Diferenciação de laticíferos articulados em plantas de Tabernaemontana catharinensis (Apocynaceae)

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Expression of cell cycle regulatory proteins in epithelial components of dental follicles

Dental follicle is a component of tooth germs, which remain adjacent to the crown of unerupted or impacted teeth. Under the influence of pathologic changes, however, dental follicles that possess reduced epithelium can proliferate into stratified squamous epithelium as far as originate dental cysts. In order to clarify the role of apoptosis and cellular proliferation herein, expression of p53 and PCNA was examined in epithelial components of dental follicles associated with impacted third molars by means of immunohistochemistry. A total of 40 cases was included in this study being 22 cases with reduced epithelium and 18 cases with stratified epithelium. Expression of p53 expression was weak or not detected in dental follicles with reduced and stratified squamous epithelium. By contrast, PCNA positive cells were evidenced in basal and supra basal layers of the stratified squamous epithelium and in reduced epithelium of dental follicles, but without any significant statistically differences between them (P > 0.05). In conclusion, these data suggest that dental follicles possess proliferative activity as depicted by PCNA-positive nuclei in some epithelial cells. However, the biological behavior of dental follicles during the late stage of dental eruptive process may not be associated with deregulation of death and/or cell proliferation.

DNA methylation patterns in bladder cancer and washing cell sediments: a perspective for tumor recurrence detection

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Rabies neutralizing antibody detection by indirect immunperoxidase serum neutralization assay performed on chicken embryo related cell line

The aim of this study was to evaluate the indirect immunoperoxidase virus neutralization (IPVN) and mouse neutralization test (MNT) to detect antibodies against rabies virus from vaccinated dogs and cattle. The IPVN was set up for the ability to measure 0.5 International Units/ml (IU) of antibody required by the World Health Organization and the Office International des Epizooties as the minimum response for proof of rabies immunization. IPVN was developed and standardized in chicken embryo related (CER) cell line when 141 dog and 110 cattle sera were applied by serial five-fold dilutions (1:5, 1:25, 1:125) as well as the positive and negative reference controls, all added in four adjacent wells, of 96-well microplates. A 50 µl amount of CVS32 strain dilution containing 50-200 TCID50/ml was mixed to each serum dilution, and after 90 min 50 µl of 3 x 10(5) cells/mlcell suspension added to each well. After five days of incubation, the monolayers were fixed and the IPVN test performed. The correlation coefficient between the MNT and IPVN performed in CER cells was r = 0.9949 for dog sera (n = 100) and r = 0.9307 for cattle sera (n = 99), as well as good specificity (94.7%), sensitivity (87.5%), and agreement (96.6%) were also obtained. IPVN technique can adequately identify vaccinated and unvaccinated animals, even from low-responding vaccinated animals, with the advantage of low cost and faster then MNT standard test.

Effects of light-curing time on the cytotoxicity of a restorative resin composite applied to an immortalized odontoblast-cell line

