Biological effects of insulin on murine melanoma cells and fish erythrophoroma cells: A comparative study

Insulin is the hormone that plays an essential role in metabolism and mitosis of normal and tumor cells, exerting its pleiotropic effects through binding to specific membrane receptors and promoting the phosphorylation of tyrosine residues of the receptor itself and of other components of the signaling pathway. The aim of this study was to investigate the effects of insulin on melanogenesis and cell growth in three different cell lines: the goldfish GEM-81 erythrophoroma cells (undifferentiated and differentiated with 1.5% dimethylsulfoxide-DMSO), and the murine B16F10 and Cloudman S91 melanoma cells. Undifferentiated GEM-81 and B16F10 cells responded to insulin with a small increase of cell proliferation, whereas S91 cells responded with a decrease of growth. In the two mammalian cell lines, and in DMSO-differentiated GEM-81 cells, the hormone strongly inhibited melanogenesis, by decreasing tyrosinase activity. In undifferentiated GEM-81 cells, insulin had no effect on tyrosinase activity. An increase in the tyrosine phosphorylation status of pp 185 (insulin receptor substrate 1 and 2-IRS-1/2) phosphorylation degree was observed in S91 mouse melanoma and in differentiated GEM-81 erythrophoroma cells, suggesting that this specific protein was maintained during transformation process and participates in insulin signaling. Our results imply an ancient and diverse history of the insulin signaling system in vertebrate pigment cells. (C) 2008 Elsevier Inc. All rights reserved.

Seasonal variation of peptidase activities in the reproductive tract of Crotalus durissus terrificus

Seasonal quantitative patterns of acid (APA), basic (APB), puromycin-sensitive (APN-PS) and puromycin-insensitive neutral (APN-PI), cystyl (CAP), dipeptidyl IV (DPPIV), type-1 pyroglutamyl (PAP-I) and prolylimino (PIP) aminopeptidases and prolyl oligopeptidase (POP) activities in soluble (SF) and solubilized membrane-bound (MF) fractions from ductus deferens, vagina and uterus were studied to evaluate their relationships with the reproductive cycle and the extensive long-term spermatozoa storage (LTSS) of the Neotropical rattlesnake Crotalus durissus terrificus. APB, PIP and POP were detected only in SF, while other peptidases were detected in SF and MF. APB, APN-PI and APN-PS were predominant in most tissues in all seasons. Peptidase activities had a common pattern of increment during the dry season (winter/autumn), which coincides with the mating period (autumn) and LTSS in the female (winter), as well as the reduction of spermatozoa motility and maintenance of fertilization capacity of spermatozoa. The high CAP activity in the soluble fraction of the vagina during winter, compared to summer (time of parturition) and spring, coincides with the relaxation of this tissue. In the soluble fraction, the low PAP-1 activity of the ductus deferens coincided with its high activity in the vagina during the winter; and the inverse occurred in summer, which is consistent with the physiological process of preserving spermatozoon viability. In conclusion, the studied peptidase activities had seasonal and tissue-specific characteristics, which suggest a relevant role in the reproductive physiology of C. d. terrificus. (C) 2008 Elsevier Inc. All rights reserved.

Light modulates the melanophore response to alpha-MSH in Xenopus laevis: An analysis of the signal transduction crosstalk mechanisms involved

