Red, infrared, and simultaneous laser-wavelengths irradiation effects on 5-fluorouracil-induced oral mucositis in hamsters.

The association of more than one wavelength for photobiomodulation therapy (PBMT) to treat oral mucositis (OM) is unusual in the literature. Thus, this study aims to compare the simultaneous irradiation effects with their isolated application to treat OM. In order of that, 48 male Syrian hamsters were divided into 4 groups: Chemotherapy (Ch), which received only a OM induction protocol (5-fluorouracil chemotherapy and superficial oral mucosa scratches); red laser (RL), which received the OM induction and a PBMT protocol at 660 nm; infrared laser (IRL), which received the OM induction, and a PBMT protocol at 808 nm; and the RL + IRL group, which received the simultaneous application, of 660 and 808 nm wavelengths. Clinical (OM grade classification), histological (light microscopy analysis with H&E and collagen staining), immunohistochemical (TNF-α expression), and biochemical (TNF-α and hydroxyproline concentration) analyzes were performed after 7 and 10 days. Mainly on the 10th day, the RL and IRL groups showed lower OM grades and faster microscopic repair process, with greater expression of collagen fibers and lower TNF-α levels, besides the higher hydroxyproline concentrations, mainly in comparison with the Ch group. In conclusion, in this study, the simultaneous protocol did not present superior results than the isolated irradiations.

The Biochemical Mechanisms of Antimicrobial Photodynamic Therapy†.

The unbridled dissemination of multidrug-resistant pathogens is a major threat to global health and urgently demands novel therapeutic alternatives. Antimicrobial photodynamic therapy (aPDT) has been developed as a promising approach to treat localized infections regardless of drug resistance profile or taxonomy. Even though this technique has been known for more than a century, discussions and speculations regarding the biochemical mechanisms of microbial inactivation have never reached a consensus on what is the primary cause of cell death. Since photochemically generated oxidants promote ubiquitous reactions with various biomolecules, researchers simply assumed that all cellular structures are equally damaged. In this study, biochemical, molecular, biological and advanced microscopy techniques were employed to investigate whether protein, membrane or DNA damage correlates better with dose-dependent microbial inactivation kinetics. We showed that although mild membrane permeabilization and late DNA damage occur, no correlation with inactivation kinetics was found. On the other hand, protein degradation was analyzed by three different methods and showed a dose-dependent trend that matches microbial inactivation kinetics. Our results provide a deeper mechanistic understanding of aPDT that can guide the scientific community toward the development of optimized photosensitizing drugs and also rationally propose synergistic combinations with antimicrobial chemotherapy.

Inhibition of osteoclastogenesis after bisphosphonate therapy discontinuation: an in vitro approach.

Osteoclasts are specialized cells that degrade and resorb bone. Bisphosphonates (BPs) are drugs with well-known capacity to inhibit the resorption of mineralized tissues. Nitrogen-containing BPs, like alendronate (ALN) and zoledronic acid (ZA), inactivate osteoclast activity mostly by alterations on the cytoskeleton architecture of the cell. In this study, we used an in vitro model to test the hypothesis that bisphosphonates may have inhibitory effects on the osteoclastogenesis and osteoclast activity after the therapy was discontinued. Primary osteoclasts were generated from mouse bone marrow in media supplemented with 1,25-dihydroxyvitamin D3 and cultivated over bones pre-treated with ALN and ZA. The pre-saturation of the bone slices with bisphosphonates did not affect cell viability. We found, however, that by disrupting the gene expression of RANKL and OPG the osteoclastogenesis and resorption activity of osteoclasts was significantly disturbed. These inhibitory effects were confirmed by scanning electron microscopy resorption assay, assessment of osteoclast ultrastructure, and by gene expression analysis of TRAP and Cathepsin K. In conclusion, ALN and ZA adhered to the bone matrix reduced the osteoclast activity in vitro.

Effects of Alendronate and Dexamethasone on Osteoclast Gene Expression and Bone Resorption in Mouse Marrow Cultures.

