Citations

In vitro studies have proposed a tumor suppressor role for sulfatase 1 (SULF1) in hepatocellular carcinoma (HCC); however, high expression in human HCC has been associated with poor prognosis. The reason underlying this paradoxical observation remains to be explored. Using a transgenic (Tg) mouse model overexpressing Sulf1 (Sulf1‐Tg), we assessed the effects of SULF1 on the diethylnitrosamine model of liver carcinogenesis. Sulf1‐Tg mice show a higher incidence of large and multifocal tumors with diethylnitrosamine injection compared to wild‐type mice. Lung metastases were found in 75% of Sulf1‐Tg mice but not in wild‐type mice. Immunohistochemistry, immunoblotting, and reporter assays all show a significant activation of the transforming growth factor‐β (TGF‐β)/SMAD transcriptional pathway by SULF1 both in vitro and in vivo . This effect of SULF1 on the TGF‐β/SMAD pathway is functional; overexpression of SULF1 promotes TGF‐β‐induced gene expression and epithelial–mesenchymal transition and enhances cell migration/invasiveness. Mechanistic analyses demonstrate that inactivating mutation of the catalytic site of SULF1 impairs the above actions of SULF1 and diminishes the release of TGF‐β from the cell surface. We also show that SULF1 expression decreases the interaction between TGF‐β1 and its heparan sulfate proteoglycan sequestration receptor, TGFβR3. Finally, using gene expression from human HCCs, we show that patients with high SULF1 expression have poorer recurrence‐free survival (hazard ratio 4.1, 95% confidence interval 1.9‐8.3; P = 0.002) compared to patients with low SULF1. We also found strong correlations of SULF1 expression with TGF‐β expression and with several TGF‐β‐related epithelial–mesenchymal transition genes in human HCC. Conclusion : Our study proposes a novel role of SULF1 in HCC tumor progression through augmentation of the TGF‐β pathway, thus defining SULF1 as a potential biomarker for tumor progression and a novel target for drug development for HCC. (H epatology 2015;61:1269–1283)

AbstractObjective The high level of stress associated with caring for others with medical conditions has been recognized for some time. Reducing caregiver stress can improve caregiver quality of life as well as improve the care they provide to loved ones. This systematic review assesses the effectiveness of internet-based interventions to decrease caregiver stress.Materials and methods A comprehensive literature search of Ovid MEDLINE (1946–2013), Embase (1988–2013), PsycINFO (1987–2013), and CINAHL was conducted using terms related to caregiver and internet-based interventions. Internet-based interventions involving informal caregivers in an open-label or randomized controlled trial setting were included. A pair of raters independently reviewed all published abstracts. Data regarding participants, interventions, and outcomes were extracted and, for randomized trials, methodology quality was also reviewed.Results Eight open-label trials met the review criteria: three showed positive benefit in reducing caregiver stress, four were partially positive (some outcomes positive, others negative), and one was a negative study. Sixteen randomized trials met the review criteria: six showed positive benefit, five were partially positive, and five were negative. There were no clear patterns as to the variables (such as study duration and complexity of intervention) associated with better outcomes, although earlier studies typically had more negative outcomes.Discussion Internet-based interventions were mostly effective in reducing aspects of caregiver stress and improving their well-being. Further studies to assess outcomes for caregivers and their recipients’ health, different technology delivery methods, and the cost of such interventions are needed.

Abstract Background/Aim: Glioma-associated oncogene 1 (GLI1) is one of the most important transcription factors in the hedgehog (Hh) signaling pathway and has been shown to be involved in embryonic development and tumor progression. The epithelial-to-mesenchymal transition (EMT) is an important cellular program which contributes to invasion and metastasis of cancer. We have previously shown that GLI1 may promote HCC metastasis via EMT. Here, we demonstrate that GLI1 upregulates SNAI1 expression, induces EMT, and promotes HCC recurrence in vivo and in vitro. Methods: Gli1 mRNA was assessed in 139 HCCs by microarray and its correlation with HCC recurrence and SNAI1 mRNA expression was analyzed. In vitro, the mRNA and protein expression of GLI1 was measured in Gli1-negative Huh7 cells transfected with GU1 and GU1-expressing SNU398 cells transfected with antisense oligonucleotides targeting GU1 (GLI1 ASO). GLI1-dependent transcriptional activity was assessed with GU reporter luciferase assay and electrophoretic mobility shift assay (EMSA). The interaction between Gli1, SNAI1 and EMT was assessed by real-time PCR, Western immunoblotting and immunofluorescence. Effects of GLI1 on proliferation, viability, migration, and invasion of tumor cells were also assessed. Results: GLI1 mRNA was detected in 74 HCC tumors and found to be upregulated in 43 (58.1 %) HCCs. Increased Gli1 mRNA in HCCs was associated with tumor recurrence after resection. There was a positive correlation between GLI1 and SNAI1 mRNA expression. In HCC cells, GLI1 overexpression increased Gli1-dependent transcriptional activity, proliferation, viability, migration, and invasion of tumor cells. GU1 also increased SNAI1 expression and induced the EMT. Opposite effects were observed in GU1-knockdown models. Conclusion: Together, these findings identify a novel mechanism mediating HCC recurrence that includes GLI1-mediated activation of EMT via upregulation of SNAI1. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the Second AACR International Conference on Frontiers in Basic Cancer Research; 2011 Sep 14-18; San Francisco, CA. Philadelphia (PA): AACR; Cancer Res 2011;71(18 Suppl):Abstract nr B20.

