RESUMO
Gelsolin is a cytoskeletal protein which participates in actin filament dynamics and promotes cell motility and plasticity. Although initially regarded as a tumor suppressor, gelsolin expression in certain tumors correlates with poor prognosis and therapy-resistance. In vitro, gelsolin has anti-apoptotic and pro-migratory functions and is critical for invasion of some types of tumor cells. We found that gelsolin was highly expressed at tumor borders infiltrating into adjacent liver tissues, as examined by immunohistochemistry. Although gelsolin contributes to lamellipodia formation in migrating cells, the mechanisms by which it induces tumor invasion are unclear. Gelsolin's influence on the invasive activity of colorectal cancer cells was investigated using overexpression and small interfering RNA knockdown. We show that gelsolin is required for invasion of colorectal cancer cells through matrigel. Microarray analysis and quantitative PCR indicate that gelsolin overexpression induces the upregulation of invasion-promoting genes in colorectal cancer cells, including the matrix-degrading urokinase-type plasminogen activator (uPA). Conversely, gelsolin knockdown reduces uPA levels, as well as uPA secretion. The enhanced invasiveness of gelsolin-overexpressing cells was attenuated by treatment with function-blocking antibodies to either uPA or its receptor uPAR, indicating that uPA/uPAR activity is crucial for gelsolin-dependent invasion. In summary, our data reveals novel functions of gelsolin in colorectal tumor cell invasion through its modulation of the uPA/uPAR cascade, with potentially important roles in colorectal tumor dissemination to metastatic sites.
Assuntos
Neoplasias Colorretais/patologia , Gelsolina/metabolismo , Transdução de Sinais , Ativador de Plasminogênio Tipo Uroquinase/metabolismo , Linhagem Celular Tumoral , Fibrinolisina/metabolismo , Gelsolina/deficiência , Gelsolina/genética , Regulação Neoplásica da Expressão Gênica , Inativação Gênica , Humanos , Invasividade Neoplásica , RNA Interferente Pequeno/genética , Receptores de Ativador de Plasminogênio Tipo Uroquinase/metabolismoRESUMO
Gastrointestinal endocrine cell tumors are a heterogeneous population of lesions believed to arise from neuroendocrine cells of the gastrointestinal mucosa. The current classification of these tumors is based on tumor size, microscopic features and clinical evidence of metastasis. Although diagnostic categories generally correlate with prognosis, molecular prognostic markers will be clinically useful adjuncts. Cofilin has been implicated in tumor invasion, and its immunolocalisation was studied in gastrointestinal endocrine cell tumors. The immunolocalisation of cofilin was studied by immunohistochemistry in 34 formalin-fixed, paraffin wax-embedded gastrointestinal endocrine cell tumors using a tissue microarray platform. A significant correlation was found between high cofilin immunolabelling and the depth of invasion (p<0.05). Our findings suggest that cofilin might be useful clinically as a molecular prognostic adjunct in the evaluation of gastrointestinal endocrine cell tumors.
Assuntos
Fatores de Despolimerização de Actina/metabolismo , Neoplasias das Glândulas Endócrinas/metabolismo , Neoplasias das Glândulas Endócrinas/patologia , Células Enteroendócrinas/metabolismo , Células Enteroendócrinas/patologia , Adulto , Idoso , Biomarcadores Tumorais/análise , Carcinoma/metabolismo , Carcinoma/patologia , Feminino , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Neoplasias Retais/metabolismo , Neoplasias Retais/patologia , Neoplasias Gástricas/metabolismo , Neoplasias Gástricas/patologia , Adulto JovemRESUMO
Butyrate, a 4-carbon fatty acid, has been shown to cause growth arrest and apoptosis of cancer cells in vitro and in vivo. The signaling pathways leading to changes in cell growth are unclear. We used a functional proteomics approach to delineate the pathways and mediators involved in butyrate action in HT-29 cells at 24 hr posttreatment. Using 2-dimensional gel electrophoresis, we showed that butyrate treatment resulted in alterations in the proteome of HT-29 cells. MALDI-TOF mass spectrometry was used to identify butyrate-regulated spots. First, our results revealed that the expression of various components of the ubiquitin-proteasome system was altered with butyrate treatment. This suggests that, in addition to the regulation of gene expression through the histone deacetylase pathway, proteolysis could be a means by which butyrate may regulate the expression of key proteins in the control of cell cycle, apoptosis and differentiation. Second, we found that both proapoptotic proteins (capase-4 and cathepsin D) and antiapoptotic proteins (hsp27, antioxidant protein-2 and pyruvate dehydrogenase E1) were simultaneously upregulated in butyrate-treated cells. Western blotting was carried out to confirm butyrate regulation of the spots. Both cathepsin D and hsp27 showed a time-dependent increase in expression with butyrate treatment in HT-29 cells. However, in HCT-116 cells, which were 5-fold more sensitive to butyrate-induced apoptosis, the upregulation of cathepsin D with time was not accompanied by a similar increase in hsp27 levels. Thus, the simultaneous upregulation of both proapoptotic and antiapoptotic proteins in HT-29 cells may account for their relative resistance to butyrate-induced apoptosis.
Assuntos
Butiratos/farmacologia , Células HT29/efeitos dos fármacos , Proteoma/análise , Eletroforese em Gel Bidimensional , Células HT29/metabolismo , Humanos , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Células Tumorais Cultivadas/efeitos dos fármacos , Células Tumorais Cultivadas/metabolismoRESUMO
BACKGROUND AND AIMS: Mutations on the a-determinant of hepatitis B virus surface antigen (HBsAg), capable of escaping detection and vaccination, are identified in HBsAg-positive/anti-HBs-positive vaccinated infants. We studied the prevalence of these mutants in HBsAg-negative/anti-HBc-positive chronic HBV carriers and patients with hepatocellular carcinoma (HCC). METHODS: DNA sequence coding for the antigenic a-determinant of HBsAg was amplified from either HCC genomic DNA or serum samples of the selected patients and sequenced. The replicative mutant genomes were reconstituted in vitro and their reactivity to commercial kits measured. RESULTS: Mutations within and/or outside the a-determinant were identified in patients seronegative for HBsAg. They were then reconstituted in vitro and transiently transfected into HepG2 cells. Culture medium containing secreted HBV viral particles was collected and assayed for their binding to commercial kits. Drastic decrease of reactivity to these kits was seen with most of the identified mutations, including those located outside the a-determinant. CONCLUSION: The existence of a more complex antigenic structure of HBsAg is indicated by the decreased reactivity to detection of mutations, some of which are outside the a-determinant, escape vaccination and may persist in seronegative patients. The high proportion of HBsAg mutants that are integrated in HCC genomes suggests a role of these mutants in hepatocarcinogenesis, possibly leading to mutant HBV-related HCC.