Clinical microscopic analysis of protaper retreatment system efficacy considering root canal thirds using three endodontic sealers

To evaluate the efficacy of ProTaper Universal rotary retreatment system and the influence of sealer type on the presence of filling debris in the reinstrumented canals viewed in an operative clinical microscope. Forty-five palatal root canals of first molars were filled with gutta-percha and one of the following sealers: G1, EndoFill; G2, AH Plus; G3, Sealapex. The canals were then reinstrumented with ProTaper Universal rotary system. Roots were longitudinally sectioned and examined under an operative clinical microscope (10x), and the amount of filling debris on canal walls was analyzed using the AutoCAD 2004 software. A single operator used a specific software tool to outline the canal area and the filling debris area in each third (cervical, middle, and apical), as well as the total canal area. Data were analyzed by Kruskal-Wallis test and Tukey test at P < 0.05. Sealapex demonstrated significant differences in the average of filling debris area/canal among the 3 thirds. This group revealed that apical third showed more debris than the both cervical and middle third (P < 0.0001). Endofill presented significantly more filling debris than Sealapex in the cervical third (P < 0.05). In the middle (P = 0.12) and apical third (P = 0.10), there were no differences amongst groups. Debris was left in all canal thirds, regardless of the retreatment technique. The greatest differences between techniques and sealers were found in the cervical third. Microsc. Res. Tech. 75:12331236, 2012. (C) 2012 Wiley Periodicals, Inc.

Push-out strength of root fillings with or without thermomechanical compaction

Carneiro SMBS, Sousa-Neto MD, Rached-Junior FA, Miranda CES, Silva SRC, Silva-Sousa YTC. Push-out strength of root fillings with or without thermomechanical compaction. International Endodontic Journal, 45, 821828, 2012. Abstract Aim To evaluate the influence of thermomechanical compaction (Taggers hybrid technique THT) on the push-out strength of several root filling materials to root dentine. Methodology Root canals of eighty roots in human canines were prepared with the ProTaper system and filled with one of the following materials, using either lateral compaction (LC) (n = 40) or THT (n = 40): AH Plus/gutta-percha (GP) (n = 10), Sealer 26/GP (n = 10), Epiphany SE/Resilon (n = 10) and Epiphany SE/GP (n = 10). Three 2-mm-thick dentine slices were obtained from each third of each root. The root filling in the first slice was subjected to a push-out test to evaluate the bond strength of the materials to intraradicular dentine. Data (in MPa) were analysed using anova and post hoc Tukeys test (P < 0.05). Failure mode was determined at x25 magnification. The other two slices were prepared for scanning electron microscopy (SEM) to examine the surface of the filling materials. Results Lateral compaction (1.34 +/- 1.14 MPa) was associated with a significantly higher bond strength (P < 0.05) than the THT (0.97 +/- 0.88 MPa). AH Plus/GP (2.23 +/- 0.83 MPa) and Sealer 26/GP (1.86 +/- 0.50 MPa) had significantly higher bond strengths than the other materials and differed significantly from each other (P < 0.05). There was a significant difference (P < 0.05) between the coronal (1.36 +/- 1.15 MPa), middle (1.14 +/- 1.05 MPa) and apical thirds (0.95 +/- 0.83 MPa). Considering the technique and root filling material interaction, AH Plus/GP-LC was associated with the highest mean values (2.65 +/- 0.66 MPa) (P < 0.05). Sealer 26/GP-LC (2.10 +/- 0.46 MPa), AH Plus/GP-THT (1.81 +/- 0.78 MPa) and Sealer 26/GP-TH (1.63 +/- 0.44 MPa) had intermediate values that were not significantly different from each other (P > 0.05). Epiphany SE was associated with the lowest mean values (3.70 +/- 0.86 MPa) (P < 0.05), regardless of the root filling technique and type of solid material (cone). Adhesive failures predominated in the specimens filled with Epiphany SE, whilst mixed and cohesive failures were more frequent in those filled with AH Plus and Sealer 26, regardless of the root filling technique. SEM analysis revealed that LC produced a dense and well-compacted filling whilst the use of a hybrid thermomechanical technique resulted in the solid material (GP or Resilon) intermingled within sealer to form a nonhomogenous mass. Conclusion Lateral compaction was associated with higher bond strengths of the materials to intraradicular dentine than a hybrid technique using thermomechanical compaction. The greatest push-out strengths were obtained when the canals were filled with LC of AH Plus and GP cones.