Melanin granule (melanosome) dispersion within Xenopus laevis melanophores is evoked either by light or alpha-MSH. We have previously demonstrated that the initial biochemical steps of light and alpha-MSH signaling are distinct, since the increase in cAMP observed in response to alpha-MSH was not seen after light exposure. cAMP concentrations in response to alpha-MSH were significantly lower in cells pre-exposed to light as compared to the levels in dark-adapted melanophores. Here we demonstrate the presence of an adenylyl cyclase (AC) in the Xenopus melanophore, similar to the mammalian type IX which is inhibited by Ca(2+)-calmodulin-activated phosphatase. This finding supports the hypothesis that the cyclase could be negatively modulated by a light-promoted Ca(2+) increase. In fact, the activity of calcineurin PP2B phosphatase was increased by light, which could result in AC IX inhibition, thus decreasing the response to alpha-MSH. St-Ht31, a disrupting agent of protein kinase A (PKA)-anchoring kinase A protein (AKAP) complex totally blocked the melanosome dispersing response to alpha-MSH, but did not impair the photo-response in Xenopus melanophores. Sequence comparison of a melanophore AKAP partial clone with GenBank sequences showed that the anchoring protein was a gravin-like adaptor previously sequenced from Xenopus non-pigmentary tissues. Co-immunoprecipitation of Xenopus AKAP and the catalytic subunit of PKA demonstrated that PKA is associated with AKAP and it is released in the presence of alpha-MSH. We conclude that in X laevis melanophores, AKAP12 (gravin-like) contains a site for binding the inactive PKA thus compartmentalizing PKA signaling and also possesses binding sites for PKC. Light diminishes alpha-MSH-induced increase of cAMP by increasing calcineurin (PP2B) activity, which in turn inhibits adenylyl cyclase type IX, and/or by activating PKC, which phosphorylates the gravin-like molecule, thus destabilizing its binding to the cell membrane. (C) 2009 Elsevier Inc. All rights reserved.

Annual profile of fecal androgen and glucocorticoid levels in free-living male American kestrels from southern mid-latitude areas

Fecal samples and behavioral data were collected at a fortnightly basis during 11 months period from free-living male American kestrels living in southeast Brazil (22 degrees S latitude). The aim was to investigate the seasonal changes in testicular and adrenal steroidogenic activity and their correlation to reproductive behaviors and environmental factors. The results revealed that monthly mean of fecal glucocorticoid metabolites in May and June were higher than those estimated in November. in parallel, monthly mean of androgen metabolites in September was higher than those from January to April and from October to November. Molt took place from January to March, whereas copulation was observed from June to October but peaked in September. Nest activity and food transfer to females occurred predominantly in October, and parental behavior was noticed only in November. Territorial aggressions were rare and scattered throughout the year. Multiple regression analysis revealed that fecal androgen levels are predicted by photoperiod and copulation, while fecal glucocorticoid levels are only predicted by photoperiod. Bivariate correlations showed that fecal androgen metabolites were positively correlated with fecal glucocorticoid metabolites and copulation, but negatively correlated with molt. Additionally, copulation was positively correlated with food transfer to females and nest activity, but negatively correlated with molt. These findings suggest that male American kestrels living in southeast Brazil exhibit significant seasonal changes in fecal androgen and glucocorticoid concentrations, which seem to be stimulated by decreasing daylength but not by rainfall or temperature. (C) 2009 Elsevier Inc. All rights reserved.

Increase of core temperature induced by corticotropin-releasing factor and urocortin: A comparative study

This study compared the ability of CRF and UCN1 to induce a thermoregulatory response when centrally injected into rats. The effects of antipyretic drugs and CRF receptor antagonists (CRF(1) and CRF(2)) on the temperature (T) changes induced by these peptides were also investigated. Rectal (rT) and tail skin (T(sk)) temperatures were measured with a thermistor probe while body (bT) temperature was measured with a battery-operated biotelemetry transmitter in male Wistar rats (200 g) every 30 min over a period of 6 h, after intracerebroventricular (i.c.v.) injection of 1 nmol of either CRF or UCN1. Rats were pre-treated with indomethacin (2 mg kg(-1), i.p.) or celecoxib (5 mg kg(-1), p.o.), dexamethasone (0.5 mg kg(-1), s.c.), astressin (a CRF(1)/CRF(2) antagonist, 7 nmol, icy.) or antalarmin (a CRF(1) antagonist, 20 mg kg 1, i.p.). The increase in body temperature induced by CRF was accompanied by a reduction in T(sk) while the response induced by UCN1 was accompanied by an elevation in T(sk). Indomethacin or celecoxib did not change the increases in rT caused by either CRF or UCN1. Although dexamethasone attenuated the increase in rectal temperature in response to CRF, dexamethasone did not modify the response induced by UCN1. Astressin blocked the UCN1-induced hyperthermia and reduced CRF-induced fever. Antalarmin did not modify the hyperthermia in response to UCN1, but reduced the fever evoked by CRF. This study demonstrated that CRF by acting on the CRF(1) receptor induces a prostaglandin-independent fever which seems to depend, at least in part, on the synthesis of other mediators while UCN1 acts on the CRF(2) receptor, promoting a hyperthermic response which seems to be independent on synthesis/release of any mediator. (C) 2010 Elsevier B.V. All rights reserved.

