Publications by authors named "Guanqun Liu"

29 Publications

  • Page 1 of 1

SCU-Net: Semantic Segmentation Network for Learning Channel Information on Remote Sensing Images.

Comput Intell Neurosci 2022 10;2022:8469415. Epub 2022 Apr 10.

School of Computer and Communication Engineering, Changsha University of Science and Technology, Changsha 410114, China.

Extracting detailed information from remote sensing images is an important direction in semantic segmentation. Not only the amounts of parameters and calculations of the network model in the learning process but also the prediction effect after learning must be considered. This paper designs a new module, the upsampling convolution-deconvolution module (CDeConv). On the basis of CDeConv, a convolutional neural network (CNN) with a channel attention mechanism for semantic segmentation is proposed as a channel upsampling network (SCU-Net). SCU-Net has been verified by experiments. The mean intersection-over-union (MIOU) of the SCU-Net-102-A model reaches 55.84%, the pixel accuracy is 91.53%, and the frequency weighted intersection-over-union (FWIU) is 85.83%. Compared with some of the state-of-the-art methods, SCU-Net can learn more detailed information in the channel and has better generalization capabilities.
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http://dx.doi.org/10.1155/2022/8469415DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC9013575PMC
April 2022

SARS-CoV-2 learned the 'Alpha'bet of immune evasion.

Nat Immunol 2022 03;23(3):351-353

Cleveland Clinic Florida Research and Innovation Center, Port St. Lucie, FL, USA.

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http://dx.doi.org/10.1038/s41590-022-01148-8DOI Listing
March 2022

NLRP3 Inflammasome Activation Enhanced by TRIM25 is Targeted by the NS1 Protein of 2009 Pandemic Influenza A Virus.

Front Microbiol 2021 12;12:778950. Epub 2021 Nov 12.

Vaccine and Infectious Disease Organization (VIDO), University of Saskatchewan, Saskatoon, SK, Canada.

Nucleotide-binding domain and leucine-rich repeat-containing protein 3 (NLRP3) inflammasome-mediated interleukin-1 beta (IL-1β) production is one of the crucial responses in innate immunity upon infection with viruses including influenza A virus (IAV) and is modulated by both viral and host cellular proteins. Among host proteins involved, we identified tripartite motif-containing protein 25 (TRIM25) as a positive regulator of porcine NLRP3 inflammasome-mediated IL-1β production. TRIM25 achieved this function by enhancing the pro-caspase-1 interaction with apoptosis-associated speck-like protein containing caspase recruitment domain (ASC). The N-terminal RING domain, particularly residues predicted to be critical for the E3 ligase activity of TRIM25, was responsible for this enhancement. However, non-structural protein 1 (NS1) C-terminus of 2009 pandemic IAV interfered with this action by interacting with TRIM25, leading to diminished association between pro-caspase-1 and ASC. These findings demonstrate that TRIM25 promotes the IL-1β signaling, while it is repressed by IAV NS1 protein, revealing additional antagonism of the NS1 against host pro-inflammatory responses.
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http://dx.doi.org/10.3389/fmicb.2021.778950DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8633893PMC
November 2021

Lactate induces aberration in the miR-30a-DBF4 axis to promote the development of gastric cancer and weakens the sensitivity to 5-Fu.

Cancer Cell Int 2021 Nov 10;21(1):602. Epub 2021 Nov 10.

Department of Clinical Laboratory, Qilu Hospital, Shandong University, 107 Wenhuaxi Road, Jinan, 250012, Shandong, P.R. China.

Background: Gastric cancer (GC) is one of the most common malignancies, molecular mechanism of which is still not clear. Aberrant expression of tumor-associated genes is the major cause of tumorigenesis. DBF4 is an important factor in cancers, although there is yet no report on its function and molecular mechanism in GC.

Methods: The expression of DBF4 in tumor tissues or cells of GC was detected by qRT-PCR and western blotting. Gastric cancer cell line MGC-803 and AGS were transfected with DBF4 siRNA or overexpression vector to detect the function of DBF4 in proliferation, migration and the sensitivity to 5-Fu with CCK-8 assay, colony formation assay, transwell assay, and wound healing assay. miR-30a was found to be the regulator of DBF4 by online bioinformatics software and confirmed with qRT-PCR, western blot and dual-luciferase reporter assays.