Osteoclasts are cells whose main function is the resorption of bone matrix. However, several factors, including medications, can interfere with the resorption process. Alendronate (ALN), a nitrogen-containing type of bisphosphonate, and dexamethasone (DEX), a glucocorticoid, are drugs that may affect the resorption activity. The aim of this study is to investigate the effects of ALN, and/or DEX on osteoclast gene expression and resorption activity in primary mouse marrow cultures stimulated with 1,25-dihydroxyvitamin D3, a model for the bone microenvironment. Cultures were treated only with ALN (10-5 M), DEX (10-6 M), and with a combination of both agents. Viability assays performed at days 5, 7, and 9 showed the highest number of viable cells at day 7. All the following assays were then performed at day 7 of cell culture: tartrate resistant acid phosphatase (TRAP) histochemistry, receptor activator of nuclear factor kappa B ligand (RANKL) immunofluorescence, osteoprotegerin (OPG), and RANKL gene expression by qPCR and resorption analysis by scanning electron microscopy. Treatment with ALN, DEX, and the combination of both did not promote significant changes in the number of TRAP+ cells, although larger giant cells were detected in groups treated with DEX. DEX treatment increased the gene expression of RANKL and reduced OPG. The treatment with ALN reduced the depth of the resorption pits, but their inhibitory effect was less effective when administered with DEX.

Wound healing process with different photobiomodulation therapy protocols to treat 5-FU-induced oral mucositis in hamsters.

OBJECTIVES: The aim of this study was to analyze the effects of three different Photobiomodulation Therapy (PBMT) protocols in the treatment of 5-fluorouracil-induced oral mucositis in hamsters. DESIGN: 60 hamsters were divided into five groups: group "C", which did not receive oral mucosa scratching, 5-fluorouracil (5-FU) or PBMT; group "Ch", which received anesthesia, superficial oral mucosa scratching and 5-FU (oral mucositis induction); and three groups that received oral mucositis induction and a PBMT protocol: groups ChLI, ChLII and ChLIII that received 0.24 J (one point), 1 J (one point) and 1.2 J (five points of 0.24 J) of energy, respectively. The laser equipment used had λ = 660 nm and 0.04 cm2 of spot area (0.226 cm diameter). The animals were euthanized on days 7 and 10 of the experiment, and their oral mucosas were removed for histological (light microscopy and collagen staining), immunohistochemical (NF-kB and TNF-α), and biochemical (TNF-α, NF-kB and hydroxyproline) analysis. RESULTS: Group ChLI (less energy), showed the most accelerated repair rates and a lower concentration of inflammatory biomarkers than group Ch. Comparing the three PBMT protocols for treatment of 5-FU-induced oral mucositis in hamsters, the one with low energy (0.24 J) showed better results, regarding reduction of inflammatory biomarkers and tissue repair, than the ones with higher energy (1 and 1.2 J).

Transmission Electron Microscopy (TEM) and Scanning Electron Microscopy (SEM) for the Examination of Dental Hard Tissues.

This chapter describes laboratory protocols for TEM and SEM approaches allowing the examination of the dental hard tissues' constituents at the ultrastructural level. TEM has the highest resolution power to examine the cellular and extracellular matrix ultrastructure inside a given sample, detecting the presence, location, and quantification of organelles related to the metabolism of the cell type as well as membrane specializations. SEM allows the observation of the sample surface, for examining dimensional topography and distribution of exposed features.

Ion-releasing dental restorative composites containing functionalized brushite nanoparticles for improved mechanical strength.

