Pam2CSK4 (TLR2 agonist) induces periodontal destruction in mice.

Lipoproteins are important bacterial immunostimulating molecules capable of inducing receptor activator of nuclear factor-κB (RANKL) and osteoclast formation in vitro and in vivo . Although these molecules are present in periodontopathogenic bacteria, their role in periodontitis is not known. In this study, we used Pam2CSK4 (PAM2), a synthetic molecule that mimics bacterial lipoprotein, to investigate the effects of lipoproteins on periodontitis in mice. C57BL/6 male mice were randomly divided into three experimental groups: 1) Negative control group: animals received vehicle injection; 2) Positive control group: animals received injection of Escherichia coli lipopolysaccharide (LPS); 3) PAM2 group: animals received PAM2 injection. All the injections were performed bilaterally every other day into the palatal mucosa between first and second molars. After twenty-four days, the animals were euthanized to assess alveolar bone volume (micro-CT), cellular and extracellular composition in the gingiva (stereometric analysis), and osteoclast numbers (TRAP staining). Treatment with either PAM2 or LPS induced gingival inflammation, as demonstrated by increased infiltration of inflammatory cells and enhanced angiogenesis, associated with a smaller number of fibroblasts and decreased extracellular matrix. Importantly, treatment not only with LPS but also with PAM2 resulted in a larger number of TRAP+ multinucleated osteoclasts and significant loss of alveolar bone. Collectively, our data demonstrate that PAM2 can induce gingival inflammation and bone loss in mice, broadening the avenues of investigation into the role of lipoproteins in the pathogenesis of periodontal disease.

Pam2CSK4 (TLR2 agonist) induces periodontal destruction in mice

Abstract Lipoproteins are important bacterial immunostimulating molecules capable of inducing receptor activator of nuclear factor-κB (RANKL) and osteoclast formation in vitro and in vivo . Although these molecules are present in periodontopathogenic bacteria, their role in periodontitis is not known. In this study, we used Pam2CSK4 (PAM2), a synthetic molecule that mimics bacterial lipoprotein, to investigate the effects of lipoproteins on periodontitis in mice. C57BL/6 male mice were randomly divided into three experimental groups: 1) Negative control group: animals received vehicle injection; 2) Positive control group: animals received injection of Escherichia coli lipopolysaccharide (LPS); 3) PAM2 group: animals received PAM2 injection. All the injections were performed bilaterally every other day into the palatal mucosa between first and second molars. After twenty-four days, the animals were euthanized to assess alveolar bone volume (micro-CT), cellular and extracellular composition in the gingiva (stereometric analysis), and osteoclast numbers (TRAP staining). Treatment with either PAM2 or LPS induced gingival inflammation, as demonstrated by increased infiltration of inflammatory cells and enhanced angiogenesis, associated with a smaller number of fibroblasts and decreased extracellular matrix. Importantly, treatment not only with LPS but also with PAM2 resulted in a larger number of TRAP+ multinucleated osteoclasts and significant loss of alveolar bone. Collectively, our data demonstrate that PAM2 can induce gingival inflammation and bone loss in mice, broadening the avenues of investigation into the role of lipoproteins in the pathogenesis of periodontal disease.

Influence of obesity on experimental periodontitis in rats: histopathological, histometric and immunohistochemical study.