Results: In our study, increased expression of DBF4 in GC tissues was first identified through The Cancer Genome Atlas (TCGA) and later confirmed using specimens from GC patients. Furthermore, functional experiments were applied to demonstrate that DBF4 promotes cell proliferation and migration in GC cell lines, moreover weakens the sensitivity of MGC803 and AGS cells to 5-Fu. We further demonstrated that miR-30a showed significantly lower expression in GC cells and inhibited the expression of DBF4 through 3'-UTR suppression. Furthermore, rescue experiments revealed that the miR-30a-DBF4 axis regulated the GC cell proliferation, migration and the sensitivity to 5-Fu. The important composition in tumor microenvironment, lactate, may be the primary factor that suppressed miR-30a to strengthen the expression of DBF4.

Conclusions: Taken together, our study was the first to identify DBF4 as a regulator of cell proliferation and migration in GC. Furthermore, our study identified the lactate-miR-30a-DBF4 axis as a crucial regulator of tumor progression and the tumor sensitivity to 5-Fu, which maybe serve useful for the development of novel therapeutic targets.
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http://dx.doi.org/10.1186/s12935-021-02291-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8582204PMC
November 2021

An improved method of recharge sources analysis and its application in an unconfined aquifer.

J Environ Manage 2021 Jul 28;290:112582. Epub 2021 Apr 28.

Key Lab of Marine Environmental Science and Ecology, Ministry of Education, College of Environmental Science and Engineering, Ocean University of China, Qingdao, 266100, PR China; College of Environmental Science and Engineering, Ocean University of China, Qingdao, 266100, China.

Groundwater recharge sources analysis, including identification of the recharge sources and calculation of the mixing ratios, is of great importance for hydrogeological research and water resources management. In this research, a new approach, multivariate mixing and mass-balance calculations (M3) model combined with MIX calculations (M3-MIX calculations), was proposed to overcome shortcomings and limitations of existing methods and to accurately describe aquifer systems with more than three groundwater sources and get more accurate mixing ratios. A synthetic case with random sources were applied to evaluate the effectiveness of M3-MIX calculations. The results of both mixing ratios and composition of recharge sources show that M3-MIX calculations is superior to traditional methods such as least squares, and is also superior to the results obtained by using M3 model or MIX calculations alone. The approach is then applied to analyze groundwater recharge sources of the Huangshui River groundwater reservoir, China. Three recharge sources were calculated based on M3-MIX calculations: brackish groundwater affected by seawater intrusion, atmospheric precipitation, and groundwater from upstream affected by agricultural activities. The mixing ratios of the three recharge sources are 3.3%, 19.3%, and 77.4%, respectively. In addition, ion concentrations deviate from the mixing line indicates that hydrochemical processes, such as mineral dissolution/precipitation and ion exchange processes, may occur in the study area. The results of this work indicate that M3-MIX calculations has the potential to provide the accurate understanding of groundwater recharge, and thus providing useful information for the exploitation, utilization, and protection of groundwater in unsaturated and saturated zones.
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http://dx.doi.org/10.1016/j.jenvman.2021.112582DOI Listing
July 2021

Mutations derived from horseshoe bat ACE2 orthologs enhance ACE2-Fc neutralization of SARS-CoV-2.

PLoS Pathog 2021 04 9;17(4):e1009501. Epub 2021 Apr 9.

Department of Immunology and Microbiology, The Scripps Research Institute, Jupiter, FL, United States of America.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike (S) protein mediates infection of cells expressing angiotensin-converting enzyme 2 (ACE2). ACE2 is also the viral receptor of SARS-CoV (SARS-CoV-1), a related coronavirus that emerged in 2002-2003. Horseshoe bats (genus Rhinolophus) are presumed to be the original reservoir of both viruses, and a SARS-like coronavirus, RaTG13, closely related to SARS-CoV-2, has been identified in one horseshoe-bat species. Here we characterize the ability of the S-protein receptor-binding domains (RBDs) of SARS-CoV-1, SARS-CoV-2, pangolin coronavirus (PgCoV), RaTG13, and LyRa11, a bat virus similar to SARS-CoV-1, to bind a range of ACE2 orthologs. We observed that the PgCoV RBD bound human ACE2 at least as efficiently as the SARS-CoV-2 RBD, and that both RBDs bound pangolin ACE2 efficiently. We also observed a high level of variability in binding to closely related horseshoe-bat ACE2 orthologs consistent with the heterogeneity of their RBD-binding regions. However five consensus horseshoe-bat ACE2 residues enhanced ACE2 binding to the SARS-CoV-2 RBD and neutralization of SARS-CoV-2 pseudoviruses by an enzymatically inactive immunoadhesin form of human ACE2 (hACE2-NN-Fc). Two of these mutations impaired neutralization of SARS-CoV-1 pseudoviruses. An hACE2-NN-Fc variant bearing all five mutations neutralized both SARS-CoV-2 pseudovirus and infectious virus more efficiently than wild-type hACE2-NN-Fc. These data suggest that SARS-CoV-1 and -2 originate from distinct bat species, and identify a more potently neutralizing form of soluble ACE2.
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http://dx.doi.org/10.1371/journal.ppat.1009501DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8059821PMC
April 2021