OBJECTIVES: This study describes the synthesis of brushite nanoparticles (CaHPO4·2H2O) functionalized with triethylene glycol dimethacrylate (TEGDMA) and their application in dental restorative composites with remineralizing capabilities. METHODS: Nanoparticles were synthesized, with TEGDMA being added to one of the precursor solutions at three different molar ratios (0:1, 0.5:1 and 1:1, in relation to the ammonium phosphate precursor). Then, they were added (10 vol%) to a photocurable dimethacrylate matrix containing 50 vol% of reinforcing glass particles. The resulting composites were tested for degree of conversion, biaxial flexural strength and elastic modulus (after 24h and 28days in water), and ion release (over a 28-day period). Commercial composites (one microhybrid and one microfilled) were tested as controls. RESULTS: The final TEGDMA content in the functionalizing layer was modulated by the molar ratio added to the precursor solution. Functionalization reduced nanoparticle size, but did not reduce agglomeration. Improved mechanical properties were found for the composite containing nanoparticles with higher TEGDMA level in comparison to the composite containing non-functionalized nanoparticles or those with a low TEGDMA level. All brushite composites presented statistically significant reductions in strength after 28 days in water, but only the material with high-TEGDMA nanoparticles retained strength similar to the microhybrid commercial control. Overall, ion release was not affected by functionalization and presented steady levels for 28 days. SIGNIFICANCE: Though agglomeration was not reduced by functionalization, the improvement in the matrix-nanoparticle interface allowed for a stronger material, without compromising its remineralizing potential.

Low-power laser irradiation decreases lipid droplet accumulation in the parotid glands of diabetic rats.

Lipid droplet accumulation has been related to salivary gland hypofunction in diabetes. In this study, the effect of laser irradiation on the parotid glands (PGs) of diabetic rats was analyzed with regard to its effect on lipid droplet accumulation, intracellular calcium concentration and calmodulin expression. The animals were distributed into 6 groups: D0, D5, D20 and C0, C5, C20, for diabetic (D) and control animals (C), respectively. Twenty-nine days following diabetes induction, PGs of groups D5 and C5; D20 and C20 were irradiated with 5 and 20 J/cm2 of a red diode laser at 100 mW, respectively. After 24 hours, PGs were removed for histological, biochemical, and western blotting analysis. The diabetic animals showed lipid droplet accumulation, which was decreased after irradiation. Ultrastructurally, the droplets were nonmembrane bound and appeared irregularly located in the cytoplasm. Moreover, diabetic animals showed an increased intracellular calcium concentration. In contrast, after laser irradiation a progressive decrease in the concentration of this ion was observed, which would be in agreement with the results found in the increased expression of calmodulin in D20. These data are promising for using laser to decrease lipid droplet accumulation in PGs, however, more studies are necessary to better understand its mechanisms. Micrographs showing decreased lipid accumulation after laser irradiation in light micrographs (LM), and morphology of lipid droplet in transmission electron microscopic (TEM). LM: (A) PGs from nondiabetic rats that did not receive Laser irradiation (LI), (B) PGs from nondiabetic rats that received a dose of 20 J/cm2 , (C) lipid accumulation (arrows) in the secretory cells from diabetic rats that did not receive irradiation, (D) reduction of lipid accumulation in the secretory cells from diabetic rats that received a dose of 20 J/cm2 and TEM: (E) scale bar = 5 µm, (F) scale bar = 1 µm, and (G) scale bar = 0.5 µm.

Avaliação histológica e histomorfométrica da regeneração óssea a partir da utilização de biomateriais em tíbias de ovinos / Histological and histomorphometric evaluation of bone regeneration with biomaterial in the tibia of sheep

Falhas ósseas são comumente encontradas na medicina veterinária em diferentes enfermidades, principalmente por traumas em grandes animais. Frequentemente faz-se necessário auxiliar este reparo tecidual para melhor correção desta falha. Os biomateriais foram desenvolvidos para cumprir esta função. A fim de avaliar o comportamento celular perante estes substitutos ósseos, a realização de testes in vivo é fundamental. A partir da observação da unidade morfofuncional do tecido é possível avaliar a quantidade e a qualidade do tecido neoformado. Atualmente, em estudos ortopédicos, os ovinos são considerados animais de escolha como modelo experimental. Falhas ósseas foram realizadas experimentalmente em ovinos, onde foram implantados biomateriais à base de quitosana, hidroxiapatita e colágeno. Após 60 dias foi realizada biopsia no local do reparo tecidual e por meio da histomorfometria da matriz orgânica, da análise descritiva da microscopia de luz e da microscopia eletrônica de transmissão, o tecido neoformado foi avaliado e comparado. Na histomorfometria, não houve diferença significativa quanto à quantidade de tecido neorformado do grupo controle e do grupo com biomaterial, porém quando comparado ao osso pré-existente, retirado no momento da confecção da falha, a área de matriz óssea foi menor. Na descrição da microscopia de luz e em maiores detalhes na análise ultra estrutural, observou-se o biomaterial em contato íntimo com o tecido neoformado, sugerindo boa biocompatibilidade. Foi possível observar o padrão da reparação tecidual, onde o membro controle e o membro com biomaterial não diferiram quanto à arquitetura tecidual, porém nas amostras com o implante do biomaterial foi identificada a presença do mesmo. Todas estas informações sugerem que o biomaterial não prejudicou a regeneração óssea, apresentou boa biocompatibilidade e denota potencial para auxiliar na rotina ortopédica de grandes animais.(AU)

