Shin-Ru Shih

Chang Gung University, Hsin-chu-hsien, Taiwan, Taiwan

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Publications (120)432.69 Total impact

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    ABSTRACT: Background Highly pathogenic influenza viruses cause high levels of morbidity, including excessive infiltration of leukocytes into the lungs, high viral loads and a cytokine storm. However, the details of how these pathological features unfold in severe influenza infections remain unclear. Accumulation of Gr1¿+¿CD11b¿+¿myeloid cells has been observed in highly pathogenic influenza infections but it is not clear how and why they accumulate in the severely inflamed lung. In this study, we selected this cell population as a target to investigate the extreme inflammatory response during severe influenza infection.ResultsWe established H1N1 IAV-infected mouse models using three viruses of varying pathogenicity and noted the accumulation of a defined Gr1¿+¿CD11b¿+¿myeloid population correlating with the pathogenicity. Herein, we reported that CCR2+ inflammatory monocytes are the major cell compartments in this population. Of note, impaired clearance of the high pathogenicity virus prolonged IFN expression, leading to CCR2+ inflammatory monocytes amplifying their own recruitment via an interferon-¿/ß receptor 1 (IFNAR1)-triggered chemokine loop. Blockage of IFNAR1-triggered signaling or inhibition of viral replication by Oseltamivir significantly suppresses the expression of CCR2 ligands and reduced the influx of CCR2+ inflammatory monocytes. Furthermore, trafficking of CCR2+ inflammatory monocytes from the bone marrow to the lung was evidenced by a CCR2-dependent chemotaxis. Importantly, leukocyte infiltration, cytokine storm and expression of iNOS were significantly reduced in CCR2¿/¿ mice lacking infiltrating CCR2+ inflammatory monocytes, enhancing the survival of the infected mice.Conclusions Our results indicated that uncontrolled viral replication leads to excessive production of inflammatory innate immune responses by accumulating CCR2+ inflammatory monocytes, which contribute to the fatal outcomes of high pathogenicity virus infections.
    Journal of biomedical science. 11/2014; 21(1):99.
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    ABSTRACT: The roles of virus-derived small RNAs (vsRNAs) have been studied in plants and insects. However, the generation and function of small RNAs from cytoplasmic RNA viruses in mammalian cells remain unexplored. This study describes four vsRNAs that were detected in enterovirus 71-infected cells using next-generation sequencing and northern blots. Viral infection produced substantial levels (>10(5) copy numbers per cell) of vsRNA1, one of the four vsRNAs. We also demonstrated that Dicer is involved in vsRNA1 generation in infected cells. vsRNA1 overexpression inhibited viral translation and internal ribosomal entry site (IRES) activity in infected cells. Conversely, blocking vsRNA1 enhanced viral yield and viral protein synthesis. We also present evidence that vsRNA1 targets stem-loop II of the viral 5' untranslated region and inhibits the activity of the IRES through this sequence-specific targeting. Our study demonstrates the ability of a cytoplasmic RNA virus to generate functional vsRNA in mammalian cells. In addition, we also demonstrate a potential novel mechanism for a positive-stranded RNA virus to regulate viral translation: generating a vsRNA that targets the IRES.
    Nucleic Acids Research 10/2014; · 8.81 Impact Factor
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    ABSTRACT: Enterovirus 71 (EV71) infections can cause hand, foot, and mouth disease with severe neurological complications. Because no clinical drug is available for treating EV71 infections, developing an efficient antiviral medication against EV71 infection is crucial. This study indicated that 6-bromo-2-[1-(2,5-dimethylphenyl)-5-methyl-1H-pyrazol-4-yl] quinoline-4-carboxylic acid (BPR-3P0128) exhibits excellent antiviral activity against EV71 (EC50=0.0029μM). BPR-3P0128 inhibits viral replication during the early post infection stage, targets EV71 RNA-dependent RNA polymerase and VPg uridylylation, and also reduces viral RNA accumulation levels and inhibits viral replication of EV71. Copyright © 2014 Elsevier B.V. All rights reserved.
