Two-Year Survival of Ceramic Endocrowns and Partial Coverage Ceramic Restorations with Fiber Post: A 2-Year Double-Blind Randomized Clinical Trial.

PURPOSE: This double-blind randomized clinical trial (RCT) aimed to evaluate the 2-year survival rates of endocrowns and partial coverage ceramic restorations (PCCR) with fiber posts. MATERIAL AND METHODS: Forty (40) participants fulfilled the elegibility criteria, and they were randomly allocated in 2 groups: Endocrown or PCCR+post. The survival rates were assessed based on USPHS modified and radiographic examinations. A Chi-square test was used to assess the distribution of characteristics between groups. Kaplan-Meier and Log-rank tests were used to estimate the survival rate. To evaluate the association between survival of the restorations and the explanatory variables, the Multivariate Cox regression model was used. Only variables presenting p⟨0.20 were maintained in final model (α= 0.05). RESULTS: The highest 2-year survival rates were recorded for the Endocrown group (100%), whereas the PCCR+post group exhibited the lowest performance (66.7%). Most of the restoration failures was due to lack of marginal adaption, fracture, and recurrent caries. Cox Regression unadjusted analysis showed that only type of restoration presented a significant effect (p⟨0.20). Thus, adjusted analysis was not performed. CONCLUSIONS: Endocrowns appear to be a promising conservative restorative option and to be feasible and reliable approach restoring endodontically.

Giomer composite compared to glass ionomer in occlusoproximal ART restorations of primary molars: 24-month RCT.

BACKGROUND: Occlusoproximal restorations of primary molars usually fail, so it is necessary to investigate new materials that may overcome this challenge. Thus, this trial aimed to evaluate the longevity of occlusoproximal ART restorations in primary molars using a glass ionomer cement - GIC (Equia Forte® - GC Corp) and a Giomer resin composite - GCR (Beautifil Bulk Restorative® - Shofu Inc) after 24 months. METHODS: One hundred and eighty-two (182) children aged from 4 to 8 years were selected and randomly assigned to GIC or GCR. A paediatric dentist treated them in the school setting in Cerquilho, Brazil, and the restorations were assessed after 3, 6, 12, 18 and 24 months. The primary outcome was the restoration survival, evaluated using the Kaplan-Meier and superiority Cox regression analyses. Intention to treat (ITT) was performed as a sensitivity analysis using superiority test P value and confidence interval (CI = 95%). Independent variables included gender, age, molar, jaw, cavity volume and caries experience. RESULTS: The restoration survival after 24 months was GIC = 58.1% and GCR = 49.1% (HR = 1.24; CI = 0.97-1.59). ITT analysis showed a success of GIC = 61.1% and GCR = 52.2% (RR = 1.17; CI = 0.91-1.52). The superiority hypothesis was not proved in both analyses (P > 0.05). CONCLUSION: GCR does not have superior longevity than GIC in occlusoproximal ART restorations of primary molars.

Noninvasive prenatal testing: benefits and limitations of the available tests.

BACKGROUND: The „gold standard“ for prenatal diagnosis of aneuploidies is provided by the karyotype, which has high accuracy, but is dependent on invasive procedures, which generate risk of fetal loss. Different methodologies of development of noninvasive prenatal genetic tests (NIPT) for tracking aneuploidies, including sex chromosomes, have been made available for clinical use, for some microdeletions and triploids and for exclusion of paternity. These exams make use of three methodological tools: s-MPS, t-MPS and SNP. Genetic tests, despite the high cost, cover a broader range of clinical applications, have the advantage that can be performed early, with high accuracy, and low false positive rate. Type of article: Review. SETTING: Department of Obstetrics and Gynecology, Science College of Santa Casa of São Paulo (FSMSCSP), São Paulo-SP, Brazil. DESIGN AND METHODS: This study was a non-asystematic review, which searched PubMed / MEDLINE as a research source and aimed at the compilation of data, which allowed approaching the evolution, the technical and methodological advances of the available tests, the recognition of its benefits, limitations and future perspectives on NIPT. CONCLUSION: NIPT stand out for being applied earlier during the pregnancy with high accuracy and low false-positive rates, including a broad spectrum of clinical applications. The t-MPS is a recent technique used to evaluate aneuploidy that shows greater accuracy and lower cost than the s-MPS, but that is limited to being applied only to the most common aneuploidies. The SNP technique can search for more genetic conditions, besides presenting better accuracy.

