RESUMO Aproximadamente 80% da população mundial sofre algum tipo de dor nas costas em alguma fase de vida, sendo que em 10% dessa população, as dores acarretam incapacidade crônica, deflagrando alto custo de tratamento desses pacientes, além de comprometer as habilidades de trabalho e convívio social desses indivíduos. As estratégias de tratamento atuais incluem o procedimento cirúrgico por ressecção do disco intervertebral degenerado, bloqueio de raízes nervosas e fisioterapia. Entretanto, tais tratamentos apenas aliviam os sintomas e não impedem que ocorra a degeneração de discos intervertebrais. Portanto, novas estratégias terapêuticas têm surgido e incluem a manipulação de células com o objetivo de recuperar o disco degenerado. No presente artigo, serão discutidas as diferentes possibilidades alternativas de terapias celulares no processo de regeneração discal, caracterizando um estudo descritivo da medicina translacional que envolve aspectos clínicos de novas alternativas de tratamento e o conhecimento de áreas básicas de pesquisa como biologia celular e molecular. Nível de evidência V; Opinião do Especialista.

ABSTRACT Approximately 80% of the world population experiences some type of back pain at some point in their life, and in 10% of this population the pain causes chronic disability resulting in a high cost for the treatment of these patients, in addition to compromising their work and social interaction abilities. Current treatment strategies include the surgical procedure for degenerated intervertebral disc resection, the nerve root block and physiotherapy. However, such treatments only relieve symptoms and do not prevent the degeneration of intervertebral discs. Therefore, new therapeutic strategies have emerged and include manipulating cells to recover the degenerated disc. This article will discuss the possible cell therapy alternatives used in the disc regeneration process, featuring a descriptive study of translational medicine that involves clinical aspects of new treatment alternatives and knowledge of basic research areas, such as cellular and molecular biology. Level of evidence V; Expert Opinion.

RESUMEN Aproximadamente 80% de la población mundial sufre algún tipo de dolor de espalda en alguna etapa de la vida, y en 10% de esa población, los dolores causan incapacidad crónica, deflagrando alto costo de tratamiento de esos pacientes, además de comprometer las habilidades laborales y convivencia social de esos individuos. Las estrategias de tratamiento actuales incluyen el procedimiento quirúrgico para la resección del disco intervertebral degenerado, bloqueo de las raíces nerviosas y fisioterapia. Entretanto, tales tratamientos solo alivian los síntomas y no impiden que ocurra la degeneración de discos intervertebrales. Por lo tanto, han surgido nuevas estrategias terapéuticas e incluyen la manipulación de células con el objetivo de recuperar el disco degenerado. En el presente artículo se discutirán las diferentes posibilidades alternativas de las terapias celulares en el proceso de regeneración discal, caracterizando un estudio descriptivo de la medicina traslacional que involucra aspectos clínicos de nuevas alternativas de tratamiento y conocimiento de áreas básicas de investigación como biología celular y molecular. Nivel de evidencia V; Opinión del especialista.

RESUMO Objetivo Investigar os genes que podem estar relacionados aos mecanismos que modulam a heparanase-1. Métodos A análise foi realizada na Universidade Federal de São Paulo, utilizando dados fornecidos por: The Cancer Genome Atlas, University of California Santa Cruz Genome Browser, Kyoto Encyclopedia of Genes and Genomes Pathway Database, Database for Annotation, Visualization and Integrated Discovery Bioinformatics Database e os softwares cBioPortal e Ingenuity Pathway Analysis. Resultados Usando o perfil de expressão de RNA mensageiro de diferentes subtipos moleculares de câncer de mama, propusemos que a heparanase-1 esteve correlacionada com a progressão tumoral. Além disso, os genes analisados apresentaram coexpressão com heparanase-1. Os resultados mostraram que a heparanase-1 coexpressa com proteína adaptadora 1 da fosfoinositídeo 3-quinase, lectina 7 tipo Ig de ligação ao ácido siálico e receptor 1 do tipo imunoglobulina associado a leucócitos, estes genes estão diretamente relacionados à evasão do sistema imune durante a progressão do câncer de mama. Além disso, a catepsina L foi coexpressa com a heparanase-1 e transformou a forma inativa da heparanase-1 em heparanase-1 ativa, desencadeando o remodelamento da matriz extracelular, o que contribuiu para a interação do tumor com o ambiente tumoral. Conclusão A análise utilizando bioinformática fornece evidências de possíveis mecanismos relacionados ao desenvolvimento do câncer de mama. Genes de evasão do sistema imune foram coexpressos com a heparanase-1, uma enzima relacionada à progressão tumoral.

