Papel da proteína de reparo XPC na regulação das proteínas de reparo APE1, OGG1 e PARP-1 em células humanas e de camundongos

studies using UV as a source of DNA damage. However, even though unrepaired UV-induced DNA damages are related to mutagenesis, cell death and tumorigenesis, they do not explain phenotypes such as neurodegeneration and internal tumors observed in patients with syndromes like Xeroderma Pigmentosum (XP) and Cockayne Syndrome (CS) that are associated with NER deficiency. Recent evidences point to a role of NER in the repair of 8-oxodG, a typical substrate of Base Excision Repair (BER). Since deficiencies in BER result in genomic instability, neurodegenerative diseases and cancer, it was investigated in this research the impact of XPC deficiency on BER functions in human cells. It was analyzed both the expression and the cellular localization of APE1, OGG1 e PARP-1, the mainly BER enzymes, in different NER-deficient human fibroblasts. The endogenous levels of these enzymes are reduced in XPC deficient cells. Surprisingly, XP-C fibroblasts were more resistant to oxidative agents than the other NER deficient fibroblasts, despite presenting the highest of 8-oxodG. Furthermore, subtle changes in the nuclear and mitochondrial localization of APE1 were detected in XP-C fibroblasts. To confirm the impact of XPC deficiency in the regulation of APE1 and OGG1 expression and activity, we constructed a XPC-complemented cell line. Although the XPC complementation was only partial, we found that XPC-complemented cells presented increased levels of OGG1 than XPC-deficient cells. The extracts from XPC-complemented cells also presented an elevated OGG1 enzimatic activity. However, it was not observed changes in APE1 expression and activity in the XPCcomplemented cells. In addition, we found that full-length APE1 (37 kDa) and OGG1- α are in the mitochondria of XPC-deficient fibroblasts and XPC-complemented fibroblasts before and after induction of oxidative stress. On the other hand, the expression of APE1 and PARP-1 are not altered in brain and liver of XPC knockout mice. However, XPC deficiency changed the APE1 localization in hypoccampus and hypothalamus. We also observed a physical interaction between XPC and APE1 proteins in human cells. In conclusion, the data suggest that XPC protein has a role in the regulation of OGG1 expression and activity in human cells and is involved mainly in the regulation of APE1 localization in mice. Aditionally, the response of NER deficient cells under oxidative stress may not be only associated to the NER deficiency per se, but it may include the new functions of NER enzymes in regulation of expression and cell localization of BER proteins

Investigação do papel modulador do Haloperidol sobre os níveis de proteínas relacionadas à plasticidade e preferência por objetos novos em camudongos

Dopamine (DA) is known to regulate both sleep and memory formations, while sleep plays a critical role in the consolidation of different types of memories. We believe that pharmacological manipulation of dopaminergic pathways might disrupt the sleep-wake cycle, leading to mnemonic deficits, which can be observed in both behavioral and molecular levels. Therefore, here we investigated how systemic injections of haloperidol (0.3 mg/kg), immediately after training in dark and light periods, affects learning assessed in the novel object preference test (NOPT) in mice. We also investigated the hippocampal levels of the plasticity-related proteins Zif-268, brain-derived neurotrophic factor (BDNF) and phosphorylated Ca2+/calmodulin-dependent protein kinases II (CaMKII-P) in non-exposed (naïve), vehicle-injected controls and haloperidol-treated mice at 3, 6 and 12 hours after training in the light period. Haloperidol administration during the light period led to a subsequent impairment in the NOPT. In contrast, preference was not observed during the dark period neither in mice injected with haloperidol, nor in vehicle-injected animals. A partial increase of CaMKII-P in the hippocampal field CA3 of vehicle-injected mice was detected at 3h. Haloperidol-treated mice showed a significant decrease in the dentate gyrus of CaMKII-P levels at 3, 6 and 12h; of Zif-268 levels at 6h, and of BDNF levels at 12h after training. Since the mnemonic effects of haloperidol were only observed in the light period when animals tend to sleep, we suggest that these effects are related to REM sleep disruption after haloperidol injection

