Effect of culture media on porcine embryos produced by in vitro fertilization or parthenogenetic activation after oocyte maturation with cycloheximide

This study evaluated the effects of reversible meiotic inhibition and different culture media (PZM3 or NCSU23) on production of porcine embryos by either in vitro fertilization (IVF) or parthenogenetic activation (PA). Oocytes from abattoir-derived ovaries were allocated into two groups for maturation: CHX (5 mu g/ml cycloheximide for 10 h) or Control (no CHX). The percentage of metaphase II (MII) oocytes was determined at 36, 40 or 44 h of in vitro maturation. For IVF and PA, denuded oocytes were fertilized with purified sperm for 6 h or activated by electric stimuli. Zygotes were then subdivided into two culture groups: NCSU23 or PZM3. No effect of treatment with CHX and culture media was observed on cleavage (D3) and blastocyst (D7) rates in IVF and PA groups. There are no differences of quality or development rates between IVF-derived embryos cultured in NCSU23 or PZM3. However, we observed high quality PA embryos in PZM3 compared with NCSU23. Maturation arrest with CHX decreased the average blastocyst cell number in IVF while it was increased in PA embryos. As older oocytes are more effectively activated, CHX-blocked oocytes reached the mature stage faster than the control group. In conclusion, the CHX treatment for 10 h, followed by oocyte maturation for 40 h, is an efficient protocol to produce high quality parthenote embryos, especially when they are cultured in PZM3. However, this protocol is not satisfactory for IVF embryos production. In this case, a shorter maturation period could provide better embryo quality.

Exercise training accelerates the removal from plasma of LDL-like nanoemulsion in moderately hypercholesterolemic subjects

Objective: Exercise training improves plasma lipid profile and diminishes risk of coronary heart disease. Previously, we showed that training increases LDL plasma clearance, as tested by an artificial LDL-like nanoemulsion method, presumably by increasing LDL receptor activity. In this study, we investigated whether training could also improve LDL clearance in hypercholesterolemic subjects (HCh) that are exposed to increased risk of cardiovascular events. Methods: Twenty sedentary HCh and 20 normolipidemic (NL) sedentary volunteers were divided into four groups: 12 HCh submitted to 4-month training program, 8 HCh with no exercise program, 12 NL submitted to 4-month training and 8 NL with no exercise program. An LDL-like nanoemulsion labeled with 14C-cholesteryl ester was injected intravenously into all subjects and plasma samples were collected during 24h after injection to determine the fractional clearance rate (FCR, in h-1) by compartmental analysis. The study was performed on the first and on the last day of the 4-month study period. Results: In both, trained HCh and NL groups, training increased nanoemulsion FCR by 36% (0.0443 +/- 0.0126; 0.0602 +/- 0.0187, p=0.0187 and 0.0503 +/- 0.0203; 0.0686 +/- 0.0216, p=0.0827, respectively). After training, LDL cholesterol diminished in both HCh and NL groups. In HCh, but not in NL group, LDL susceptibility to oxidation decreased, but oxidized LDL was unchanged. In both non-trained groups FCR was the same for the last and the 4-month previous evaluation. Conclusion: In HCh, exercise training increased the removal of LDL as tested by the nanoemulsion, and this probably accounted for decreased LDL cholesterol and diminished LDL susceptibility to oxidation. (C) 2010 Elsevier Ireland Ltd. All rights reserved.

Detection of Herpesviruses and Periodontal Pathogens in Subgingival Plaque of Patients With Chronic Periodontitis, Generalized Aggressive Periodontitis, or Gingivitis