This in vitro study evaluated the cytotoxic effects of a restorative resin composite applied to an immortalized odontoblast-cell line (MDPC-23). Seventy-two round resin discs (2-mm thick and 4 mm in diameter) were light-cured for 20 or 40 seconds and rinsed, or not, with PBS and culture medium. The resin discs were divided into four experimental groups: Group 1: Z-100/20 seconds; Group 2: Z-100/20 seconds/rinsed; Group 3: Z100/40 seconds; Group 4: Z-100/40 seconds/rinsed. Circular filter paper was used as a control material (Group 5). The round resin discs and filter papers were placed in the bottom of wells of four 24-well dishes (18 wells for each experimental and control group). MDPC-23 cells (30,000 cells/cm(2)) were plated in the wells and allowed to incubate for 72 hours. The zone of inhibition around the resin discs was measured under inverted light microscopy; the MTT assay was carried out for mitochondrial respiration and cell morphology was measured under SEM. The scores obtained from inhibition zone and MTT assay were analyzed with the Kruskal-Wallis followed by Dunnett tests. In Groups 1, 2, 3 and 4, the thickness of the inhibition zone was 1,593 +/- 12.82 mum, 403 +/- 15.49 mum, 1,516 +/- 9.81 mum and 313 +/- 13.56 mum, respectively. There was statistically significant difference among the experimental and control groups at the 0.05 level of significance. The MTT assay demonstrated that the resin discs of the experimental groups 1, 2, 3 and 4 reduced the cell metabolism by 83%, 40.1%, 75.5% and 24.5%. Only between the Groups 2 and 4 was there no statistically significant difference for mitochondrial respiration. Close to the resin discs, the MDPC-23 cells exhibited rounded shapes, with only a few cellular processes keeping the cells attached to the substrate or, even disruption of plasma membrane. Adjacent to the inhibition zone, the cultured cells exhibited multiple fine cellular processes on the cytoplasmic membrane organized in epithelioid nodules, similar to the morphology observed to the control group. Based on the results, the authors may conclude that the Z-100 resin composite light cured for 20 seconds was more cytopathic to MDPC-23 cells than Z-100 light cured for 40 seconds. The cytotoxic effects of the resin discs decreased after rinsing them with PBS and culture medium. This was confirmed by MTT assay and upon evaluation of the inhibition zone, which was narrower following rinsing of the resin discs.

Cholangiocarcinoma and Squamous Cell Carcinoma of the Stratified Epithelial Portion of the Stomach in a Horse: A Case Report

Both cholangiocarcinoma and squamous cell carcinoma of the stomach stratified squamous epithelial portion are rarely found in equine medicine despite being more common in human beings, dogs, and cats. The objective of the present article was to report the simultaneous occurrences of these two types of neoplasias in an 11-year-old mare. Numerous firm, whitish nodules were distributed throughout the liver parenchyma and those protruding over its surface were umbilicated in their appearance. It was verified that the nodules adhered to the peritoneum and omentum, diaphragm, spleen, and stomach serosa compressed the adjacent structures. The stomach stratified squamous epithelial portion, particularly originating in the margo plicatus toward the cardia, was covered by numerous smooth, whitish spherules. Microscopic examination allowed the liver, tumor, and the abdominal implants to be identified as a cholangiocarcinoma, and the stomach neoplasia as a carcinoma of its stratified squamous epithelial portion. Considering this as an uncommon finding, although when considered individually, the presence of a cholangiocarcinoma and a squamous cell carcinoma of the stratified squamous epithelial portion of the equine stomach in the same specimen is worthy of reporting. (C) 2011 Elsevier B.V. All rights reserved.

Cell death in ovarioles causes permanent sterility in Frieseomelitta varia worker bees

Frieseomelitta varia worker bees do not lay eggs even when living in queenless colonies, a condition that favors ovary development and oviposition in the majority of highly social bees. The permanent sterility of these worker bees was initially attributed to a failure in ovary morphogenesis and differentiation. Using transmission electron microscopy we found that at the beginning of the pupal phase the ovaries of F. varia workers are formed by four ovarioles, each of them composed of 1) a terminal filament at the apex of the ovarioles, containing juxtaposed and irregularly shaped cells, 2) a germarium with clusters of cystocytes and prefollicular cells showing long cytoplasmic projections that envelop the cystocyte clusters, 3) fusiform interfollicular and basal stalk precursor cells, and 4) globular, irregularly contoured basal cells with large nuclei. However, during the pupal phase an accentuated and progressive process of cell death takes place in the ovarioles. The dying cells are characterized by large membrane bodies, electron-dense apoptotic bodies, vacuoles, vesiculation, secondary lysosomes, enlarged rough endoplasmic reticulum cisternae, swollen mitochondria, pycnotic nuclei, masses of chromatin adjacent to the convoluted nuclear envelope, and nucleoli showing signs of fragmentation. Cell death continues in ovarioles even after the emergence of the workers. Once they become nurse bees, the ovaries have become transformed into a cell mass in which structurally organized ovarioles can no longer be identified. In F. varia workers, ovariole cell death most certainly is part of the program of caste differentiation.

