaped specimens were prepared using a split polytetrafluoroethylene matrix (4.0 mm diameter x 8.0 mm hight) in which the materials were inserted incrementally. The curing time of each increment was of 40 s with the QTH and 10 s with the LED. The specimens were randomly assigned to 4 groups (n=10), according to the light source and the restorative material. After storage in distilled water at 37 degrees C +/- 2 degrees C for 24 h, the specimens was tested in compressive strength in a universal testing machine with load cell of 500 kgf running at a crosshead speed of 0.5 mm/min. Data (in MPa) were analyzed statistically by ANOVA and Student-Newman-Keuls test (p<0.05). For the composite resin, light curing with the QTH source did not produce statistically significant difference (p>0.05) in the compressive strength when compared to light curing with the LED source. However, light curing of the compomer with the QTH source resulted in significantly higher compressive strength than the use of the LED unit (p>0.05). The composite resin presented significantly higher (p>0.05) compressive strength than the compomer, regardless of the light source. In conclusion, the compressive strength of the tested materials photoactivated with a QTH and a LED light source was influenced by the energy density employed and the chemical composition of the esthetic restorative materials.

This study compared the thickness of the hybrid layer formed using Scotchbond Multi-Purpose Plus, Single Bond 2, Prime & Bond 2.1 and Xeno III on a dentin surface prepared with a diamond bur in a high speed handpiece or prepared with an Er:YAG laser used with two parameters of pulse energy (200 and 400 mJ) and two parameters of frequency (4 and 6 Hz). Flat dentin surfaces obtained from 20 human third molars were treated with the two methods and were then prepared with the dentin adhesive systems according to the manufacturers' instructions. After a layer of composite was applied, the specimens were sectioned, flattened, polished and prepared for Scanning Electronic Microscopy observation. Five different measurements of the hybrid layer thickness were obtained along the bonded surface in each specimen. The results were statistically analyzed using Analysis of Variance and Student-Newman-Keuls tests (p < or = 0.05). When analyzing the hybrid layer thickness and comparing the cavity preparation method, four groups were formed: Group I (diamond bur) > Group II (Laser 200 mJ/4 Hz) = Group III (Laser 200 mJ/6 Hz) > Group IV (Laser 400 mJ/4 Hz) > Group V (Laser 400 mJ/6 Hz). When comparing the dentin adhesive systems, there were no statistically significant differences. These results showed that the four tested dentin adhesive systems produced a 2.90 +/- 1.71 microm hybrid layer in dentin prepared with a diamond bur. This hybrid layer was regular and routinely found. In the laser groups, the dentin adhesive systems produced hybrid layers ranging from 0.41 +/- 1.00 microm to 2.06 +/- 2.49 microm, which were very irregular and not routinely found. It was also concluded that the Er:YAG laser, with the parameters used in this experiment, has a negative influence on the formation of a hybrid layer and cavity preparation methods influence formation of the hybrid layer.

The aim of this study was to evaluate the remaining dentine/cementum thickness using Gates-Glidden burs in serial and crown-down sequences and to observe which of the two sequences is the safest for preparing mesial roots of molars. Thirty-six left and right human mandibular first molars were selected. Standard access cavities were made and initially explored with Flexofiles sizes 10 and 15 until the tip was visible at the apex. The teeth were embedded in a muffle specially developed for this study using a PVC tube with two parallel metal rods in its lid. Each tooth-block was sectioned 3 mm apically to the furcation using a low-speed saw with a diamond disc. The tooth-block was examined under a microscope and an initial image was captured by a digital video system with 8 X and 12 X magnifications. Finally, the tooth-blocks were reassembled in the muffle so that the canals could be instrumented. After instrumentation the area of each mesial canal as well as the smallest distance to the root furcation were measured again. The mesio-buccal canals (crown-down order) and the mesio-lingual canals (serial sequence) presented an average area of 0.46 +/- 0.16 mm(2) and 0.88 +/- 0.27 mm(2) (P < 0.01), respectively. The mean values of the smallest distance to the furcation for the mesio-buccal and mesio-lingual canals were 0.66 +/- 0.19 mm and 0.39 +/- 0.13 mm (P < 0.01), respectively. The remaining dentine/cementum thickness using Gates-Glidden burs was greater in the crown-down sequence than in the serial sequence.

The concepts of health promotion, self-care and community participation emerged during the 1970s and, since then, their application has grown rapidly in the developed world, showing evidence of effectiveness. In spite of this, a major part of the population in the developing countries still has no access to specialized dental care such as endodontic treatment, dental care for patients with special needs, minor oral surgery, periodontal treatment and oral diagnosis. This review focuses on a program of the Brazilian Federal Government named CEOs (Dental Specialty Centers), which is an attempt to solve the dental care deficit of a population that is suffering from oral diseases and whose oral health care needs have not been addressed by the regular programs offered by the SUS (Unified National Health System). Literature published from 2000 to the present day, using electronic searches by Medline, Scielo, Google and hand-searching was considered. The descriptors used were Brazil, Oral health, Health policy, Health programs, and Dental Specialty Centers. There are currently 640 CEOs in Brazil, distributed in 545 municipal districts, carrying out dental procedures with major complexity. Based on this data, it was possible to conclude that public actions on oral health must involve both preventive and curative procedures aiming to minimize the oral health distortions still prevailing in developing countries like Brazil.

The aim of this study was to compare two self-etching and a total-etch adhesive systems by assessing their shear bond strength to bovine enamel and the microleakage on class V composite restorations prepared on bovine enamel. Bovine teeth selected and allocated in three groups: Group 1: Scothbond Multi-Purpose; Group 2: Clearfil Liner Bond 2V; Group 3: Etch & Prime 3.0. For the microleakage test, each group was composed of ten class V restorations on the buccal surface. Two examiners attributed scores ranging from 0 (without leakage) to 3 (maximum leakage) to determine silver nitrate penetration at enamel-composite interface. Microleakage data were analyzed statistically by Kruskal-Wallis and Mann-Whitney tests at 5% significance level. For the bond strength test, ten teeth of each group were included, had their buccal surfaces flattened in order to obtain a 3-mm-diameter area to which a resin cylinder was bonded. After one week, the specimens were tested in shear strength at a crosshead speed of 0.5 mm/min. Bond strength data were treated by ANOVA and LSD tests at 5% significance level. The debonded interfaces were examined under scanning electron microscopy. No leakage was observed along enamel margins. Means (+/- SD) in MPa were: 18.75 (+/-5.83), 22.17 (+/-4.95) and 14.93 (+/-6.7) for Groups 1, 2, and 3, respectively. According to the results of this study, the self-etching primer systems presented statistically similar behavior (p>0.05) to that of the total-etch adhesive system (used as a control), not only regarding marginal leakage at bovine enamel-composite resin interface, but also regarding the shear bond strength of the bovine enamel. However, the self-etching primer systems differed significantly (p>0.05) to each other, with better results for Clearfil Liner Bond 2V. In conclusion, the self-etching primer systems had a performance comparable to that of the total-etch adhesive system.