Metformin and soybean-derived bioactive molecules attenuate the expansion of stem cell-like epithelial subpopulation and confer apoptotic sensitivity in human colon cancer cells

Colorectal cancer (CRC) is a disease whose genesis may include metabolic dysregulation. Cancer stem cells are attractive targets for therapeutic interventions since their aberrant expansion may underlie tumor initiation, progression, and recurrence. To investigate the actions of metabolic regulators on cancer stem cell-like cells (CSC) in CRC, we determined the effects of soybean-derived bioactive molecules and the anti-diabetes drug metformin (MET), alone and together, on the growth, survival, and frequency of CSC in human HCT116 cells. Effects of MET (60 μM) and soybean components genistein (Gen, 2 μM), lunasin (Lun, 2 μM), β-conglycinin (β-con, 3 μM), and glycinin (Gly, 3 μM) on HCT116 cell proliferation, apoptosis, and mRNA/protein expression and on the frequency of the CSC CD133(+)CD44(+) subpopulation by colonosphere assay and fluorescence-activated cell sorting/flow cytometry were evaluated. MET, Gen, and Lun, individually and together, inhibited HCT116 viability and colonosphere formation and, conversely, enhanced HCT116 apoptosis. Reductions in frequency of the CSC CD133(+)CD44(+) subpopulation with MET, Gen, and Lun were found to be associated with increased PTEN and reduced FASN expression. In cells under a hyperinsulinemic state mimicking metabolic dysregulation and without and with added PTEN-specific inhibitor SF1670, colonosphere formation and frequency of the CD133(+)CD44(+) subpopulation were decreased by MET, Lun and Gen, alone and when combined. Moreover, MET + Lun + Gen co-treatment increased the pro-apoptotic and CD133(+)CD44(+)-inhibitory efficacy of 5-fluorouracil under hyperinsulinemic conditions. Results identify molecular networks shared by MET and bioavailable soy food components, which potentially may be harnessed to increase drug efficacy in diabetic and non-diabetic patients with CRC.

Aspectos biológicos e bioatividade de materiais a base de MTA em cultura de células

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Biologia pulpar: da agressão à reparação

The study of the dental pulp can be extended from factors related to its aggression to those related to new concepts of regeneration. The purpose of this compilation of studies is to present the evolution of a research subject from damage to repair. Innitially, studies will demonstrate the ability of dental procedures to generate heat and consequently affect the dental pulp. In sequence, studies will also present some effects of different pulp capping materials on dental pulp cells, related to the cytotoxicity of these materials and inflammatory potential. Finally, as the subject is emmerging and gaining importance in the literature, this compilation will present data from recent studies on the role of dental pulp progenitor cells in the regeneration and repair of dental pulp, as well as an alternative for a scaffold that could be used for clinical translation of research in the field. In summary, dentists must be aware of these different aspects and that the knowledge on factors and mechanisms involved in the aggression of the dental pulp can also serve as basis for understanding aspects for regeneration.

Evaluation of two mineral trioxide aggregate compounds as pulp-capping agents in human teeth

Aim: The present randomized, controlled prospective study evaluated the histomorphological response of human dental pulps capped with two grey mineral trioxide aggregate (MTA) compounds. Methodology: Pulp exposures were performed on the occlusal floor of 40 human permanent pre-molars. The pulp was capped either with ProRoot (Dentsply) or MTA-Angelus (Angelus) and restored with zinc oxide eugenol cement. After 30 and 60 days, teeth were extracted and processed for histological examination and the effects on the pulp were scored. The data were subjected to Kruskal-Wallis and Conover tests (α = 0.05). Results: In five out of the 40 teeth bacteria were present in pulp tissue. No significant difference was observed between the two materials (P > 0.05) in terms of overall histological features (hard tissue bridge, inflammatory response, giant cells and particles of capping materials). Overall, 94% and 88% of the specimens capped with MTA-Angelus and ProRoot, respectively, showed either total or partial hard tissue bridge formation (P > 0.05). Conclusions: Both commercial materials ProRoot (Dentsply) and MTA-Angelus (Angelus) produced similar responses in the pulp when used for pulp capping in intact, caries-free teeth. © 2009 International Endodontic Journal.