    Antiviral research. 10/2014; 112C:18-25.
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    ABSTRACT: Neural progenitor cells (NPCs) are stem cells that can differentiate into various neural lineage cells. The damage and loss of NPCs are associated with neurological conditions such as cognitive deficits and memory impairment. In a long-term study of patients with EV71, cognitive disorders were observed. Therefore, we hypothesized that NPCs may be permissive to EV71 infection. We demonstrated that NPCs are prone to EV71 infection and that these stem cells can support the active replication of this virus. Furthermore, EV71 infection triggers apoptosis, resulting in significant cell death in infected NPCs. However, EV71 did not replicate in the differentiated cell types that were tested. Our findings suggest that EV71 can infect NPCs and cause the depletion of these cells.
    Virology. 10/2014; 468-470C:592-600.
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    ABSTRACT: Enterovirus 71 (EV71) infections can cause hand, foot, and mouth disease with severe neurological complications. Because no clinical drug is available for treating EV71 infections, developing an efficient antiviral medication against EV71 infection is crucial. This study indicated that 6-bromo-2-[1-(2, 5-dimethylphenyl)-5-methyl-1H-pyrazol-4-yl] quinoline-4-carboxylic acid (BPR-3P0128) exhibits excellent antiviral activity against EV71 (EC50 = 0.0029 lM). BPR-3P0128 inhibits viral replication during the early post infection stage, targets EV71 RNA-dependent RNA polymerase and VPg uridylylation, and also reduces viral RNA accumulation levels and inhibits viral replication of EV71.
    Antiviral Research 08/2014; 112(2014):18-25. · 3.93 Impact Factor
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    ABSTRACT: Infections of the novel avian influenza A H7N9 virus cause severe respiratory diseases and death. In this study, to develop highly sensitive methods for differentially detecting the H7N9 virus, multiplex and singular real-time reverse transcription polymerase chain reaction (RT-PCR) assays were established and examined by targeting the H7 and N9 genes of the H7N9 virus. Furthermore, an additional multiplex assay combining previous real-time RT-PCR designs was established to subtype the pandemic H1N1, H3, and H5 influenza viruses. Applying the proposed assay system to analyze 100 clinical specimens collected from respiratory infection cases identified influenza A viruses (pandemic H1N1 and H3) in 23 samples. It has been demonstrated that other common respiratory viruses will not be detected by using this platform.
    Journal of virological methods. 07/2014;
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    ABSTRACT: Introduction: Enterovirus 71 (EV71) is an etiological agent that causes severe neurological complications in children. EV71 outbreaks have occurred throughout the Asia-Pacific region, posing a severe global public health threat; however, no specific therapeutic strategy exists for treating EV71-infected children. Areas covered: Five manufacturers have produced inactivated EV71 whole virus vaccines in mainland China, Taiwan, and Singapore, which have completed Phase III (mainland China) and Phase I (Taiwan and Singapore) clinical trials. Various EV71 vaccine candidates are being researched in animal models, including live-attenuated virus vaccine, recombinant VP1 vaccine, VP1-based DNA vaccine, synthetic peptide vaccine and virus-like particle vaccine. In this review, the present situation is summarized, and feasible improvements to the EV71 vaccine are explored. Expert opinion: Although inactivated EV71 vaccines are safe, efficient and elicit strong immune responses to protect adults, children and infants against infection, the quality control of production is critical.