Bone defects are commonly found in veterinary medicine through different disease such as trauma, especially in large animals. Frequently it is necessary to assist this tissue repair for better regeneration. Biomaterials in general are designed to this function. In order to understand the bone cells in the presence of bone substitutes, tests in vivo are essential for these studies. The quantity and quality of new tissue formation can be assessed by observation of tissue morphofunctional unit. Currently, in orthopedic studies sheep are considerated animals of choice for experimental development. Bone defects were performed experimentally in sheep and were implanted chitosan, hydroxyapatite and collagen biomaterials. After 60 days, biopsy was performed at the site of tissue repair and the new tissue formation was described, evaluated and compared by organic matrix bone histomorphometry and electronic transmission microscopy. After hystomorfometry analysis there was no difference between control group and biomaterial group, but when compare with pre-existent tissue, the values are smaller. In descriptive analysis was observed intimate contact with biomaterial and new tissue formation suggesting biocompatibility. It observed the pattern of tissue repair in control group and biomaterial group did not differ in the architecture tissue. However, in the biomaterial group was identified presence of the biomaterial in the specimen. All these data suggest that the biomaterial did not impair bone regeneration, had good biocompatibility and potential to assistant in orthopedic routine of large animals.(AU)

Avaliação histológica e histomorfométrica da regeneração óssea a partir da utilização de biomateriais em tíbias de ovinos / Histological and histomorphometric evaluation of bone regeneration with biomaterial in the tibia of sheep

Falhas ósseas são comumente encontradas na medicina veterinária em diferentes enfermidades, principalmente por traumas em grandes animais. Frequentemente faz-se necessário auxiliar este reparo tecidual para melhor correção desta falha. Os biomateriais foram desenvolvidos para cumprir esta função. A fim de avaliar o comportamento celular perante estes substitutos ósseos, a realização de testes in vivo é fundamental. A partir da observação da unidade morfofuncional do tecido é possível avaliar a quantidade e a qualidade do tecido neoformado. Atualmente, em estudos ortopédicos, os ovinos são considerados animais de escolha como modelo experimental. Falhas ósseas foram realizadas experimentalmente em ovinos, onde foram implantados biomateriais à base de quitosana, hidroxiapatita e colágeno. Após 60 dias foi realizada biopsia no local do reparo tecidual e por meio da histomorfometria da matriz orgânica, da análise descritiva da microscopia de luz e da microscopia eletrônica de transmissão, o tecido neoformado foi avaliado e comparado. Na histomorfometria, não houve diferença significativa quanto à quantidade de tecido neorformado do grupo controle e do grupo com biomaterial, porém quando comparado ao osso pré-existente, retirado no momento da confecção da falha, a área de matriz óssea foi menor. Na descrição da microscopia de luz e em maiores detalhes na análise ultra estrutural, observou-se o biomaterial em contato íntimo com o tecido neoformado, sugerindo boa biocompatibilidade. Foi possível observar o padrão da reparação tecidual, onde o membro controle e o membro com biomaterial não diferiram quanto à arquitetura tecidual, porém nas amostras com o implante do biomaterial foi identificada a presença do mesmo. Todas estas informações sugerem que o biomaterial não prejudicou a regeneração óssea, apresentou boa biocompatibilidade e denota potencial para auxiliar na rotina ortopédica de grandes animais.(AU)