Biodegradação de hidrocarbonetos aromáticos policíclicos: prospecção metagenômica e modelagem computacional 3-D de proteínas

Knowledge of the native prokaryotes in hazardous locations favors the application of biotechnology for bioremediation. Independent strategies for cultivation and metagenomics contribute to further microbiological knowledge, enabling studies with non-cultivable about the "native microbiological status and its potential role in bioremediation, for example, of polycyclic aromatic hydrocarbons (HPA's). Considering the biome mangrove interface fragile and critical bordering the ocean, this study characterizes the native microbiota mangrove potential biodegradability of HPA's using a biomarker for molecular detection and assessment of bacterial diversity by PCR in areas under the influence of oil companies in the Basin Petroleum Geology Potiguar (BPP). We chose PcaF, a metabolic enzyme, to be the molecular biomarker in a PCR-DGGE detection of prokaryotes that degrade HPA s. The PCR-DGGE fingerprints obtained from Paracuru-CE, Fortim-CE and Areia Branca-RN samples revealed the occurrence of fluctuations of microbial communities according to the sampling periods and in response to the impact of oil. In the analysis of microbial communities interference of the oil industry, in Areia Branca-RN and Paracuru-CE was observed that oil is a determinant of microbial diversity. Fortim-CE probably has no direct influence with the oil activity. In order to obtain data for better understanding the transport and biodegradation of HPA's, there were conducted in silico studies with modeling and simulation from obtaining 3-D models of proteins involved in the degradation of phenanthrene in the transport of HPA's and also getting the 3-D model of the enzyme PcaF used as molecular marker in this study. Were realized docking studies with substrates and products to a better understanding about the transport mechanism and catalysis of HPA s

Proteínas SAG1, SAG2, SAG3 de toxoplasma gondii como perspectivas para o desenvolvimento de protótipo vacinal contra a toxoplasmose

Toxoplasmosis is a zoonosis of worldwide distribution caused by the protozoan Toxoplasma gondii, triggering dangerous complications in immunocompromised patients and pregnant women, as well as having great economic impact for the livestock. So far the control of toxoplasmosis is made primarily by chemotherapy. However, most drugs used routinely have some limitations. In order to control this disease, several research groups, including ours, has been working to develop a medical-veterinary vaccine based on parasite antigens, vectors and protocols of immunization. In this study were implemented and standardized methodologies for amplification and cloning of recombinant immunogens in the system for the development of a prototype vaccine, based on the surface antigens of T. gondii and recombinant adenovirus encoding these antigens. Genes encoding BAG1, GRA2 and SAG1 proteins were amplified. We established a strategy for cloning SAG1, SAG2, SAG3 and TgAMA1- genes in recombinant system. The genes encoding SAG1 and SAG2 were cloned and their sequences showed high similarity with sequences from GenBank. The virtual translation of these proteins showed polymorphisms in the amino acid sequence, which can be correlated with levels of antigenicity. Simultaneously, the adenovirus encoding the SAGs (HAdSAGs) were expanded, purificated and characterizated. Immunization of C57bl/6 mice, using viral supernatant was not enought to elicit immune responses at high levels, being required HAdSAGs titration for future immunizations. Therefore, this study allowed the cloning of the two genes important for the development of a prototype vaccine. Besides, implementations methodologies that permit advancements in the development of a vaccine against toxoplasmosis using adenovirus to express proteins of the parasite

Extração, caracterização e atividades biológicas de proteínas da espécie cnidoscolus urens (L.) Arthur