Expression of tlr4, md2 and cd14 in equine blood leukocytes during endotoxin infusion and in intestinal tissues from healthy horses

The expression of tlr4, md2 and cd14 was studied in equine blood leukocytes and in intestinal samples using real time PCR. The stability of three commonly used reference genes, glyceraldehyde-3P-dehydrogenase (GAPDH), hypoxantine ribosyltransferase (HPRT) and succinate dehydrogenase complex subunit A (SDHA), was evaluated using qbase(PLUS). The equine peripheral blood mononuclear cells (eqPBMC) examined were either stimulated in vitro with Phorbol 12-myristate 13-acetate (PMA) and ionomycin or with the CpG oligodeoxynuclotide 2216 (CpG-ODN 2216) or obtained from horses before, during and after infusion of endotoxin. Intestinal tissue from healthy horses was sampled at ileum, right dorsal colon and rectum. Ranking of the three reference genes used for normalisation identified the combination HPRT/SDHA as most suitable both when determined ex vivo in leukocytes obtained from experimentally induced endotoxaemia and in eqPBMC activated in vitro while HPRT/GAPDH were most appropriate for the intestinal samples. The relative amounts of mRNA for TLR4 and MD-2 increased threefold during in vitro activation of the cells with CpG-ODN 2216 but was decreased in cultures stimulated with PMA/ionomycin. A transient elevation in the transcription of tlr4 and md2 was also evident for equine blood leukocytes following endotoxaemia. The levels of mRNA for CD14 on the other hard remained unaffected both during the induction of endotoxaemia and in the in vitro stimulated PBMCs. A low steady expression of TLR4, MD-2 and CD14 mRNA was demonstrated for the intestinal samples with no variation between the intestinal segments analysed. Thus, the foundation for real time PCR based levels of analysis of mRNA for all three components in the equine LPS receptor complex in different intestinal segments was set, making it possible to carry out future expression studies on clinical material. (C) 2012 Elsevier B.V. All rights reserved.

Long-term amphetamine treatment exacerbates inflammatory lung reaction while decreases airway hyper-responsiveness after allergic stimulus in rats

Asthma is an allergic lung disease can be modulated by drugs that modify the activity of central nervous system (CNS) such as amphetamine (AMPH). AMPH is a highly abused drug that exerts potent effects on behavior and immunity. In this study we investigated the mechanism involved in the effects of long-term AMPH treatment on the increased magnitude of allergic lung response. We evaluated mast cells degranulation, cytokines release, airways responsiveness and, expression of adhesion molecules. Male Wistar rats were treated with AMPH or vehicle (PBS) for 21 days and sensitized with ovalbumin (OVA) one week after the first injection of vehicle or AMPH. Fourteen days after the sensitization, the rats were challenged with an OVA aerosol, and 24 h later their parameters were analyzed. In allergic rats, the treatment with AMPH exacerbated the lung cell recruitment due increased expression of ICAM-1, PECAM-1 and Mac-1 in granulocytes and macrophages recovered from bronchoalveolar lavage. Elevated levels of IL-4, but decreased levels of IL-10 were also found in samples of lung explants after AMPH treatment. Conversely, the ex-vivo tracheal hyper-responsiveness to methacholine (MCh) was reduced by AMPH treatment, whereas the force contraction of tracheal segments due to in vitro antigen challenge remained unaltered. Our findings suggest that lung inflammation and airway hyper-responsiveness due to OVA challenge are under the distinct control of AMPH during long-term treatment. Our data strongly indicate that AMPH positively modulates allergic lung inflammation via the increase of ICAM-1, PECAM-1, Mac-1 and IL-4. AMPH also abrogates the release of the anti-inflammatory cytokine IL-10. (c) 2012 Elsevier B.V. All rights reserved.