    Expert opinion on biological therapy. 07/2014;
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    ABSTRACT: The primary role of cytoplasmic viral RNA-dependent RNA polymerase (RdRp) is viral genome replication in the cellular cytoplasm. However, picornaviral RdRp denoted 3D polymerase (3Dpol) also enters the host nucleus, where its function remains unclear. In this study, we describe a novel mechanism of viral attack in which 3Dpol enters the nucleus through the nuclear localization signal (NLS) and targets the pre-mRNA processing factor 8 (Prp8) to block pre-mRNA splicing and mRNA synthesis. The fingers domain of 3Dpol associates with the C-terminal region of Prp8, which contains the Jab1/MPN domain, and interferes in the second catalytic step, resulting in the accumulation of the lariat form of the splicing intermediate. Endogenous pre-mRNAs trapped by the Prp8-3Dpol complex in enterovirus-infected cells were identified and classed into groups associated with cell growth, proliferation, and differentiation. Our results suggest that picornaviral RdRp disrupts pre-mRNA splicing processes, that differs from viral protease shutting off cellular transcription and translation which contributes to the pathogenesis of viral infection.
    PLoS Pathogens 06/2014; 10(6):e1004199. · 8.14 Impact Factor
  • Peng-Nien Huang, Shin-Ru Shih
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    ABSTRACT: Human enterovirus type 71 (EV71) has emerged as a major cause of viral encephalitis in children worldwide. The identified EV71 receptors provide useful information for understanding EV71replication and tissue tropism. Host factors interact with the internal ribosome entry site (IRES) of EV71 to regulate viral translation. However, the specific molecular features of the EV71 genome that determine virulence remain unclear. The EV71 capsid protein VP1 region might contribute to virulence and neurotropism. Transgenic mice expressing the EV71 receptor that were infected with the virus exhibited a disease similar to that observed in infected humans. Antiviral drug and vaccine development is urgently required to prevent EV71 epidemics. Delineating viral host interactions and identifying specific mechanisms that might control the neural tropism of EV71 pathogenesis would be substantial advances.
    Current opinion in virology. 04/2014; 5C:98-104.
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    Jing-Yi Lin, Shin-Ru Shih
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    ABSTRACT: Enterovirus 71 (EV71) is a member of Picornaviridae that causes mild and self-limiting hand, foot, and mouth disease (HFMD). However, EV71 infections can progress to polio-like paralysis, neurogenic pulmonary edema, and fatal encephalitis in infants and young children. Large EV71 outbreaks have been reported in Taiwan, China, Japan, Malaysia, Singapore, and Australia. This virus is considered a critical emerging public health threat. EV71 is an important crucial neurotropic enterovirus for which there is currently no effective antiviral drug or vaccine. The mechanism by which EV71 causes severe central nervous system complications remains unclear. The interaction between the virus and the host is vital for viral replication, virulence, and pathogenicity. SCARB2 or PSGL-1 receptor binding is the first step in the development of viral infections, and viral factors (e.g., 5[prime] UTR, VPI, 3A, 3C, 3D), host factors and environments (e.g., ITAFs, type I IFN) are also involved in viral infections. The tissue tropism and pathogenesis of viruses are determined by a combination of several factors. This review article provides a summary of host and virus factors affecting cell and tissue tropism and the pathogenesis of enteroviruses.
    Journal of Biomedical Science 03/2014; 21(1):18. · 2.46 Impact Factor
  • Peng-Nien Huang, Shin-Ru Shih
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    ABSTRACT: Human enterovirus type 71 (EV71) has emerged as a major cause of viral encephalitis in children worldwide. The identified EV71 receptors provide useful information for understanding EV71replication and tissue tropism. Host factors interact with the internal ribosome entry site (IRES) of EV71 to regulate viral translation. However, the specific molecular features of the EV71 genome that determine virulence remain unclear. The EV71 capsid protein VP1 region might contribute to virulence and neurotropism. Transgenic mice expressing the EV71 receptor that were infected with the virus exhibited a disease similar to that observed in infected humans. Antiviral drug and vaccine development is urgently required to prevent EV71 epidemics. Delineating viral host interactions and identifying specific mechanisms that might control the neural tropism of EV71 pathogenesis would be substantial advances.
    Current Opinion in Virology. 01/2014; 5:98–104.