The extraction, chemical and structural characterization of a wide variety of compounds derived from plants has been a major source of bioactive molecules. Several proteases have been isolated in the plant kingdom, with numerous pharmacological and biotechnological applications. Among the proteases isolated from plants, are the fibrinogenolytic, with relevant application in the treatment of disorders in the coagulation cascade, in addition to potential use as a tool in clinical laboratories. In this study, in addition to evaluating the effects of the protein extract of Cnidoscolus urens (L.) Arthur (Euphorbiaceae) in the coagulation cascade also investigates the presence of antimicrobial activity and characterizes the proteolytic activity detected in this extract, aiming to determine their potential pharmacological and biotechnological application. In this way, crude protein extracts obtained from the leaves of C. urens in Tris-HCl 0.05M, NaCl 0.15M, pH 7.5, were precipitated in different concentrations of acetone, and assessed for the presence of proteolytic activity in azocaseína and fibrinogen. The most active fraction (F1.0) in these tests was chosen for assessment of biological activity and biochemical characterization. The Aα chain and Bβ of fibrinogen were completely cleaved at a concentration of 0.18 μg/μL of protein fraction in 4 minutes. Fibrinogenolytic activity presented total inhibition in the presence of E-64 and partial in the presence of EDTA. The fraction demonstrated coagulant activity in plasm and reduced the APTT, demonstrating acting on the factors coagulation of the intrinsic pathway and common, not exerting effects on the PT. Fibrinolytic activity on plasma clot was detected only in SDS-PAGE in high concentrations of fraction, and there were no defibrinating. Although several proteases isolated from plants and venomous animals are classically toxic, the fraction F1.0 of C. urens not expressed hemorrhagic nor hemolytic activities. Fraction F1.0 also showed no antimicrobial activity. In proteolytic activity on the azocasein, the optimal pH was 5.0 and optimum temperature of 60ºC. The enzyme activity has been shown to be sensitive to the presence of salts tested, with inhibition for all compounds. The surfactant triton did not influence the enzyme activity, but the tween-20 and SDS inhibited the activity. In the presence of reducing agents increase in enzyme activity occurred, a typical feature of enzymes belonging to the class of cysteine proteases. Several bands with proteolytic activity were detected in zymogram, in the region of high-molecular-weight, which were inhibited by E-64. In this study, we found that C. urens presents in its constitution cysteine proteases with fibrinogenolytic and procoagulant activity, which may be isolated, with potential application in treatment of bleeding disorders, thrombolytic and clinical laboratory

Aplicação de técnicas de aprendizado de máquina no reconhecimento de classes estruturais de proteínas

Nowadays, classifying proteins in structural classes, which concerns the inference of patterns in their 3D conformation, is one of the most important open problems in Molecular Biology. The main reason for this is that the function of a protein is intrinsically related to its spatial conformation. However, such conformations are very difficult to be obtained experimentally in laboratory. Thus, this problem has drawn the attention of many researchers in Bioinformatics. Considering the great difference between the number of protein sequences already known and the number of three-dimensional structures determined experimentally, the demand of automated techniques for structural classification of proteins is very high. In this context, computational tools, especially Machine Learning (ML) techniques, have become essential to deal with this problem. In this work, ML techniques are used in the recognition of protein structural classes: Decision Trees, k-Nearest Neighbor, Naive Bayes, Support Vector Machine and Neural Networks. These methods have been chosen because they represent different paradigms of learning and have been widely used in the Bioinfornmatics literature. Aiming to obtain an improvment in the performance of these techniques (individual classifiers), homogeneous (Bagging and Boosting) and heterogeneous (Voting, Stacking and StackingC) multiclassification systems are used. Moreover, since the protein database used in this work presents the problem of imbalanced classes, artificial techniques for class balance (Undersampling Random, Tomek Links, CNN, NCL and OSS) are used to minimize such a problem. In order to evaluate the ML methods, a cross-validation procedure is applied, where the accuracy of the classifiers is measured using the mean of classification error rate, on independent test sets. These means are compared, two by two, by the hypothesis test aiming to evaluate if there is, statistically, a significant difference between them. With respect to the results obtained with the individual classifiers, Support Vector Machine presented the best accuracy. In terms of the multi-classification systems (homogeneous and heterogeneous), they showed, in general, a superior or similar performance when compared to the one achieved by the individual classifiers used - especially Boosting with Decision Tree and the StackingC with Linear Regression as meta classifier. The Voting method, despite of its simplicity, has shown to be adequate for solving the problem presented in this work. The techniques for class balance, on the other hand, have not produced a significant improvement in the global classification error. Nevertheless, the use of such techniques did improve the classification error for the minority class. In this context, the NCL technique has shown to be more appropriated