ABSTRACT Objective To investigate the possible genes that may be related to the mechanisms that modulate heparanase-1. Methods The analysis was conducted at Universidade Federal de São Paulo, on the data provided by: The Cancer Genome Atlas, University of California Santa Cruz Genome Browser, Kyoto Encyclopedia of Genes and Genomes Pathway Database, Database for Annotation, Visualization and Integrated Discovery Bioinformatics Database and the softwares cBioPortal and Ingenuity Pathway Analysis. Results Using messenger RNA expression pattern of different molecular subtypes of breast cancer, we proposed that heparinase-1 was co-related with its progression. In addition, genes that were analyzed presented co-expression with heparanase-1. The results that showed that heparanase-1 co-expressed with phosphoinositide 3-kinase adapter protein 1, sialic acid-binding immunoglobulin-like lectin 7, and leukocyte-associated immunoglobulin-like receptor 1 are directed related with immune system evasion during breast cancer progression. Furthermore, cathepsin L was co-expressed with heparanase-1 and transformed inactive heparanase-1 form into active heparanase-1, triggering extracellular matrix remodeling, which contributes to enhanced tumor-host interaction of the tumor. Conclusion The signaling pathway analysis using bioinformatics tools gives supporting evidence of possible mechanisms related to breast cancer development. Evasion genes of the immune system co-expressed with heparanase-1, a enzyme related with tumor progression.

Resumo Objetivo Investigar a ação da testosterona (T) isolada ou associada ao benzoato de estradiol (EB) na proliferação e apoptose de mamas de ratas ovariectomizadas. Métodos Um total de 48 ratas Wistar castradas foram divididas em 6 grupos, e cada um foi submetido a um dos seguintes tratamentos durante 5 semanas: 1) controle; 2) BE 50 mcg/dia + T 50mcg/dia; 3) T 50 mcg/dia; 4) BE 50 mcg + T 300mcg/dia; e) T 300 mcg/dia; e f) BE 50 mcg/dia. Após o tratamento, o tecido mamário foi submetido a análise histológica e avaliação de imunoexpressão de marcadores de proliferação (antígeno nuclear de células proliferantes, PCNA) e apoptose (caspase-3). Resultados Houve diferença estatisticamente significante entre os grupos com relação às microcalcificações e à atividade secretora, com maior prevalência nos grupos tratados com BE. Não houve diferença entre os grupos quanto à atrofia, mas houve maior prevalência de atrofia nos grupos que receberam T versus os que receberam BE+ T. Houve diferença entre os grupos quanto ao ANCP (p= 0,028), com maior expressão no grupo BE+ T 300 mcg/dia. Com relação à caspase-3, não houve diferença entre os grupos, mas, no grupo BE+ T 300 mcg/dia, a expressão foi maior do que no grupo de T isolada. Conclusão A T isolada não apresentou efeito proliferativo do tecido mamário, contrariamente ao EB. A T em associação ao EB pode diminuir ou não a proliferação, a depender da dose de T.

Abstract Objective To investigate the action of testosterone (T), isolated or associated with estradiol benzoate (EB), on the proliferation markers and apoptosis of breasts of ovariectomized rats. Methods A total of 48 castrated female Wistar rats were divided into 6 groups, and each of them were submitted to one of the following treatments for 5 weeks: 1) control; 2) EB 50 mcg/day + T 50 mcg/day; 3) T 50mcg/day; 4) EB 50 mcg +T 300 mcg/day; 5) T 300 mcg/day; and 6) EB 50 mcg/day. After the treatment, the mammary tissue was submitted to a histological analysis and immunoexpression evaluation of proliferation markers (proliferating cell nuclear antigen, PCNA) and apoptosis (caspase-3). Results There was a statistically significant difference among the groups regarding microcalcifications and secretory activity, with higher prevalence in the groups treated with EB. There was no difference among the groups regarding atrophy, but a higher prevalence of atrophy was found in the groups that received T versus those that received EB +T. There was a difference among the groups regarding the PCNA (p = 0.028), with higher expression in the group submitted to EB +T 300 mcg/day. Regarding caspase-3, there was no difference among the groups; however, in the group submitted to EB +T 300 mcg/day, the expression was higher than in the isolated T group. Conclusion Isolated T did not have a proliferative effect on the mammary tissue, contrary to EB. Testosterone in combination with EB may or may not decrease the proliferation, depending on the dose of T.

