Publications by authors named "David M Owen"

11 Publications

  • Page 1 of 1

Loss of Selenoprotein Iodothyronine Deiodinase 3 Expression Correlates with Progression of Complete Hydatidiform Mole to Gestational Trophoblastic Neoplasia.

Reprod Sci 2021 Jun 15. Epub 2021 Jun 15.

Division of Gynecologic Oncology, Department of Obstetrics and Gynecology, Brigham and Women's Hospital, 75 Francis St., Boston, MA, 02115, USA.

To investigate if differences in imprinting at tropho-microRNA (miRNA) genomic clusters can distinguish between pre-gestational trophoblastic neoplasia cases (pre-GTN) and benign complete hydatidiform mole (CHM) cases at the time of initial uterine evacuation. miRNA sequencing was performed on frozen tissue from 39 CHM cases including 9 GTN cases. DIO3, DLK1, RTL1, and MEG 3 mRNA levels were assessed by qRT-PCR. Protein abundance was assessed by Western blot for DIO3, DLK1, and RTL1. qRT-PCR and Western blot were performed for selenoproteins and markers of oxidative stress. Immunohistochemistry (IHC) was performed for DIO3 on an independent validation set of clinical samples (n = 42) and compared to normal placenta controls across gestational ages. Relative expression of the 14q32 miRNA cluster was lower in pre-GTN cases. There were no differences in protein abundance of DLK1 or RTL1. Notably, there was lower protein expression of DIO3 in pre-GTN cases (5-fold, p < 0.03). There were no differences in mRNA levels of DIO3, DLK1, RTL1 or MEG 3. mRNA levels were higher in all CHM cases compared to normal placenta. IHC showed syncytiotrophoblast-specific DIO3 immunostaining in benign CHM cases and normal placenta, while pre-GTN cases of CHM lacked DIO3 expression. We describe two new biomarkers of pre-GTN CHM cases: decreased 14q32 miRNA expression and loss of DIO3 expression by IHC. Differences in imprinting between benign CHM and pre-GTN cases may provide insight into the fundamental development of CHM.
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http://dx.doi.org/10.1007/s43032-021-00634-yDOI Listing
June 2021

Prosecuting child abusers, respecting victims: a contrast of approaches in the UK, Tajikistan and Tanzania.

Authors:
David M Owen

Paediatr Int Child Health 2013 Nov 13;33(4):253-8. Epub 2013 Sep 13.

Crown Prosecution Service, Liverpool, UK.

This article summarises child abuse as a global problem of increasing breadth and complexity. It reviews the development of procedures for prosecuting alleged abusers and treating complainants appropriately in the course of investigations, medical examinations and court hearings. It contrasts the diverse environments of the UK, Tajikistan and Tanzania. The author draws on his experience of practising criminal law since 1984 in England and Wales and working as a consultant in 2011 with the Girls Support Service in Tajikistan, a European-and UN-funded NGO, and with UNICEF in Tanzania in 2012.
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http://dx.doi.org/10.1179/2046905513Y.0000000094DOI Listing
November 2013

Multiple effects of silymarin on the hepatitis C virus lifecycle.

Hepatology 2010 Jun;51(6):1912-21

Department of Laboratory Medicine, University of Washington, Seattle, WA 98104-2499, USA.

Unlabelled: Silymarin, an extract from milk thistle (Silybum marianum), and its purified flavonolignans have been recently shown to inhibit hepatitis C virus (HCV) infection, both in vitro and in vivo. In the current study, we further characterized silymarin's antiviral actions. Silymarin had antiviral effects against hepatitis C virus cell culture (HCVcc) infection that included inhibition of virus entry, RNA and protein expression, and infectious virus production. Silymarin did not block HCVcc binding to cells but inhibited the entry of several viral pseudoparticles (pp), and fusion of HCVpp with liposomes. Silymarin but not silibinin inhibited genotype 2a NS5B RNA-dependent RNA polymerase (RdRp) activity at concentrations 5 to 10 times higher than required for anti-HCVcc effects. Furthermore, silymarin had inefficient activity on the genotype 1b BK and four 1b RDRPs derived from HCV-infected patients. Moreover, silymarin did not inhibit HCV replication in five independent genotype 1a, 1b, and 2a replicon cell lines that did not produce infectious virus. Silymarin inhibited microsomal triglyceride transfer protein activity, apolipoprotein B secretion, and infectious virion production into culture supernatants. Silymarin also blocked cell-to-cell spread of virus.