Screening of hydrogel-based scaffolds for dental pulp regeneration-A systematic review.

OBJECTIVE: The aim of this systematic review was to evaluate the most appropriate hydrogel scaffold type (natural, synthetic or hybrid) to be applied with stem cells for dental pulp regeneration. The findings should help clinicians make an informed choice about the appropriate scaffold to be applied for this approach. DESIGN: Three electronic databases were searched (Medline, Web of Science and Scopus). The review was conducted based on the Preferred Reporting Items for Systematic Reviews and Meta-analysis (PRISMA). RESULTS: From 4990 potentially relevant studies initially identified, 18 papers fulfilled the eligibility criteria and were considered for this review. Natural scaffolds were applied in most studies. Collagen was the most studied scaffold. In 5 of 10 studies, only growth factors were added to the constructs. Even without growth factors, these scaffolds containing stem cells were able to support the formation of dentin. The synthetic scaffolds were the least studied. Only 4 studies were selected, and in 3 of them, the same scaffold (Puramatrix) was evaluated. Puramatrix by itself was unable to form dental pulp when dental pulp stem cells were not present. Synthetic and hybrid hydrogels were unable to attract stem cells from the host. The presence of growth factors in these constructs seems to be of relevance since dental pulp tissue formation was achieved only when the hybrid scaffold was applied with growth factors. CONCLUSION: All types of hydrogel-based scaffolds, when containing mesenchymal stem cells, are able to form connective tissue with different degrees of similarity to dental pulp. However, current data is too heterogeneous to compare and identify the advantages of any specific scaffold.

Correlation between terminal restriction fragments and flow-FISH measures in samples over wide range telomere lengths.

OBJECTIVES: Terminal restriction fragment (TRF) analysis of human telomeres was used to calibrate flow-fluorescence in situ hybridization (FF) measures of telomere lengths to expand the range of measures and increase power of resolution of our previously published protocol. TRF data used as the gold standard should be obtained by electrophoresis with suitable resolution applied to appropriately isolated genomic DNA. When we considered TRF attained by correct methods, we found our method to be insufficiently accurate, thus we have reviewed our previously published FF protocol to obtain the best coefficient of determination (r(2)) between our experimental results and valid TRF lengths. MATERIALS AND METHODS: Using human telomere-specific PNA probe, Cy5-OO-(CCCTAA)3 , we measured telomere lengths of continuous cell line and of peripheral blood lymphocytes by FF. We modified hybridization, stringency, negative control handling, stoichiometric DNA staining and telomere fluorescence assessment of the protocol. RESULTS: We realized a procedure with increased power of resolution, improved TRF versus FF r(2) values that allowed simultaneous analysis of DNA and telomere duplication. Notwithstanding multiple steps in formamide sampling, recovery was satisfactory. DISCUSSION: The reviewed FF protocol appeared at least as suitable as the TRF method. Measures obtained by TRF can be affected by chromosome end variability, DNA fragmentation, incomplete digestion and unsuitable electrophoresis. In contrast, the FF technique analyses telomeric sequences confined to preserved nuclei thus overcome most previous limitations. As yet, however, the FF telomere measure cannot be performed together with immunophenotyping and/or generation study by the dye dilution method.

Comparison of validation methods for the diagnosis of occlusal caries in primary molars.

