A New Sugarcane Cystatin Strongly Binds to Dental Enamel and Reduces Erosion.

Cystatin B was recently identified as an acid-resistant protein in acquired enamel pellicle; it could therefore be included in oral products to protect against caries and erosion. However, human recombinant cystatin is very expensive, and alternatives to its use are necessary. Phytocystatins are reversible inhibitors of cysteine peptidases that are found naturally in plants. In plants, they have several biological and physiological functions, such as the regulation of endogenous processes, defense against pathogens, and response to abiotic stress. Previous studies performed by our research group have reported high inhibitory activity and potential agricultural and medical applications of several sugarcane cystatins, including CaneCPI-1, CaneCPI-2, CaneCPI-3, and CaneCPI-4. In the present study, we report the characterization of a novel sugarcane cystatin, named CaneCPI-5. This cystatin was efficiently expressed in Escherichia coli, and inhibitory assays demonstrated that it was a potent inhibitor of human cathepsins B, K, and L ( Ki = 6.87, 0.49, and 0.34 nM, respectively). The ability of CaneCPI-5 to bind to dental enamel was evaluated using atomic force microscopy. Its capacity to protect against initial enamel erosion was also tested in vitro via changes in surface hardness. CaneCPI-5 showed a very large force of interaction with enamel (e.g., compared with mucin and casein) and significantly reduced initial enamel erosion. These results suggest that the inclusion of CaneCPIs in dental products might confer protection against enamel erosion.

Clinicopathological prognostic factors of oral tongue squamous cell carcinoma: a retrospective study of 202 cases.

Although several histopathological parameters and grading systems have been described as predictive of the treatment response and outcome of oral squamous cell carcinoma (OSCC), none is universally accepted. A new scoring system, the histological risk model, was recently described to be a powerful predictive tool for recurrence and overall survival in OSCC. The aim of this study was to verify the predictive role of the histological risk model in a cohort of 202 patients at all stages of oral/mobile tongue squamous cell carcinoma (OTSCC). Demographic and clinical data were collected from the medical records and the tumours were evaluated using the histological risk model. Statistical analyses were performed using the χ(2) test, the Kaplan-Meier method, and the Cox regression model. The histological risk model showed no statistical correlation with demographic or clinical parameters and did not Predict the outcome of the OTSCC patients. However, multivariate regression analysis revealed a significant correlation of the clinical disease stage with the disease outcome. Despite major efforts to identify new predictive parameters and histological systems, clinical features are still the most reliable prognostic factors for patients with OTSCC.

Analysis of the frequency and nature of hyaline ring granulomas in inflammatory odontogenic cysts.

AIM: To determine the prevalence of hyaline ring granulomas (HRGs) in a large case series of inflammatory odontogenic cysts, and to investigate the nature of these structures. METHODOLOGY: All records from the patients diagnosed with inflammatory odontogenic cysts between January 1970 and April 2009 were reviewed. Histologic sections were evaluated by light microscopy and cases with HRGs for which sufficient biological material was available were submitted to histochemical analysis (Masson's trichrome) and immunohistochemistry (CD34, CD68 and collagen IV). RESULTS: Twenty-two (3.3%) of the 661 cases of inflammatory odontogenic cysts diagnosed during the study period presented HRGs. The relative frequency of HRGs was higher amongst residual radicular cysts (6.1%), followed by paradental cysts (5.6%) and radicular cysts (3.0%). HRGs appeared as roughly circular homogeneous/fibrillar masses in 14 (63.6%) cases and as round structures enclosing amorphous material in 3 (13.6%) cases. Most (77.8%) roughly circular homogeneous/fibrillar masses were positive for collagen, whereas all (100.0%) round structures enclosing amorphous material were negative for this protein. Immunohistochemistry showed that most mononucleated cells and all multinucleated giant cells were positive for CD68, but negative for CD34, in all cases. In addition, collagen IV immunostaining was negative in amorphous structures and weakly positive in homogeneous/fibrillar masses. CONCLUSIONS: The present results suggest a very low frequency of HRGs in inflammatory odontogenic cysts and support the hypothesis that these structures arise from the implantation of foreign material, most likely food particles of plant or vegetable origin. The diverse microscopic features of HRG possibly represent different developmental stages of this structure.