The erosion and abrasion-inhibiting effect of TiF4 and NaF varnishes and solutions on enamel in vitro

Objective. Previous in vitro study has shown that TiF(4) varnish might reduce enamel erosion. No data regarding the effect of this experimental varnish on enamel erosion plus abrasion, however, are available so far. Thus, this in vitro study aimed to analyse the effect of TiF4 compared with NaF varnishes and solutions, to protect against enamel erosion with or without abrasion. Methods. Enamel specimens were pre-treated with experimental-TiF4 (2.45% F), experimentalNaF (2.45% F), NaF-Duraphat (2.26% F), and placebo varnishes; NaF (2.26% F) and TiF4 (2.45% F) solutions. Controls remained untreated. The erosive challenge was performed using a soft drink (pH 2.6) 4 u 90 s / day (ERO) and the toothbrushing abrasion (ERO+ ABR) 2 u 10 s / day, for 5 days. Between the challenges, the specimens were exposed to artificial saliva. Enamel loss was measured profilometrically (lm). Results. Kruskal-Wallis / Dunn tests showed that all fluoridated varnishes (TiF4-ERO: 0.53 +/- 0.20, ERO+ ABR: 0.65 +/- 0.19/ NaF-ERO: 0.94 +/- 0.18, ERO+ ABR: 1.74 +/- 0.37 / Duraphat-ERO: 1.00 +/- 0.37, ERO+ ABR: 1.72 +/- 0.58) were able to significantly reduce enamel loss when compared with placebo varnish (ERO: 3.45 +/- 0.41 / ERO+ ABR: 3.20 +/- 0.66) (P < 0.0001). Placebo varnish, control (ERO: 2.68 +/- 0.53 / ERO+ ABR: 3.01 +/- 0.34), and fluoridated (NaF-ERO: 2.84 +/- 0.09 / ERO+ ABR: 2.40 +/- 0.21 / TiF4-ERO: 3.55 +/- 0.59 / ERO+ ABR: 4.10 +/- 0.38) solutions did not significantly differ from each other. Conclusion. Based on the results, it can be concluded that the TiF4 varnish seems to be a promising treatment to reduce enamel loss under mild erosive and abrasive conditions in vitro.

Mechanical Performance of Cylindrical and Dual-Core Pedicle Screws After Repeated Insertion

Study Design. Ex vivo study of the mechanical performance of cylindrical and dual-core pedicle screws after insertion, removal, and reinsertion in the same hole. Objective. To evaluate the effect of repeated use of same screw hole on the insertion torque and the retentive strength of the cylindrical and dual-core screws. Summary of Background Data. Insertion and removal of pedicle screws is sometimes necessary during surgical procedure to assess the integrity of the pilot-hole wall. However, this maneuver may compromise the implant-holding capacity. Methods. Sixty thoracolombar vertebrae (T13-L5), harvested from 10 healthy calves, were used to insert 2 different designs of pedicle screws: cylindrical (5.0-mm outer diameter) and dual-core screws (5.2-mm outer diameter). Three experimental groups were created on the basis of the number of insertions of the screws and 2 subgroups were established according to the core pedicle screw design (dual-core and cylindrical). The insertion torque was measured during initial insertion, second insertion, and third insertion. Pullout screw tests were performed using a universal testing machine to evaluate the pullout strength after initial insertion, second insertion, and third insertion. Results. Significant reductions of 38% in mean insertion torque and 30% in mean pullout strength of dual-core screw were observed between the initial insertion and the third insertion. The cylindrical screw observed significant reductions of 52.5% in mean insertion torque and 42.3% in mean pullout strength between the initial insertion and the third insertion. A reduction of mean insertion torque and pullout strength between the first insertion and the second insertion but without significance was also observed for both types of screws. Conclusion. Insertions and reinsertion of either cylindrical or dual-core pedicle screws have compromised insertion torque and pullout strength of the implants as measured by mechanical tests.