OBJECTIVES: This study assessed the influence of obesity on the progression of ligature-induced periodontitis in rats. MATERIALS AND METHODS: Forty-eight adult Wistar rats were randomly divided into two groups: the HL group (n = 24) was fed high-fat animal food to induce obesity, and the NL group (n = 24) was fed normolipidic animal food. Obesity was induced within a period of 120 days, and the induction of experimental periodontitis (EP) was subsequently performed for 30 days. The animals were euthanized after 7, 15, and 30 days, and the jaws were removed for histopathological, histometric, and immunohistochemical analyses. Tartrate-resistant acid phosphatase (TRAP), receptor activator of nuclear factor kappa beta ligand (RANKL), and osteoprotegerin (OPG) were analyzed via immunolabeling. RESULTS: Histological findings indicated that the inflammation was more extensive and lasted longer in the HL/EP; however, advanced destruction also occurred in the NL/EP. Greater bone loss was verified in the HL/EP group (2.28 ± 0.35) in the period of 7 days than in the NL/EP group (1.2 ± 0.29). High immunolabeling was identified in the HL/EP group in the initial periods for RANKL and TRAP, whereas the NL/EP group presented with moderate immunolabeling for both factors. The HL/EP and NL/EP groups showed low immunolabeling for OPG. CONCLUSIONS: Obesity induced by a high-fat diet influenced alveolar bone metabolism when associated with experimental periodontitis and caused a more severe local inflammatory response and alveolar bone loss. CLINICAL RELEVANCE: Obesity is related to greater alveolar bone loss and an accentuated local inflammatory response, which may be reflected in the clinical severity of periodontitis and dental loss.

Effect of a probiotic beverage consumption (Enterococcus faecium CRL 183 and Bifidobacterium longum ATCC 15707) in rats with chemically induced colitis.

BACKGROUND: Some probiotic strains have the potential to assist in relieving the symptoms of inflammatory bowel disease. The impact of daily ingestion of a soy-based product fermented by Enterococcus faecium CRL 183 and Lactobacillus helveticus 416 with the addition of Bifidobacterium longum ATCC 15707 on chemically induced colitis has been investigated thereof within a period of 30 days. METHODS: Colitis was induced by dextran sulfate sodium. The animals were randomly assigned into five groups: Group C: negative control; Group CL: positive control; Group CLF: DSS with the fermented product; Group CLP: DSS with the non-fermented product (placebo); Group CLS: DSS with sulfasalazine. The following parameters were monitored: disease activity index, fecal microbial analyses, gastrointestinal survival of probiotic microorganisms and short-chain fatty acids concentration in the feces. At the end of the protocol the animals' colons were removed so as to conduct a macroscopical and histopathological analysis, cytokines and nitrite quantification. RESULTS: Animals belonging to the CLF group showed fewer symptoms of colitis during the induction period and a lower degree of inflammation and ulceration in their colon compared to the CL, CLS and CLP groups (p<0.05). The colon of the animals in groups CL and CLS presented severe crypt damage, which was absent in CLF and CLP groups. A significant increase in the population of Lactobacillus spp. and Bifidobacterium spp. at the end of the protocol was verified only in the CLF animals (p<0.05). This group also showed an increase in short-chain fatty acids (propionate and acetate). Furthermore, the intestinal survival of E. faecium CRL 183 and B. longum ATCC 15707 in the CLF group has been confirmed by biochemical and molecular analyzes. CONCLUSIONS: The obtained results suggest that a regular intake of the probiotic product, and placebo to a lesser extent, can reduce the severity of DSS-induced colitis on rats.

Low-level laser therapy (670 nm) on viability of random skin flap in rats.

This study investigated the effects of 670 nm laser, at different fluences, on the viability of skin flap in rats. One hundred male animals were used. The animals were divided into control group; group treated with 3 J/cm(2); group treated with 6 J/cm(2); group treated with 12 J/cm(2) and group treated with 24 J/cm(2). The skin flap was made on the backs of all animals studied, with a plastic sheet interposed between the flap and the donor site. Laser irradiation was done immediately after the surgery and on days 1, 2, 3 and 4 after surgery. The percentage of necrosis of the flap was calculated at the 7th postoperative day. Additionally, a sample of each flap was collected to enable us to count the blood vessels. Treated animals showed a statistically significant smaller area of necrosis than did the control group. The necrosis in the treated groups was 41.82% (group 2), 36.51% (group 3), 29.45% (group 4) and 20.37% (group 5). We also demonstrated that laser irradiation at 670 nm, at all doses used, had a stimulatory effect on angiogenesis. Our study showed that the 670 nm laser was efficient to increase the viability of the skin flap, at all fluences used, with a tendency of reaching better results at higher doses.