Evaluation of mineral trioxide aggregate and calcium hydroxide cement as pulp-capping agents in human teeth

This study evaluated the histomorphologic response of human dental pulps capped with mineral trioxide aggregate (MTA) and Ca(OH)(2) cement (CH). Pulp exposures were performed on the occlusal floor of 40 human permanent premolars. After that, the pulp was capped either with CH or MTA and restored with composite resin. After 30 and 60 days, teeth were extracted and processed for histologic exam and categorized in a histologic score system. The data were subjected to Kruskal-Wallis and Conover tests (alpha = .05). All groups performed well in terms of hard tissue bridge formation, inflammatory response, and other pulpal findings. However, a lower response of CH30 was observed for the dentin bridge formation, when compared with MTA30 and MTA60 groups. Although the pulp healing with calcium hydroxide was slower than that of MTA, both materials were successful for pulp capping in human teeth.

Penetration of 38% hydrogen peroxide into the pulp chamber in bovine and human teeth submitted to office bleach technique

This study evaluated the pulp chamber penetration of peroxide bleaching agent in human and bovine teeth after office bleach technique. All the teeth were sectioned 3 mm apical of the cement-enamel junction and were divided into 2 groups, A (70 third human molars) and B (70 bovine lateral incisors), that were subdivided into A1 and B1 restored by using composite resin, A2 and B2 by using glass ionomer cement, and A3 and B3 by using resin-modified glass ionomer cement; A4, A5, B4, and B5 were not restored. Acetate buffer was placed in the pulp chamber, and the bleaching agent was applied for 40 minutes as follows: A1-A4 and B1-B4, 38% hydrogen peroxide exposure and A5 and B5, immersion into distilled water. The buffer solution was transferred to a glass tube in which leuco crystal violet and horseradish peroxidase were added, producing a blue solution. The optical density of the blue solution was determined by spectrophotometer and converted into microgram equivalents of hydrogen peroxide. Data were submitted to analysis of variance and Dunnett, Kruskal-Wallis, and Tukey tests (5%). A higher level of hydrogen peroxide penetrated into the pulp chamber in resin-modified glass ionomer cements in bovine (0.79 +/- 0.61 mu g) and human (2.27 +/- 0.41 mu g) groups. The bleaching agent penetration into the pulp chamber was higher in human teeth for any experimental situation. The penetration of the hydrogen peroxide depends on restorative materials, and under the conditions of this study human teeth are more susceptible to penetration of bleaching agent into the pulp chamber than bovine teeth.

Response of human pulps capped with a self-etching adhesive system

Objective: The aim of this study was to evaluate the human pulp response following direct pulp capping with a current self-etching bonding agent and calcium hydroxide (CH). Methods: Thirty-three sound human premolars had their pulp tissue mechanically exposed. Sterile distilled water was used to control the hemorrhage and exudation from the pulp exposure site. The pulps were capped with Clearfil Liner Bond 2 (CLB-2) or CH and the cavities were filled with a resin composite (Z-100) according to the manufacturer's instructions. After 5, 30 and 120-300 days, the teeth were extracted and processed for microscopic examination. Results: At short-term, CLB-2 elicited a mild to moderate inflammatory pulp response with dilated and congested blood vessels adjacent to pulp exposure site. With time, macrophages and giant cells engulfing globules and particulates of resinous material displaced into the pulp space were observed. This chronic inflammatory pulp response triggered by fragments of bonding agent displaced into the pulp space did not allow pulp repair interfering with the dentin bridging. On the other hand, pulps capped with CH exhibited an initial organization of elongated pulp cells underneath the coagulation necrosis. Pulp repair and complete dentin bridge formation was observed at long-term evaluation. Significance: The present study demonstrated that CH remains the pulp capping agent of choice for mechanically exposed human pulps. CLB-2 did not allow complete connective tissue repair adjacent to the pulp exposure site. Consequently, this bonding agent cannot be recommended for pulp therapy of sound human teeth. © 2001 Academy of Dental Materials. Published by Elsevier Science Ltd. All rights reserved.