Assessment of swim-up and discontinuous density gradient in sperm sex preselection for bovine embryo production / Avaliação do swim-up e do gradiente descontinuo de densidade na pré-seleção do sexo de espermatozoides na produção de embriões bovinos

The purpose of this work was to associate the modified swim-up method with centrifugation in density gradient for the separation of X-bearing spermatozoa. Sperm viability and integrity were evaluated through the Trypan Blue/Giemsa staining method. Quality control of centrifuged spermatozoa was performed in in vitro produced embryos. The results were validated by the sex ratio of in vitro produced embryos using PCR by Y- specific sequences present in bovine male genomic DNA. After determining genetic sex of in vitro produced embryos, the results showed difference (P<0.05) in deviation of sex ratio when comparing the control group (45.2% females) with the other spermatozoa selection procedures (60.6% females) (P<0.05). The sperm selection methods are capable of selecting X-bearing spermatozoa without compromising the spermatozoa fertility (cleavage and blastocyst rates, 70% and 26%, respectively) and were considered relevant methods to be introduced in bovine in vitro produced embryo programs.(AU)

O objetivo do presente trabalho foi associar o método de swim-up modificado à centrifugação em gradiente de densidade para a separação de espermatozoides portadores do cromossomo X. A viabilidade e a integridade espermática foram avaliadas pelo método de coloração Azul de Tripan e Giemsa. O controle de qualidade dos espermatozoides centrifugados foi realizado por meio da produção in vitro de embriões bovinos. Os resultados foram validados pela técnica de PCR para verificar a proporção sexual dos embriões produzidos in vitro, com o uso de sequências Y especificas presente no DNA genômico de machos bovinos. Após determinar o sexo genético dos embriões produzidos in vitro, os resultados não mostraram diferença (P<0,05) no desvio da proporção do sexo quando comparou o grupo controle (45,2% de fêmeas) com os outros processos de seleção de espermatozoides (60,6% de fêmeas) (P<0,05). Os métodos de seleção de espermatozoides são capazes de selecionar espermatozoides portadores do cromossomo X sem comprometer a fertilidade, medida pelas taxas de clivagem e blastocisto de 70% e 26%, respectivamente, e foram considerados métodos de relevância para serem introduzidos nos programas de produção in vitro de embriões bovinos.(AU)

Assessment of swim-up and discontinuous density gradient in sperm sex preselection for bovine embryo production / Avaliação do swim-up e do gradiente descontinuo de densidade na pré-seleção do sexo de espermatozoides na produção de embriões bovinos

The purpose of this work was to associate the modified swim-up method with centrifugation in density gradient for the separation of X-bearing spermatozoa. Sperm viability and integrity were evaluated through the Trypan Blue/Giemsa staining method. Quality control of centrifuged spermatozoa was performed in in vitro produced embryos. The results were validated by the sex ratio of in vitro produced embryos using PCR by Y- specific sequences present in bovine male genomic DNA. After determining genetic sex of in vitro produced embryos, the results showed difference (P<0.05) in deviation of sex ratio when comparing the control group (45.2% females) with the other spermatozoa selection procedures (60.6% females) (P<0.05). The sperm selection methods are capable of selecting X-bearing spermatozoa without compromising the spermatozoa fertility (cleavage and blastocyst rates, 70% and 26%, respectively) and were considered relevant methods to be introduced in bovine in vitro produced embryo programs.

O objetivo do presente trabalho foi associar o método de swim-up modificado à centrifugação em gradiente de densidade para a separação de espermatozoides portadores do cromossomo X. A viabilidade e a integridade espermática foram avaliadas pelo método de coloração Azul de Tripan e Giemsa. O controle de qualidade dos espermatozoides centrifugados foi realizado por meio da produção in vitro de embriões bovinos. Os resultados foram validados pela técnica de PCR para verificar a proporção sexual dos embriões produzidos in vitro, com o uso de sequências Y especificas presente no DNA genômico de machos bovinos. Após determinar o sexo genético dos embriões produzidos in vitro, os resultados não mostraram diferença (P<0,05) no desvio da proporção do sexo quando comparou o grupo controle (45,2% de fêmeas) com os outros processos de seleção de espermatozoides (60,6% de fêmeas) (P<0,05). Os métodos de seleção de espermatozoides são capazes de selecionar espermatozoides portadores do cromossomo X sem comprometer a fertilidade, medida pelas taxas de clivagem e blastocisto de 70% e 26%, respectivamente, e foram considerados métodos de relevância para serem introduzidos nos programas de produção in vitro de embriões bovinos.

Effects of discontinuous Percoll gradient centrifugation on the quality of bovine spermatozoa evaluated with computer-assisted semen analysis and fluorescent probes association.