Abstract Background: Mohs micrographic surgery is a surgical technique for the treatment of nonmelanoma skin cancer. Surgery begins by removing the visible tumor before excision of the tissue specimens for evaluation of the tumor margins. Objectives: To present a new way to evaluate the material obtained from debulking, by horizontal histological analysis of the fragment. Methods: Descriptive retrospective cross-sectional study based on the medical records and histological lamellae of patients with primary basal cell carcinomas smaller than 1.5 cm submitted to Mohs micrographic surgery and who had the visible tumor analyzed by horizontal histological sections. Results: The sample evaluated included 16 patients with lesions located on the face. Comparing the histopathological examinations of incisional biopsy in vertical sections and debulking in horizontal sections, there was agreement in seven cases. The histological analysis performed in horizontal sections allowed identification of the tumor site in 13 cases, and the relation between tumor and margin showed that in 11 cases, the lateral margin was compromised.Study limitations: The technique was better-applied in lesions smaller than 2 cm. Conclusion: Horizontal histological analysis of debulking has advantages for Mohs surgery, since it allows visualization of almost all tumor extension in the same view plane of the dermatoscopy, allowing better definition of the histological subtype, tumor site, and tumor/margin of lesions less than 1.5 cm.

Resumo Objetivo Analisar os efeitos do estrogênio isolado ou em combinação com progestogênios e tibolona (TIB) na expressão das metaloproteinases 2 e 9 da matriz extracelular (MMP-2 e MMP-9), da perlecan e da heparanase (HPSE) das paredes vasculares das artérias carótidas. Métodos Trinta ratas Wistar ovariectomizadas com 250 dias de idade foram tratadas oralmente por 5 semanas com: a) 1 mg/kg de benzoato de estradiol (EB); b) EB + 0,2 mg/kg de acetato de medroxiprogesterona (MPA); c) EB + 0,2mg/kg de acetato de noretisterona (NETA); d) EB + 2 mg/kg de didrogesterona (DI); e) 1 mg/kg de TIB; f) placebo (CTR). Após o tratamento, a expressão de mRNA para MMP-2, MMP- 9, e HPSE foi analisada por reação em cadeia da polimerase (RCP) em tempo real, e a expressão de MMP-2, MMP-9, inibidor tecidual de metaloproteinase 2 (TIMP-2), e de perlecan foi quantificado por imunohistoquímica em artérias carótidas. Resultados Os grupos apresentaram diferenças significativas na expressão do mRNA HPSE (p = 0,048), sendo maiores nos grupos EB, EB + MPA e TIB. Não houve diferença estatisticamente significativa nas expressões de mRNA MMP-2 ou MMP-9. A expressão imunohistoquímica de MMP-2, TIMP-2, MMP-9, HPSE e perlecan não mostrou diferenças entre os grupos. Conclusão O estradiol isolado ou associado ao tratamento com MPA e TIB pode aumentar a expressão de mRNA HSPE nas paredes das artérias carótidas em ratas ovariectomizadas.

Abstract Objective To analyze the effects of estrogen alone or in combination with progestogens and tibolone (TIB) on the expression of the extracellular matrix metalloproteinases 2 and 9 (MMP-2 and MMP-9), of perlecan, and of heparanase (HPSE) of the vascular walls of the carotid arteries. Methods A total of 30 250-day-old ovariectomized Wistar rats were orally treated for 5 weeks with: a) 1 mg/kg of estradiol benzoate (EB); b) EB + 0.2 mg/kg of medroxyprogesterone acetate (MPA); c) EB + 0.2mg/kg of norethisterone acetate (NETA); d) EB + 2 mg/kg of dydrogesterone (DI); e) 1 mg/kg of TIB; f) placebo (CTR). Following treatment, the expression of mRNA for MMP-2, MMP-9, and HPSE was analyzed by realtime polymerase chain-reaction (PCR), and the expression of MMP-2, of MMP-9, of tissue inhibitor of metalloproteinase 2 (TIMP-2), and of perlecan was quantified by immunohistochemistry in the carotid arteries. Results The groups showed significant differences on mRNA HPSE expression (p = 0.048), which was higher in the EB, EB + MPA, and TIB groups. There was no statistically significant difference in mRNA MMP-2 or MMP-9 expression. The immunohistochemical expression of MMP-2, of TIMP-2, of MMP-9, of HPSE, and of perlecan showed no differences between groups. Conclusion Estradiol alone or associated with MPA and TIB treatment can increase mRNA HSPE expression of the walls of the carotid arteries in ovariectomized rats.