ISG15-dependent activation of the sensor MDA5 is antagonized by the SARS-CoV-2 papain-like protease to evade host innate immunity.

Nat Microbiol 2021 04 16;6(4):467-478. Epub 2021 Mar 16.

Florida Research and Innovation Center, Cleveland Clinic, Port St Lucie, FL, USA.

Activation of the RIG-I-like receptors, retinoic-acid inducible gene I (RIG-I) and melanoma differentiation-associated protein 5 (MDA5), establishes an antiviral state by upregulating interferon (IFN)-stimulated genes (ISGs). Among these is ISG15, the mechanistic roles of which in innate immunity still remain enigmatic. In the present study, we report that ISG15 conjugation is essential for antiviral IFN responses mediated by the viral RNA sensor MDA5. ISGylation of the caspase activation and recruitment domains of MDA5 promotes its oligomerization and thereby triggers activation of innate immunity against a range of viruses, including coronaviruses, flaviviruses and picornaviruses. The ISG15-dependent activation of MDA5 is antagonized through direct de-ISGylation mediated by the papain-like protease of SARS-CoV-2, a recently emerged coronavirus that has caused the COVID-19 pandemic. Our work demonstrates a crucial role for ISG15 in the MDA5-mediated antiviral response, and also identifies a key immune evasion mechanism of SARS-CoV-2, which may be targeted for the development of new antivirals and vaccines to combat COVID-19.
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http://dx.doi.org/10.1038/s41564-021-00884-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8103894PMC
April 2021

Viral Evasion of RIG-I-Like Receptor-Mediated Immunity through Dysregulation of Ubiquitination and ISGylation.

Viruses 2021 01 26;13(2). Epub 2021 Jan 26.

Florida Research and Innovation Center, Cleveland Clinic, Port Saint Lucie, FL 34987, USA.

Viral dysregulation or suppression of innate immune responses is a key determinant of virus-induced pathogenesis. Important sensors for the detection of virus infection are the RIG-I-like receptors (RLRs), which, in turn, are antagonized by many RNA viruses and DNA viruses. Among the different escape strategies are viral mechanisms to dysregulate the post-translational modifications (PTMs) that play pivotal roles in RLR regulation. In this review, we present the current knowledge of immune evasion by viral pathogens that manipulate ubiquitin- or ISG15-dependent mechanisms of RLR activation. Key viral strategies to evade RLR signaling include direct targeting of ubiquitin E3 ligases, active deubiquitination using viral deubiquitinating enzymes (DUBs), and the upregulation of cellular DUBs that regulate RLR signaling. Additionally, we summarize emerging new evidence that shows that enzymes of certain coronaviruses such as SARS-CoV-2, the causative agent of the current COVID-19 pandemic, actively deISGylate key molecules in the RLR pathway to escape type I interferon (IFN)-mediated antiviral responses. Finally, we discuss the possibility of targeting virally-encoded proteins that manipulate ubiquitin- or ISG15-mediated innate immune responses for the development of new antivirals and vaccines.
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http://dx.doi.org/10.3390/v13020182DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7910861PMC
January 2021

ISG15-dependent Activation of the RNA Sensor MDA5 and its Antagonism by the SARS-CoV-2 papain-like protease.

bioRxiv 2020 Oct 27. Epub 2020 Oct 27.

Florida Research and Innovation Center, Cleveland Clinic, FL 34987, USA.