Computer-assisted analysis of cell proliferation markers in oral lesions.

Abnormalities in any component of the cell cycle regulatory machine may result in oral cancer, and markers of cell proliferation have been used to determine the prognosis of tumor progression. The aim of this study was to determine whether silver-stained nucleolar organizer region (AgNOR) and Ki-67 measurements could improve the assessment of growth rates in oral lesions. Eighty-three oral biopsies were studied, 20 of which were classified as fibrous inflammatory hyperplasia (FIH), 40 as leukoplakia (LKP) and 23 as oral squamous cell carcinoma (OSCC). Within the LKP group, 22 out of 29 biopsies were diagnosed as non-dysplastic leukoplakia (LK) and 18 as dysplastic leukoplakia (DLK), presenting discrete, moderate and severe dysplasia. Ki-67 immunolabeling of the lesions increased steadily in the following order: FIH, DLK, LK and OSCC, indicating that Ki-67 is a good marker for predicting the proliferative fraction among benign, premalignant and malignant oral lesions. The median values of AgNOR parameters indicate that the morphometric index gives better results regarding the proliferative rate than the numerical one. A series of linear regressions between AgNOR parameters and Ki-67 showed positive associations. We conclude that a combination of Ki-67 and morphometric AgNOR analyses could be used as an aid in the determination of the proliferative status of oral epithelial cells in oral cancer.

Os efeitos de um programa de atividade física de carga progressiva nas propriedades físicas e na força óssea de ratas osteopênicas / The effects of a progressive loading exercise program on femoral physical properties and strength of osteopenic rats

Muitos estudos demonstram que programas de atividade física são eficazes para estimular o metabolismo ósseo, sendo utilizados como uma modalidade terapêutica em caso de perda óssea devido a osteoporose. Entretanto, vários pontos relacionados a intensidade e freqüência ideal para do exercício físico ainda não estão esclarecidos. Com isso, o objetivo deste estudo foi determinar os efeitos de um programa de exercícios físicos de alta intensidade na morfometria, na força óssea e no conteúdo mineral do fêmur de ratas osteopênicas. Foram utilizadas 40 ratas, distribuídas em 4 grupos: grupo intacto sedentário (SS); grupo osteopênico sedentário (OS); grupo intacto treinamento (ST) e grupo osteopênico treinamento (OT). O programa de exercício foi iniciado 8 semanas após a ovariectomia e foi realizado 3 vezes por semana, durante 8 semanas. Cada sessão foi constituída por 4 séries de 10 saltos cada. Após a eutanásia dos animais, os fêmures foram retirados e processados para as analises. RESULTADOS: os animais osteopênicos sedentários demonstraram uma diminuição da força óssea e uma diminuição dos pesos ósseos, da densidade óssea e do conteúdo de cálcio. As ratas osteopênicas exercitadas mostraram maiores valores na avaliação dos pesos ósseos, da força óssea, da densidade óssea e do conteúdo mineral, evidenciando o efeito positivo deste protocolo no metabolismo ósseo. Os resultados deste estudo indicam que a intensidade e a duração do programa de exercícios utilizado foi eficaz para produzir modificações nas propriedades geométricas e na forca óssea, nos fêmures das ratas osteopênicas, o que contribuiu para reverter as perdas ósseas após a ovariectomia.