RESUMO Objetivo: Avaliar os níveis de fatores inflamatórios nos discos intervertebrais (interleucina 6) e proteinase (catepsina B) em pacientes com doença degenerativa de disco intervertebral, além de verificar os níveis séricos de interleucina 6, ácido hialurônico e atividade sérica da catepsina B. Métodos: Foi realizado exame imuno-histoquímica dos discos intervertebrais de pacientes com doença degenerativa e fratura da coluna (Grupo Controle) e análise do plasma de pacientes com doença degenerativa de disco intervertebral. Como controle, foram utilizados plasma de pacientes com fraturas, além de indivíduos saudáveis. Resultados: Interleucina 6 e catepsina B sugerem relação com a fisiopatologia da doença degenerativa de disco intervertebral, uma vez que os níveis de ambos foram maiores nos discos de pacientes com doença degenerativa de disco intervertebral. Interleucina 6 e catepsina B não representam bons biomarcadores da doença degenerativa do disco intervertebral, já que também encontram níveis aumentados em plasma de pacientes com fratura. Conclusão: O ácido hialurônico é um possível biomarcador de doença degenerativa de disco intervertebral, porque os níveis de ácido hialurônico foram maiores apenas em plasma de pacientes com doença degenerativa de disco intervertebral.

ABSTRACT Objective: To evaluate intervertebral disc levels of inflammatory factor (interleukin 6) and proteinase activity (cathepsin B) in patients with a degenerative disease and serum levels of interleukin 6, serum cathepsin B activity and hyaluronic acid biomarkers. Methods: We conducted immunohistochemistry studies of intervertebral discs to analyze interleukin 6 and cathepsin B levels of patients with degenerative disease and spine fracture (Control Group) and to measure hyaluronic acid, interleukin 6 and cathepsin B activity from sera of intervertebral disc degeneration patients, fracture patients, and healthy individuals. Results: Interleukin 6 and cathepsin B seem to be related with physiopathology of intervertebral disc degeneration, since the levels of both were higher in discs of patients with intervertebral disc degeneration. Interleukin 6 and cathepsin B do not represent good biomarkers of degenerative intervertebral disc disease, since the level of such compounds is increased in the plasma of patients with fractures. Conclusion: Hyaluronic acid can be a biomarker for intervertebral disc degeneration, because hyaluronic acid levels were higher only in sera of patients with intervertebral disc degeneration.

ABSTRACT Purpose: The present study evaluates chondroitin sulfate (CS) and heparan sulfate (HS) in the urine and hyaluronic acid (HA) in the plasma of patients with prostate cancer before and after treatment compared to a control group. Materials and Methods: Plasma samples were used for HA dosage and urine for quantification of CS and HS from forty-four cancer patients and fourteen controls. Clinical, laboratory and radiological information were correlated with glycosaminoglycan quantification by statistical analysis. Results: Serum HA was significantly increased in cancer patients (39.68 ± 30.00 ng/ mL) compared to control group (15.04 ± 7.11 ng/mL; p=0.004) and was further increased in high-risk prostate cancer patients when compared to lower risk patients (p = 0.0214). Also, surgically treated individuals had a significant decrease in seric levels of heparan sulfate after surgical treatment, 31.05 ± 21.01 μg/mL (before surgery) and 23.14 ± 11.1 μg/mL (after surgery; p=0.029). There was no difference in the urinary CS and HS between prostate cancer patients and control group. Urinary CS in cancer patients was 27.32 ± 25.99 μg/mg creatinine while in the men unaffected by cancer it was 31.37 ± 28.37 μg/mg creatinine (p=0.4768). Urinary HS was 39.58 ± 32.81 μg/ mg creatinine and 35.29 ± 28.11 μg/mg creatinine, respectively, in cancer patients and control group (p=0.6252). Conclusions: Serum HA may be a useful biomarker for the diagnosis and prognosis of prostate cancer. However, urinary CS and HS did not altered in the present evaluation. Further studies are necessary to confirm these preliminary findings.

