A Novel Synaptobrevin/VAMP Homologous Protein (VAMP5) Is Increased during In Vitro Myogenesis and Present in the Plasma Membrane

cDNA clones encoding a novel protein (VAMP5) homologous to synaptobrevins/VAMPs are detected during database searches. The predicted 102–amino acid VAMP5 harbors a 23-residue hydrophobic region near the carboxyl terminus and exhibits an overall amino acid identity of 33% with synaptobrevin/VAMP1 and 2 and cellubrevin. Northern blot analysis reveals that the mRNA for VAMP5 is preferentially expressed in the skeletal muscle and heart, whereas significantly lower levels are detected in several other tissues but not in the brain. During in vitro differentiation (myogenesis) of C2C12 myoblasts into myotubes, the mRNA level for VAMP5 is increased ∼8- to 10-fold. Immunoblot analysis using antibodies specific for VAMP5 shows that the protein levels are also elevated ∼6-fold during in vitro myogenesis of C2C12 cells. Indirect immunofluorescence microscopy and immunoelectron microscopy reveal that VAMP5 is associated with the plasma membrane as well as intracellular perinuclear and peripheral vesicular structures of myotubes. Epitope-tagged versions of VAMP5 are similarly targeted to the plasma membrane.

Intrapulpar temperature during continuous CO2 laser irradiation in human molars: An in vitro study

To establish safety parameters, we in vitro studied the increase in intrapulpal temperature caused by the use of a cw CO2 laser. A thermistor was implanted in the inner part of the pulpal chamber of 25 human lower third molars to measure the intrapulpal temperature produced by laser powers between 2-10 W and exposure times of 0.5-25.0 s. The Pearson linear correlation factor applied to the measured values showed there is a direct relationship between the independent variable and the applied power. A variance analysis produced the linear regression equation: T=1.10+(0.127)E where T is the temperature and E the energy. The results showed that, with a power of 4 W and maximum exposure time of 2.5 s (10 J) and a power density of 12738.85 W cm-2, there will be no damaging reactions affecting the pulpal tissues.

Chemotaxis of T-cells after infection of human choroid plexus papilloma cells with Echovirus 30 in an in vitro model of the blood-cerebrospinal fluid barrier

Enterovirus is the most common pathogen causing viral meningitis especially in children. Besides the blood-brain barrier (BBB) the choroid plexus, which forms the blood-cerebrospinal-fluid (CSF) barrier (BCSFB), was shown to be involved in the pathogenesis of enteroviral meningitis. In a human in vitro model of the BCSFB consisting of human choroid plexus papilloma cells (HIBCPP), the permissiveness of plexus epithelial cells for Echovirus 30 (EV30) was analyzed by immunoblotting and quantitative real-time PCR (Q-PCR). HIBCPP could be directly infected by EV30 from the apical as well as from the physiological relevant basolateral side. During an infection period of 5h no alterations of barrier function and cell viability could be observed. Analysis of the cytokine/chemokine-profile following enteroviral infection with a cytometric bead array (CBA) and Q-PCR revealed an enhanced secretion of PanGRO (CXCL1, CXCL2 and CXCL3), IL8 and CCL5. Q-PCR showed a significant upregulation of CXCL1, CXCL2 and CXCL3 in a time dependant manner. However, there was only a minor effect of HIBCPP-infection with EV30 on transepithelial T lymphocyte migration with or without the chemoattractant CXCL12. Moreover, CXCL3 did not significantly enhance T cell migrations. Therefore additional factors must be involved for the in vivo reported enhanced T cell migration into the CNS in the context of enteroviral meningitis. As HIBCPP are permissive for infection with EV30, they constitute a valuable human in vitro model to study viral infection at the BCSFB.

ANALISE FAMILIAL DO COMPORTAMENTO IN VITRO DOS MACROFAGOS HUMANOS FRENTE AO MYCOBACTERIUM LEPRAE

The lysogenic capacity of human macrophages facing M. leprae in vitro may be dependent on an important genetic component. Although the familial aggregation of the trait is demonstrated, this is a necessary but not sufficient condition to prove genetic influence. The data do not fit some simple genetic models (autosomal dominant or incompletely dominant gene; dominant or recessive sex-linked gene). The results obtained are consistent with the hypothesis that the macrophages' lysogenic capacity is mainly due to a major gene with variable expressivity. This hypothesis may be too simple to account for the whole variability detected and therefore must be considered a working hypothesis.

In vitro evaluation of marginal leakage in bonded restorations, with mechanical or chemical-mechanical (Carisolv) removal of carious tissue

This study carried out an in vitro evaluation and comparison of the occurrence of marginal leakage in bonded restorations using mechanical or chemical-mechanical (Carisolv) removal of carious tissue. For that purpose, 40 extracted decayed human molars were divided into 4 groups: GI (burs + Prime & Bond NT + TPH), GII (Carisolv + Prime & Bond NT + TPH), GIII (burs + SBMP + Z100) and GIV (Carisolv + SBMP + Z100). After accomplishment of the restorations and thermal cycling, the teeth were exposed to dye, sectioned and qualitatively evaluated. The results demonstrated that the system of removal of carious tissue did not influence the results of microleakage at any of the cavity margins. At dentinal margins, use of the Prime & Bond NT + TPH restorative system allowed the occurrence of less microleakage than the SBMP + Z100 system.

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.

The in vitro antimicrobial activity of natural infant fluoride-free toothpastes on oral micro-organisms

Purpose: The objective of this study was to evaluate the antimicrobial activity of six toothpastes for infants: 3 fluoride-free experimental toothpastes - cashew-based, mango-based and without plant extract and fluoride compared with 2 commercially fluoride-free toothpastes and 1 fluoridated toothpastes. Methods: Six toothpastes for infants were evaluated in this study: (1) experimental cashew-based toothpaste; (2) experimental mango-based toothpaste; (3) experimental toothpaste without plant extract and fluoride (negative control); (4) First Teeth brand toothpaste; (5) Weleda brand toothpaste; and (6) Tandy brand toothpaste (positive control). The antimicrobial activity was recorded against Streptococcus mutans, Streptococcus sobrinus, Lactobacillus acidophilus, and Candida albicans using the agar plate diffusion test. Results: First Teeth, Weleda, mango-based toothpaste, and toothpaste without plant extract presented no antimicrobial effect against any of the tested micro-organisms. Cashew toothpaste had antimicrobial activity against S mutans, S sobrinus, and L acidophilus, but it showed no antimicrobial activity against C albicans. There was no statistical difference between the inhibition halo of cashew and Tandy toothpastes against S mutans and L acidophilus. Conclusions: Cashew fluoride-free toothpaste had inhibitory activity against Streptococcus mutans and Lactobacillus acidophilus, and these results were similar to those obtained for fluoridated toothpaste.