Background: Herpesviruses may be related to the etiology of aggressive periodontitis (AgP) and chronic periodontitis (CP) by triggering periodontal destruction or by increasing the risk for bacterial infection. This case-control study evaluated the presence of herpes simplex virus type 1 (HSV-1), Epstein-Barr virus type 1 (EBV-1), human cytomegalovirus (HCMV), Aggregatibacter actinomycetemcomitans (previously Actinobacillus actinomycetemcomitans), Porphyromonas gingivalis, Prevotella intermedia, and Tannerella forsythia (previously T. forsythensis) in patients with generalized AgP (AgP group), CP (CP group), or gingivitis (G group) and in healthy individuals (C group). Methods: Subgingival plaque samples were collected with paper points from 30 patients in each group. The nested polymerase chain reaction (PCR) method was used to detect HSV-1, EBV-1, and HCMV. Bacteria were identified by 16S rRNA-based PCR. Results: HSV-1, HCMV, and EBV-1 were detected in 86.7%, 46.7%, and 33.3% of the AgP group, respectively; in 40.0%, 50.0%, and 46.7% of the CP group, respectively; in 53.3%, 40.0%, and 20.0% of the G group, respectively; and in 20.0%, 56.7%, and 0.0% of the C group, respectively. A. actinomycetemcomitans was detected significantly more often in the AgP group compared to the other groups (P<0.005). P. gingivalis and T. forsythia were identified more frequently in AgP and CP groups, and AgP, CP, and G groups had higher frequencies of P. intermedia compared to the C group. Conclusion: In Brazilian patients, HSV-1 and EBV-1, rather than HCMV, were more frequently associated with CP and AgP. J Periodontol 2008;79:2313-2321.

Antimicrobial Effects of Calcium Hydroxide and Chlorhexidine on Enterococcus faecalis

Introduction: Endodontic treatment is commonly based on nonspecific elimination of intraradicular micro-organisms. Although some authors prefer single-visit root canal operations for endodontic treatment, several studies have shown the importance of intracanal medication between sessions to kill microorganisms that biomechanical preparations alone cannot achieve. The purpose of this study was to evaluate the efficacy of calcium hydroxide Ca(OH)2 and chlorhexidine gel on the elimination of intratubular Enterococcus faecalis. Methods: Human uniradicular teeth contaminated with E. faecalis were treated with Ca(OH)(2), 2% chlorhexidine gel, Ca(OH)(2) plus 2% chlorhexidine gel, or saline (0.9% NaCl) as a negative control. Samples obtained at a depth of 0 to 100 mu m and 100 to 200 mu m from these root canal preparations were analyzed for bacterial load by counting the number of colonyforming units (CFUs) and bacterial viability using fluorescence microscopy. Results: A significant decrease in the number of CFUs and the percentage of viable E. faecalis was observed after treatment with either Ca(OH)(2) or chlorhexidine when compared with the control group. Additionally, chlorhexidine gel had a significantly higher antimicrobial efficacy as measured by the number of CFUs and the percentage of viable cells than Ca(OH)(2). No differences were observed between the antimicrobial properties of chlorhexidine gel with and without the addition of Ca(OH)(2). Conclusion: Both Ca(OH)(2) and chlorhexidine have antimicrobial effects on E. faecalis. Chlorhexidine had increased antimicrobial activity when compared with Ca(OH)(2.) Ca(OH)(2) combined with chlorhexidine showed similar antimicrobial activity to chlorhexidine alone. (J Endod 2010;36:1389-1393)

Heat-killed Enterococcus faecalis Alters Nitric Oxide and CXCL12 Production but not CXCL8 and CCL3 Production by Cultured Human Dental Pulp Fibroblasts

Introduction: Fibroblasts are the most abundant cells in dental pulp. To investigate their capacity to produce the chemokines CCL3, CXCL8, and CXCL12 as well as nitric oxide (NO), we evaluated the production of these mediators in supernatants of cultured human dental pulp fibroblasts (HDPF) stimulated by heat-killed Enterococcus faecalis (HKEF). Methods: Primary cultures of HDPF were stimulated with medium alone or HKEF (1:1, 10:1, or 100:1 bacteria:fibroblast) for 1, 6, and 24 hours. Chemokines and NO were assessed through enzyme-linked immunosorbent assay and Griess reaction, respectively. Statistical analysis was performed by using analysis of variance and Tukey post test. Results: CCL3 was not detected, whereas constitutive CXCL8 was not affected. Production of CXCL12 was increased at 1 and 6 hours, and NO was increased at the concentration of 1:1 bacteria:fibroblast at 24 hours. Viability and proliferation assays did not reveal cell number differences. Conclusions: These findings demonstrate that heat-killed E. faecalis is able to increase production of CXCL12 and NO by HDPF. (J Endod 2010;36:91-94)