The objective of this study was to evaluate the quality of bovine frozen-thawed sperm cells after Percoll gradient centrifugation. Frozen semen doses were obtained from six bulls of different breeds, including three taurine and three Zebu animals. Four ejaculates per bull were evaluated before and after discontinuous Percoll gradient centrifugation. Sperm motility was assessed by computer-assisted semen analysis and the integrity of the plasma and acrosomal membranes, as well as mitochondrial function, were evaluated using a combination of fluorescent probes propidium iodide, fluorescein isothiocyanate-conjugated Pisum sativum agglutinin and 5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolcarbocyanine iodide. The procedure of Percoll gradient centrifugation increased the percentage of total and progressive sperm motility, beat frequency, rectilinear motility, linearity and rapidly moving cells. In addition, the percentage of cells with intact plasma membrane and mitochondrial membrane potential was increased in post-centrifugation samples. However, the percentage of sperm cells with intact acrosomal membrane was markedly reduced. The method used selected the motile cells with intact plasma membrane and higher mitochondrial functionality in frozen-thawed bull semen, but processing, centrifugation and/or the Percoll medium caused damage to the acrosomal membrane.

Induction of the acrosome reaction test to in vitro estimate embryo production in Nelore cattle / Teste de indução da reação acrossomal na estimativa de produção in vitro de embriões Nelore

The effectiveness of induction of the acrosome reaction (AR) test as a parameter to in vitro estimate embryo production (IVP) in Nelore breed and the AR pattern by the Trypan Blue/Giemsa (TB) stain were evaluated. Frozen semen samples from ten Nelore bulls were submitted to AR induction and were also evaluated for cleavage and blastocyst rates. The treatments utilized for AR induction were: control (TALP medium), TH (TALP medium + 10μg heparin), TL (TALP medium + 100μg lysophosphatidylcholine) and THL (TALP medium + 10μg heparin + 100μg lysophosphatidylcholine). Sperm acrosomal status and viability were evaluated by TB staining at 0 and after 4h incubation at 38ºC. The results obtained for AR presented a significant difference (P<0.05) in the percentage of acrosome reacted live sperm after 4h of incubation in the treatments that received heparin. The cleavage and blastocyst rates were 60 percent and 38 percent respectively and a significant difference was observed among bulls (P<0.05). It was founded a satisfactory model to estimate the cleavage and blastocyst rates by AR induction test. Therefore, it can be concluded that the induction of the AR test is a valuable tool to predict the IVP in Nelore breed.(AU)

Avaliou-se a eficiência da técnica de indução da reação acrossomal (RA) como parâmetro para estimar a produção in vitro (PIV) de embriões Nelore e analisou-se o padrão de RA pela técnica de coloração Azul de Tripan/Giemsa (TB). Amostras de sêmen congelado de dez touros foram submetidas à indução da RA e avaliadas quanto a taxa de clivagem e blastocisto. Os tratamentos utilizados para indução da RA foram: controle (meio TALP), TH (meio TALP + 10μg heparina), TL (meio TALP + 100μg lisofosfatidilcolina) e THL (meio TALP + 10μg heparina + 100μg lisofosfatidilcolina). Avaliou-se viabilidade espermática e acrossomal pela coloração TB a zero e após 4h de incubação a 38ºC. Os resultados obtidos para RA mostram uma diferença significativa (P<0,05) na porcentagem de espermatozoides vivos com acrossoma reagindo após 4h de incubação nos tratamentos que receberam heparina. As taxas de clivagem e blastocisto obtidas foram 60 por cento e 38 por cento respectivamente e observou-se uma diferença significativa entre touros (P<0,05). Delineou-se um modelo satisfatório para estimar as taxas de clivagem e blastocisto. Desta forma, conclui-se que o teste de indução da RA é uma ferramenta valiosa para predizer a PIV na raça Nelore.(AU)

Induction of the acrosome reaction test to in vitro estimate embryo production in Nelore cattle / Teste de indução da reação acrossomal na estimativa de produção in vitro de embriões Nelore

The effectiveness of induction of the acrosome reaction (AR) test as a parameter to in vitro estimate embryo production (IVP) in Nelore breed and the AR pattern by the Trypan Blue/Giemsa (TB) stain were evaluated. Frozen semen samples from ten Nelore bulls were submitted to AR induction and were also evaluated for cleavage and blastocyst rates. The treatments utilized for AR induction were: control (TALP medium), TH (TALP medium + 10μg heparin), TL (TALP medium + 100μg lysophosphatidylcholine) and THL (TALP medium + 10μg heparin + 100μg lysophosphatidylcholine). Sperm acrosomal status and viability were evaluated by TB staining at 0 and after 4h incubation at 38ºC. The results obtained for AR presented a significant difference (P<0.05) in the percentage of acrosome reacted live sperm after 4h of incubation in the treatments that received heparin. The cleavage and blastocyst rates were 60 percent and 38 percent respectively and a significant difference was observed among bulls (P<0.05). It was founded a satisfactory model to estimate the cleavage and blastocyst rates by AR induction test. Therefore, it can be concluded that the induction of the AR test is a valuable tool to predict the IVP in Nelore breed.