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    ABSTRACT: Enterovirus 71 (EV-A71) is a neurotropic virus that can cause severe complications involving the central nervous system. No effective antiviral therapeutics are available for treating EV-A71 infection and drug discovery efforts are rarely focused to target this disease. Thus, the main goal of this study was to discover existing drugs with novel indications that may effectively inhibit EV-A71 replication and the inflammatory cytokines elevation. In this study, we showed that LiCl, a GSK3β inhibitor, effectively suppressed EV-A71 replication, apoptosis and inflammatory cytokines production (Interleukin 6, Interleukin-1β) in infected cells. Furthermore, LiCl and an immunomodular agent were shown to strongly synergize with each other in suppressing EV-A71 replication. The results highlighted potential new treatment regimens in suppressing sequelae caused by EV-A71 replication.
    PLoS ONE 01/2014; 9(11):e111331. · 3.53 Impact Factor
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    ABSTRACT: Neuraminidase (NA) is a homotetramer viral surface glycoprotein that is essential for virus release during influenza virus infections. Previous studies have not explored why influenza NA forms a tetramer when the bacterial monomer NA already exhibits excellent NA enzymatic activity levels. In this study, we focused on 28 highly conserved residues among all NA subtypes, identifying 21 of 28 positions as crucial residues for viral survival by using reverse genetics. Maintaining NA enzymatic activity levels is critical and numerous conserved residues were located at the oligomerization interface; however, these mutations did not affect NA enzymatic activity levels or NA cellular localization, but rather affected the stability of NA oligomerization, suggesting that the oligomerization of NA is essential for viral viability. An increased understanding of the biological functions of NA, in particular NA oligomerization, could facilitate an alternative design for antivirals to combat influenza virus infections.
    Virology 12/2013; 447(1-2):32-44. · 3.35 Impact Factor
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    ABSTRACT: Background. The cutaneous manifestations of human enterovirus (HEV) infection are usually limited, such as hand-foot-mouth disease. By comparison, Stevens-Johnson syndrome (SJS) is a life-threatening severe cutaneous adverse reaction (SCAR), mainly caused by drugs. During the HEV outbreaks in 2010 to 2012 in Taiwan, we identified 21 patients developed widespread blistering mucocutaneous reactions without any suspected drug causality.Methods. We screened possible pathogen(s) for detecting human herpes virus (HHV1∼HHV7), HEV, or Mycoplasma pneumoniae infections using throat swab virus cultures, real-time PCR, DNA sequencing, immunochemistry and electron microscopy analyses.Results. Coxsackievirus A6 (CVA6) DNA was identified in the blistering skin lesions in 6 of 21 patients. Cytotoxic T lymphocytes and natural killer cells expressing granulysin predominantly infiltrated into the skin lesions, sharing the histopathological features with SJS. Intact CVA6 viral particles were identified in the blister fluids and skin lesions by electron microscopy. The phylogenetic analysis of the viral genome showed the CVA6 DNA sequence sharing higher similarity (97.6-98.1%) to CVA6 strains reported from Finland at 2008.Conclusions. This study identifies a new variant of CVA6 as the causative agent for severe mucocutaneous blistering reactions mimicking SCAR. An awareness of this unusual presentation of HEV infection is needed in the epidemic area.
    The Journal of Infectious Diseases 07/2013; · 5.85 Impact Factor
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    ABSTRACT: Background. Reassortment within polymerase genes causes changes in the pathogenicity of influenza A viruses. We previously reported that the 2009 pH1N1 PA enhanced the pathogenicity of seasonal H1N1. We examined the effects of the PA gene from the HPAI H5N1 following its introduction into currently circulating seasonal influenza viruses.Methods. To evaluate the role of H5N1 PA in altering the virulence of seasonal influenza viruses, we generated a recombinant seasonal H3N2 (3446) that expressed the H5N1 PA protein (VPA), and evaluated the RNP activity, growth kinetics, and pathogenicity of the reassortant virus in mice.Results. Compared with the wild type 3446 virus, the substitution of the H5N1 PA gene into the 3446 virus (VPA/3446) resulted in increased RNP activity, and an increased replication rate in A549 cells. The recombinant VPA/3446 virus also caused more severe pneumonia in Casp 1(-/-) mice than in IL1β(-/-) and wild type B6 mice.Conclusions. Although the PA from H5N1 is incidentally compatible with a seasonal H3N2 backbone, the H5N1 PA affected the virulence of seasonal H3N2, particularly in inflammasome-related innate immunity deficient mice. These findings highlight the importance of monitoring PA reassortment in seasonal flu, and confirm the role of the Caspase-1 gene in influenza pathogenesis.