Bone defects are commonly found in veterinary medicine through different disease such as trauma, especially in large animals. Frequently it is necessary to assist this tissue repair for better regeneration. Biomaterials in general are designed to this function. In order to understand the bone cells in the presence of bone substitutes, tests in vivo are essential for these studies. The quantity and quality of new tissue formation can be assessed by observation of tissue morphofunctional unit. Currently, in orthopedic studies sheep are considerated animals of choice for experimental development. Bone defects were performed experimentally in sheep and were implanted chitosan, hydroxyapatite and collagen biomaterials. After 60 days, biopsy was performed at the site of tissue repair and the new tissue formation was described, evaluated and compared by organic matrix bone histomorphometry and electronic transmission microscopy. After hystomorfometry analysis there was no difference between control group and biomaterial group, but when compare with pre-existent tissue, the values are smaller. In descriptive analysis was observed intimate contact with biomaterial and new tissue formation suggesting biocompatibility. It observed the pattern of tissue repair in control group and biomaterial group did not differ in the architecture tissue. However, in the biomaterial group was identified presence of the biomaterial in the specimen. All these data suggest that the biomaterial did not impair bone regeneration, had good biocompatibility and potential to assistant in orthopedic routine of large animals.(AU)

Synthesis and characterization of silver phosphate/calcium phosphate mixed particles capable of silver nanoparticle formation by photoreduction.

Silver phosphate is a semi-conductor sensitive to UV-Vis radiation (<530nm). Exposure to radiation removes electrons from the oxygen valence shell, which are scavenged by silver cations (Ag+), forming metallic silver (Ag0) nanoparticles. The possibility of silver nanoparticle formation in situ by a photoreduction process was the basis for the application of mixed calcium phosphate/silver phosphate particles as remineralizing and antibacterial fillers in resin-based dental materials. Mixed phosphate particles were synthesized, characterized and added to a dimethacrylate resin in 20% or 30% mass fractions to investigate their efficacy as ion-releasing fillers for dental remineralization and antibacterial activity. The formation of metallic silver nanoparticles after exposure to visible radiation from a dental curing unit (peak emission: 470nm) was demonstrated by particle X-ray diffraction and scanning electron microscopy analysis of the composite fractured surface. Calcium and phosphate release from materials containing the mixed particles were similar to those containing pure CaP particles, whereas Streptococcus mutans colonies were reduced by three orders of magnitude in relation to the control, which can be attributed to silver release. As expected, the optical properties of the materials containing mixed phosphate particles were compromised by the presence of silver. Nevertheless, materials containing mixed phosphate particles presented higher fracture strength and elastic modulus than those with pure CaP particles.

Effects of bisphosphonates on osteogenesis and osteoclastogenesis signaling during the endochondral ossification of growing rats.

Osteoclasts and chondroclasts are necessary, during endochondral ossification, for the resorption of primary bone and calcified cartilage septa, respectively. The bisphosphonates inhibit mineralized tissue resorption by various mechanisms according to the different types of this drug, which can affect bone remodeling during skeletal growth. The objective of the present study is to analyze the way that alendronate (ALN) and etidronate (ETN) can affect osteoclastogenesis and bone formation during endochondral ossification of the long bones of growing rats. Newborn Wistar rats were treated daily with ETN, ALN, or sterile saline solution (control) for 21 days. Their femur and tibiae epiphyses were radiographed and analyzed by light, scanning and transmission electron microscopy. The expression of genes related to osteogenesis and to osteoclast differentiation and activity were analyzed by real-time quantitative polymerase chain reaction. The ETN group presented reduced body weight, disorganized growth plate and an extended area of cartilage in the ossification zone with little bone matrix; in the ALN group, this area was not altered. The ALN presented latent TRAP-positive cells, whereas in the ETN group, they were activated. The expression of NFκB1 and 2, OPG, Spp1 and Runx2 in the ossification zone was reduced by both bisphosphonates. RANKL expression was reduced by ETN, whereas ALN decreased the expression of RANK. The results also indicated that, in addition to the anti-resorptive effect of the drugs, disturbances in bone deposition occurred concomitantly with the reduced expression of osteogenesis-related genes.

Polymer-based material containing calcium phosphate particles functionalized with a dimethacrylate monomer for use in restorative dentistry.