Abstract: Surgical excision of basal cell carcinoma with minimum margins requires serial assessment of layers by frozen histopathology in the case of Mohs micrographic surgery. Evaluation of presurgical tumor margins by in vivo reflectance confocal microscopy is a potential alternative. We selected 12 basal cell carcinoma lesions that were analyzed by confocal microscopy to define margins. The lesions were excised by Mohs surgery. Six tumors showed negative margins in the first phase of Mohs micrographic surgery. We concluded that reflectance confocal microscopy can be useful in the preoperative definition of basal cell carcinoma margins.

Abstract: Background: Heparanase is an enzyme that cleaves heparan sulfate chains. Oligosaccharides generated by heparanase induce tumor progression. Basal cell carcinoma and squamous cell carcinoma comprise types of nonmelanoma skin cancer. Objectives: Evaluate the glycosaminoglycans profile and expression of heparanase in two human cell lines established in culture, immortalized skin keratinocyte (HaCaT) and squamous cell carcinoma (A431) and also investigate the expression of heparanase in basal cell carcinoma, squamous cell carcinoma and eyelid skin of individuals not affected by the disease (control). Methods: Glycosaminoglycans were quantified by electrophoresis and indirect ELISA method. The heparanase expression was analyzed by quantitative RT-PCR (qRTPCR). Results: The A431 strain showed significant increase in the sulfated glycosaminoglycans, increased heparanase expression and decreased hyaluronic acid, comparing to the HaCaT lineage. The mRNA expression of heparanase was significantly higher in Basal cell carcinoma and squamous cell carcinoma compared with control skin samples. It was also observed increased heparanase expression in squamous cell carcinoma compared to the Basal cell carcinoma. Conclusion: The glycosaminoglycans profile, as well as heparanase expression are different between HaCaT and A431 cell lines. The increased expression of heparanase in Basal cell carcinoma and squamous cell carcinoma suggests that this enzyme could be a marker for the diagnosis of such types of non-melanoma cancers, and may be useful as a target molecule for future alternative treatment.

ABSTRACT Objective: To evaluate the structural and molecular changes in the extracellular matrix (ECM) during the process of intervertebral disc degeneration, using animal model. Methods: Wistar rats underwent intervertebral disc degeneration through 20-gauge needle puncture, and 360° rotation applied for 30 sec, representing the degenerated group, while control group was not submitted to this procedure. Histological parameters and expression of extracellular matrix molecules were evaluated in the 15th and 28th days after degenerative induction. Results: Fifteen days after the induction of intervertebral disc degeneration, significant changes were observed, such as reduction in the expression metalloprotease-9 (MMP9) and interleukins (IL-6 and IL-10). There was a significant increase in the expression of vascular endothelial growth factor (VEGF) and caspase-3. However, different alterations in the ECM were observed at 28 days, the level of collagen I, metalloprotease-2 (MMP2) and caspase-3 were enhanced. Furthermore, expression of heparanase isoforms (HPSE1 and HPSE2) mRNA were increased in the degenerative intervertebral disc. Conclusion: The different profiles of ECM molecules observed during the intervertebral disc degeneration suggest that molecular processes such as ECM remodeling, neovascularization, apoptosis and inflammation occur. Experimental Study.

ABSTRACT Objective: To evaluate the evolution of injuries of the supraspinatus muscle by immunohistochemistry (IHC) and anatomopathological analysis in animal model (Wistar rats). Methods: Twenty-five Wistar rats were submitted to complete injury of the supraspinatus tendon, then subsequently sacrificed in groups of five animals at the following periods: immediately after the injury, 24h after the injury, 48h after, 30 days after and three months after the injury. All groups underwent histological and IHC analysis. Results: Regarding vascular proliferation and inflammatory infiltrate, we found a statistically significant difference between groups 1(control group) and 2 (24h after injury). IHC analysis showed that expression of vascular endothelial growth factor (VEGF) showed a statistically significant difference between groups 1 and 2, and collagen type 1 (Col-1) evaluation presented a statistically significant difference between groups 1 and 4. Conclusion: We observed changes in the extracellular matrix components compatible with remodeling and healing. Remodeling is more intense 24h after injury. However, VEGF and Col-1 are substantially increased at 24h and 30 days after the injury, respectively. Level of Evidence I, Experimental Study.