    The Journal of Infectious Diseases 07/2013; · 5.85 Impact Factor
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    ABSTRACT: BACKGROUND/PURPOSE: Human infections by a new avian influenza A (H7N9) virus have been reported. As of April 23, 2013, there were 108 confirmed cases including 22 deaths in China. MATERIALS AND METHODS: Influenza protein sequences were downloaded from the Influenza Virus Resource and GISAID EpiFlu databases. Pairwise nucleotide identities were computed for assessing the evolutionary distance of H7N9 to other known avian and human viruses, and multiple sequence alignments with their position-specific entropy values were used in discussing how mutations on species-associated signature positions were introduced in the new H7N9 which may steer its way to human infection. RESULTS: This report analyzed the genomic characteristics of this new H7N9 virus. Nucleotide sequence analysis clearly reveals its origin from avian viruses. In this article, we particularly focus on its internal genes that are found to derive from H9N2-another subtype of avian influenza A virus which has been circulating in birds for years. Amino acid sequences at species-specific genomic positions were examined. Although the new virus contains mostly avian-like residues at these signature positions, it does contain several human-like signatures. For instance, at the position 627 of PB2, the new virus has human-characteristic K instead of avian-characteristic E; in addition, PB2-627K, PA-100A, PA-356R, and PA-409N are also human-like signatures in the new H7N9 virus. CONCLUSION: The new H7N9 is an avian influenza A virus; however, it does harbor several human virus-like signatures, which raises great concern that it may have a higher probability to cross species barriers and infect humans.
    Journal of the Formosan Medical Association 06/2013; · 1.00 Impact Factor
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    ABSTRACT: Molecular chaperones are reported to be crucial for virus propagation, but are not yet addressed in Human Enterovirus 71 (EV71). Here we describe the specific association of heat shock protein-90-beta (Hsp90β), but not alpha form (Hsp90α), with EV71 viral particles by the co-purification with virions using sucrose density gradient ultracentrifugation, and by the colocalization with viral particles, as assessed by immunogold electron microscopy. The reduction of the Hsp90β protein using RNA interference decreased the correct assembly of viral particles, without affecting EV71 replication levels. Tracking ectopically expressed Hsp90β protein associated with EV71 virions revealed that Hsp90β protein was transmitted to new host cells through its direct association with infectious viral particles. Our findings suggest a new antiviral strategy in which extracellular Hsp90β protein is targeted to decrease the infectivity of EV71 and other enteroviruses, without affecting the broader functions of this constitutively expressed molecular chaperone.
    Virology 05/2013; · 3.35 Impact Factor
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    ABSTRACT: BACKGROUND: Although human papillomavirus (HPV) infections have been causally linked to oral cavity squamous cell carcinoma (OSCC), the potential role of low-risk HPV (LR-HPV) types in the pathogenesis of this malignancy remains unclear. OBJECTIVES: We sought to investigate the distribution of HPV genotypes and their prognostic significance in OSCC patients treated by radical surgery, either with or without adjuvant therapy. STUDY DESIGN: We studied two non-overlapping OSCC cohorts for the periods 2005-2006 (2005 cohort, n=204) and 2010-2011 (2010 cohort, n=206). Paraffin-embedded tissue blocks were collected, and the HPV genotype was determined using PCR plus HPV blot tests. The primary study endpoint was the prevalence of HPV genotypes. The secondary endpoints were the 2-year therapeutic outcomes. RESULTS: The overall prevalence of HPV infections did not differ significantly in the two study cohorts. However, the prevalence of LR-HPV was significantly higher in the 2010 cohort than in the 2005 cohort (p=0.002). The overall prevalence of HPV infections was not significantly associated with the 2-year outcomes. However, multivariate analysis demonstrated that LR-HPV infection was a predictor of poor 2-year disease-free survival (p=0.036, hazard ratio [HR]=3.1), disease-specific survival (p=0.014, HR=3.8), and overall survival (p=0.016, HR=3.2) in the subgroups of OSCC patients with poor differentiation, pN2 lymph node metastases, or extracapsular spread (n=150). CONCLUSIONS: LR-HPV infections may have an important role in determining the clinical outcomes of certain OSCC patients bearing specific risk factors.