Recuperação e purificação de proteínas do soro de queijo tipo coalho usando cromatografia de troca iônica e interação hidrofóbica em leito na forma expandida

Expanded Bed Adsorption plays an important role in the downstream processing mainly for reducing costs as well as steps besides could handling cells homogenates or fermentation broth. In this work Expanded Bed Adsorption was used to recover and purify whey proteins from coalho cheese manufacture using Streamline DEAE and Streamline SP both ionic resins as well as a hydrophobic resin Streamline Phenyl. A column of 2.6 cm inner diameter with 30 cm in height was coupled to a peristaltic pump. Hydrodynamics study was carried out with the three resins using Tris-HCl buffer in concentration of 30, 50 and 70 mM, with pH ranging from 7.0 to 8.0. In this case, assays of the expansion degree as well as Residence Time Distribution (RTD) were carried out. For the recovery and purification steps, a whey sample of 200 mL, was submitted to a column with 25mL of resin previously equilibrated with Tris/HCl (50 mM, pH 7.0) using a expanded bed. After washing, elution was carried out according the technique used. For ionic adsorption elution was carried out using 100 mL of Tris/HCl (50 mM, pH 7.0 in 1M NaCl). For Hydrophobyc interaction elution was carried out using Tris/HCl (50 mM, pH 7.0). Adsorption runs were carried out using the three resins as well as theirs combination. Results showed that for hydrodynamics studies a linear fit was observed for the three resins with a correlation coefficient (R2) about 0.9. In this case, Streamline Phenyl showed highest expansion degree reaching an expansion degree (H0/H) of 2.2. Bed porosity was of 0.7 when both resins Streamline DEAE and Streamline SP were used with StremLine Phenyl showing the highest bed porosity about 0.75. The number of theorical plates were 109, 41.5 and 17.8 and the axial dipersion coefficient (Daxial) were 0.5, 1.4 and 3.7 x 10-6 m2/s, for Streamline DEAE, Streamline SP and Streamline Phenyl, respectively. Whey proteins were adsorved fastly for the three resins with equilibrium reached in 10 minutes. Breakthrough curves showed that most of proteins stays in flowthrough as well as washing steps with 84, 77 and 96%, for Streamline DEAE, Streamline SP and Streamline Phenyl, respectively. It was observed protein peaks during elution for the three resins used. According to these peaks were identified 6 protein bands that could probably be albumin (69 KDa), lactoferrin (76 KDa), lactoperoxidase (89 KDa), β-lactoglobulin (18,3 KDa) e α-lactoalbumin (14 KDa), as well as the dimer of beta-lactoglobulin. The combined system compound for the elution of Streamline DEAE applied to the Streamline SP showed the best purification of whey proteins, mainly of the α-lactoalbumina

Expressão imuno-histoquímica das proteínas MMP-9, VEGF e FVW em lesões centrais e periféricas de células gigantes

Central giant cell lesion (CGCL) and peripheral giant cell lesion (PGCL) of the jaws have a distinct clinical behavior, although they share histopathologic features. It is still unclear whether these clinical differences are supported by a distinct pattern of immunoexpression of markers for multinucleated giant cells (GC) and mononuclear cells (MC). The purpose of this study was to compare the immunohistochemical expression of VEGF, MMP-9 in CG and MC and measure the vascularization by vWF to check whether there are differences in expression of these biomarkers between CGCL and PGCL. Paraffin wax blocks of 20 cases of LCCG and 20 LPCG were retrieved. MMP-9 immunoreactivity was greater in the CM of PGCL compared to VEGF (p<0.05). VEGF expression was greater in the CM of CGCL compared to PGCL (p<0.05) and it was greater in the overall expression of CGCL compared to PGCL (p<0.05). Vascularity was quantified by microvascular counting (MVC). MVC was greater in the PGCL compared CGCL (p<0.05). MMP-9 showed a greater tendency of expression in CGCL, though was not significant (p>0.05). We tested correlation between the proteins studied in each group and found a significant negative correlation between VEGF and vWF in CGCL (p<0.05). These results suggest that there are differences in the expression of VEGF in CM and overall expression between the lesions, although no statistically significant difference in the overall expression of the MMP-9. Then, there was a trend in increased expression of MMP-9 and VEGF in CGCL, possibly by the involvement of both proteins in osteoclastogenesis. Additionally, the results of this study indicate a higher degree of vascularization in PGCL compared to CGCL, fact that can be directly linked to the reactive nature of the PGCL, where the inflammatory process with its rich angiogenesis contributes significantly to these findings.