Activation of the RIG-I-like receptors, RIG-I and MDA5, establishes an antiviral state by upregulating interferon (IFN)-stimulated genes (ISGs). Among these is ISG15 whose mechanistic roles in innate immunity still remain enigmatic. Here we report that ISGylation is essential for antiviral IFN responses mediated by the viral RNA sensor MDA5. ISG15 conjugation to the caspase activation and recruitment domains of MDA5 promotes the formation of higher-order assemblies of MDA5 and thereby triggers activation of innate immunity against a range of viruses including coronaviruses, flaviviruses and picornaviruses. The ISG15-dependent activation of MDA5 is antagonized through direct de-ISGylation mediated by the papain-like protease (PLpro) of SARS-CoV-2, a recently emerged coronavirus that causes the COVID-19 pandemic. Our work demonstrates a crucial role for ISG15 in the MDA5-mediated antiviral response, and also identifies a novel immune evasion mechanism of SARS-CoV-2, which may be targeted for the development of new antivirals and vaccines to combat COVID-19.
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http://dx.doi.org/10.1101/2020.10.26.356048DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7605552PMC
October 2020

Distinct and Orchestrated Functions of RNA Sensors in Innate Immunity.

Immunity 2020 07;53(1):26-42

Department of Microbiology, The University of Chicago, Chicago, IL 60637, USA. Electronic address:

Faithful maintenance of immune homeostasis relies on the capacity of the cellular immune surveillance machinery to recognize "nonself", such as the presence of pathogenic RNA. Several families of pattern-recognition receptors exist that detect immunostimulatory RNA and then induce cytokine-mediated antiviral and proinflammatory responses. Here, we review the distinct features of bona fide RNA sensors, Toll-like receptors and retinoic-acid inducible gene-I (RIG-I)-like receptors in particular, with a focus on their functional specificity imposed by cell-type-dependent expression, subcellular localization, and ligand preference. Furthermore, we highlight recent advances on the roles of nucleotide-binding oligomerization domain (NOD)-like receptors and DEAD-box or DEAH-box RNA helicases in an orchestrated RNA-sensing network and also discuss the relevance of RNA sensor polymorphisms in human disease.
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http://dx.doi.org/10.1016/j.immuni.2020.03.017DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7367493PMC
July 2020

Dysregulation of type I interferon responses in COVID-19.

Nat Rev Immunol 2020 07 26;20(7):397-398. Epub 2020 May 26.

Department of Microbiology, The University of Chicago, Chicago, IL, USA.

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http://dx.doi.org/10.1038/s41577-020-0346-xDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7249038PMC
July 2020

Acquisition of Avian-Origin PB1 Facilitates Viral RNA Synthesis by the 2009 Pandemic H1N1 Virus Polymerase.

Viruses 2020 02 28;12(3). Epub 2020 Feb 28.

Vaccine and Infectious Disease Organization - International Vaccine Centre (VIDO-InterVac), University of Saskatchewan, Saskatoon, SK S7N 5E3, Canada.

The constant crosstalk between the large avian reservoir of influenza A viruses (IAV) and its mammalian hosts drives viral evolution and facilitates their host switching. Direct adaptation of an avian strain to human or reassortment between avian-origin gene segments with that of human strains are the two mechanisms for the emergence of pandemic viruses. While it was suggested that the 1918 pandemic virus is of avian origin, reassortment of 1918 human isolates and avian influenza viruses led to the generation of 1957 and 1968 pandemic viruses. Interestingly, the avian PB1 segment, which encodes the catalytic subunit of IAV polymerase, is present in the 1957 and 1968 pandemic viruses. The biological consequence and molecular basis of such gene exchange remain less well understood. Using the 2009 pandemic H1N1 virus as a model, whose polymerase contains a human-origin PB1 subunit, we demonstrate that the acquisition of an avian PB1 markedly enhances viral RNA synthesis. This enhancement is also effective in the absence of PB2 adaptive mutations, which are key determinants of host switching. Mechanistically, the avian-origin PB1 does not appear to affect polymerase assembly but imparts the reassorted pandemic polymerase-augmented viral primary transcription and replication. Moreover, compared to the parental pandemic polymerase, the reassorted polymerase displays comparable complementary RNA (cRNA)-stabilizing activity but is specifically enhanced in progeny viral RNA (vRNA) synthesis from cRNA in a trans-activating manner. Overall, our results provide the first insight into the mechanism via which avian-origin PB1 enhances viral RNA synthesis of the 2009 pandemic virus polymerase.
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http://dx.doi.org/10.3390/v12030266DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7150768PMC
February 2020

Innate immunemodulator containing adjuvant formulated HA based vaccine protects mice from lethal infection of highly pathogenic avian influenza H5N1 virus.