BACKGROUND: Many studies have shown that physical exercises are able to stimulate bone formation and increase bone mass, constituting a therapeutic modality for the treatment of bone loss due to osteoporosis. However, some points about the intensity, duration and frequency of the exercises remain confusing and contradictory. Thus, the aim of this study was to determine the effects of a progressive loading exercise program on femur of osteopenic rats. To induce osteopenia, we used the animal model of ovariectomy (OVX). Forty animals were studied and divided into 4 groups: sham-operated sedentary (SS); ovariectomy-sedentary (OS); sham-operated training (ST) and ovariectomy training (OT). The trained groups performed jumps into water: 4 series of 10 jumps each, with an overload of 50 percent to 80 percent of the animal's body weight, during 8 weeks. Femora were submitted to an evaluation of physical properties, a biomechanical test, calcium and phosphorus content measurement and a morphometric histological evaluation. RESULTS: osteopenic animals showed a decrease of bone strength and lower values of bone weights, bone density and calcium content. The exercised osteopenic rats showed higher values of geometrical, physical properties, bone strength and calcium content compared to controls. The results of the present study indicate that the progressive loading exercise program has stimulatory effects on osteopenic rats' femora. It seems that the intensity and duration of the used protocol produced bone structural adaptations, which contributed to revert bone loss due to ovariectomy.

Análise da superfície de esmalte de dentes decíduos submetidos a diferentes tempos de desmineralização / Deciduous enamel surface analysis after different periods of demineralization

É objetivo deste trabalho avaliar as alterações superficiais apresentadas pelo esmalte de dentes decíduos após diferentes períodos de desmineralização em ácido láctico (pH=4,5). Utilizou-se 25 dentes decíduos divididos nos seguintes grupos: I-Controle; II-Desmineralização por 12 horas; III-Desmineralização por 24 horas; IV-Desmineralização por 48 horas; V-Desmineralização por 72 horas. Logo após, as amostras foram analisadas macroscópica e microcospicamente. Observou-se que houve alterações estruturais superficiais no esmalte, o qual exibiu macroscopicamente perda de brilho e microscopicamente grande perda de mineral, sendo estas mais evidentes a partir de 48 horas de desmineralização. Os autores concluíram que as alterações subclínicas, devem ser de conhecimento do profissional para que ele possa educar e conscientizar seus pacientes quanto ao desenvolvimento de medidas preventivas efetivas, para evitar o desenvolvimento de futuras lesões de cárie

Presença do papilomavirus humano em lesöes malignas de mucosa oral / Presence of human papillomavirus in malignant oral lesions

O objetivo deste estudo foi investigar a prevalência do papilomavírus humano 6/11 e 16/18 em pacientes, com lesöes orais clínicamente diagnosticadas como leucoplasias, atendidas na Faculdade de Odontologia de Araraquara, UNESP, Brasil. Após a inclusäo em parafina, os cortes corados com H&E, foram selecionadas 30 biópsias e separadas em 3 grupos: lesöes sem displasia (n=10), lesöes com diferentes graus de displasia (n=10) e carcinoma espinocelular invasivo(n=10). As lesöes que apresentaram displasia epitelial foram classificadas de acordo com os critérios histopatológicos propostos por Van Der Waal. As lesöes foram investigadas para a presença de HPV por hibridizaçäo in situ com sondas biotiniladas de amplo espectro, 6/11 e 16/18. HPV 16/18 foi detectado em 20 por cento (n=2) das biópsias com displasia severa. A presença de HPV 16/18 em lesöes malignas sugere sua importância como fator de risco na carcinogênese oral

[Presence of human papillomavirus in malignant oral lesions]. / Presença do papilomavirus humano em lesões malignas de mucosa oral.

The purpose of this study was to investigate the prevalence of human papillomavirus 6/11 and 16/18 in patients, with oral lesions clinically diagnosed as leucoplakia, attending the School of Dentistry, University of São Paulo State/UNESP, Brazil. After paraffin embedded process, in the sections staining with H&E, 30 biopsies were screened and separated on 3 groups: 10 oral lesions without dysplasia, 10 with dysplasia, and 10 with invasive squamous cell carcinoma. The lesions with dysplasia were classified in agreement with Van Der Wall's histopathological standard method. Oral lesions were investigated for the presence of human papillomavirus (HPV) by in situ hybridization with wide-spectrum, 6/11 and 16/18 biotinylated probes. HPV 16/18 was found in 20% (n = 2) of the leucoplakia with severe-degree dysplasia. The presence of HPV 16/18 in malignant lesions suggests its importance as a risk factor for oral carcinogenesis.