Biocompatibility of resin-based dental materials applied as liners in deep cavities prepared in human teeth

Background: Since only a few data have been published concerning the effects of resinous dental materials on the pulp-dentin complex, the aim of this study was to evaluate the biocompatibility of resin-based materials applied as liners in deep cavities prepared in duman teeth. Methods: After preparing class V cavities, the following dental materials were applied on the axial walls: group 1, Vitrebond™ (VIT; 3M ESPE); group 2, Ultra-Blend® Plus™ (UBP; Untradent); and group 3, Clearfil™ SE Bond (CSEB; Kuraray). In group 4 (control), the hard-setting calcium hydroxide cement Dycal (CH; Caulk/Dentsply) was used. The teeth extracted at 7 days or between 30 and 85 days after the clinical procedures were processed for histological evaluation. Results: For all the experimental and control groups, most of specimens exhibited no pulpal response or slight inflammatory reaction associated with slight tissue disorganization at 7-day period. Moderate inflammatory pulpal response occurred only in one tooth (RDT = 262 μm) of group 3 in which transdentinal diffusion of resin components was observed. Conclusion: The resin-based dental cements VIT and UBP as well as the bonding agent CSEB presented acceptable biocompatibility when applied in deep cavities prepared in sound human teeth. © 2006 Wiley Periodicals, Inc.

In vitro assessment of pulp chamber temperature of different teeth submitted to dental bleaching associated with LED/laser and halogen lamp appliances

This study sought to assess the pulp chamber temperature in different groups of human teeth that had been bleached using hydrogen peroxide gel activated with halogen lamps or hybrid LED/laser appliances. Four groups of ten teeth (maxillary central incisors, mandibular incisors, mandibular canines, and maxillary canines) were used. A digital thermometer with a K-type thermocouple was placed inside pulp chambers that had been filled with thermal paste. A 35% hydrogen peroxide-based red bleaching gel was applied to all teeth and photocured for a total of three minutes and 20 seconds (five activations of 40 seconds each), using light from an LED/laser device and a halogen lamp. The temperatures were gauged every 40 seconds and the data were analyzed by three-way ANOVA, followed by Tukey's test. Regardless of the light source, statistically significant differences were observed between the groups of teeth. The mean temperature values (±SD) were highest for maxillary central incisors and lowest for mandibular canines. The halogen lamp appliance produced more pulp chamber heating than the LED/laser appliance. The increase in irradiation time led to a significant increase in temperature.

Influence of the coloring agent concentration on bleaching gel and pulp chamber temperatures during dental bleaching

This study evaluated the Influence of the coloring agent concentration on the temperature of the gel layer and pulp chamber during dental bleaching with an LED/laser light source. Ten human incisors and a digital thermometer with K-type thermocouples were used. Using a high-speed spherical diamond bur, endodontic access was gained through openings on the lingual faces until pulp chamber was exposed. One end of the thermocouple was placed on the labial surface (immersed in bleaching gel) and the other end in the pulp chamber. The same 10 specimens were used in the 12 groups, according to the type and concentration of bleaching gel. Each bleaching gel was used in four different concentrations: manipulated without coloring, with normal quantity recommended by the manufacturer, with double the recommended amount of coloring, and with triple the recommended amount of coloring. The temperature rise was measured every 30 seconds for three minutes with a K-type thermocouple. The data were analyzed by ANOVA to examine the concentration and type of bleaching gel. This test was followed by Tukey's test, which was performed Independently for the gel at the labial surface and the pulp chamber (a = 5%). For both surfaces, values of p = 0.00 were obtained for all factors and for the Interaction between them. The varying concentrations of coloring agent produced statistically significant differences in terms of temperature increase for both the gel layer and the pulp chamber during activation.