Vaccine 2020 02 31;38(10):2387-2395. Epub 2020 Jan 31.

Vaccine and Infectious Disease Organization - International Vaccine Center (VIDO-InterVac), University of Saskatchewan, Saskatoon, Canada; Department of Veterinary Microbiology, Western College of Veterinary Medicine, University of Saskatchewan, Saskatoon, Canada. Electronic address:

The highly pathogenic avian influenza (HPAI) H5N1 viruses and their spillover into the human population pose substantial economic and public health threats. Although antiviral drugs have some effect in treating influenza infection, vaccination is still the most effective intervention to prevent possible pandemic outbreaks. We have developed a novel H5 influenza vaccine to improve the world's pandemic preparedness. We produced a hemagglutinin (HA) of HPAI H5N1 virus A/Alberta/01/2014 (AB14) using both mammalian (m) and bacterial (b) expression systems. The purified recombinant proteins were formulated with a proprietary adjuvant (TriAdj) and their efficacy as vaccine candidates was evaluated in mice. Intramuscular delivery of two doses of TriAdj formulated mammalian expressed HA (m-HA/TriAdj) was shown to provide full protection against a lethal challenge of AB14 in mice. In contrast, bacterially expressed HA with TriAdj (b-HA/TriAdj), b-HA without adjuvant, and m-HA without adjuvant resulted in no protection in immunized mice. Furthermore, m-HA/TriAdj elicited significantly higher levels of balanced Th1 and Th2 responses and neutralizing antibody titres. All the mice in the m-HA/TriAdj group survived a lethal AB14 H5N1 challenge and showed no signs of disease or infection as demonstrated by no loss of body weight or detectable virus in the lungs. Our results suggest that m-HA formulated with TriAdj has potential to protect against pandemic H5N1 in the event of its cross over to the human host.
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http://dx.doi.org/10.1016/j.vaccine.2020.01.051DOI Listing
February 2020

MiR-135b protects cardiomyocytes from infarction through restraining the NLRP3/caspase-1/IL-1β pathway.

Int J Cardiol 2020 05 17;307:137-145. Epub 2019 Nov 17.

Department of Pharmacology (State-Province Key Laboratories of Biomedicine- Pharmaceutics of China, Key Laboratory of Cardiovascular Medicine Research, Ministry of Education), College of Pharmacy, Harbin Medical University, Harbin 150081, PR China; Translational Medicine Research and Cooperation Center of Northern China, Heilongjiang Academy of Medical Sciences, Harbin 150081, PR China. Electronic address:

Background: Myocardial infarction (MI) is the most common cause of cardiovascular morbidity and mortality worldwide. Despite the identification of many pathogenic genes associated with MI, the underlying molecular mechanisms remain poorly understood. MicroRNAs (miRNAs, miRs), which regulate target genes at the post-transcriptional level, play a significant role in the regulation of cardiovascular diseases such as MI. Pyroptosis is a caspase-1-dependent pro-inflammatory programmed cell death (PCD) mechanism. The role of pyroptosis in several diseases associated with various miRNAs has been studied extensively. Meanwhile, the role of NOD-like receptor-containing pyrin 3 (NLRP3)/caspase-1/interleukin-1β (IL-1β) pathway in cardiac diseases has also been more recognized.

Methods: We established a mice MI model which ligated with the left anterior descending coronary artery and a cardiomyocytes injury model treated by hydrogen peroxide (HO) to detect the expressions of miR-135b and NLRP3/caspase-1/IL-1β pathway. Then miR-135b mimic, agomir-135b, and α-MHC-miR-135b transgenic mice were used to evaluate the effects of miR-135b overexpression.

Result: We demonstrated that miR-135b was downregulated after cardiomyocytes injury both in vivo and in vitro. Pyroptosis pathway was also activated. MiR-135b overexpression remarkably restored impaired cardiac function and attenuated the upregulation of NLRP3/caspase-1/IL-1β pathway.

Conclusions: The present findings shed light on the protective role of miR-135b in MI mediated by the inhibition of the NLRP3/caspase-1/IL-1β pathway.
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http://dx.doi.org/10.1016/j.ijcard.2019.09.055DOI Listing
May 2020

Zika Virus NS3 Mimics a Cellular 14-3-3-Binding Motif to Antagonize RIG-I- and MDA5-Mediated Innate Immunity.