Expressão imuno-hitoquímica das proteínas RANK, RANKL e OPG em cistos radiculares e cistos dentígeros

Receptor ativador nuclear κappa B (RANK), ligante do receptor ativador nuclear κappa B (RANKL) e osteoprotegerina (OPG) são membros da família do fator de necrose tumoral relacionados com o metabolismo ósseo. A formação, diferenciação e atividade dos osteoclastos são reguladas por estas três proteínas. RANK é um receptor transmembrana presente em diversos tipos celulares, principalmente em células de linhagem macrofágica, linfócitos, células dendríticas e fibroblastos e quando ativado pelo seu ligante, RANKL, promove a diferenciação e ativação de células osteoclásticas responsáveis pelo processo de reabsorção óssea. A OPG impede a ligação RANK/RANKL atuando como um receptor inibitório para a atividade osteolítica. O objetivo deste estudo foi comparar a expressão imuno-histoquímica destes biomarcadores em cistos radiculares (n=20) e cistos dentígeros (n=20). A expressão imuno-histoquímica destes marcadores foi avaliada no epitélio e na cápsula dos cistos por escores e percentuais médios de imunomarcação. Para o epitélio, a análise semi-quantitativa revelou um padrão similar dos escores de imunomarcação de RANK, RANKL e OPG nas lesões, não havendo diferença estatística significante (p=0.589, p=0.688, p=0.709, respectivamente). Para a cápsula cística a análise quantitativa, mostrou diferença estatística significante entre os percentuais médios de imunomarcação do RANK e RANKL (p=0,001 e p=0,005, respectivamente) nos cistos. A correlação dos escores de imunomarcação de RANKL e OPG no epitélio do CR e do CD revelou diferença estatística significante (p=0,029, p=0,003, respectivamente). No epitélio dos CRs e dos CDs observou-se uma maior imunoexpressão da OPG comparada a do RANKL. Os resultados apontam a presença de RANK, RANKL e OPG nos cistos radiculares e cistos dentígeros, sugerindo a atuação destas proteínas no desenvolvimento e expansão das lesões no osso adjacente

Análise comparativa da imunoexpressão das proteínas hmlh1 e hmsh2 em carcinomas epidermóides de lábio inferior e queilites actínicas com graus variados de displasia