AIM: To evaluate the reproducibility of 7 validation methods used for caries diagnosis in primary teeth. METHODS: Seventy-two occlusal sites were selected on 40 primary molars. The sites were evaluated independently by 3 experienced examiners using validation methods that involved direct assessment, i.e. by using a (1) magnifying glass (8×) and (2) stereomicroscope (35×), or indirect assessment i.e. by using (3) photographs, (4) slide projections of photographs, (5) stereomicroscope (35×) photographs, (6) stereomicroscope (35×) slide projections, and (7) projections of polarised light microscope slides. Cohen's kappa coefficients were calculated and subjected to the Kruskal-Wallis test at a significance level of 5%. RESULTS: The mean inter-examiner kappa values for the validation methods were 0.31-0.51. There were statistically significant differences (p<0.05) between methods 1 and 3, 1 and 4, 2 and 4, 4 and 5, 4 and 6, and 4 and 7. Moderate agreement was observed for all methods except methods 1 and 4, for which the agreement was fair. CONCLUSIONS: The inter-examiner agreement for all validation methods for caries diagnosis was moderate, except for the method based on indirect assessment by slide projection, which showed low agreement.

Erosive effects of beverages in the presence or absence of caries simulation by acidogenic challenge on human primary enamel: an in vitro study.

AIM: To evaluate in vitro the erosive effects of beverages in the presence or absence of caries simulation (acidogenic challenge) on the microhardness of primary enamel. METHODS: Forty human primary teeth were submitted to the erosive effects: 3 × 20-min-long daily immersion in fresh orange juice (orange group), strawberry yogurt drink (yog group), or cola soft drink (cola group) separately or in combination with acidogenic challenge (pH cycling for 10 days). Specimens were also submitted to acidogenic challenge alone, and in the negative control group specimens were not submitted to any treatment. Mineral loss was evaluated by cross-sectional microhardness determination. The data (Knoop hardness numbers, KHN) were subjected to 2-way analysis of variance and Tukey's post hoc test (α = 0.05%). RESULTS: All the test beverages significantly reduced the sample cross-sectional enamel hardness (KHN ± SD, 235.93 ± 18.15, 257.23 ± 21.79, and 253.23 ± 13.86 in the orange, yog, and cola groups, respectively) compared to samples in the negative control group (290.27 ± 3.92). In vitro acidogenic challenge exacerbated the mineral loss induced by all beverages (166.02 ± 4.28, 190.43 ± 17.55, and 198.39 ± 21.39 in the orange, yog, and cola groups combined to acidogenic challenge, respectively) compared to acidogenic challenge alone. CONCLUSIONS: All beverages exhibited erosive effects on primary enamel. Simulated caries challenge considerably exacerbated the enamel softening of primary teeth.

A unified procedure for conservative (morphology) and integral (DNA and immunophenotype) cell staining for flow cytometry.

BACKGROUND: Current methods for multiparameter DNA flow cytometry suffer from several limitations. These include significant modifications of cell morphological parameters, the impossibility to counterstain cells with certain fluorochromes, and laborious tuning of the instrument that, for some procedures, must be equipped with an ultraviolet (UV) laser. To overcome these problems, we developed a novel method for the simultaneous analysis of morphological parameters, four-color immunophenotyping, and stoichiometric DNA labeling using a bench-top flow cytometer. METHODS: The method consists of a mild permeabilization/fixation treatment at room temperature, followed by labeling with fluorochrome-conjugated monoclonal antibodies (mAbs) and with the DNA dye 7-aminoactinomycin D (7-AAD) at 56 degrees C. RESULTS: Using this method, we analyzed resting peripheral blood mononucleated cells (PBMC), proliferating T cells cultured in the presence of interleukin-2 (IL-2), and lymphoblastoid B cells. Lymphocytes, monocytes, and lymphoblasts treated by this procedure retained differential light scattering (DLS) characteristics virtually identical to those of untreated cells. This allowed regions to be drawn on forward scatter (FSC) and side scatter (SSC) cytograms resolving different cell populations. DLS were preserved well enough to distinguish large lymphoblasts in the S or G2/M phases from small G0/G1 cells. Also, stainability with fluorescein-isothiocyanate (FITC), R-phycoerythrin (PE), allophycocyanin (APC)-conjugated mAbs was generally preserved. DNA labeling with 7-AAD was of quality good enough to permit accurate cell cycle analysis. CONCLUSIONS: The method described here, which we called integral hot staining (IHS), represents a very simple, reproducible, and conservative assay for multiparameter DNA analysis using a bench-top flow cytometer. Last but not least, the cytometer tuning for multiparameter acquisition is straightforward.