ABSTRACT Target cell overexpression of the integrase binding domain (IBD) of LEDGF/p75 (LEDGF) inhibits HIV-1 replication. The mechanism and protein structure requirements for this dominant interference are unclear. More generally, how and when HIV-1 uncoating occurs postentry is poorly defined, and it is unknown whether integrase within the evolving viral core becomes accessible to cellular proteins prior to nuclear entry. We used LEDGF dominant interference to address the latter question while characterizing determinants of IBD antiviral activity. Fusions of green fluorescent protein (GFP) with multiple C-terminal segments of LEDGF inhibited HIV-1 replication substantially, but minimal chimeras of either polarity (GFP-IBD or IBD-GFP) were most effective. Combining GFP-IBD expression with LEDGF depletion was profoundly antiviral. CD4 + T cell lines were rendered virtually uninfectable, with single-cycle HIV-1 infectivity reduced 4 logs and high-input (multiplicity of infection = 5.0) replication completely blocked. We restricted GFP-IBD to specific intracellular locations and found that antiviral activity was preserved when the protein was confined to the cytoplasm or directed to the nuclear envelope. The life cycle block triggered by the cytoplasm-restricted protein manifested after nuclear entry, at the level of integration. We conclude that integrase within the viral core becomes accessible to host cell protein interaction in the cytoplasm. LEDGF dominant interference and depletion impair HIV-1 integration at distinct postentry stages. GFP-IBD may trigger premature or improper integrase oligomerization.

ABSTRACT HIV-1 and certain other retroviruses initiate plus-strand synthesis in the center of the genome as well as at the standard retroviral 3′ polypurine tract. This peculiarity of reverse transcription results in a central DNA “flap” structure that has been of controversial functional significance. We mutated both HIV-1 flap-generating elements, the central polypurine tract (cPPT) and the central termination sequence (CTS). To avoid an ambiguity of previous studies, we did so without affecting integrase coding. DNA flap formation was disrupted but single-cycle infection was unaffected in all target cells tested, regardless of cell cycle status. Spreading HIV-1 infection was also normal in most T cell lines, and flap mutant viruses replicated equivalently to the wild type in nondividing cells, including macrophages. However, spreading infection of flap mutant HIV-1 was impaired in non- vif -permissive cells (HuT78, H9, and primary human peripheral blood mononuclear cells [PBMCs]), suggesting APOBEC3G (A3G) restriction. Single-cycle infections confirmed that vif -intact flap mutant HIV-1 is restricted by producer cell A3G/F. Combining the Δ vif and cPPT-CTS mutations increased A3G restriction synergistically. Moreover, RNA interference knockdown of A3G in HuT78 cells released the block to flap mutant HIV-1 replication. Flap mutant HIV-1 also accrued markedly increased A3G-mediated G→A hypermutation compared to that of wild-type HIV-1 (a full log 10 in the 0.36 kb downstream of the mutant cPPT). We suggest that the triple-stranded DNA structure, the flap, is not the consequential outcome. The salient functional feature is central plus-strand initiation, which functions as a second line of defense against single-stranded DNA editing by A3 proteins that survive producer cell degradation by Vif.

ABSTRACT Measles virus (MV) entry requires at least 2 viral proteins, the hemagglutinin (H) and fusion (F) proteins. We describe the rescue and characterization of a measles virus with a specific mutation in the stalk region of H (I98A) that is able to bind normally to cells but infects at a lower rate than the wild type due to a reduction in fusion triggering. The mutant H protein binds to F more avidly than the parent H protein does, and the corresponding virus is more sensitive to inhibition by fusion-inhibitory peptide. We show that after binding of MV to its receptor, H-F dissociation is required for productive infection.

ABSTRACT Nonprimate animal models of HIV-1 infection are prevented by missing cellular cofactors and by antiviral actions of species-specific host defense factors. These blocks are profound in rodents but may be less abundant in certain Carnivora. Here, we enabled productive, spreading replication and passage of HIV-1 in feline cells. Feline fibroblasts, T-cell lines, and primary peripheral blood mononuclear cells supported early and late HIV-1 life cycle phases in a manner equivalent to that of human cells, except that produced virions had low infectivity. Stable expression of feline immunodeficiency virus (FIV) Vif-green fluorescent protein (GFP) in HIV-1 entry receptor-complemented feline (CrFK) cells enabled robust spreading HIV-1 replication. FIV Vif colocalized with feline APOBEC3 (fA3) proteins, targeted them for degradation, and prevented G→A hypermutation of the HIV-1 cDNA by fA3CH and fA3H. HIV-1 Vif was inactive against fA3s as expected and even paradoxically augmented restriction in some assays. In an interesting contrast, simian immunodeficiency virus SIVmac Vif had substantial anti-fA3 activities, which were complete against fA3CH and partial against fA3H. Moreover, both primate lentiviral Vifs colocalized with fA3s and could be pulled down from cell lysates by fA3CH. HIV-1 molecular clones that encode FIV Vif or SIVmac Vif (HIV-1 VF and HIV-1 VS ) were then constructed. These viruses replicated productively in HIV-1 receptor-expressing CrFK cells and could be passaged serially to uninfected cells. Thus, with the exception of entry receptors, the cat genome can supply the dependency factors needed by HIV-1, and a main restriction can be countered by vif chimerism. The results raise the possibility that the domestic cat could yield an animal model of HIV-1 infection.

ABSTRACT Enveloped viruses trigger membrane fusion to gain entry into cells. The receptor affinities of their attachment proteins vary greatly, from 10 −4 M to 10 −9 M, but the significance of this is unknown. Using six retargeted measles viruses that bind to Her-2/neu with a 5-log range in affinity, we show that receptor affinity has little impact on viral attachment but is nevertheless a key determinant of infectivity and intercellular fusion. For a given cell surface receptor density, there is an affinity threshold above which cell-cell fusion proceeds efficiently. Suprathreshold affinities do not further enhance the efficiency of membrane fusion.