    Journal of clinical virology: the official publication of the Pan American Society for Clinical Virology 05/2013; · 3.12 Impact Factor
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    ABSTRACT: Recombination plays an important role in shaping the genetic diversity of a number of DNA and RNA viruses. Although some recent studies have reported bioinformatic evidence of mosaic sequences in a variety of influenza A viruses, it remains controversial as to whether these represent bona fide natural recombination events or laboratory artifacts. Importantly, mosaic genome structures can create significant topological incongruence during phylogenetic analyses, which can mislead additional phylogeny-based molecular evolutionary analyses such as molecular clock dating, the detection of selection pressures and phylogeographic inference. As a result, there is a strong need for systematic screenings for mosaic structures within the influenza virus genome database. We used a combination of sequence-based and phylogeny-based methods to identify 388 mosaic influenza genomic segments, of which 332 are previously unreported and are significantly supported by phylogenetic methods. It is impossible, however, to ascertain whether these represent natural recombinants. To facilitate the future identification of recombinants, reference sets of non-recombinant sequences were selected for use in an automatic screening protocol for detecting mosaic sequences. Tests using real and simulated mosaic sequences indicate that our screening protocol is both sensitive (average >90%) and accurate (average >77%) enough to identify a range of different mosaic patterns. The relatively high prevalence of mosaic influenza virus sequences implies that efficient systematic screens, such as that proposed here, should be performed routinely to detect natural recombinant strains, potential laboratory artifacts, and sequencing contaminants either prior to sequences being deposited in GenBank or before they are used for phylogenetic analyses.
    Infection, genetics and evolution: journal of molecular epidemiology and evolutionary genetics in infectious diseases 03/2013; · 3.22 Impact Factor

Publication Stats

2k Citations
432.69 Total Impact Points

Institutions

  • 2000–2014
    • Chang Gung University
      • • Center for Emerging Viral Infections Research
      • • Department of Medical Biotechnology and Laboratory Science
      • • Graduate Institute of Clinical Medicine Sciences
      • • Department of Pediatrics
      Hsin-chu-hsien, Taiwan, Taiwan
  • 2010–2012
    • National Institutes of Health
      Maryland, United States
  • 2004–2012
    • Chang Gung Memorial Hospital
      • • Department of Laboratory Medicine
      • • Division of Pediatric Infectious Diseases
      T’ai-pei, Taipei, Taiwan
    • Robert Wood Johnson University Hospital
      New Brunswick, New Jersey, United States
  • 2010–2011
    • Mackay Memorial Hospital
      • Department of Pediatrics
      Taipei, Taipei, Taiwan
  • 2002–2011
    • National Health Research Institutes
      • Institute of Biotechnology and Pharmaceutical Research
      Miaoli, Taiwan, Taiwan
  • 2009
    • National Tsing Hua University
      • Institute of Molecular Medicine
      Hsin-chu-hsien, Taiwan, Taiwan
  • 2008
    • National Chiao Tung University
      • Department of Biological Science and Technology
      Hsinchu, Taiwan, Taiwan
  • 2007
    • National Chung Cheng University
      Chia-i-hsien, Taiwan, Taiwan