Human T cells with a type-2 cytokine profile are resistant to apoptosis induced by primary activation: consequences for immunopathogenesis.

The mechanisms leading to a relative dominance of T cells producing type 2 cytokines in certain human immune disorders are still unclear. We investigated the relative susceptibility to apoptosis induced by primary in vitro activation of human type 1 (producing interferon-gamma (IFN-gamma)) or type 2 (producing IL-4) T cells. Peripheral blood lymphocytes were isolated from patients with immune disorders characterized by expansion of type 2 cells (four with AIDS and hyper-IgE/hypereosinophilia, one with Churg-Strauss syndrome, and one with idiopathic hypereosinophilic syndrome) or from individuals with normal cytokine balances. Cells were stimulated for 16 h with ionomycin and phorbol ester, and apoptosis of cytokine-producing cells was assessed by flow cytometry. T cells with a type-2 cytokine profile, i.e. producing IL-4 alone, were significantly more resistant to activation-induced apoptosis than those producing IFN-gamma alone. This was observed in AIDS patients, whose type 2 cells were mostly CD8+, as well as in the patients with Churg-Strauss and with hypereosinophilic syndrome. CD4+ and CD8+ IL-4-producing cells were equally resistant to apoptosis. Lower susceptibility to apoptosis of type-2 T cells was also observed in subjects with normal cytokine balances. Bcl-2 expression was high in type-2 cells and in viable type-1 cells, whereas it was low in apoptotic type-1 cells. Resistance to activation-induced apoptosis may explain the expansion of cells producing type-2 cytokines in certain immune disorders.

Novel mutations in 13 probands with galactokinase deficiency.

Galactokinase is an essential enzyme in the metabolism of galactose. Patients with deficiencies in galactokinase exhibit early-onset cataracts. We examined the sequence of the human galactokinase gene (GK1) from 13 patients exhibiting galactokinase deficiency and identified 12 novel mutations. One of the mutations occurred in six of the 13 probands examined, and the remaining 11 were unique mutations. Expression of each of the mutant GK1 genes in Xenopus oocytes resulted in very low galactokinase activity levels. These results provide important information regarding the types of GK1 mutations that occur in the human population.

Lymphocyte activation gene-3 (LAG-3) expression and IFN-gamma production are variably coregulated in different human T lymphocyte subpopulations.

We evaluated the relationship between cytokine profile and the expression of the lymphocyte activation gene-3 (LAG-3) in both T cell clones and polyclonal T cell lines; LAG-3 is a CD4-like protein whose expression is reportedly restricted to Th1/0 cells and dependent upon IFN-gamma. We found that, while LAG-3 was expressed only by CD4+ T cell clones producing IFN-gamma, most CD8+ clones producing IL-4 but not IFN-gamma (i.e., with a T cytotoxic-2-like profile) were LAG-3+. The intensity of LAG-3 expression by CD8+ clones correlated with the amount of released IFN-gamma, suggesting that this cytokine is not required for expression but rather for the up-regulation of LAG-3. Flow cytometric analyses of polyclonal T cell lines confirmed that LAG-3 could be expressed by both CD4+ and CD8+ cells that did not contain cytoplasmic IFN-gamma. In these cell lines, large proportions of CD4+ and CD8+ cells coexpressed LAG-3 and CD30, a putative marker of Th2-like cells. Overall, our data do not support the earlier suggestion that LAG-3 and CD30 are selective markers of T cells with type-1 and type-2 cytokine profiles, respectively.

Identification of genetic alterations associated with the process of human experimental colon cancer liver metastasis in the nude mouse.