Dicalcium phosphate dihydrate particles functionalized with triethyleneglycol dimethacrylate were synthesized and added to a photocurable mixture of bisphenol-A glycidyl dimethacrylate and triethyleneglycol dimethacrylate with the purpose of developing a resin composite capable of releasing calcium and phosphate ions to foster dental remineralization. Particle functionalization would minimize the deleterious effect of adding low cohesive strength nano-structured particles with no chemical interaction with the organic matrix on the material's mechanical properties. The results showed that calcium release over 28 days was not impaired by particle functionalization. A statistically significant 32% increase in strength was recorded with the use of functionalized dicalcium phosphate dihydrate in comparison to the material containing non-functionalized particles. However, the strength of the unfilled resin was not matched by the composite with functionalized particles. Elastic modulus increased with particle incorporation, regardless of functionalization. Degree of conversion and optical properties (total transmittance and color change/ΔE) of the resin-based materials were not affected by the addition of dicalcium phosphate dihydrate particles (functionalized or not).

Bioactive composites containing TEGDMA-functionalized calcium phosphate particles: Degree of conversion, fracture strength and ion release evaluation.

OBJECTIVE: To evaluate the strength and ion release of experimental composites containing TEGDMA-functionalized calcium phosphate particles. METHODS: Seven composites containing equal parts (in mols) of BisGMA and TEGDMA and 60vol% of fillers were manipulated. Filler phase was constituted by silanized barium glass and 0% (control), 10% or 20% (volume) of dicalcium phosphate dihydrate (DPCD) particles, either non-functionalized or functionalized with two different TEDGMA contents. DCPD particles were synthesized and characterized by X-ray diffraction (XRD), elemental analysis, surface area and dynamic light scattering. Composites were tested for degree of conversion (DC) by near-FTIR. Biaxial flexural strength (BFS) was determined after 24h and 28days in water. Calcium and phosphate release after 7days was assessed using inductively coupled plasma optical emission spectrometry (ICP-OES). Data were analyzed by ANOVA/Tukey test (alpha:5%). RESULTS: XRD confirmed the crystalline structure corresponding to DCPD. Elemental analysis revealed particles with zero, 14% or 22% TEGDMA, with similar D50 (around 19µm) and surface areas from 3.5 to 11.4m2/g. The presence of DCPD did not reduce DC. After 24h, functionalization (both 14% and 22% TEGDMA) improved composite strength in comparison to non-functionalized DCPD, both at 10% and 20% levels. After 28days, BFS of materials containing 10% functionalized DCPD were statistically similar to the control containing only barium glass. Among composites containing 10% DCPD, particle functionalization with 14% TEGDMA did not jeopardize ion release. SIGNIFICANCE: At 10vol%, the use of TEGDMA-functionalized CaP particles improved composite strength in relation to non-functionalized particles, while maintaining similar ion release levels.

Ureaplasma diversum Genome Provides New Insights about the Interaction of the Surface Molecules of This Bacterium with the Host.

Whole genome sequencing and analyses of Ureaplasma diversum ATCC 49782 was undertaken as a step towards understanding U. diversum biology and pathogenicity. The complete genome showed 973,501 bp in a single circular chromosome, with 28.2% of G+C content. A total of 782 coding DNA sequences (CDSs), and 6 rRNA and 32 tRNA genes were predicted and annotated. The metabolic pathways are identical to other human ureaplasmas, including the production of ATP via hydrolysis of the urea. Genes related to pathogenicity, such as urease, phospholipase, hemolysin, and a Mycoplasma Ig binding protein (MIB)-Mycoplasma Ig protease (MIP) system were identified. More interestingly, a large number of genes (n = 40) encoding surface molecules were annotated in the genome (lipoproteins, multiple-banded antigen like protein, membrane nuclease lipoprotein and variable surface antigens lipoprotein). In addition, a gene encoding glycosyltransferase was also found. This enzyme has been associated with the production of capsule in mycoplasmas and ureaplasma. We then sought to detect the presence of a capsule in this organism. A polysaccharide capsule from 11 to 17 nm of U. diversum was observed trough electron microscopy and using specific dyes. This structure contained arabinose, xylose, mannose, galactose and glucose. In order to understand the inflammatory response against these surface molecules, we evaluated the response of murine macrophages J774 against viable and non-viable U. diversum. As with viable bacteria, non-viable bacteria were capable of promoting a significant inflammatory response by activation of Toll like receptor 2 (TLR2), indicating that surface molecules are important for the activation of inflammatory response. Furthermore, a cascade of genes related to the inflammasome pathway of macrophages was also up-regulated during infection with viable organisms when compared to non-infected cells. In conclusion, U. diversum has a typical ureaplasma genome and metabolism, and its surface molecules, including the identified capsular material, represent major components of the organism immunopathogenesis.