Conclusion: Although inhibition of in vitro NS5B polymerase activity is demonstrable, the mechanisms of silymarin's antiviral action appear to include blocking of virus entry and transmission, possibly by targeting the host cell.
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http://dx.doi.org/10.1002/hep.23587DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2909978PMC
June 2010

Apolipoprotein E on hepatitis C virion facilitates infection through interaction with low-density lipoprotein receptor.

Virology 2009 Nov 13;394(1):99-108. Epub 2009 Sep 13.

Department of Immunology, University of Washington School of Medicine, Seattle, WA, USA.

Hepatitis C virus (HCV) infection is a major cause of liver disease. HCV associates with host apolipoproteins and enters hepatocytes through complex processes involving some combination of CD81, claudin-1, occludin, and scavenger receptor BI. Here we show that infectious HCV resembles very low density lipoprotein (VLDL) and that entry involves co-receptor function of the low-density lipoprotein receptor (LDL-R). Blocking experiments demonstrate that beta-VLDL itself or anti-apolipoprotein E (apoE) antibody can block HCV entry. Knockdown of the LDL-R by treatment with 25-hydroxycholesterol or siRNA ablated ligand uptake and reduced HCV infection of cells, whereas infection was rescued upon cell ectopic LDL-R expression. Analyses of gradient-fractionated HCV demonstrate that apoE is associated with HCV virions exhibiting peak infectivity and dependence upon the LDL-R for cell entry. Our results define the LDL-R as a cooperative HCV co-receptor that supports viral entry and infectivity through interaction with apoE ligand present in an infectious HCV/lipoprotein complex comprising the virion. Disruption of HCV/LDL-R interactions by altering lipoprotein metabolism may therefore represent a focus for future therapy.
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http://dx.doi.org/10.1016/j.virol.2009.08.037DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2767442PMC
November 2009

Fighting the flu with inflammasome signaling.

Immunity 2009 Apr;30(4):476-8

Department of Immunology, University of Washington, Seattle, WA 98195, USA.

A wide variety of stimuli induce the inflammasome, but little is known about its role in immune protection against viruses. In this issue of Immunity, Allen et al. (2009) and Thomas et al. (2009) describe a critical role for NLRP3 induction of the inflammasome and protection against influenza virus infection.
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http://dx.doi.org/10.1016/j.immuni.2009.03.011DOI Listing
April 2009

Evasion and disruption of innate immune signalling by hepatitis C and West Nile viruses.

Cell Microbiol 2009 Jun 31;11(6):880-8. Epub 2009 Mar 31.

Department of Immunology, University of Washington School of Medicine, Seattle, WA 98195, USA.

Signalling pathways leading to type I interferon production are the first line of defence employed by the host to combat viruses, and represent a barrier that an invading virus must overcome in order to establish infection. In this review we highlight the ability of two members of the Flaviviridae, a globally distributed family of RNA viruses that represent a significant public health concern, to disrupt and evade these defences. Hepatitis C virus is a hepatotropic virus, infecting greater than 170 million people worldwide, while West Nile virus is a neurotropic virus that causes encephalitis in humans and horses. While these viruses cause distinct disease phenotypes, the ability of pathogenic strains to modulate the innate immune response is a key factor in influencing disease outcome. Both viruses have evolved unique strategies to target various aspects of type I interferon induction and signalling in order to prevent viral clearance and to promote virus replication.
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http://dx.doi.org/10.1111/j.1462-5822.2009.01311.xDOI Listing
June 2009

Innate immunity induced by composition-dependent RIG-I recognition of hepatitis C virus RNA.

Nature 2008 Jul 11;454(7203):523-7. Epub 2008 Jun 11.

Department of Immunology, University of Washington School of Medicine, Seattle, Washington 98195-7650, USA.