Presence of arsenic in different types of MTA and white and gray Portland cement

Objective. The presence of arsenic in various types of mineral trioxide aggregate (MTA) and Portland cements were evaluated to verify if they comply with the ISO-recommended limit for water-based cements of 2 mg arsenic/kg material. Study design. An amount of 5 mL of hydrochloric acid was added to 2 g each of MTA and Portland cement to be analyzed. After 15 minutes, the material was filtered and the volume of supernatant was diluted with reagent-grade water up to 40 mL. Atomic absorption spectrophotometry readings were performed in triplicate. Results. The following mean values were obtained: CPM (Egeo, Buenos Aires, Argentina) 11.06 mg/kg; CPM sealer (Egeo) 10.30 mg/kg; MTA- Obtura (Angelus, Londrina, PR, Brazil) 0.39 mg/kg; Experimental MTA: 10.30 mg/kg; White MTA- Angelus (Angelus) 1.03 mg/kg; Gray MTA- Angelus (Angelus) 5.91 mg/kg; ProRoot-MTA (Dentsply/Tulsa Dental Specialties, Tulsa, OK) 5.25 mg/kg; Gray Portland cement (Votorantim Cimentos, Cubatao, SP, Brazil): 34.27 mg/kg; and White Portland cement (Cimento Rio Branco, Rio de Janeiro, RJ, Brazil) 0.52 mg/kg. Conclusion. All tested materials presented arsenic in their composition. The form of arsenic was not analyzed nor the toxicity of the arsenic found. Only MTA- Obtura, White MTA- Angelus, and White Portland cement presented arsenic levels below the limit set in the ISO 9917-1 standard. (Oral Surg Oral Med Oral Pathol Oral Radiol Endod 2008; 106: 909-913)

The 434(G > C) polymorphism in the eosinophil cationic protein gene and its association with tissue eosinophilia in oral squamous cell carcinomas

Objective: The aim of this study was to investigate the prevalence of the Eosinophil cationic protein (ECP)-gene polymorphism 434(G > C) in oral squamous cell carcinoma (OSCC) patients and its association with tumor-associated tissue eosinophilia (TATE), demographic, clinical, and microscopic variables. Methods: The ECP genotypes of 165 healthy individuals and 157 OSCC patients were detected by PCR-RFLP analysis after cleavage of the amplified DNA sequence with enzyme PstI. TATE was obtained by morphometric analysis. Chi-square test or Fisher`s exact test was used to analyze the association of ECP-gene polymorphism 434(G > C) with TATE, demographic, clinical, and microscopic variables in OSCC patients. Disease-free survival and overall survival were calculated by the Kaplan-Meier product-limit actuarial method and the comparison of the survival curves were performed using log rank test. Results: Most of healthy individuals (53.33%) and OSCC patients (57.97%) were heterozygous for the ECP 434(G > C) polymorphism. Based on numerical differences, our results showed that OSCC patients with intense TATE and at least one C allele had a higher frequency of bilateral neck dissection, local recurrence, vascular embolization, involved resection margins, and postoperative radiotherapy. No statistically significant differences on survival rates were found in OSCC patients presenting different ECP 434(G > C) genotypes. Conclusions: These results suggest a tendency towards a poor clinical outcome in OSCC patients with intense TATE and 434GC/CC genotypes, probably due to an ECP genetic variant with altered cytotoxic activity.