Comparative study of dental arch width in plaster models, photocopies and digitized images

The aim of this study was to comparatively assess dental arch width, in the canine and molar regions, by means of direct measurements from plaster models, photocopies and digitized images of the models. The sample consisted of 130 pairs of plaster models, photocopies and digitized images of the models of white patients (n = 65), both genders, with Class I and Class II Division 1 malocclusions, treated by standard Edgewise mechanics and extraction of the four first premolars. Maxillary and mandibular intercanine and intermolar widths were measured by a calibrated examiner, prior to and after orthodontic treatment, using the three modes of reproduction of the dental arches. Dispersion of the data relative to pre- and posttreatment intra-arch linear measurements (mm) was represented as box plots. The three measuring methods were compared by one-way ANOVA for repeated measurements (α = 0.05). Initial / final mean values varied as follows: 33.94 to 34.29 mm / 34.49 to 34.66 mm (maxillary intercanine width); 26.23 to 26.26 mm / 26.77 to 26.84 mm (mandibular intercanine width); 49.55 to 49.66 mm / 47.28 to 47.45 mm (maxillary intermolar width) and 43.28 to 43.41 mm / 40.29 to 40.46 mm (mandibular intermolar width). There were no statistically significant differences between mean dental arch widths estimated by the three studied methods, prior to and after orthodontic treatment. It may be concluded that photocopies and digitized images of the plaster models provided reliable reproductions of the dental arches for obtaining transversal intra-arch measurements.

Comparative study of chemical and mechanical retentive systems for bonding of indirect composite resin to commercially pure titanium

This study evaluated the effect of chemical and mechanical surface treatments for cast metal alloys on the bond strength of an indirect composite resin (Artglass) to commercially pure titanium (cpTi). Thirty cylindrical metal rods (3 mm diameter x 60 mm long) were cast in grade-1 cpTi and randomly assigned to 6 groups (n=5) according to the received surface treatment: sandblasting; chemical treatment; mechanical treatment - 0.4 mm beads; mechanical treatment - 0.6 mm beads; chemical/mechanical treatment - 0.4 mm; and chemical/mechanical treatment - 0.6 mm beads. Artglass rings (6.0 mm diameter x 2.0 mm thick) were light cured around the cpTi rods, according manufacturer's specifications. The specimens were invested in hard gypsum and their bond strength (in MPa) to the rods was measured at fracture with a universal testing machine at a crosshead speed of 2.0 mm/min and 500 kgf load cell. Data were analyzed statistically by one-way ANOVA and Tukey test (a=5%). The surface treatments differed significantly from each other (p<0.05) regarding the recorded bond strengths. Chemical retention and sandblasting showed statistically similar results to each other (p=0.139) and both had significantly lower bond strengths (p<0.05) than the other treatments. In conclusion, mechanical retention, either associated or not to chemical treatment, provided higher bond strength of the indirect composite resin to cpTi.