Effects of TGF-ß1 on mineralization mediated by rat calvaria-derived osteogenic cells.

In this study, we have analyzed the viability and cell growth, as well as, the mineralization of extracellular matrix (ECM) by alizarin red and von Kossa staining of calvaria-derived osteogenic cultures, treated with TGF-ß1 alone or associated with Dex comparing with acid ascorbic (AA) + ß-glicerophosphate (ßGP) (positive mineralization control). The expression of the noncollagenous proteins bone sialoprotein (BSP), osteopontin (OPN) and fibronectin (FN) were evaluated by indirect immunofluorescence. In addition, the main ultrastructural morphological findings were assessed by transmission electron microscopy. Osteogenic cells were isolated of calvaria bone from newborn (2-day-old) Wistar rats were treated with TGF-ß1 alone or with dexamethasone for 7, 10, and 14 days. As positive mineralization control, the cells were supplemented only with AA+ ßGP. As negative control, the cells were cultured with basal medium (α-MEM + 10%FBS + 1%gentamicin). The treatment with TGF-ß1, even when combined with Dex, decreased the viability and cell growth when compared with the positive control. Osteoblastic cell cultures were positive to alizarin red and von Kossa stainings after AA + ßGP and Dex alone treatments. Positive immunoreaction was found for BSP, OPN and FN in all studied treatments. Otherwise, when the cell cultures were supplemented with TGF-ß1 and TGF-ß1 + Dex, no mineralization was observed in any of the studied periods. These present findings suggest that TGF-ß1, in the studied in vitro doses, inhibits the proliferation and differentiation of osteoblastic cells by impairment of nodule formation.

Comparative study among three different phototherapy protocols to treat chemotherapy-induced oral mucositis in hamsters.

In this study, clinical, biochemical and histological analysis were used to compare different phototherapies, including LED, low and high-power laser (HPL) for the treatment of chemotherapy (CT)-induced oral mucositis (OM). One-hundred-fifty hamsters were divided into five groups: C: control; CH: CT/OM induction; L: CT/OM induction and treatment with LED (635 nm, 1.2 J), HL: CT/OM induction and treatment with HPL (808 nm, 10 J), LL: CT/OM induction and treatment with low-level laser therapy (LLLT) (660 nm, 1.2 J). OM was induced by scratches performed on check pouch mucosa after two injections of 5-Fluorouracil. The experiment lasted 10 days and OM was analyzed by specific clinical scales on days 5, 7 and 10. The animals were euthanized and the cheek pouch mucosa removed for biochemical (TNF-α concentration) and histological (light microscopy) analysis. After statistical analysis, the authors' results showed LED and LLLT therapies were efficient treatments for OM, decreasing TNF-α concentration on day 7 (p < 0.05) and completely healing the mucosa on day 10. HPL showed no interference in final healing of OM. According to the methodology used and the results obtained in the present study, LLLT and LED therapies were the best choices to decrease the severity of OM, accelerating tissue repair and decreasing the inflammatory process. Clinical evaluation of OM in Groups CH, LL, L and HL and their respective arrangement of phototherapy treatments at different time intervals (5, 7 and 10 days).

Mechanical properties and ion release from bioactive restorative composites containing glass fillers and calcium phosphate nano-structured particles.

