PPARgamma inhibits hepatocellular carcinoma metastases in vitro and in mice

Background: We have previously demonstrated that peroxisome proliferator-activated receptor (PPARγ) activation inhibits hepatocarcinogenesis. We aim to investigate the effect of PPARγ on hepatocellular carcinoma (HCC) metastatic potential and explore its underlying mechanisms. Methods: Human HCC cells (MHCC97L, BEL-7404) were infected with adenovirus-expressing PPARγ (Ad-PPARγ) or Ad-lacZ and treated with or without PPARγ agonist (rosiglitazone). The effects of PPARγ on cell migration and invasive activity were determined by wound healing assay and Matrigel invasive model in vitro, and in an orthotopic liver tumour metastatic model in mice.Results:Pronounced expression of PPARγ was demonstrated in HCC cells (MHCC97L, BEL-7404) treated with Ad-PPARγ, rosiglitazone or Ad-PPARγ plus rosiglitazone, compared with control (Ad-LacZ). Such induction markedly suppressed HCC cell migration. Moreover, the invasiveness of MHCC97L and BEL-7404 cells infected with Ad-PPARγ, or treated with rosiglitazone was significantly diminished up to 60%. Combination of Ad-PPARγ and rosiglitazone showed an additive effect. Activation of PPARγ by rosiglitazone significantly reduced the incidence and severity of lung metastasis in an orthotopic HCC mouse model. Key mechanisms underlying the effect of PPARγ in HCC include upregulation of cell adhesion genes, E-cadherin and SYK (spleen tyrosine kinase), extracellular matrix regulator tissue inhibitors of metalloproteinase (TIMP) 3, tumour suppressor gene retinoblastoma 1, and downregulation of pro-metastatic genes MMP9 (matrix metallopeptidase 9), MMP13, HPSE (heparanase), and Hepatocyte growth factor (HGF). Direct transcriptional regulation of TIMP3, MMP9, MMP13, and HPSE by PPARγ was shown by ChIP-PCR. Conclusion: Peroxisome proliferator-activated receptor-gamma exerts an inhibitory effect on the invasive and metastatic potential of HCC in vitro and in vivo, and is thus, a target for the prevention and treatment of HCC metastases. © 2012 Cancer Research UK All rights reserved.published_or_final_versio

Analysis and characterization of heparin impurities

This review discusses recent developments in analytical methods available for the sensitive separation, detection and structural characterization of heparin contaminants. The adulteration of raw heparin with oversulfated chondroitin sulfate (OSCS) in 2007–2008 spawned a global crisis resulting in extensive revisions to the pharmacopeia monographs on heparin and prompting the FDA to recommend the development of additional physicochemical methods for the analysis of heparin purity. The analytical chemistry community quickly responded to this challenge, developing a wide variety of innovative approaches, several of which are reported in this special issue. This review provides an overview of methods of heparin isolation and digestion, discusses known heparin contaminants, including OSCS, and summarizes recent publications on heparin impurity analysis using sensors, near-IR, Raman, and NMR spectroscopy, as well as electrophoretic and chromatographic separations

Vav proteins maintain epithelial traits in breast cancer cells using miR-200c-dependent and independent mechanisms