Comparative study of two commercially pure titanium casting methods

The interest in using titanium to fabricate removable partial denture (RPD) frameworks has increased, but there are few studies evaluating the effects of casting methods on clasp behavior. OBJECTIVE: This study compared the occurrence of porosities and the retentive force of commercially pure titanium (CP Ti) and cobalt-chromium (Co-Cr) removable partial denture circumferential clasps cast by induction/centrifugation and plasma/vacuum-pressure. MATERIAL AND METHODS: 72 frameworks were cast from CP Ti (n=36) and Co-Cr alloy (n=36; control group). For each material, 18 frameworks were casted by electromagnetic induction and injected by centrifugation, whereas the other 18 were casted by plasma and injected by vacuum-pressure. For each casting method, three subgroups (n=6) were formed: 0.25 mm, 0.50 mm, and 0.75 mm undercuts. The specimens were radiographed and subjected to an insertion/removal test simulating 5 years of framework use. Data were analyzed by ANOVA and Tukey's to compare materials and cast methods (α=0.05). RESULTS: Three of 18 specimens of the induction/centrifugation group and 9 of 18 specimens of plasma/vacuum-pressure cast presented porosities, but only 1 and 7 specimens, respectively, were rejected for simulation test. For Co-Cr alloy, no defects were found. Comparing the casting methods, statistically significant differences (p<0.05) were observed only for the Co-Cr alloy with 0.25 mm and 0.50 mm undercuts. Significant differences were found for the 0.25 mm and 0.75 mm undercuts dependent on the material used. For the 0.50 mm undercut, significant differences were found when the materials were induction casted. CONCLUSION: Although both casting methods produced satisfactory CP Ti RPD frameworks, the occurrence of porosities was greater in the plasma/vacuum-pressure than in the induction/centrifugation method, the latter resulting in higher clasp rigidity, generating higher retention force values.

Comparative study of two autogenous graft techniques using piezosurgery for sinus lifting

PURPOSE: Maxillary sinus lifting is a technique, in which, a possible complication is sinus membrane perforation. The aim of this study was to compare two techniques using ultrasound surgery to perform autogenous graft for maxillary sinus lifting. METHODS: Ten rabbits were used in the study, one of them did not undergo surgery. The other nine rabbits had their maxillary sinuses filled with autogenous bone grafts collected from the external skull diploe in particulate form on the right side, and shaved on the left side, both with ultrasonic device. Data on bone density in left and right maxillary sinus, obtained by computed tomography in transverse and longitudinal sections, recorded 90 days after the grafts, were statistically compared. RESULTS: There were no statistically significant differences between the two techniques that used shaved and particulate bone collected by means of ultrasonic device from rabbit skulls. CONCLUSION: Assessment of operative procedures led to the conclusion that piezoelectric ultrasound was shown to be a safe tool in the surgical approach to the maxillary sinus of rabbits, allowing sinus membrane integrity to be maintained during surgical procedures.

Comparative study of intermaxillary relationships of manual and swallowing methods

This study compared the mandibular displacement from three methods of centric relation record using an anterior jig associated with (A) chin point guidance, (B) swallowing (control group) and (C) bimanual manipulation. Ten patients aged 25-39 years were selected if they met the following inclusion criteria: complete dentition (up to the second molars), Angle class I and absence of signs and symptoms of temporomandibular disorders and diagnostic casts showing stability in the maximum intercuspation (MI) position. Impressions of maxillary and mandibular arches were made with an irreversible hydrocolloid impression material. Master casts of each patient were obtained, mounted on a microscope table in MI as a reference position and 5 records of each method were made per patient. The mandibular casts were then repositioned with records interposed and new measurements were obtained. The difference between the two readings allowed measuring the displacement of the mandible in the anteroposterior and lateral axes. Data were analyzed statistically by ANOVA and Tukey's test at 5% significance level. There was no statistically significant differences (p>0.05) among the three methods for measuring lateral displacement (A=0.38 ± 0.26, B=0.32 ± 0.25 and C=0.32 ± 0.23). For the anteroposterior displacement (A=2.76 ± 1.43, B=2.46 ± 1.48 and C=2.97 ± 1.51), the swallowing method (B) differed significantly from the others (p<0.05), but no significant difference (p>0.05) was found between chin point guidance (A) and bimanual manipulation (C). In conclusion, the swallowing method produced smaller mandibular posterior displacement than the other methods.