Innate immune defences are essential for the control of virus infection and are triggered through host recognition of viral macromolecular motifs known as pathogen-associated molecular patterns (PAMPs). Hepatitis C virus (HCV) is an RNA virus that replicates in the liver, and infects 200 million people worldwide. Infection is regulated by hepatic immune defences triggered by the cellular RIG-I helicase. RIG-I binds PAMP RNA and signals interferon regulatory factor 3 activation to induce the expression of interferon-alpha/beta and antiviral/interferon-stimulated genes (ISGs) that limit infection. Here we identify the polyuridine motif of the HCV genome 3' non-translated region and its replication intermediate as the PAMP substrate of RIG-I, and show that this and similar homopolyuridine or homopolyriboadenine motifs present in the genomes of RNA viruses are the chief feature of RIG-I recognition and immune triggering in human and murine cells. 5' terminal triphosphate on the PAMP RNA was necessary but not sufficient for RIG-I binding, which was primarily dependent on homopolymeric ribonucleotide composition, linear structure and length. The HCV PAMP RNA stimulated RIG-I-dependent signalling to induce a hepatic innate immune response in vivo, and triggered interferon and ISG expression to suppress HCV infection in vitro. These results provide a conceptual advance by defining specific homopolymeric RNA motifs within the genome of HCV and other RNA viruses as the PAMP substrate of RIG-I, and demonstrate immunogenic features of the PAMP-RIG-I interaction that could be used as an immune adjuvant for vaccine and immunotherapy approaches.
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http://dx.doi.org/10.1038/nature07106DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2856441PMC
July 2008

Interferon regulatory factor-3 activation, hepatic interferon-stimulated gene expression, and immune cell infiltration in hepatitis C virus patients.

Hepatology 2008 Mar;47(3):799-809

Liver Center, Department of Medicine, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA, USA.

Unlabelled: Interferon regulatory factor-3 (IRF-3) activation directs alpha/beta interferon production and interferon-stimulated gene (ISG) expression, which limits virus infection. Here, we examined the distribution of hepatitis C virus (HCV) nonstructural 3 protein, the status of IRF-3 activation, and expression of IRF-3 target genes and ISGs during asynchronous HCV infection in vitro and in liver biopsies from patients with chronic HCV infection, using confocal microscopy and functional genomics approaches. In general, asynchronous infection with HCV stimulated a low-frequency and transient IRF-3 activation within responsive cells in vitro that was associated with cell-to-cell virus spread. Similarly, a subset of HCV patients exhibited the nuclear, active form of IRF-3 in hepatocytes and an associated increase in IRF-3 target gene expression in hepatic tissue. Moreover, ISG expression profiles formed disease-specific clusters for HCV and control nonalcoholic fatty liver disease patients, with increased ISG expression among the HCV patients. We identified the presence of T cell and plasmacytoid dendritic cell infiltrates within all biopsy specimens, suggesting they could be a source of hepatic interferon in the setting of hepatitis C and chronic inflammatory condition.

Conclusion: These results indicate that HCV can transiently trigger IRF-3 activation during virus spread and that in chronic HCV, IRF-3 activation within infected hepatocytes occurs but is limited.
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http://dx.doi.org/10.1002/hep.22076DOI Listing
March 2008

Hepatitis C virus production by human hepatocytes dependent on assembly and secretion of very low-density lipoproteins.

Proc Natl Acad Sci U S A 2007 Apr 21;104(14):5848-53. Epub 2007 Mar 21.

Department of Molecular Genetics, University of Texas Southwestern Medical Center, Dallas, TX 75390-9046, USA.

Hepatitis C virus (HCV) and triglyceride-rich very low-density lipoproteins (VLDLs) both are secreted uniquely by hepatocytes and circulate in blood in a complex. Here, we isolated from human hepatoma cells the membrane vesicles in which HCV replicates. These vesicles, which contain the HCV replication complex, are highly enriched in proteins required for VLDL assembly, including apolipoprotein B (apoB), apoE, and microsomal triglyceride transfer protein. In hepatoma cells that constitutively produce infectious HCV, HCV production is reduced by two agents that block VLDL assembly: an inhibitor of microsomal triglyceride transfer protein and siRNA directed against apoB. These results provide a possible explanation for the restriction of HCV production to the liver and suggest new cellular targets for treatment of HCV infection.
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http://dx.doi.org/10.1073/pnas.0700760104DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1829327PMC
April 2007

Functional and therapeutic analysis of hepatitis C virus NS3.4A protease control of antiviral immune defense.