Retroviral transfer of the p16INK4a cDNA inhibits C6 glioma formation in Wistar rats

Abstract Background The p16INK4A gene product halts cell proliferation by preventing phosphorylation of the Rb protein. The p16INK4a gene is often deleted in human glioblastoma multiforme, contributing to unchecked Rb phosphorylation and rapid cell division. We show here that transduction of the human p16INK4a cDNA using the pCL retroviral system is an efficient means of stopping the proliferation of the rat-derrived glioma cell line, C6, both in tissue culture and in an animal model. C6 cells were transduced with pCL retrovirus encoding the p16INK4a, p53, or Rb genes. These cells were analyzed by a colony formation assay. Expression of p16INK4a was confirmed by immunohistochemistry and Western blot analysis. The altered morphology of the p16-expressing cells was further characterized by the senescence-associated β-galactosidase assay. C6 cells infected ex vivo were implanted by stereotaxic injection in order to assess tumor formation. Results The p16INK4a gene arrested C6 cells more efficiently than either p53 or Rb. Continued studies with the p16INK4a gene revealed that a large portion of infected cells expressed the p16INK4a protein and the morphology of these cells was altered. The enlarged, flat, and bi-polar shape indicated a senescence-like state, confirmed by the senescence-associated β-galactosidase assay. The animal model revealed that cells infected with the pCLp16 virus did not form tumors. Conclusion Our results show that retrovirus mediated transfer of p16INK4a halts glioma formation in a rat model. These results corroborate the idea that retrovirus-mediated transfer of the p16INK4a gene may be an effective means to arrest human glioma and glioblastoma.

Cellular and humoral immune responses against the Plasmodium vivax MSP-119 malaria vaccine candidate in individuals living in an endemic area in north-eastern Amazon region of Brazil

Abstract Background Plasmodium vivax merozoite surface protein-1 (MSP-1) is an antigen considered to be one of the leading malaria vaccine candidates. PvMSP-1 is highly immunogenic and evidences suggest that it is target for protective immunity against asexual blood stages of malaria parasites. Thus, this study aims to evaluate the acquired cellular and antibody immune responses against PvMSP-1 in individuals naturally exposed to malaria infections in a malaria-endemic area in the north-eastern Amazon region of Brazil. Methods The study was carried out in Paragominas, Pará State, in the Brazilian Amazon. Blood samples were collected from 35 individuals with uncomplicated malaria. Peripheral blood mononuclear cells were isolated and the cellular proliferation and activation was analysed in presence of 19 kDa fragment of MSP-1 (PvMSP-119) and Plasmodium falciparum PSS1 crude antigen. Antibodies IgE, IgM, IgG and IgG subclass and the levels of TNF, IFN-γ and IL-10 were measured by enzyme-linked immunosorbent assay. Results The prevalence of activated CD4+ was greater than CD8+ T cells, in both ex-vivo and in 96 h culture in presence of PvMSP-119 and PSS1 antigen. A low proliferative response against PvMSP-119 and PSS1 crude antigen after 96 h culture was observed. High plasmatic levels of IFN-γ and IL-10 as well as lower TNF levels were also detected in malaria patients. However, in the 96 h supernatant culture, the dynamics of cytokine responses differed from those depicted on plasma assays; in presence of PvMSP-119 stimulus, higher levels of TNF were noted in supernatant 96 h culture of malaria patient’s cells while low levels of IFN-γ and IL-10 were verified. High frequency of malaria patients presenting antibodies against PvMSP-119 was evidenced, regardless class or IgG subclass.PvMSP-119-induced antibodies were predominantly on non-cytophilic subclasses. Conclusions The results presented here shows that PvMSP-119 was able to induce a high cellular activation, leading to production of TNF and emphasizes the high immunogenicity of PvMSP-119 in naturally exposed individuals and, therefore, its potential as a malaria vaccine candidate.