Micromorphological and hardness analyses of human and bovine sclerotic dentin: a comparative study

The purpose of this study was to test the hypothesis that both human and bovine sclerotic dentin have similar hardness properties, in addition to similar micromorphological characteristics. Sixteen teeth (8 human and 8 bovine) exhibiting exposed dentin in the incisal edge and showing characteristics typical of sclerosis were used. Vickers surface microhardness testing was conducted. Three areas of the dentin surface of each specimen were selected. All teeth were processed for scanning electron microscopy in order to estimate the amount (in percentage) of solid dentin on the sclerotic dentin surface. The data were compared by Student's t test (α = 0.05). The micromorphological and microhardness data were compared by Pearson's linear correlation test (α = 0.05). The mean percentages of solid dentin of human and bovine sclerotic dentin were similar (human 90.71 ± 0.83 and bovine 89.08 ± 0.81, p = 0.18). The mean microhardness value (VHN) of human sclerotic dentin was significantly higher than that of bovine sclerotic dentin (human 45.26 ± 2.92 and bovine 29.93 ± 3.83, p = 0.006). No correlation was found between the microhardness values and the amount of solid dentin in the sclerotic dentin, irrespective of the species considered (human R² = 0.0240, p = 0.714; bovine R² = 0.0017, p = 0.923; and combined R² = 0.038, p = 0.46). We concluded that although both bovine and human sclerotic dentin present a similar amount of solid tissue, human sclerotic dentin presents higher microhardness than bovine sclerotic dentin.

Comparative analysis of microsatellite variability in five macaw species (Psittaciformes, Psittacidae): application for conservation

Cross-amplification was tested and variability in microsatellite primers (designed for Neotropical parrots) compared, in five macaw species, viz., three endangered blue macaws (Cyanopsitta spixii [extinct in the wild], Anodorhynchus leari [endangered] and Anodorhynchus hyacinthinus [vulnerable]), and two unthreatened red macaws (Ara chloropterus and Ara macao). Among the primers tested, 84.6% successfully amplified products in C. spixii, 83.3% in A. leari, 76.4% in A. hyacinthinus, 78.6% in A. chloropterus and 71.4% in A. macao. The mean expected heterozygosity estimated for each species, and based on loci analyzed in all the five, ranged from 0.33 (A. hyacinthinus) to 0.85 (A. macao). As expected, the results revealed lower levels of genetic variability in threatened macaw species than in unthreatened. The low combined probability of genetic identity and the moderate to high potential for paternity exclusion, indicate the utility of the microsatellite loci set selected for each macaw species in kinship and population studies, thus constituting an aid in planning in-situ and ex-situ conservation.

Tensile bond strength and SEM analysis of enamel etched with Er:YAG laser and phosphoric acid: a comparative study In vitro

Er:YAG laser has been studied as a potential tool for restorative dentistry due to its ability to selectively remove oral hard tissue with minimal or no thermal damage to the surrounding tissues. The purpose of this study was to evaluate in vitro the tensile bond strength (TBS) of an adhesive/composite resin system to human enamel surfaces treated with 37% phosphoric acid, Er:YAG laser (lambda=2.94 mum) with a total energy of 16 J (80 mJ/pulse, 2Hz, 200 pulses, 250 ms pulse width), and Er:YAG laser followed by phosphoric acid etching. Analysis of the treated surfaces was performed by scanning electron microscopy (SEM) to assess morphological differences among the groups. TBS means (in MPa) were as follows: Er:YAG laser + acid (11.7 MPa) > acid (8.2 MPa) > Er:YAG laser (6.1 MPa), with the group treated with laser+acid being significantly from the other groups (p=0.0006 and p= 0.00019, respectively). The groups treated with acid alone and laser alone were significantly different from each other (p=0.0003). The SEM analysis revealed morphological changes that corroborate the TBS results, suggesting that the differences in TBS means among the groups are related to the different etching patterns produced by each type of surface treatment. The findings of this study indicate that the association between Er:YAG laser and phosphoric acid can be used as a valuable resource to increase bond strength to laser-prepared enamel.