Tooth abnormalities and soft tissue alterations in patients with G/BBB syndrome

OBJECTIVE: The G/BBB syndrome is an X-linked recessive disorder characterized by eye anomalies, laryngotracheoesophageal cleft, congenital heart disease, genitourinary anomalies and gastrointestinal disorders. Patients may also present cleft lip and palate, high-arched palate and thin upper lip. This study aimed to investigate the occurrence of tooth abnormalities and soft tissue changes in patients with G/BBB syndrome. DESIGN: Cross-sectional. SUBJECTS AND METHODS: Twenty-one patients with G/BBB syndrome were analyzed as to the presence of tooth abnormalities and soft tissue alterations. MAIN OUTCOME MEASURES: The prevalence of tooth agenesis and supernumerary teeth was compared to patients without morphofunctional alterations, matched for gender and age. RESULTS: All patients had complete cleft lip and palate; 95.23% of patients presented tooth abnormalities, mainly hypoplastic alterations, with predominance of alterations of number, followed by alterations of structure, shape and position. The frequency of tooth agenesis and supernumerary teeth was significantly higher compared with the control group; 11 patients presented incisiform supernumerary teeth in the mandibular anterior region. Ankyloglossia was observed in 11 of 21 patients. CONCLUSION: The presence of mandibular anterior supernumerary teeth and ankyloglossia should be investigated in the clinical evaluation of patients with suspected diagnosis of the G/BBB syndrome. Oral Diseases (2008) 14, 747-753

Relational learning in children with deafness and cochlear implants

This four-experiment series sought to evaluate the potential of children with neurosensory deafness and cochlear implants to exhibit auditory-visual and visual-visual stimulus equivalence relations within a matching-to-sample format. Twelve children who became deaf prior to acquiring language (prelingual) and four who became deaf afterwards (postlingual) were studied. All children learned auditory-visual conditional discriminations and nearly all showed emergent equivalence relations. Naming tests, conducted with a subset of the: children, showed no consistent relationship to the equivalence-test outcomes.. This study makes several contributions: to the literature on stimulus equivalence. First; it demonstrates that both pre- and postlingually deaf children-can: acquire auditory-visual equivalence-relations after cochlear implantation, thus demonstrating symbolic functioning. Second, it directs attention to a population that may be especially interesting for researchers seeking to analyze the relationship. between speaker and listener repertoires. Third, it demonstrates the feasibility of conducting experimental studies of stimulus control processes within the limitations of a hospital, which these children must visit routinely for the maintenance of their cochlear implants.

Differential Production of Macrophage Inflammatory Protein-1 alpha, Stromal-Derived Factor-1, and IL-6 by Human Cultured Periodontal Ligament and Gingival Fibroblasts Challenged With Lipopolysaccharide From P. gingivalis

Background: Fibroblasts are considered important cells in periodontitis. When challenged by different agents, they respond through the release of cytokines that participate in the inflammatory process. The aim of this study is to evaluate and compare the expression and production of macrophage inflammatory protein (MIP)-1 alpha, stromal-derived factor (SDF)-1, and interleukin (IL)-6 by human cultured periodontal ligament and gingival fibroblasts challenged with lipopolysaccharide (LPS) from Porphyromonas gingivalis. Methods: Fibroblasts were cultured from biopsies of gingival tissue and periodontal ligament of the same donors and used on the fourth passage. After confluence in 24-well plates, the culture medium alone (control) or with 0.1 to 10 mu g/ml of LPS from P. gingivalis was added to the wells, and after 1, 6, and 24 hours, the supernatant and the cells were collected and analyzed by enzyme-linked immunosorbent assay and real-time polymerase chain reaction, respectively. Results: MIP-1 alpha, SDF-1, and IL-6 protein production was significantly greater in gingival fibroblasts compared to periodontal ligament fibroblasts. IL-6 was upregulated in a time-dependent manner, mainly in gingival fibroblasts (P<0.05), which secreted more MIP-1 alpha in the lowest concentration of LPS used (0.1 mu g/ml). In contrast, a basal production of SDF-1 that was inhibited with the increase of LPS concentration was detected, especially after 24 hours (P<0.05). Conclusion: The distinct ability of the gingival and periodontal ligament fibroblasts to secrete MIP-1 alpha, SDF-1, and IL-6 emphasizes that these cells may differently contribute to the balance of cytokines in the LPS-challenged periodontium. J Periodontol 2010;81:310-317.