Tomografia de coerência óptica broncoscópica

OBJETIVO: Avaliar a viabilidade e o potencial do uso da tomografia de coerência óptica em conjunto com um broncoscópio convencional na avaliação das vias aéreas. MÉTODOS: Estudo piloto baseado em um modelo experimental ex vivo com três animais: um coelho adulto da raça Nova Zelândia e dois suínos da raça Landrace. Um cateter de imagem de tomografia de coerência óptica foi inserido no canal de trabalho de um broncoscópio flexível para alcançarmos a traqueia distal dos animais. As imagens foram obtidas sistematicamente em toda a traqueia ao longo das paredes, partindo da porção distal para a proximal. RESULTADOS: O cateter de imagem se adaptou com facilidade ao canal de trabalho do broncoscópio. Imagens em alta resolução de cortes transversais da traqueia foram obtidas em tempo real, sendo delineadas microestruturas, tais como epitélio, submucosa, cartilagem e camada adventícia nas paredes anteriores e laterais da traqueia. As camadas correspondentes do epitélio, mucosa e cartilagens foram claramente diferenciadas. Na parede posterior, foi possível identificar mucosa, submucosa e musculatura traqueal. CONCLUSÕES: O uso de tomógrafo de coerência óptica em conjunto com um broncoscópio flexível é viável. A tomografia de coerência óptica produz imagens de alta resolução que permitem visualizar a microanatomia da traqueia, inclusive estruturas que normalmente são visualizadas somente na histologia convencional.

Acute inhibition of excessive mitochondrial fission after myocardial infarction prevents long-term cardiac dysfunction

BACKGROUND: Ischemia and reperfusion (IR) injury remains a major cause of morbidity and mortality and multiple molecular and cellular pathways have been implicated in this injury. We determined whether acute inhibition of excessive mitochondrial fission at the onset of reperfusion improves mitochondrial dysfunction and cardiac contractility postmyocardial infarction in rats. METHODS AND RESULTS: We used a selective inhibitor of the fission machinery, P110, which we have recently designed. P110 treatment inhibited the interaction of fission proteins Fis1/Drp1, decreased mitochondrial fission, and improved bioenergetics in three different rat models of IR, including primary cardiomyocytes, ex vivo heart model, and an in vivo myocardial infarction model. Drp1 transiently bound to the mitochondria following IR injury and P110 treatment blocked this Drp1 mitochondrial association. Compared with control treatment, P110 (1 μmol/L) decreased infarct size by 28 ± 2% and increased adenosine triphosphate levels by 70+1% after IR relative to control IR in the ex vivo model. Intraperitoneal injection of P110 (0.5 mg/kg) at the onset of reperfusion in an in vivo model resulted in improved mitochondrial oxygen consumption by 68% when measured 3 weeks after ischemic injury, improved cardiac fractional shortening by 35%, reduced mitochondrial H2O2 uncoupling state by 70%, and improved overall mitochondrial functions. CONCLUSIONS: Together, we show that excessive mitochondrial fission at reperfusion contributes to long-term cardiac dysfunction in rats and that acute inhibition of excessive mitochondrial fission at the onset of reperfusion is sufficient to result in long-term benefits as evidenced by inhibiting cardiac dysfunction 3 weeks after acute myocardial infarction.

The effect of bone allografts combined with bone marrow stromal cells on the healing of segmental bone defects in a sheep model

Background: The repair of large bone defects is a major orthopedic challenge because autologous bone grafts are not available in large amounts and because harvesting is often associated with donor-site morbidity. Considering that bone marrow stromal cells (BMSC) are responsible for the maintenance of bone turnover throughout life, we investigated bone repair at a site of a critically sized segmental defect in sheep tibia treated with BMSCs loaded onto allografts. The defect was created in the mid-portion of the tibial diaphysis of eight adult sheep, and the sheep were treated with ex-vivo expanded autologous BMSCs isolated from marrow aspirates and loaded onto cortical allografts (n = 4). The treated sheep were compared with control sheep that had been treated with cell-free allografts (n = 4) obtained from donors of the same breed as the receptor sheep. Results: The healing response was monitored by radiographs monthly and by computed tomography and histology at six, ten, fourteen, and eighteen weeks after surgery. For the cell-loaded allografts, union was established more rapidly at the interface between the host bone and the allograft, and the healing process was more conspicuous. Remodeling of the allograft was complete at 18 weeks in the cell-treated animals. Histologically, the marrow cavity was reestablished, with intertrabecular spaces being filled with adipose marrow and with evidence of focal hematopoiesis. Conclusions: Allografts cellularized with AOCs (allografts of osteoprogenitor cells) can generate great clinical outcomes to noncellularized allografts to consolidate, reshape, structurally and morphologically reconstruct bone and bone marrow in a relatively short period of time. These features make this strategy very attractive for clinical use in orthopedic bioengineering