Lip squamous cell carcinoma (SCC) may develop from a premalignant condition, actinic cheilitis (AC) in 95% of the cases. Both premalignant and neoplastic lip diseases are caused mainly by chronic exposure to the ultraviolet component of solar radiation, especially UVB. This exposure causes disruption of the cell cycle and damage to DNA repair systems, like mismatch repair, altering proteins repair as hMLH1 and hMSH2. This research aimed to investigate the immunohistochemical expression of hMLH1 and hMSH2 proteins in lower lip SCCs and ACs, providing additional information about carcinogenesis of the lower lip. The sample consisted 40 cases of ACs and 40 cases of lower lip SCCs. Histological sections of 3 μm were submitted to immunoperoxidase method, for immunohistochemical analysis of lesions were counted in 1000 cells (positive and negative), data were evaluated both in absolute numbers and percentage of immunostained cells, the latter by assigning scores. Associations of the variables and comparative analysis of biomarker expression were performed by Fisher s exact and Pearson s chi-square, "t" student, one-way ANOVA, Mann- Whitney e Kruskal-Wallis tests. The level of significance was 5%. It was found that, in lower lip SCC, the mean of the proteins was higher in female patients (hMLH1= 369,80 + 223,98; hMHS2 = 534,80 + 343,62), less than 50 years old (hMLH1 = 285,50 + 190,65; hMHS2 = 540,00 + 274,79) and classified as low-grade malignancy (hMLH1 = 264,59 + 179,21; hMHS2 = 519,32 + 302,58), in these data only to sex, for hMLH1 protein, was statistically significant (p=0.034). Comparing the different lesions, we observed that for both hMLH1 and hMSH2 protein, the average of positive epithelial cells decreased as the lesion was graded at later stages. The ACs classified without dysplasia or mild dysplasia had the highest average of immunostained cells (hMLH1 = 721.23 + 88.116; hMHS2 = 781.50 + 156.93). The ACs classified as moderate or severe dysplasia had intermediate values (hMLH1 = 532,86 + 197,72; hMHS2 = 611,14 + 172,48) and SSCs of the lower lip had the lowest averages (hMLH1 = 255,03 + 199,47; hMHS2 = 518,38 + 265,68). There was a statistically significant difference between groups (p<0.001). In conclusion, our data support the hypothesis that changes in immunoexpression of these proteins is related to the process of carcinogenesis of the lower lip

Expressão Imuno-histoquímica das proteínas GLUT-1 e HIF-1α em lesões vasculares de mucosa oral

The correct histological diagnosis of vascular lesions in the oral mucosa is critical, especially in defining the treatment and prognosis, as some vascular lesions show spontaneous involution and others do not show such behavior. This study analyzed the expression immunohistochemistry of human glucose transporter protein (GLUT-1), in oral benign vascular tumors and to reclassify such lesions according to with his immunoexpression. In addition, we evaluated the immunohistochemical expression of hypoxia-inducible factor 1 alpha (HIF-1α), the main transcription factor involved in cellular adaptation to hypoxia. We analyzed 60 cases of benign oral vascular tumors: 30 cases with histological diagnosis of HEM and 30 cases of oral pyogenic granuloma (PG). The results of this research showed that of the 30 lesions initially classified as HEM, only 7 showed immuno-positivity for GLUT-1, remaining with the initial diagnosis. The remaining 23 were reclassified as vascular malformation (VM) (13 cases) and PG (10 cases). All cases in the sample with an initial diagnosis of PG were negative for GLUT-1, demonstrating the accuracy of histological diagnosis of these lesions. Concerning to the immunoexpression of HIF-1α, the Mann-Whitney test revealed a statistically significant difference between the cases of GP and MV (p = 0.002), where the median of GP (m=78) was higher than the MV (m=53). Based on these results, this study showed that a histological diagnosis alone is not always sufficient for the correct diagnosis of oral HEM and that HIF-1α participates in the pathogenesis of vascular lesions

Análise imuno-histoquímica das proteínas VEGF e HIF-1α na doença periodontal

Periodontal disease is a complex inflammatory and infectious condition that immune host, front of the microbial aggressions, can lead to disease progression, resulting in tissue destruction. The tissue damage induces the release of regulatory molecules, which protective roles and / or destructive, including proteins VEGF (vascular endothelial growth factor vascular) and HIF-1 α (hypoxia-induced factor α -1). Thus, this study investigated, quantitatively and comparatively, the immunohistochemical expression of VEGF (vascular endothelial growth factor) and HIF-1 α (hypoxia induced factor 1-α), proteins involved in inflammation, angiogenesis and hypoxia, in human gingival tissues. Therefore, 75 samples of gingival tissues were examined. Thirty samples were chronic periodontitis, 30 with chronic gingivitis and 15 healthy gingival. After sections analysis, positives and negatives inflammatory and endothelial cells in the connective tissue were counted and converted into percentage. Data were statistically analyzed using Kruskal-Wallis test and Spearman correlation. The results showed that both proteins marked. It was observed higher immunoreactivity for HIF-1 α at chronic gingivitis and periodontitis specimens compared to healthy sites, however, no statistically significant differences were observed among them (p>0.05). The VEGF immunostaining showed similarity among the cases of periodontitis, gingivitis and healthy gingiva. Moderate and positive correlation statistically significant differences were verified for the expressions of VEGF and HIF-1α in gingival health (r = 0,529, p = 0.04). Thus, it can be conclude that possibly the route of HIF-1 α, is activated in periodontal disease may have involvement of the protein in pathogenesis and progression of disease, and that activation of VEGF, can be in addition to being triggered transcription by HIF-1 α may be also influenced by other additional factors such as the action of periodontal microorganisms endotoxins