RESUMO Objetivo Determinar a presença de glicosaminoglicanos na matriz extracelular do tecido conjuntivo colorretal neoplásico e não neoplásico, tendo em vista seu papel central no desenvolvimento e na progressão dos tumores. Métodos Amostras de tecidos colorretais neoplásicos e não neoplásicos foram obtidas de 64 pacientes operados com carcinoma colorretal sem metástases a distância. As expressões de heparan sulfato, sulfato de condroitina e sulfato de dermatan e seus fragmentos foram analisadas por espectrometria de massa por ionização por electrospray, com técnica de extração e quantificação de glicosaminoglicanos após proteólise e eletroforese. Para análise estatística, utilizaram-se média, desvio padrão e teste t de Student. Resultados Em gel de agarose, os glicosaminoglicanos extraídos de tecido colorretal mostraram três bandas eletroforéticas. A espectrometria de massa por ionização por electrospray mostrou fragmentos de dissacarídeos característicos de glicosaminoglicanos e indicou sua característica estrutural. Alguns picos na espectrometria de massa por ionização por electrospray não foram caracterizados como fragmentos de açúcares, sugerindo a presença de fragmentos de proteínas estruturais dos proteoglicanos, formadas durante a purificação dos glicosaminoglicanos. A quantidade média de condroitina e dermatan aumentou no tecido neoplástico em relação ao tecido normal (p=0,01). Por outro lado, a quantidade média de heparan foi menor no tecido neoplásico em relação ao tecido normal (p=0,03). Conclusão O método empregado permitiu determinar o perfil estrutural dos glicosaminoglicanos nas amostras. Tecidos neoplásicos apresentaram maiores quantidades de sulfato de condroitina e sulfato de dermatan em comparação com os não neoplásicos, enquanto o sulfato de heparan foi encontrado em menores quantidades nos tecidos neoplásicos.

ABSTRACT Objective To determine the presence of glycosaminoglycans in the extracellular matrix of connective tissue from neoplastic and non-neoplastic colorectal tissues, since it has a central role in tumor development and progression. Methods Tissue samples from neoplastic and non-neoplastic colorectal tissues were obtained from 64 operated patients who had colorectal carcinoma with no distant metastases. Expressions of heparan sulphate, chondroitin sulphate, dermatan sulphate and their fragments were analyzed by electrospray ionization mass spectrometry, with the technique for extraction and quantification of glycosaminoglycans after proteolysis and electrophoresis. The statistical analysis included mean, standard deviation, and Student’st test. Results The glycosaminoglycans extracted from colorectal tissue showed three electrophoretic bands in agarose gel. Electrospray ionization mass spectrometry showed characteristic disaccharide fragments from glycosaminoglycans, indicating their structural characterization in the tissues analyzed. Some peaks in the electrospray ionization mass spectrometry were not characterized as fragments of sugars, indicating the presence of fragments of the protein structure of proteoglycans generated during the glycosaminoglycan purification. The average amount of chondroitin and dermatan increased in the neoplastic tissue compared to normal tissue (p=0.01). On the other hand, the average amount of heparan decreased in the neoplastic tissue compared to normal tissue (p= 0.03). Conclusion The method allowed the determination of the glycosaminoglycans structural profile in colorectal tissue from neoplastic and non-neoplastic colorectal tissue. Neoplastic tissues showed greater amounts of chondroitin sulphate and dermatan sulphate compared to non-neoplastic tissues, while heparan sulphate was decreased in neoplastic tissues.

Objetivo Avaliar os efeitos da transferência gênica do VEGF165 no processo de remodelamento da matriz extracelular após infarto agudo do miocárdio. Métodos Ratos Wistar foram submetidos ao infarto do miocárdio por ligação da artéria coronária descendente esquerda, e a fração de ejeção de ventrículo esquerdo foi utilizada para classificar os infartos em grandes e pequenos. Os animais foram divididos em grupos de dez animais, de acordo com o tamanho do infarto (grande ou pequeno), e receberam ou não tratamento com o VEGF165. A avaliação dos diferentes marcadores foi realizada por imuno-histoquímica e quantificação digital. Os anticorpos primários utilizados foram antifibronectina, antivimentina, anti- CD44, anti-E-caderina, anti-CD24, anti-alfa-1-actina e anti-PCNA. Os resultados foram representados como média e erro padrão, e analisados por ANOVA, sendo considerado estatisticamente significativo se p≤0,05. Resultados Houve aumento significativo da expressão de marcadores de células indiferenciadas, como fibronectina (proteína presente na matriz extracelular) e CD44 (glicoproteína presente nas células endoteliais). Entretanto, houve diminuição da expressão de vimentina e PCNA, indicando possível diminuição do processo de proliferação celular após o tratamento com VEGF165. Os marcadores de células diferenciadas, E-caderina (proteína de adesão entre as células do miocárdio), CD24 (proteína presente nos vasos sanguíneos) e alfa-1-actina (marcador especifico de miócitos) também apresentaram maior expressão nos grupos submetidos à terapia gênica, comparativamente com o grupo não tratado. O valor obtido pela relação entre alfa-1-actina e vimentina foi aproximadamente três vezes maior nos grupos tratados com VEGF165, indicando maior diferenciação tecidual. Conclusão O papel dos miócitos se mostrou importante no processo de remodelamento tecidual, confirmando que o VEGF165 parece conferir um efeito protetor no tratamento do infarto agudo do miocárdio.