Understanding the genetic elements controlling the process of tumor metastasis to distant organ sites such as the liver may be the key to improving survivorship from colon cancer. By using standard cytogenetic techniques in combination with comparative genomic hybridization, multiple genetic imbalances within three human colon cancer cell lines previously selected for differences in liver-metastatic behavior were identified. The entire genome of one poorly metastatic cell line (KM12C) was compared directly with that of two highly metastatic cell lines (KM12SM, KM12L4A) derived from it. A number of chromosomal gains (8q, 12q15, 20q11.2) and losses (5p13, 6p21.3, 18) were common to all three cell lines and are likely related to early tumor development rather than to the selection process used to generate cell lines of increased metastatic potential. Chromosomal imbalances detected only in the highly metastatic cell lines were also observed. KM12SM showed losses of portions of 2p22, 2q24.3--> 2q32.2, 4p15.3--> cen, 4q24 without the 13q and 15q22.3 gains noted for KM12C. Both gains (1p31.3--> 1p21, 2q22--> 2q33, 3cen--> 3q26.2, 5q14--> 5q23, 6cen--> 6q23) and losses (16p, 17p, 17q 19p, 19q 22q) were observed for KM12L4A but not for the other two cell lines. Identification of these alterations provides valuable insight into the process of experimental liver metastasis and is a first step towards mapping genes linked to the terminal phases of human colon cancer progression.

Lowe oculocerebrorenal syndrome in a female with a balanced X;20 translocation: mapping of the X chromosome breakpoint.

A Hispanic girl with Lowe oculocerebrorenal syndrome (OCRL), an X-linked recessive condition characterized by cataracts, glaucoma, mental retardation, and proteinuria, is reported. A balanced X;20 chromosomal translocation with the X chromosome breakpoint at q26.1 was found with high-resolution trypsin-Giemsa banding. Somatic cell hybridization was used to separate the X chromosome derivative and the chromosome 20 derivative in order to position, with respect to the translocation breakpoint, several DNA loci that are linked to the Lowe syndrome locus (Xq24-q26). DXS10 and DXS53 were found to be distal to the breakpoint, whereas DXS37 and DXS42 were located proximal to it. These studies suggest that the OCRL locus lies in the region between these probes. The translocation chromosome originated from an unaffected male without a visible translocation, indicating that the most likely cause of OCRL in this patient is the de novo translocation that disrupted the OCRL locus.

Cell line segregation in a 45,X/46,XY mosaic child with asymmetric leg growth.

Clinical evaluation of a 13 1/2-year-old male revealed a 4.4-cm leg length discrepancy and a small penis with a normal endocrine evaluation. Cytogenetic analysis of peripheral blood lymphocytes and skin fibroblasts derived from the back showed 45,X/46,XY mosaicism with similar percentages of 45,X cells, 36% and 30% respectively. However, two separate skin fibroblast cultures derived from the thigh and calf of the short (right) leg showed significant lack of Y-bearing cells with 100% and 80% 45,X, respectively. In contrast, skin biopsies of the thigh and calf of the normal (left) leg both showed 100% 46,XY. Similar evidence for differences in the percentages of Y-bearing cells in the left versus right leg fibroblast cultures was obtained using densitometric scanning of dot blots following DNA hybridization with a Y-specific probe at the DYZ4 locus. Asymmetric limb growth in cases of X/XY lymphocyte mosaicism warrants further cytogenetic investigation to substantiate possible genotype-phenotype correlations which may help uncover the fundamental growth deficiency in Turner syndrome.

Florida newborn screening for galactosemia.

Galactosemia, an inborn error of metabolism characterized by the inability to transform galactose-1-phosphate into glucose-1-phosphate, occurs in 1:50,000 live births. If not diagnosed and treated within the newborn period, it can lead to severe morbidity and mortality within a few weeks of life. All children in Florida are screened for this disorder by a fluorescence assay system to measure galactose-1-phosphate uridyltransferase (GALT) activity in a dried blood spot. Genetic factors and external forces can affect the activity of the GALT enzyme and lead to confusing results. Parents of infants heterozygous for galactosemia should be offered the opportunity for carrier detection. If both are carriers, genetic counseling should be provided.