Homeobox gene expression profile indicates HOXA5 as a candidate prognostic marker in oral squamous cell carcinoma

The search for molecular markers to improve diagnosis, individualize treatment and predict behavior of tumors has been the focus of several studies. This study aimed to analyze homeobox gene expression profile in oral squamous cell carcinoma (OSCC) as well as to investigate whether some of these genes are relevant molecular markers of prognosis and/or tumor aggressiveness. Homeobox gene expression levels were assessed by microarrays and qRT-PCR in OSCC tissues and adjacent non-cancerous matched tissues (margin), as well as in OSCC cell lines. Analysis of microarray data revealed the expression of 147 homeobox genes, including one set of six at least 2-fold up-regulated, and another set of 34 at least 2-fold down-regulated homeobox genes in OSCC. After qRT-PCR assays, the three most up-regulated homeobox genes (HOXA5, HOXD10 and HOXD11) revealed higher and statistically significant expression levels in OSCC samples when compared to margins. Patients presenting lower expression of HOXA5 had poorer prognosis compared to those with higher expression (P=0.03). Additionally, the status of HOXA5, HOXD10 and HOXD11 expression levels in OSCC cell lines also showed a significant up-regulation when compared to normal oral keratinocytes. Results confirm the presence of three significantly upregulated (>4-fold) homeobox genes (HOXA5, HOXD10 and HOXD11) in OSCC that may play a significant role in the pathogenesis of these tumors. Moreover, since lower levels of HOXA5 predict poor prognosis, this gene may be a novel candidate for development of therapeutic strategies in OSCC.

An Integrative Genomic and Transcriptomic Analysis Reveals Potential Targets Associated with Cell Proliferation in Uterine Leiomyomas

Background: Uterine Leiomyomas (ULs) are the most common benign tumours affecting women of reproductive age. ULs represent a major problem in public health, as they are the main indication for hysterectomy. Approximately 40-50% of ULs have non-random cytogenetic abnormalities, and half of ULs may have copy number alterations (CNAs). Gene expression microarrays studies have demonstrated that cell proliferation genes act in response to growth factors and steroids. However, only a few genes mapping to CNAs regions were found to be associated with ULs. Methodology: We applied an integrative analysis using genomic and transcriptomic data to identify the pathways and molecular markers associated with ULs. Fifty-one fresh frozen specimens were evaluated by array CGH (JISTIC) and gene expression microarrays (SAM). The CONEXIC algorithm was applied to integrate the data. Principal Findings: The integrated analysis identified the top 30 significant genes (P<0.01), which comprised genes associated with cancer, whereas the protein-protein interaction analysis indicated a strong association between FANCA and BRCA1. Functional in silico analysis revealed target molecules for drugs involved in cell proliferation, including FGFR1 and IGFBP5. Transcriptional and protein analyses showed that FGFR1 (P = 0.006 and P<0.01, respectively) and IGFBP5 (P = 0.0002 and P = 0.006, respectively) were up-regulated in the tumours when compared with the adjacent normal myometrium. Conclusions: The integrative genomic and transcriptomic approach indicated that FGFR1 and IGFBP5 amplification, as well as the consequent up-regulation of the protein products, plays an important role in the aetiology of ULs and thus provides data for potential drug therapies development to target genes associated with cellular proliferation in ULs. © 2013 Cirilo et al.