Avaliou-se a eficiência da técnica de indução da reação acrossomal (RA) como parâmetro para estimar a produção in vitro (PIV) de embriões Nelore e analisou-se o padrão de RA pela técnica de coloração Azul de Tripan/Giemsa (TB). Amostras de sêmen congelado de dez touros foram submetidas à indução da RA e avaliadas quanto a taxa de clivagem e blastocisto. Os tratamentos utilizados para indução da RA foram: controle (meio TALP), TH (meio TALP + 10μg heparina), TL (meio TALP + 100μg lisofosfatidilcolina) e THL (meio TALP + 10μg heparina + 100μg lisofosfatidilcolina). Avaliou-se viabilidade espermática e acrossomal pela coloração TB a zero e após 4h de incubação a 38ºC. Os resultados obtidos para RA mostram uma diferença significativa (P<0,05) na porcentagem de espermatozoides vivos com acrossoma reagindo após 4h de incubação nos tratamentos que receberam heparina. As taxas de clivagem e blastocisto obtidas foram 60 por cento e 38 por cento respectivamente e observou-se uma diferença significativa entre touros (P<0,05). Delineou-se um modelo satisfatório para estimar as taxas de clivagem e blastocisto. Desta forma, conclui-se que o teste de indução da RA é uma ferramenta valiosa para predizer a PIV na raça Nelore.

Oral foreign body granuloma: unusual presentation of a rare adverse reaction to permanent injectable cosmetic filler.

A variety of injectable permanent fillers have been used in orofacial tissues for cosmetic purposes. Most of these substances seem to be well tolerated but adverse reactions have been reported. Foreign body granulomas are a rare adverse reaction to injectable permanent fillers. The authors report the unusual case of a 56-year-old woman with a foreign body granuloma located exclusively in the oral cavity that was due to injection of a permanent filler.

Hyperplasia of myoepithelial cells expressing calponin during atrophy of the rat parotid gland induced by duct ligation.

The number and location of myoepithelial cells in the rat parotid gland submitted to surgical ligation of its main excretory duct were studied through immunohistochemical labelling for calponin. These cells were labelled by the streptavidin-biotin method using anti-calponin primary antibody, and their number was determined during each step of glandular atrophy, i.e., at time zero (control) and 1, 7, 15, 21, 30 and 60 days after ligation. Morphological analysis showed a gradual decrease and fibrosis of the glandular lobules accompanied by disappearance of the acini and the occurrence of duct-like structures. The expression of calponin was observed in all specimens analysed, being restricted to myoepithelial cells. Labelling revealed the distribution of myoepithelial cells around the acini and intercalated ducts in the control group and around duct-like structures later during the course of atrophy. Quantitative analysis demonstrated significant increase in the number of myoepithelial cells up to day 7 post-ligation, followed by gradual increases which, however, were not statistically significant. These results suggest that myoepithelial cells proliferate intensely up to day 7 post-ligation, an event that coincides with a higher rate of disappearance of acinar cells. After this period, the elevatednumber of cells observed at the end of the previous period is maintained.

Chondroma in rufescent tiger-heron (Tigrisoma leneatum): case report / Condroma em socó-boi (Tigrisoma leneatum): relato de caso

The paper describes a chondroma affecting the left metacarpus of a free-ranging rufescent tiger-heron (Tigrisoma lineatum). There are very few reports of such neoplastic process in this avian species.(AU)

Relata-se a ocorrência de neoplasia em Tigrisoma lineatum (socó-boi), localizada na região metacarpiana esquerda, caracterizada histologicamente como condroma. Este é um dos primeiros relatos desta patologia em indivíduos desta espécie.(AU)

Chondroma in rufescent tiger-heron (Tigrisoma leneatum): case report

The paper describes a chondroma affecting the left metacarpus of a free-ranging rufescent tiger-heron (Tigrisoma lineatum). There are very few reports of such neoplastic process in this avian species