Objective Evaluate the effects of VEGF165 gene transfer in the process of remodeling of the extracellular matrix after an acute myocardial infarct. Methods Wistar rats were submitted to myocardial infarction, after the ligation of the left descending artery, and the left ventricle ejection fraction was used to classify the infarcts into large and small. The animals were divided into groups of ten, according to the size of infarcted area (large or small), and received or not VEGF165 treatment. Evaluation of different markers was performed using immunohistochemistry and digital quantification. The primary antibodies used in the analysis were anti-fibronectin, anti-vimentin, anti-CD44, anti-E-cadherin, anti-CD24, anti-alpha-1-actin, and anti-PCNA. The results were expressed as mean and standard error, and analyzed by ANOVA, considering statistically significant if p≤0.05. Results There was a significant increase in the expression of undifferentiated cell markers, such as fibronectin (protein present in the extracellular matrix) and CD44 (glycoprotein present in the endothelial cells). However, there was decreased expression of vimentin and PCNA, indicating a possible decrease in the process of cell proliferation after treatment with VEGF165. Markers of differentiated cells, E-cadherin (adhesion protein between myocardial cells), CD24 (protein present in the blood vessels), and alpha-1-actin (specific myocyte marker), showed higher expression in the groups submitted to gene therapy, compared to non-treated group. The value obtained by the relation between alpha-1-actin and vimentin was approximately three times higher in the groups treated with VEGF165, suggesting greater tissue differentiation. Conclusion The results demonstrated the important role of myocytes in the process of tissue remodeling, confirming that VEGF165 seems to provide a protective effect in the treatment of acute myocardial infarct.

CONTEXTO E OBJETIVO: A heparanase-1 degrada heparam sulfato e está relacionada à progressão de tumor. Apesar de a isoforma heparanase-2 não possuir atividade catalítica, parece ser importante para modular a atividade da heparanase-1. A catepsina B é uma proteinase envolvida na metástase de tumores. O objetivo deste estudo foi analisar a expressão das isoformas de heparanase e atividade da catepsina B em amostras de plasma de pacientes com carcinomas gastrointestinais, comparando-se com indivíduos saudáveis (grupo controle). TIPO DE ESTUDO E LOCAL: Este é um estudo transversal analítico. Foram coletadas amostras de sangue periférico, em hospital público brasileiro, de 21 pacientes com diagnóstico histopatológico de carcinoma gastrointestinal e 43 indivíduos saudáveis. As análises foram realizadas em duas faculdades de medicina brasileiras. MÉTODOS: As isoformas da heparanase foram identificadas e quantificadas em amostras de plasma por Western blot. As atividades enzimáticas de heparanase-1 e catepsina B foram também mensuradas. RESULTADOS: Os resultados demonstraram que as expressões das isoformas de heparanase foram significativamente maiores nas amostras de plasma de pacientes com carcinoma gastrointestinal em comparação com o grupo controle. A análise feita por regressão logística mostra que aumento na expressão de heparanase-1 e heparanase-2 é exclusivamente dependente da presença de tumor. Houve aumento significativo na atividade da heparanase-1 e catepsina B no plasma dos pacientes. CONCLUSÃO: A superexpressão de heparanase-1 e heparanase-2, bem como o aumento da atividade de heparanase-1 e catepsina B no plasma, está associada com o diagnóstico de carcinoma gastrointestinal. Tais achados dão suporte ao uso de ensaios não invasivos (amostras de plasma) como método auxiliar para o diagnóstico de tumores gastrointestinais.