Expressão imuno-histoquímica das proteínas IFN-γ e TGF-β1 em cistos radiculares e cistos dentígeros

Odontogenic cysts are pathologic cavities covered by odontogenic epithelium and filled by liquid, desquamated cells or other materials. The intraosseous lesions, such as radicular cyst and dentigerous cyst, present a potential of expansion capable of promoting the destruction of the surrounding osseous tissue. The mechanisms related to this process of expansion are the proliferation of cystic epithelium, the increase of the osmolarity of the cystic fluid and the synthesis of reabsorption factors such as IFN-γ and TGF-β1. The aim of this study was to evaluate and compare the immunohistochemical expression of IFN-γ and TGF-β1 between radicular cysts and dentigerous cysts in order to understand the role and behavior of these proteins in the expansion of these cysts. We selected 20 cases of radicular cyst and 20 cases of dentigerous cyst chosen from the files of UFRN s Laboratory of Oral Pathology. After analyzing the clinical data, the cases underwent the routine staining technique (HE) and immunohistochemistry for the appearance of IFN-γ and TGF-β1 in the epithelium and capsule of these cysts. The statistical analysis using the Mann-Whitney test revealed no statistically significant difference in immunoexpression of IFN-γ between the epithelium (p = 0.565) and capsules (p = 0.414) of radicular cysts and dentigerous cysts. Moreover, there was no statistically significant difference of immunoexpression of TGF-β1 between the epithelium (p = 0.620) and capsules (p = 0.056) of radicular cysts and dentigerous cysts. The Wilcoxon test revealed no statistically significant difference between IFN-γ and TGF-β1 imunoexpressions in the epithelium (p = 0.225) and capsules (p = 0.370) of radicular cysts. There was no statistically significant difference between IFN-γ and TGF-β1 imunoexpressions in the epithelium (p = 0.361) of dentigerous cysts. However, there was a statistically significant difference between IFN-γ and TGF-β1 immunoexpressions in the capsule (p = 0.001) of dentigerous cysts, being TGF-β1 the factor which presented the most significant immunoexpression. Given these results, we conclude that there was no difference in immunohistochemical expression of IFN-γ and TGF-β1 between radicular and dentigerous cysts and that TGF-β1 was more significant than the IFN-γ in the capsule of dentigerous cysts

Estudo da expressão imuno-histoquímica das proteínas MMP-9, MMP-13 e TIMP-1 em ameloblastomas e tumores odontogênicos ceratocistos