Endo-PDI is required for TNFα-induced angiogenesis

Protein disulfide isomerase (PDI) and its homologs are oxidoreductases facilitating protein folding in the ER. Endo-PDI (also termed ERp46) is highly expressed in endothelial cells. It belongs to the PDI family but its physiological function is largely unknown. We studied the role of Endo-PDI in endothelial angiogenic responses. Stimulation of human umbilical vein endothelial cells (with TNFα (10ng/ml) increased ERK1/2 phosphorylation. This effect was largely attenuated by Endo-PDI siRNA, whereas JNK and p38 MAP kinase phosphorylation was Endo-PDI independent. Similarly, TNFα-stimulated NF-κB signaling determined by IκBα degradation as well as TNFα-induced ICAM expression was unaffected by Endo-PDI siRNA. The action of Endo-PDI was not mediated by extracellular thiol exchange or cell surface PDI as demonstrated by nonpermeative inhibitors and PDI-neutralizing antibody. Moreover, exogenously added PDI failed to restore ERK1/2 activation after Endo-PDI knockdown. This suggests that Endo-PDI acts intracellularly potentially by maintaining the Ras/Raf/MEK/ERK pathway. Indeed, knockdown of Endo-PDI attenuated Ras activation measured by G-LISA and Raf phosphorylation. ERK activation influences gene expression by the transcriptional factor AP-1, which controls MMP-9 and cathepsin B, two proteases required for angiogenesis. TNFα-stimulated MMP-9 and cathepsin B induction was reduced by silencing of Endo-PDI. Accordingly, inhibition of cathepsin B or Endo-PDI siRNA blocked the TNFα-stimulated angiogenic response in the spheroid outgrowth assays. Moreover ex vivo tube formation and in vivo Matrigel angiogenesis in response to TNFα were attenuated by Endo-PDI siRNA. In conclusion, our study establishes Endo-PDI as a novel, important mediator of AP-1-driven gene expression and endothelial angiogenic function

The putative role of ovary removal and progesterone when considering the effect of formaldehyde exposure on lung inflammation induced by ovalbumin

OBJECTIVE: Formaldehyde exposure during the menstrual cycle is known to affect the course of allergic lung inflammation. Because our previous data demonstrated that formaldehyde combined with an ovariectomy reduced allergic lung inflammation, we investigated the putative role of ovary removal and progesterone treatment when considering the effect of formaldehyde on allergic lung inflammation. METHOD: Ovariectomized rats and their matched controls were exposed to formaldehyde (1%, 3 days, 90 min/day) or vehicle, and immediately after exposure, the rats were sensitized to ovalbumin by a subcutaneous route. After 1 week, the rats received a booster by the same route, and after an additional week, the rats were challenged with ovalbumin (1%) by an aerosol route. The leukocyte numbers, interleukin-10 (IL-10) release, myeloperoxidase activity, vascular permeability, ex vivo tracheal reactivity to methacholine and mast cell degranulation were determined 24 h later. RESULTS: Our results showed that previous exposure to formaldehyde in allergic rats decreased lung cell recruitment, tracheal reactivity, myeloperoxidase activity, vascular permeability and mast cell degranulation while increasing IL-10 levels. Ovariectomy only caused an additional reduction in tracheal reactivity without changing the other parameters studied. Progesterone treatment reversed the effects of formaldehyde exposure on ex vivo tracheal reactivity, cell influx into the lungs and mast cell degranulation. CONCLUSION: In conclusion, our study revealed that formaldehyde and ovariectomy downregulated allergic lung inflammation by IL-10 release and mast cell degranulation. Progesterone treatment increased eosinophil recruitment and mast cell degranulation, which in turn may be responsible for tracheal hyperreactivity and allergic lung inflammation