Tissue limited mosaicism in a patient with tetrasomy 9p.

Cytogenetic analysis of an abnormal newborn girl showed an extra chromosome with the characteristics of an isodicentric 9p chromosome [idic (9)(pter----q12----pter)] in 98% of peripheral lymphocyte metaphases examined. This cytogenetic interpretation was substantiated by quantitative measurement of erythrocyte galactose-1-P-uridyltransferase (GALT) activity, which is consistent with the expression of 4 normal GALT genes. Cytogenetic results from skin fibroblasts showed mosaicism with only 11% of the metaphases having the extra chromosome. Selective genetic pressure based on a functional disadvantage of tetrasomy 9p in the skin is proposed. The in vivo establishment of cytogenetically normal cells in various tissues may be necessary for in utero survival of tetrasomy 9p infants.

Effects of human chorionic gonadotropin, prostaglandin F2 alpha and protein kinase C activators on the cyclic AMP and inositol phosphate second messenger systems in cultured human granulosa-luteal cells.

The effects of human chorionic gonadotropin (hCG) and prostaglandin F2 alpha (PGF2 alpha) on the adenylate cyclase-cAMP and inositol phospholipid-phospholipase C-inositol trisphosphate and diacylglycerol transmembrane signalling systems were evaluated in cultured human granulosa-luteal cells. Granulosa-luteal cells obtained from patients undergoing in vitro fertilization were cultured for 72 h prior to addition of hormones. During the last 24 h of culture granulosa-luteal cells were incubated with [3H]inositol. Neither hCG nor gonadotropin-releasing hormone (GnRH) stimulated the inositol phospholipid-phospholipase C signalling system. PGF2 alpha stimulated increases in inositol mono-, bis-, and trisphosphate accumulation in 30 min incubations. NaF (20 mM) mimicked the stimulatory effect of PGF2 alpha on inositol phosphate accumulation suggesting the involvement of a guanine nucleotide regulatory protein in the activation of phospholipase C. In contrast, hCG but not PGF2 alpha or NaF stimulated cAMP accumulation in 30 min incubations. Simultaneous treatment with hCG and PGF2 alpha did not alter the stimulatory effect of PGF2 alpha on inositol phosphate accumulation but reduced (37%) the stimulatory effect of hCG on cAMP accumulation. The protein kinase C activator, 12-O-tetradecanoylphorbol 13-acetate (TPA) inhibited the stimulatory effects of hCG (76%) and PGF2 alpha (62%) on cAMP and inositol phosphate accumulation, respectively. Thus, cultures of human granulosa-luteal cells possess multiple transmembrane signalling systems which may be modulated by the activation of protein kinase C.

Trisomy 3p23----pter and monosomy 11q23----qter in an infant with two translocation carrier parents.

A newborn male infant with multiple congenital abnormalities was found to be trisomic for 3p23----pter and monosomic for 11q23----qter. His parents were both carriers of a balanced reciprocal translocation. Considerable overlap in phenotype-karyotype correlations was found between the two chromosomal syndromes in the patient.

Monosomy, trisomy, fragile sites, and rearrangements of chromosome no. 1 in a mentally retarded male with multiple congenital anomalies.

A 17-year-old male was referred for evaluation because of short stature and severe mental retardation. Major clinical findings included microphthalmia, micrognathia, low-set ears, a prominent beaked nose, clubbing of digits, and premature graying of hair. Cytogenetic analysis revealed a 45,XY,-1/46,XY/47,XY,+1 mosaicism in lymphocytic culture, a 45,XY,-1/46,XY mosaicism in skin fibroblasts, and fra(1p) sites in 2% of the metaphases from lymphocyte, fibroblast and bone marrow cultures. Post-zygotic non-disjunction causing this mosaicism is believed to be responsible for the patient's phenotype.