CONTEXT AND OBJECTIVE: Heparanase-1 degrades heparan sulfate and has been correlated with tumor progression. Although the isoform heparanase-2 has no catalytic activity, it seems to be important for modulating heparanase-1 activity. Cathepsin B is a proteinase involved in tumor metastasis. The aim of this study was to analyze heparanase isoform expression and cathepsin B activity in plasma samples from patients with gastrointestinal carcinomas, compared with healthy individuals (control group). DESIGN AND SETTING: This was an analytical cross-sectional study. Peripheral blood samples were collected at a Brazilian public hospital, from 21 patients with histopathological diagnoses of gastrointestinal carcinomas and from 43 healthy individuals. The analyses were performed in two Brazilian medical schools. METHODS: Heparanase isoforms were identified and quantified in plasma samples by means of Western blot. The enzymatic activities of heparanase-1 and cathepsin B were also measured. RESULTS: The results demonstrated that the expression of both heparanase isoforms was significantly greater in plasma samples from gastrointestinal carcinoma patients, compared with the control group. Logistic regression analysis showed that increased heparanase-1 and heparanase-2 expression was exclusively dependent on the tumor. There was a significant increase in heparanase-1 and cathepsin B activity in the patients' plasma. CONCLUSION: Overexpression of heparanase-1 and heparanase-2, along with increased heparanase-1 and cathepsin B activity in plasma, is associated with the diagnosis of gastrointestinal carcinoma. These findings provide support for using non-invasive assays (plasma samples) as an auxiliary method for diagnosing gastrointestinal tumors.

OBJETIVO: Avaliar a expressão das metaloproteinases e do TGFb em pacientes com estenose do canal vertebral e em pacientes mais jovens que apresentam hérnia de disco. MÉTODOS: Foram analisadas 19 amostras de LA, sendo nove de pacientes com estenose de canal lombar e 10 de pacientes com hérnia discal. Do total, cinco pacientes tinham de 15 a 40 anos, 10 tinham de 40 a 65 anos e quatro tinham mais de 65 anos. As áreas representativas do LA foram escolhidas com base na coloração dos tecidos por hematoxilina-eosina. Os cortes de 3 µm de espessura incluídos em parafina e fixados em formalina foram desparafinizados e reidratados. Todos os ligamentos foram incubados overnight a 4 ºC com os anticorpos primários. RESULTADOS: Constatou-se aumento do TGFb em indivíduos mais velhos, embora sem significância estatística. CONCLUSÃO: As metaloproteinases não apresentaram diferença importante entre os grupos tanto com relação à idade quanto ao tipo de alteração da coluna vertebral.

OBJECTIVE: To evaluate the expression of matrix metalloproteinases and TGFb in patients with spinal stenosis and in younger patients who have herniated disc. METHODS: 19 samples of LA were analyzed, nine of them with lumbar canal stenosis and 10 with disc herniation. Of the total, five patients were aged between 15 and 40 years, 10 were between 40 and 65 years and four had more than 65 years. Representative areas of LF were chosen based on the staining of tissues with hematoxylin-eosin. The 3µm-thick sections embedded in paraffin and fixed in formalin were deparaffinized and rehydrated. All ligaments were incubated overnight at 4 °C with primary antibodies. RESULTS: An increase of TGFb was verified in older individuals, although without statistical significance. CONCLUSION: Metalloproteinases showed no significant difference between both groups with respect to age and type of abnormality of the spine.

OBJETIVO: Evaluar la expresión de metaloproteinasas de la matriz y del TGFb en pacientes con estenosis espinal y en pacientes más jóvenes que tienen una hernia de disco. MÉTODOS: Diecinueve muestras de LA fueron enviadas, de nueve pacientes con estenosis del canal lumbar y diez pacientes con hernia de disco. Del total de pacientes, cinco tenían de 15 a 40 años, 10 tenían de 40 a 65 años y cuatro tenían más de 65 años. Áreas representativas de LA se eligieron sobre la base de la tinción de los tejidos con hematoxilina-eosina. Las secciones de 3 µm de espesor, incluidas en parafina y fijadas en formalina fueron desparafinadas y rehidratadas. Todos los ligamentos se incubaron durante la noche a 4 °C con anticuerpos primarios. RESULTADOS: Se encontró un aumento de TGFb en personas mayores, aunque sin significación estadística. CONCLUSIÓN: Las metaloproteinasas no mostraron diferencias significativas entre ambos grupos con respecto a la edad y el tipo de anomalía de la columna vertebral.