Effects of Orthopedic Rapid Maxillary Expansion on Internal Nasal Dimensions in Children With Cleft Lip and Palate Assessed by Acoustic Rhinometry

The objective of the current study was to characterize the internal nasal dimensions of children with repaired cleft lip and palate and transverse maxillary deficiency, using acoustic rhinometry and analyze the changes caused by rapid maxillary expansion (RME). A convenience sampling of 19 cleft lip and palate individuals, aged 14 to 18 years, of both sexes, previously submitted to primary surgeries and referred for RME were analyzed prospectively at the Hospital for Rehabilitation of Craniofacial Anomalies, University of Sao Paulo, Bauru, Sao Paulo, Brazil. All patients underwent acoustic rhinometry before installation of the expansor and at 30 and 180 days after the active expansion phase. Nasal cross-sectional areas and volumes corresponding to the nasal valve (CSA(1) and V(1)) and the turbinates (CSA(2), CSA(3), and V(2)) regions were determined before and after nasal decongestion. Rapid maxillary expansion led to a statistically significant increase (P < 0.05) in mean CSA(1), CSA(2), V(1), and V(2) (without nasal decongestion) and in CSA(1) and V(1) (with decongestion) in the group as a whole. Individual data analysis showed that 58% of the patients responded positively to RME, with an average increase in CSA(1) of 26% (with decongestion), whereas 37% of the patients had no significant change. Only 1 patient (5%) showed a decrease. The findings contribute toward the characterization of nasal deformities determined by the cleft and demonstrate the positive effect RME had on nasal morphophysiology in a significant number of the patients who underwent this procedure.

Influence of curing protocol on selected properties of light-curing polymers: Degree of conversion, volume contraction, elastic modulus, and glass transition temperature

Objectives. The purpose of this study was to investigate the effect of light-curing protocol on degree of conversion (DC), volume contraction (C), elastic modulus (E), and glass transition temperature (T(g)) as measured on a model polymer. It was a further aim to correlate the measured values with each other. Methods. Different light-curing protocols were used in order to investigate the influence of energy density (ED), power density (PD), and mode of cure on the properties. The modes of cure were continuous, pulse-delay, and stepped irradiation. DC was measured by Raman micro-spectroscopy. C was determined by pycnometry and a density column. E was measured by a dynamic mechanical analyzer (DMA), and T(g) was measured by differential scanning calorimetry (DSC). Data were submitted to two-and three-way ANOVA, and linear regression analyses. Results. ED, PD, and mode of cure influenced DC, C, E, and T(g) of the polymer. A significant positive correlation was found between ED and DC (r = 0.58), ED and E (r = 0.51), and ED and T(g) (r = 0.44). Taken together, ED and PD were significantly related to DC and E. The regression coefficient was positive for ED and negative for PD. Significant positive correlations were detected between DC and C (r = 0.54), DC and E (r = 0.61), and DC and T(g) (r = 0.53). Comparisons between continuous and pulse-delay modes of cure showed significant influence of mode of cure: pulse-delay curing resulted in decreased DC, decreased C, and decreased T(g). Influence of mode of cure, when comparing continuous and step modes of cure, was more ambiguous. A complex relationship exists between curing protocol, microstructure of the resin and the investigated properties. The overall performance of a composite is thus indirectly affected by the curing protocol adopted, and the desired reduction of C may be in fact a consequence of the decrease in DC. (C) 2009 Academy of Dental Materials. Published by Elsevier Ltd. All rights reserved.

