Bone demineralization with citric acid enhances adhesion and spreading of preosteoblasts.

BACKGROUND: Previous studies have demonstrated that bone demineralization can improve consolidation in bone grafts. The biologic mechanisms underlying this phenomenon remain unclear. METHODS: Twelve adult male guinea pigs were used in this experiment. Forty-five bone samples removed from the calvaria of nine animals were divided in groups (n = 9) according to the time of demineralization with citric acid (50%, pH 1): 15, 30, 90, and 180 seconds and non-demineralized samples (control). Preosteoblasts (MC3T3-E1) were cultured on the bone samples for 24, 48, and 72 hours (n = 3). Fifteen samples removed from the remaining three animals were analyzed by scanning electron microscopy/energy dispersive spectrometry (SEM/EDS) after demineralization (n = 3). RESULTS: The number of preosteoblasts increased significantly with time in all groups. The bone surface area covered by these cells increased with time, except in the control group. Intragroup differences occurred between 24 and 72 hours (P < 0.05). Samples demineralized for 30 seconds showed greater area covered by preosteoblast cells than for the other times of demineralization in all periods of cell culture (P < 0.05) without a statistically significant difference compared with 15 seconds. SEM/EDS showed diminished content of calcium (Ca) after 15 seconds of demineralization, but the Ca content increased after 180 seconds of demineralization (P < 0.05). The phosphorus (P) amount increased significantly only after 30 seconds of demineralization (P < 0.5). The sulfur (S) content was increased in demineralized samples in relation to non-demineralized ones, reaching the highest level after 90 seconds, when the difference became significant in relation to all the other times of demineralization (P < 0.05). Magnesium (Mg) content did not differ significantly between demineralized and non-demineralized samples. CONCLUSIONS: Bone surfaces demineralized for 30 seconds increased the spreading of preosteoblasts as well as the surface area covered by these cells. Bone demineralization deserves to be studied in periodontal and maxillofacial regenerative procedures.

Laser therapy as an effective method for implant surface decontamination: a histomorphometric study in rats.

BACKGROUND: To the best of the authors' knowledge, a standard protocol for treating peri-implantitis is not yet established. METHODS: A total of 150 titanium disks with smooth or rough surfaces contaminated with microbial biofilm were implanted subcutaneously in rats after undergoing one of three treatments: 1) low-intensity laser (LIL); 2) antimicrobial photodynamic therapy (aPDT); or 3) toluidine blue O (TBO). Sterile and contaminated disks served as negative (NC) and positive (C) control groups, respectively. After days 7, 28, and 84, tissue inflammation was evaluated microscopically by measuring the density of collagen fibers (degree of fibrosis) and concentration of polymorphonuclear neutrophils. RESULTS: Surface texture did not affect the degree of inflammation, but the area of reactive tissue was significantly greater for rough implants (2.6 ± 3.7 × 10(6) µm(2)) than for smooth ones (1.9 ± 2.6 × 10(6) µm(2); P = 0.0377). Group C presented the lowest and group NC presented the highest degree of fibrosis with significance only after day 7; these groups had the highest and lowest scores, respectively, for degree of inflammation. Group C showed the largest area of reactive tissue (9.11 ± 2.10 × 10(6) µm(2)), but it was not significantly larger than group LIL (P = 0.3031) and group TBO (P = 0.1333). Group aPDT showed the smallest area (4.34 ± 1.49 × 10(6) µm(2)) of reactive tissue among the treatment groups. After day 28, groups LIL, aPDT, TBO, and C resembled group NC in all the studied parameters. CONCLUSION: Group aPDT showed more favorable results in parameter area of reactive tissue than the other methods after day 7, but over longer time periods all methods produced outcomes equivalent to sterile implants.

Fluoride modulates preosteoblasts viability and matrix metalloproteinases-2 and -9 activities.

This study evaluated the influence of fluoride on cell viability and activity of matrix metalloproteinases (MMP) -2 and -9 secreted by preosteoblasts. Preosteoblasts (MC3T3-E1 murine cell line) were cultured in MEM medium supplement with 10% Fetal Bovine Serum (FBS) and nucleosides/ribonucleosides without ascorbic acid. Adherent cells were treated with different concentrations of F (as sodium fluoride-NaF) in medium (5 x 10(-6) M, 10(-5) M, 10(-4) M and 10(-3) M) for 24, 48, 72 and 96 h at 37ºC, 5% CO(2). Control cells were cultivated in MEM only. After each period, preosteoblast viability was assessed by MTT assay. MMP-2 and -9 activities were performed by gel zymography. Also, alkaline phosphatase (ALP) activity was quantified by colorimetry in all experimental groups. It was shown that cultured cells with the highest dose of F (10(-3) M) for 96 h decreased preosteoblast viability while lower doses of F did not alter it, when compared to untreated cells. No differences were observed in ALP activity among groups. Moreover, compared to control, the treatment of cells with F at low dose slightly increased MMP-2 and -9 activities after 24 h. It was concluded that F modulates preosteoblast viability in a dose-dependent manner and also may regulate extracellular matrix remodeling.