J Biol Chem 2007 Apr 8;282(14):10792-803. Epub 2007 Feb 8.

Department of Microbiology, University of Texas Southwestern Medical Center, Dallas, Texas 75390, USA.

Chronic hepatitis C virus (HCV) infection is a major global public health problem. HCV infection is supported by viral strategies to evade the innate antiviral response wherein the viral NS3.4A protease complex targets and cleaves the interferon promoter stimulator-1 (IPS-1) adaptor protein to ablate signaling of interferon alpha/beta immune defenses. Here we examined the structural requirements of NS3.4A and the therapeutic potential of NS3.4A inhibitors to control the innate immune response against virus infection. The structural composition of NS3 includes an amino-terminal serine protease domain and a carboxyl-terminal RNA helicase domain. NS3 mutants lacking the helicase domain retained the ability to control virus signaling initiated by retinoic acid-inducible gene-I (RIG-I) or melanoma differentiation antigen 5 and suppressed the downstream activation of interferon regulatory factor-3 (IRF-3) and nuclear factor kappaB (NF-kappaB) through the targeted proteolysis of IPS-1. This regulation was abrogated by truncation of the NS3 protease domain or by point mutations that ablated protease activity. NS3.4A protease control of antiviral immune signaling was due to targeted proteolysis of IPS-1 by the NS3 protease domain and minimal NS4A cofactor. Treatment of HCV-infected cells with an NS3 protease inhibitor prevented IPS-1 proteolysis by the HCV protease and restored RIG-I immune defense signaling during infection. Thus, the NS3.4A protease domain can target IPS-1 for cleavage and is essential for blocking RIG-I signaling to IRF-3 and NF-kappaB, whereas the helicase domain is dispensable for this action. Our results indicate that NS3.4A protease inhibitors have immunomodulatory potential to restore innate immune defenses to HCV infection.
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http://dx.doi.org/10.1074/jbc.M610361200DOI Listing
April 2007

Viral and therapeutic control of IFN-beta promoter stimulator 1 during hepatitis C virus infection.

Proc Natl Acad Sci U S A 2006 Apr 3;103(15):6001-6. Epub 2006 Apr 3.

Department of Microbiology, University of Texas Southwestern Medical Center, Dallas, TX 75390, USA.

Viral signaling through retinoic acid-inducible gene-I (RIG-I) and its adaptor protein, IFN promoter-stimulator 1 (IPS-1), activates IFN regulatory factor-3 (IRF-3) and the host IFN-alpha/beta response that limits virus infection. The hepatitis C virus (HCV) NS3/4A protease cleaves IPS-1 to block RIG-I signaling, but how this regulation controls the host response to HCV is not known. Moreover, endogenous IPS-1 cleavage has not been demonstrated in the context of HCV infection in vitro or in vivo. Here, we show that HCV infection transiently induces RIG-I- and IPS-1-dependent IRF-3 activation. This host response limits HCV production and constrains cellular permissiveness to infection. However, HCV disrupts this response early in infection by NS3/4A cleavage of IPS-1 at C508, releasing IPS-1 from the mitochondrial membrane. Cleavage results in subcellular redistribution of IPS-1 and loss of interaction with RIG-I, thereby preventing downstream activation of IRF-3 and IFN-beta induction. Liver tissues from chronically infected patients similarly demonstrate subcellular redistribution of IPS-1 in infected hepatocytes and IPS-1 cleavage associated with a lack of ISG15 expression and conjugation of target proteins in vivo. Importantly, small-molecule inhibitors of NS3/4A prevent cleavage and restore RIG-I signaling of IFN-beta induction. Our results suggest a dynamic model in which early activation of IRF-3 and induction of antiviral genes are reversed by IPS-1 proteolysis and abrogation of RIG-I signaling as NS3/4A accumulates in newly infected cells. HCV protease inhibitors effectively prevent IPS-1 proteolysis, suggesting they may be capable of restoring this innate host response in clinical practice.
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http://dx.doi.org/10.1073/pnas.0601523103DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1458687PMC
April 2006
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