OBJECTIVE: To evaluate the effect of the replacement of barium glass by dicalcium phosphate dihydrate (DCPD) particles on the mechanical properties and degree of conversion (DC) of composites. Additionally, calcium and hydrogen phosphate (HPO4(2-)) release were followed for 28 days. METHODS: Nine composites containing equal parts (in mols) of BisGMA and TEGDMA and 40, 50 or 60 vol% of total filler were manipulated. Filler phase was constituted by silanated barium glass and 0%, 10% or 20% of DCPD particles. DC was determined by near-FTIR. Biaxial flexural strength (BFS) and modulus (E) were tested using the "piston on three balls" method, while fracture toughness (KIc) used the "single edge notched beam" method. Specimens were tested after 24h and 28 days in water. Ion release was determined using inductively coupled plasma optical emission spectrometry (ICP-OES). Data were analyzed by ANOVA/Tukey (DC and ion release) or Kruskal-Wallis/Mann-Whitney (mechanical properties; alpha: 5%). RESULTS: DC was not affected by DCPD. The presence of DCPD reduced BFS for both storage times, while differences in E became evident after 28 days. After 24h, KIc increased with the addition of DCPD; after 28 days, however, KIc decreased only for DCPD-containing composites. Calcium release was similar for both DCPD contents and remained fairly constant during the 28-day period. Overall, HPO4(2-) release was higher at 7 days and did not decrease after 14 days. SIGNIFICANCE: The composite with the highest filler level and 10% DCPD represented the best compromise between mechanical properties after aging in water and ion release.

Evaluation of bone repair of critical size defects treated with simvastatin-loaded poly(lactic-co-glycolic acid) microspheres in rat calvaria.

PURPOSE: Statins are hypolipemiant drugs with osteoinductive effect. We evaluated the potential of simvastatin loaded into poly(lactic-co-glycolic acid) (PLGA) microspheres to heal critical size defects in rat calvaria. METHODS: PLGA scaffolds (50:50 ratio) were synthesized as pure membranes or as microspheres loaded with 2.5% simvastatin. Critical size defects (5-mm diameter) were created in the parietal bone of 3-month-old male Wistar rats; they were either left filled with blood clot (C group), covered with a PLGA membrane (M group) or with PLGA microspheres loaded with simvastatin (MSI group) or not (MM group), and then covered with the PLGA membrane. The defects were evaluated after 30 or 60 days by light and electron microscopy, immunohistochemistry for osteopontin (OPN), bone sialoprotein (BSP) and osteoadherin (OSAD), and immunocytochemistry for OPN. RESULTS: Scanning electron microscopy showed that the calvarial defects treated with MSI were almost completely healed after 60 days, while groups M and C presented less bone formation, whereas the bone matrix formed into the defects of MSI group was more organized and mature. The immunolabeling for OPN and BSP on the matrix in groups C and M showed typical areas of primary bone unlike the MSI that presented weak labeling at the formed area. In the MSI group, there was an intense immunostaining for OSAD in osteoid, as well as in osteocyte cytoplasm. The immunocytochemistry showed intense labeling for OPN with homogeneous distribution in the interfibrillar spaces in all groups after 30 days and after 60 days; however, while C and M groups exhibited similar aspect, the MSI specimens showed weak labeling. The ultrastructural evaluation showed the interaction between the biomaterial and the surrounding tissue where some cells established intimate contact with microspheres. CONCLUSIONS: The repair of critical size bone defects was accelerated and enhanced by the implantation of simvastatin-loaded PLGA microspheres.

Histological and radiological analyses of a maxillary sinus lift with extensive drilling of the schneider membrane using xenogeneic bone.

The objective of this study is to report a clinical case of maxillary sinus with lyophilized, xenogeneic graft, in which, despite a large perforation of the sinus membrane, the surgery was not aborted and the results of histological examinations indicate bone neoformation in the surgical area. Results. This case showed that the biomaterials evaluated in this study and the procedure used to place them proved to be biocompatible and presented high osteogenic potential, leading to a successful surgery and osseointegration implant. Conclusion. Positioning Schneider's membrane and filling it with the graft biomaterial helped to achieve the desired osteoconduction and proliferation of bone cells even though the patient had a large perforation of the sinus membrane.