Cloning, purification and comparative characterization of two digestive lysozymes from Musca domestica larvae

cDNA coding for two digestive lysozymes (MdL1 and MdL2) of the Musca domestica housefly was cloned and sequenced. MdL2 is a novel minor lysozyme, whereas MdL1 is the major lysozyme thus far purified from M. domestica midgut. MdL1 and MdL2 were expressed as recombinant proteins in Pichia pastoris, purified and characterized. The lytic activities of MdL1 and MdL2 upon Micrococcus lysodeikticus have an acidic pH optimum (4.8) at low ionic strength (μ = 0.02), which shifts towards an even more acidic value, pH 3.8, at a high ionic strength (μ = 0.2). However, the pH optimum of their activities upon 4-methylumbelliferyl N-acetylchitotrioside (4.9) is not affected by ionic strength. These results suggest that the acidic pH optimum is an intrinsic property of MdL1 and MdL2, whereas pH optimum shifts are an effect of the ionic strength on the negatively charged bacterial wall. MdL2 affinity for bacterial cell wall is lower than that of MdL1. Differences in isoelectric point (pI) indicate that MdL2 (pI = 6.7) is less positively charged than MdL1 (pI = 7.7) at their pH optima, which suggests that electrostatic interactions might be involved in substrate binding. In agreement with that finding, MdL1 and MdL2 affinities for bacterial cell wall decrease as ionic strength increases.

Comparative morphology among representatives of main taxa of Scaphopoda and basal protobranch Bivalvia (Mollusca)

This study deals with detailed morphology and anatomy of 4 species of Scaphopoda and 5 species of protobranch Bivalvia. Both classes are traditionally grouped in the taxon Diasoma, which has been questioned by different methodologies, such as molecular and developmental. This study is developed under a phylogenetic methodology with the main concern in performing it in an intelligible and testable methodology. The analyzed Scaphopoda species came from the Brazilian coast and belong to the family Dentaliidae [(1) Coccodentalium carduus; (2) Paradentalium disparile] and Gadiliidae; [(3) Polyschides noronhensis, n. sp. from Fernando de Noronha Archipelago; (4) Gadila braziliensis]. These species represent the main branches of the class Scaphopoda. From protobranch bivalves, representatives of the families Solemyidae [(5) Solemya occidentalis, from Florida; S. notialis, n. sp. from S.E. Brazil], Nuculanidae [(6) Propeleda carpentieri from Florida], and Nuculidae [(7) Ennucula puelcha, from south Brazil] are included. These species represent the main branches of the basal Bivalvia. The descriptions on the anatomy of S. occidentalis and of P. carpentieri are published elsewhere. The remaining are included here, for which a complete taxonomical treatment is performed. Beyond these species, representatives of other taxa are operationally included as part of the ingroup (indices are then shared with them), as a procedure to test the morphological monophyly of Diasoma. These taxa are: two lamellibranch bivalves [(8) Barbatia - Arcidae; (9) Serratina - Tellinidae; both published elsewhere;, and Propilidium (10) Patellogastropoda, and (11) Nautilus, basal Cephalopoda, based on basal taxa. The effective outgroups are (12) Neopilina (Monoplacophora) and (13) Hanleya (Polyplacophora). The phylogenetic analysis based on morphology revealed that the taxon Diasoma is supported by 14 synapomorphies, and is separated from Cyrtosoma (Gastropoda + Cephalopoda). Although they are not the main goal of this paper, the taxa Scaphopoda and Bivalvia are supported by 8 and by 7 synapomorphies respectively. The taxon Protobranchia resulted paraphyletic. Both scaphopod orders resulted monophyletic. The obtained cladogram is: ((((Coccodentalium carduus - Paradentalium disparile) (Polyschides noronhensis - Gadila brasiliensis)) ((Solemya occidentalis - S. notialis) (Propeleda carpenteri (Ennucula puelcha (Barbatia cancellaria - Serratina capsoides))))) (Propilidium curumim - Nautilus pompilius - Lolliguncula brevis)).