The Predominance of Alternatively Activated Macrophages Following Challenge with Cell Wall Peptide-Polysaccharide After Prior Infection with Sporothrix schenckii

Sporotrichosis is a subcutaneous mycosis that is caused by the dimorphic fungus Sporothrix schenckii. This disease generally occurs within the skin and subcutaneous tissues, causing lesions that can spread through adjacent lymphatic vessels and sometimes leading to systemic diseases in immunocompromised patients. Macrophages are crucial for proper immune responses against a variety of pathogens. Furthermore, macrophages can play different roles in response to different microorganisms and forms of activation, and they can be divided into classic or alternatively activated populations, as also known as M1 and M2 macrophages. M1 cells can lead to tissue injury and contribute to pathogenesis, whereas M2 cells promote angiogenesis, tissue remodeling, and repair. The aim of this study was to investigate the roles of M1 and M2 macrophages in a sporotrichosis model. Toward this end, we performed phenotyping of peritoneal exudate cells and evaluated the concomitant production of several immunomediators, including IL-12, IL-10, TGF-β, nitric oxide, and arginase-I activity, which were stimulated ex vivo with cell wall peptide-polysaccharide. Our results showed the predominance of the M2 macrophage population, indicated by peaks of arginase-I activity as well as IL-10 and TGF-β production during the 6th and 8th weeks after infection. These results were consistent with cellular phenotyping that revealed increases in CD206-positive cells over this period. This is the first report of the participation of M2 macrophages in sporotrichosis infections. © 2013 Springer Science+Business Media Dordrecht.

Cell therapy in experimental model of inflammatory bowel disease

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Gingival squamous cell carcinoma: a case report

Squamous cell carcinoma is a malignant epithelial neoplasm characterized by variable clinical manifestations. When located in the gingiva, this neoplasm may mimic common inflammatory lesions. The aim of this study was to report a case of atypical squamous cell carcinoma, in which the patient had no risk factors for the development of this neoplasm. A 61 year old Caucasian female was seen with a 3 month history of a rapidly growing, painful nodule in the gingiva adjacent to tooth #11. Clinical examination revealed a proliferative lesion in the vestibular marginal gingiva of teeth #11 and #12, presenting with purulent exudation. Thus, in view of the clinical symptoms and differential diagnosis of an infectious granulomatous process and malignant neoplasm, an incisional biopsy was obtained from the lesion. The diagnosis of squamous cell carcinoma was made and fourteen days after incisional biopsy, healing was found to be unsatisfactory. The patient was referred for treatment consisting of surgical excision of the tumour. A removable partial denture was fabricated for rehabilitation, one month after surgery of the maxilla; the patient was submitted to dissection of the regional lymph nodes and radiotherapy for an additional 3 months. Three years after the end of treatment, the patient continues to be followed-up and does not show any sign of recurrence. Gingival squamous cell carcinoma is a condition which chance of cure is higher when carcinomatous lesions are diagnosed and treated early. In this instance dentists play an important role in early detection of gingival squamous cell carcinoma.

Primary squamous cell carcinoma of the thyroid diagnosed as anaplastic carcinoma: failure in fine-needle aspiration cytology?

A case of primary squamous-cell carcinoma (SCC) of the thyroid which had been initially diagnosed as an anaplastic carcinoma (ATC) is described: female, 73 years old, with a fast-growing cervical nodule on the left side and hoarseness for 3 months. Ultrasonography showed a 4.5 cm solid nodule. FNA was compatible with poorly differentiated carcinoma with immunoreactivity for AE1/AE3, EMA. Thyroidectomy was performed. Histopathological examination showed a nonencapsulated tumor. Immunohistochemistry disclosed positivity for AE1/AE3, p53,p63, and Ki67. The diagnosis was ATC. A second opinion reported tumor consisting of squamous cells, with intense inflammatory infiltrate both in tumor and in the adjacent thyroid, with final diagnosis of SCC, associated with Hashimoto thyroiditis. No other primary focus of SCC was found. Patient has shown a 48-month survival period. Clinically, primary SCCs of the thyroid and ATCs are similar. The distinction is often difficult particularly when based on the cytological analysis of FNA material.