In Vitro Regulation of CCL3 and CXCL12 by Bacterial By-products Is Dependent on Site of Origin of Human Oral Fibroblasts

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Phototherapy up-regulates dentin matrix proteins expression and synthesis by stem cells from human-exfoliated deciduous teeth

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Immediate human pulp response to ethanol-wet bonding technique

To evaluate the short-term response of human pulps to ethanol-wet bonding technique. Methods Deep class V cavities were prepared on 17 sound premolars and divided into three groups. After acid-etching, the cavities from groups 1 (G1) and 2 (G2) were filled with 100% ethanol or distilled water, respectively, for 60 s before the application of Single Bond 2. In group 3 (G3, control), the cavity floor was lined with calcium hydroxide before etching and bonding. All cavities were restored with resin composite. Two teeth were used as intact control. The teeth were extracted 48 h after the clinical procedures. From each tooth serial sections were obtained and stained with haematoxylin and eosin (H/E) and Masson's trichrome. Bacteria microleakage was assessed using Brown & Brenn. All sections were blindly evaluated for five histological features. Results Mean remaining dentine thickness was 463 ± 65 μm (G1); 425 ± 184 μm (G2); and 348 ± 194 μm (G3). Similar pulp reactions followed ethanol- or water-wet bonding techniques. Slight inflammatory responses and disruption of the odontoblast layer related to the cavity floor were seen in all groups. Stained bacteria were not detected in any cavities. Normal pulp tissue was observed in G3 except for one case. Conclusions After 48 h, ethanol-wet bonding does not increase pulpal damage compared to water-wet bonding technique. Clinical significance Ethanol-wet bonding may increase resin-dentine bond durability. This study reported the in vivo response of human pulp tissue when 100% ethanol was applied previously to an etch-and-rinse simplified adhesive system.

Diabetes triggers the loss of tooth structure associated to radiographical and histological dental changes and its evolution to progressive pulp and periapical lesions in rats

The aim of this study was to evaluate the putative influence of diabetes without metabolic control in the loss of tooth structure as well as histological changes in dentin and pulp tissue in rats. Diabetes was induced in Wistar rats (n=25) by intravenous administration of alloxan (42mg/kg). Diabetic and non-diabetic control rats were evaluated at 1, 3, 6, 9 and 12 months of follow-up. In order to evaluate the presence and progression of dental caries and periapical lesions, hemimandibles were removed and submitted to radiographical, histological, and morphometrical procedures. Dental caries were detected after radiographical and histological evaluations in diabetic group from the third month of diabetes onset, increasing gradually in frequency and severity in periods. Diabetic rats dental pulps also presented significant reduction in volume density of collagen fibers and fibroblasts at third month, parallel with a trend towards the increase in inflammatory cells volume density. Diabetic rats presented a generalized pulp tissue necrosis after 6 months of diabetes induction. Moreover, periapical lesions were not detected in control group, while these lesions were observed in all rats after 3, 6, 9, and 12 months of diabetes induction. Uncontrolled diabetes seems to trigger the loss of tooth structure, associated to histological dental changes and mediates its evolution to progressive severe pulp and periapical lesions in rats. Therefore, diabetes may be considered a very important risk factor regarding alterations in dental pulp, development of dental caries, and periapical lesions.

Culture of osteogenic cells from human alveolar bone: A useful source of alkaline phosphatase

The aim of this study was to obtain membrane-bound alkaline phosphatase from osteoblastic-like cells of human alveolar bone. Cells were obtained by enzymatic digestion and maintained in primary culture in osteogenic medium until subconfluence. First passage cells were cultured in the same medium and at 7, 14, and 21 days, total protein content, collagen content, and alkaline phosphatase activity were evaluated. Bone-like nodule formation was evaluated at 21 days. Cells in primary culture at day 14 were washed with Tris-HCl buffer, and used to extract the membrane-bound alkaline phosphatase. Cells expressed osteoblastic phenotype. The apparent optimum pH for PNPP hydrolysis by the enzyme was pH 10.0. This enzyme also hydrolyzes ATP, ADP, fructose-1-phosphate, fructose-6-phosphate, pyrophosphate and beta-glycerophosphate. PNPPase activity was reduced by typical inhibitors of alkaline phosphatase. SDS-PAGE of membrane fraction showed a single band with activity of similar to 120 kDa that could be solubilized by phospholipase C or Polidocanol. (c) 2007 International Federation for Cell Biology. Published by Elsevier Ltd. All rights reserved.