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  • The Polycomb group protein Ring1b is essential for pectoral fin development. 22619390

    Polycomb group (PcG) proteins are transcriptional repressors that mediate epigenetic gene silencing by chromatin modification. PcG-mediated gene repression is implicated in development, cell differentiation, stem-cell fate maintenance and cancer. However, analysis of the roles of PcG proteins in orchestrating vertebrate developmental programs in vivo has been hampered by the early embryonic lethality of several PcG gene knockouts in mice. Here, we demonstrate that zebrafish Ring1b, the E3 ligase in Polycomb Repressive Complex 1 (PRC1), is essential for pectoral fin development. We show that differentiation of lateral plate mesoderm (LPM) cells into presumptive pectoral fin precursors is initiated normally in ring1b mutants, but fin bud outgrowth is impaired. Fgf signaling, which is essential for migration, proliferation and cell-fate maintenance during fin development, is not sufficiently activated in ring1b mutants. Exogenous application of FGF4, as well as enhanced stimulation of Fgf signaling by overactivated Wnt signaling in apc mutants, partially restores the fin developmental program. These results reveal that, in the absence of functional Ring1b, fin bud cells fail to execute the pectoral fin developmental program. Together, our results demonstrate that PcG-mediated gene regulation is essential for sustained Fgf signaling in vertebrate limb development.
    Document Type:
    Reference
    Product Catalog Number:
    07-146
    Product Catalog Name:
    Anti-Histone H2A (acidic patch) Antibody
  • 2-Amino-3-methylimidazo[4,5-f]quinoline (IQ) promotes mouse hepatocarcinogenesis by activating transforming growth factor-β and Wnt/β-catenin signaling pathways. 22094457

    The purposes of the present study were to investigate the modifying effects of 2-amino-3-methylimidazo[4,5-f]quinoline (IQ), a genotoxic carcinogen produced during cooking of protein-rich foods, and elucidate underlying mechanisms in a two-stage hepatocarcinogenesis mice model. Six-week-old B6C3F1 mice were subjected to two-thirds partial hepatectomy at the beginning of the study, followed by an intraperitoneal injection of diethylnitrosamine on day 1. Starting 1 week later, they were fed diets containing IQ at doses of 30, 100, or 300 ppm for 39 weeks. A dose-dependent trend for increase in eosinophilic altered foci as well as eosinophilic hepatocellular adenomas was observed, along with significant elevation in the incidence of hepatocellular carcinomas in the 100- and 300-ppm IQ groups as compared with initiation control group. Furthermore, IQ elevated the protein expression levels of Wnt1, transforming growth factor-β (TGF-β), TGF-β receptors 1 and 2 (TβR1 and TβR2), and phosphorylated c-Jun (p-c-Jun), while suppressing those of E-cadherin and p21(WAF1/Cip1). Moreover, translocation of β-catenin to the nuclei as well as upregulated nuclear expression of c-Myc and cyclin D1, which are downstream targets of β-catenin and p-c-Jun, were detected at 100 and 300 ppm. These findings suggest that IQ exerts dose-dependent promoting effects on mice hepatocarcinogenesis by activating TGF-β and Wnt/β-catenin signaling pathways and inhibiting cell adhesion.
    Document Type:
    Reference
    Product Catalog Number:
    07-146
    Product Catalog Name:
    Anti-Histone H2A (acidic patch) Antibody
  • In vitro 3D culture of human chondrocytes using Modified {varepsilon}-caprolactone scaffolds with varying hydrophilicity and porosity. 21586602

    Two series of 3D scaffolds based on ε-caprolactone were synthesized. The pore size and architecture (spherical interconnected pores) was the same in all the scaffolds. In one of the series of scaffolds, made of pure ε-polycaprolactone, the volume fraction of pores varied between 60% and 85% with the main consequence of varying the interconnectivity between pores since the pore size was kept constant. The other scaffolds were prepared with copolymers made of a ε-caprolactone-based hydrophobous monomer and hydroxyethyl acrylate, as the hydrophilic component. Thus, the hydrophilicity and, presumably, the adhesion properties varied monotonously in the copolymer series while porosity was kept constant. A suspension of human chondrocytes in culture medium was injected in the 3D scaffolds and cultured in static conditions up to 28 days. SEM and immunofluorescence assays allowed characterizing cells and extracellular matrix inside the scaffolds after different culture times. To do that, cross sections of the scaffolds were observed by SEM and confocal microscopy. The quantity of cells inside the scaffolds decreases with a decrease of the volume fraction of pores, due to the lack of interconnectivity between the cavities. The scaffolds up to a 30% of hydrophilicity behave in a similar way than the hydrophobous; a further increase of the hydrophilicity rapidly decreases cell viability. In all the experiments production of collagen type I, type II, and aggrecan was found, and some cells were Ki-67 positive, showing that some cells are adhered to the pore walls and maintain their dedifferentiated phenotype even when cultured in three-dimensional conditions.
    Document Type:
    Reference
    Product Catalog Number:
    AB745
    Product Catalog Name:
    Anti-Collagen Type I Antibody
  • Mybl2, downregulated during colon epithelial cell maturation, is suppressed by miR-365. 21737779

    Altered profiles of gene expression reflect the reprogramming of intestinal epithelial cells during their maturation along the crypt-luminal axis. To focus on genes important in this process, and how they in turn are regulated, we identified 14 transcripts commonly downregulated in expression during lineage-specific maturation of the immortalized cell lines Caco-2 (absorptive), HT29Cl16E (goblet), and HT29Cl19A (secretory) induced by contact inhibition of growth or the short-chain fatty acid butyrate. One such gene, Mybl2 (Myb-related protein B), has been linked to the stem cell phenotype, and we report is also markedly suppressed in maturing cells along the crypt-luminal axis in vivo. Mybl2 is not significantly downregulated transcriptionally during colon cell maturation, but we identified a potential micro-RNA (miRNA)-binding sequence in the Mybl2 3'-untranslated region that mediates reporter gene suppression in differentiating colon cells. Accordingly, miRNAs predicted to bind this functional target are upregulated in differentiating colon epithelial cells in vitro and in vivo; expression of one of these, hsa-miR-365 (but not hsa-324-5p), suppresses Mybl2 protein expression in proliferating Caco-2 cells. These data demonstrate that miRNA silencing plays an important role in regulating gene expression in maturing colon epithelial cells, and that utilizing a target-centered approach, rather than profiling global miRNA expression, can identify physiologically relevant, functional miRNAs.
    Document Type:
    Reference
    Product Catalog Number:
    07-146
    Product Catalog Name:
    Anti-Histone H2A (acidic patch) Antibody
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