Ameloblastomas and keratocystic odontogenic tumors (KOT) represent odontogenic lesions that, despite their benign nature, are distinguished by a distinct biological behavior, characterized by locally aggressive growth and recurrent episodes. The gnathic bone resorption caused by the growth of these lesions is a key to the expansion of the same, both being mediated by osteoclastic cells like enzymatic activity of various matrix metalloproteinases (MMPs) factor. The expression of stimulatory factors and inhibitors of bone resorption has been correlated with the development of these lesions, with emphasis to some MMPs such as collagenases and gelatinases and tissue inhibitors of metalloproteinases (TIMPs), among others. Based on the premise that stimulatory and inhibitory factors of osteolytic processes can be decisive for the growth rate of intraosseous odontogenic lesions, this experiment evaluated the immunoreactivity of MMP-9, -13 and TIMP-1 protein in the epithelium and mesenchyme of ameloblastoma and the KOT specimens, by a quantitative analysis of the immunoreactivity cells. Statistical analysis was performed using the Mann-Whitney and Wilcoxon tests with a significance level set at 5 %. Immunohistochemical expression of MMP-9, -13 and TIMP-1 was observed in 100% of cases both in the epithelium and in mesenchyme. The immunoreactivity in the epithelium of KOT and ameloblastomas revealed a predominance of score 3 for MMP-9 (p=0.382) and MMP-13 (p=0.069) and no statistically significance for TIMP-1, the latter being significantly higher immunoreactivity in ameloblastomas. In the mesenchyme, there was a higher score immunoreactivity of MMP-13 (p=0.031) in ameloblastomas in relation to KOT, whereas for MMP-9 and TIMP-1 no statistically significant difference (p=0.403 was observed, p=1.000). The calculation of the ratio of scores revealed expression of proteins in general, similarity of the lesions, a significant predominance of equal expression of TIMP-1 and MMP-9 was observed only in the epithelium of ameloblastoma. The marked immunostaining of MMP-9 , MMP-13 and TIMP-1 in epithelium and mesenchyme of the lesion indicate that these proteins involved in ECM remodeling required for tumor progression, however, specific differences in the expression of some of these proteins, are not sufficient to suggest differences in the biological behavior of ameloblastomas and KOTs

Estudo clínico-patológico do carcinoma epidermóide de língua e imunoistoquímico das proteínas BMP-2, BMPR-IA, BMPR-II e endoglina

Bone morphogenetic proteins (BMPs) are cytokines involved in proliferation and angiogenesis of many kind of human cancer. The present study analyzed the immunohistochemical expression of BMP-2, BMPR-II, BMPR-IA and endoglin (CD105) and their relationship with the biological behavior and local angiogenesis in tongue oral squamous cells carcinoma (SCC). The sample consisted of 25 cases of tongue SCC without metastasis, 25 tongue SCC with metastasis and 25 cases of Inflamatory Fibrous Hyperplasia (IFH).The histological grade of malignancy proposed by Bryne (1998), adapted by Miranda (2002) was used to classify all tongue SCC cases. Score 0 was attributed to absent-weak immunoexpression and score 1 for strong immunostaning and pattern of distribution was focal or diffuse. Microvessel counts (MVC) was established for CD105. Most of the patients with tongue SCC was male. The principal age in tongue SCC without metastasis was over 65 years and in tongue SCC with metastasis was between 45-65 years. There were predominance of stage II in TNM and in the specimens with high-grade, independent of studied group. For BMP-2, 56% of tongue SCC without metastasis and 72% tongue SCC with metastasis exhibited score 1 while the IFH showed secore 0 in 72% of the cases, with statistical association (p=0,007). Considering the BMPR-II, 52% of tongue SCC without metastasis exhibited score 0; 56% tongue SCC with metastasis and 60% IFH showed score 1. The majority cases of BMPR-IA demonstrated score 1 and 100% of CD105 exhibited strong immunoexpression in tongue SCC. Regarding the pattern distribution, it was noted a tendency to diffuse pattern for the proteins in all groups. The means of MVC were similar in tongue SCC without metastasis (32,91) and in tongue SCC with metastasis (32,05), however existed statistical difference with IFH (p<0,001). There was statistical association of BMP-2 expression with BMPR-II (p=0,008), BMPR-IA (p=0,006) and CD105 (p=0,046). An association between TNM and BMP-2 immunoexpression and their receptors was not detected, nevertheless this association was found with MVC (p=0,047) whose averages were higher for the stages II (35,97) e IV (35,69). No association between histological grading and these proteins was observed. This study suggests that the superexpression of BMP-2 signaling pathways acts on cell proliferation in tongue SCC and can be implicated with more invasive potential. Additionaly, the CD105 is a potent biological marker of neovascularization in this neoplasm and their association with BMP-2 and BMPR-IA receptor, showed that this type of cancer in BMP-2 is presented as pro-angiogenic in the metastatic process