The bidirectional regulation of epithelial–mesenchymal transitions (EMT) is key in tumorigenesis. Rho GTPases regulate this process via canonical pathways that impinge on the stability of cell-to-cell contacts, cytoskeletal dynamics, and cell invasiveness. Here, we report that the Rho GTPase activators Vav2 and Vav3 utilize a new Rac1-dependent and miR-200c-dependent mechanism that maintains the epithelial state by limiting the abundance of the Zeb2 transcriptional repressor in breast cancer cells. In parallel, Vav proteins engage a mir-200c-independent expression prometastatic program that maintains epithelial cell traits only under 3D culture conditions. Consistent with this, the depletion of endogenous Vav proteins triggers mesenchymal features in epithelioid breast cancer cells. Conversely, the ectopic expression of an active version of Vav2 promotes mesenchymal-epithelial transitions using E-cadherin-dependent and independent mechanisms depending on the mesenchymal breast cancer cell line used. In silico analyses suggest that the negative Vav anti-EMT pathway is operative in luminal breast tumors. Gene signatures from the Vav-associated proepithelial and prometastatic programs have prognostic value in breast cancer patients.Fil: Lorenzo Martín, L. Francisco. Universidad de Salamanca; España. Consejo Superior de Investigaciones Científicas; España. Instituto de Biología Molecular y Celular del Cáncer de Salamanca; EspañaFil: Citterio, Carmen. Consejo Superior de Investigaciones Científicas; España. Instituto de Biología Molecular y Celular del Cáncer de Salamanca; EspañaFil: Menacho Márquez, Mauricio Ariel. Universidad de Salamanca; España. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Rosario. Instituto de Investigaciones para el Descubrimiento de Fármacos de Rosario. Universidad Nacional de Rosario. Instituto de Investigaciones para el Descubrimiento de Fármacos de Rosario; Argentina. Instituto de Biología Molecular y Celular del Cáncer de Salamanca; EspañaFil: Conde, Javier. Consejo Superior de Investigaciones Científicas; España. Instituto de Biología Molecular y Celular del Cáncer de Salamanca; EspañaFil: Larive, Romain M.. Consejo Superior de Investigaciones Científicas; España. Institut Des Biomolécules Max Mousseron; FranciaFil: Rodríguez Fdez, Sonia. Consejo Superior de Investigaciones Científicas; EspañaFil: García Escudero, Ramón. Consejo Superior de Investigaciones Científicas; España. Universidad de Salamanca; EspañaFil: Robles Valero, Javier. Universidad de Salamanca; España. Consejo Superior de Investigaciones Científicas; España. Centro de Investigación del Cáncer; España. Instituto de Biología Molecular y Celular del Cáncer de Salamanca; EspañaFil: Cuadrado, Myriam. Universidad de Salamanca; España. Instituto de Biología Molecular y Celular del Cáncer de Salamanca; España. Consejo Superior de Investigaciones Científicas; EspañaFil: Fernández Pisonero, Isabel. Universidad de Salamanca; España. Consejo Superior de Investigaciones Científicas; España. Instituto de Biología Molecular y Celular del Cáncer de Salamanca; EspañaFil: Dosil, Mercedes. Universidad de Salamanca; España. Consejo Superior de Investigaciones Científicas; España. Instituto de Biología Molecular y Celular del Cáncer de Salamanca; EspañaFil: Sevilla, María A.. Universidad de Salamanca; EspañaFil: Montero, María J.. Universidad de Salamanca; EspañaFil: Fernández Salguero, Pedro. Universidad de Extremadura; EspañaFil: Paramio, Jesús M.. Universidad de Salamanca; España. Consejo Superior de Investigaciones Científicas; España. Centro de Investigaciones Energéticas, Medioambientales y Tecnológicas; EspañaFil: Bustelo, Xosé R.. Universidad de Salamanca; España. Consejo Superior de Investigaciones Científicas; Españ

Broader implications of SILAC-based proteomics for dissecting signaling dynamics in cancer

Large-scale transcriptome and epigenome analyses have been widely utilized to discover gene alterations implicated in cancer development at the genetic level. However, mapping of signaling dynamics at the protein level is likely to be more insightful and needed to complement massive genomic data. Stable isotope labeling with amino acids in cell culture (SILAC)-based proteomic analysis represents one of the most promising comparative quantitative methods that has been extensively employed in proteomic research. This technology allows for global, robust and confident identification and quantification of signal perturbations important for the progress of human diseases, particularly malignancies. The present review summarizes the latest applications of in vitro and in vivo SILAC-based proteomics in identifying global proteome/phosphoproteome and genome-wide protein-protein interactions that contribute to oncogenesis, highlighting the recent advances in dissecting signaling dynamics in cancer