Fluoride modulates preosteoblasts viability and matrix metalloproteinases-2 and -9 activities

This study evaluated the influence of fluoride on cell viability and activity of matrix metalloproteinases (MMP) -2 and -9 secreted by preosteoblasts. Preosteoblasts (MC3T3-E1 murine cell line) were cultured in MEM medium supplement with 10% Fetal Bovine Serum (FBS) and nucleosides/ribonucleosides without ascorbic acid. Adherent cells were treated with different concentrations of F (as sodium fluoride-NaF) in medium (5 x 10-6 M, 10-5 M, 10-4 M and 10-3 M) for 24, 48, 72 and 96 h at 37ºC, 5% CO2. Control cells were cultivated in MEM only. After each period, preosteoblast viability was assessed by MTT assay. MMP-2 and -9 activities were performed by gel zymography. Also, alkaline phosphatase (ALP) activity was quantified by colorimetry in all experimental groups. It was shown that cultured cells with the highest dose of F (10-3 M) for 96 h decreased preosteoblast viability while lower doses of F did not alter it, when compared to untreated cells. No differences were observed in ALP activity among groups. Moreover, compared to control, the treatment of cells with F at low dose slightly increased MMP-2 and -9 activities after 24 h. It was concluded that F modulates preosteoblast viability in a dose-dependent manner and also may regulate extracellular matrix remodeling.

Neste estudo, buscou-se avaliar a influência do fluoreto (F) na viabilidade celular e atividade das metaloproteinases de matriz (MMP) -2 e -9 secretado pelos pré-osteoblastos. Pré-osteoblastos (linhagem celular MC3T3-E1 murina) foram cultivados em meio MEM suplementado com 10% de soro fetal bovino (FBS) e nucleosídeos/ribonucleosídeos sem ácido ascórbico. Células aderidas foram tratadas com diferentes concentrações de F (na forma de fluoreto de sódio-NaF) em meio (5 x 10-6 M, 10-5 M, 10-4 M e 10-3 M) por 24, 48, 72 e 96 h a 37ºC, 5 % de CO2. Células do grupo controle foram cultivadas apenas em MEM. Após cada período, a viabilidade dos pré-osteoblastos foi avaliada por MTT. A atividade das MMP-2 e -9 foram analisadas pela zimografia. Além disso, a atividade da fosfatase alcalina (FA) foi quantificada por colorimetria em todos os grupos experimentais. Foi demonstrado que as células cultivadas com a maior dose de F (10-3 M) no período de 96 h tiveram sua viabilidade comprometida, enquanto doses mais baixas de F não a alteraram significativamente, quando comparado com células não tratadas. Não foi observada diferença na atividade da FA entre os grupos. Além disso, o tratamento de células com F em baixas doses, comparado ao grupo controle, promoveu um pequeno aumento da atividade das MMP-2 e -9 após 24 h. Pode-se concluir que o F modula a viabilidade de pré-osteoblastos de uma maneira dose-dependente e também pode regular a remodelação da matriz extracelular.

Histomorphometric study of the healing of human oral mucosa after gingivoplasty and low-level laser therapy.

BACKGROUND AND OBJECTIVES: The aim of the present study was to analyze the effects of diode laser irradiation on the healing of human oral mucosa. MATERIALS AND METHODS: After gingivoplasty, the right hemi-arch (test group) of 16 patients was irradiated with a diode laser. The left side (control group) was not irradiated. Incisional biopsies were performed on both sides at 7, 14, 21, and 60 days after surgery and morphometrically analyzed by light microscopy. RESULTS: Epithelium width ranged from 260.6 to 393.5 microm. Volume densities of basal (20.2%), prickle cell (55.6%), and cornified (24.2%) layers remained stable. The peak number of neutrophils were 6 cells/mm(2) and the mononuclear cells were 44 cells/mm(2). Collagen fibers (80%) and fibroblasts (14%) occupied the main volume of connective tissue. The one-way ANOVA and the paired Student's t-test were used for statistical analysis (P < 0.05). CONCLUSION: Low-level laser therapy did not accelerate the healing of oral mucosa after gingivoplasty.