Transcriptional response of murine macrophages upon infection with opsonized Paracoccidioides brasiliensis yeast cells

Paracoccidioides brasiliensis is the etiologic agent of the Paracoccidioidomycosis the most common systemic mycosis in Latin America. Little is known about the regulation of genes involved in the innate immune host response to P. brasiliensis. We therefore examined the kinetic profile of gene expression of peritoneal macrophage infected with P. brasiliensis. Total RNA from macrophages at 6, 24 and 48 h was extracted, hybridized onto nylon membranes and analyzed. An increase in the transcription of a number of pro-inflammatory molecules encoding membrane proteins, metalloproteases, involved in adhesion and phagocytosis, are described. We observed also the differential expression of genes whose products may cause apoptotic events induced at 24 h. In addition, considering the simultaneous analyses of differential gene expression for the pathogen reported before by our group, at six hours post infection, we propose a model at molecular level for the P. brasiliensis-macrophage early interaction. In this regard, P. brasiliensis regulates genes specially related to stress and macrophages, at the same time point, up-regulate genes related to inflammation and phagocytosis, probably as an effort to counteract infection by the fungus. (c) 2007 Elsevier Masson SAS. All fights reserved.

Evaluation of chronic omega-3 fatty acids supplementation on behavioral and neurochemical alterations in 6-hydroxydopamine-lesion model of Parkinson`s disease

Omega-3 polyunsaturated fatty acids (omega-3 PUFAs) have been widely associated to beneficial effects over different neuropathologies, but only a few studies associate them to Parkinson`s disease (PD). Rats were submitted to chronic supplementation (21-90 days of life) with fish oil, rich in omega-3 PUFAs, and were uni- or bilaterally lesioned with 4 mu g of the neurotoxin 6-hydroxydopamine (6-OHDA) in the medial forebrain bundle Although lipid incorporation was evidenced in neuronal membranes, it was not sufficient to compensate motor deficits induced by 6-OHDA. In contrast, omega-3 PUFAs were capable of reducing rotational behavior induced by apomorphine, suggesting neuroprotection over dyskinesia The beneficial effects of omega-3 PUFAs were also evident in the maintenance of thiobarbituric acid reactive substances index from animals lesioned with 6-OHDA similar to levels from SHAM and intact animals. Although omega-3 PUFAs did not modify the tyrosine hydroxylase immunoreactivity in the substantia nigra pars compacta and in the ventral tegmental area, nor the depletion of dopamine (DA) and its metabolites in the striatum, DA turnover was increased after omega-3 PUFAs chronic supplementation Therefore, it is proposed that omega-3 PUFAs action characterizes the adaptation of remaining neurons activity. altering striatal DA turnover without modifying the estimated neuronal population. (C) 2009 Elsevier Ireland Ltd and the Japan Neuroscience Society. All rights reserved

A simple, inexpensive and easily reproducible model of spinal cord injury in mice: Morphological and functional assessment

Spinal cord injury (SCI) causes motor and sensory deficits that impair functional performance, and significantly impacts life expectancy and quality. Animal models provide a good opportunity to test therapeutic strategies in vivo. C57BL/6 mice were subjected to laminectomy at T9 and compression with a vascular clip (30 g force, 1 min). Two groups were analyzed: injured group (SCI, n = 33) and laminectomy only (Sham, n = 15). Locomotor behavior (Basso mouse scale-BMS and global mobility) was assessed weekly. Morphological analyses were performed by LM and EM. The Sham group did not show any morphofunctional alteration. All SCI animals showed flaccid paralysis 24 h after injury. with subsequent improvement. The BMS score of the SCI group improved until the intermediate phase (2.037 +/- 1.198): the Sham animals maintained the highest BMS score (8.981 +/- 0.056). p < 0.001 during the entire time. The locomotor speed was slower in the SCI animals (5.581 +/- 0.871) than in the Sham animals (15.80 +/- 1.166), p < 0.001. Morphological analysis of the SCI group showed, in the acute phase, edema, hemorrhage, multiple cavities, fiber degeneration, cell death and demyelination. In the chronic phase we observed glial scarring, neuron death, and remyelination of spared axons by oligodendrocytes and Schwann cells. In conclusion, we established a simple, reliable, and inexpensive clip compression model in mice, with functional and morphological reproducibility and good validity. The availability of producing reliable injuries with appropriate outcome measures represents great potential for studies involving cellular mechanisms of primary injury and repair after traumatic SCI. (C) 2008 Elsevier B.V. All rights reserved.