Cell Host Microbe 2019 10;26(4):493-503.e6

Department of Microbiology, The University of Chicago, Chicago, IL 60637, USA. Electronic address:

14-3-3 protein family members facilitate the translocation of RIG-I-like receptors (RLRs) to organelles that mediate downstream RLR signaling, leading to interferon production. 14-3-3ϵ promotes the cytosolic-to-mitochondrial translocation of RIG-I, while 14-3-3η facilitates MDA5 translocation to mitochondria. We show that the NS3 protein of Zika virus (ZIKV) antagonizes antiviral gene induction by RIG-I and MDA5 by binding to and sequestering the scaffold proteins 14-3-3ϵ and 14-3-3η. 14-3-3-binding is mediated by a negatively charged RLDP motif in NS3 that is conserved in ZIKV strains of African and Asian lineages and is similar to the one found in dengue and West Nile viruses. ZIKV NS3 is sufficient to inhibit the RLR-14-3-3ϵ/η interaction and to suppress antiviral signaling. Mutational perturbation of 14-3-3ϵ/η binding in a recombinant ZIKV leads to enhanced innate immune responses and impaired growth kinetics. Our study provides molecular understanding of immune evasion functions of ZIKV, which may guide vaccine and anti-flaviviral therapy development.
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http://dx.doi.org/10.1016/j.chom.2019.09.012DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6922055PMC
October 2019

In Vivo Characterization of Avian Influenza A (H5N1) and (H7N9) Viruses Isolated from Canadian Travelers.

Viruses 2019 02 23;11(2). Epub 2019 Feb 23.

Vaccine and Infectious Disease Organization - International Vaccine Centre (VIDO-InterVac), University of Saskatchewan, Saskatoon, SK S7N 5E3, Canada.

Highly pathogenic avian influenza (HPAI) H5N1 and low pathogenic avian influenza (LPAI) H7N9 viruses pose a severe threat to public health through zoonotic infection, causing severe respiratory disease in humans. While HPAI H5N1 human infections have typically been reported in Asian countries, avian H7N9 human infections have been reported mainly in China. However, Canada reported a case of fatal human infection by the HPAI H5N1 virus in 2014, and two cases of human illness associated with avian H7N9 virus infection in 2015. While the genomes of the causative viruses A/Alberta/01/2014 (H5N1) (AB14 (H5N1)) and A/British Columbia/1/2015 (H7N9) (BC15 (H7N9)) are reported, the isolates had not been evaluated for their pathogenicity in animal models. In this study, we characterized the pathogenicity of AB14 (H5N1) and BC15 (H7N9) and found that both strain isolates are highly lethal in mice. AB14 (H5N1) caused systemic viral infection and erratic proinflammatory cytokine gene expression in different organs. In contrast, BC15 (H7N9) replicated efficiently only in the respiratory tract, and was a potent inducer for proinflammatory cytokine genes in the lungs. Our study provides experimental evidence to complement the specific human case reports and animal models for evaluating vaccine and antiviral candidates against potential influenza pandemics.
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http://dx.doi.org/10.3390/v11020193DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6409709PMC
February 2019

Cytoplasm and Beyond: Dynamic Innate Immune Sensing of Influenza A Virus by RIG-I.

J Virol 2019 04 3;93(8). Epub 2019 Apr 3.

Vaccine and Infectious Disease Organization-International Vaccine Centre (VIDO-InterVac), University of Saskatchewan, Saskatoon, Saskatchewan, Canada

Innate immune sensing of influenza A virus (IAV) requires retinoic acid-inducible gene I (RIG-I), a fundamental cytoplasmic RNA sensor. How RIG-I's cytoplasmic localization reconciles with the nuclear replication nature of IAV is poorly understood. Recent findings provide advanced insights into the spatiotemporal RIG-I sensing of IAV and highlight the contribution of various RNA ligands to RIG-I activation. Understanding a compartment-specific RIG-I-sensing paradigm would facilitate the identification of the full spectrum of physiological RIG-I ligands produced during IAV infection.
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http://dx.doi.org/10.1128/JVI.02299-18DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6450113PMC
April 2019

Inhibition of Ongoing Influenza A Virus Replication Reveals Different Mechanisms of RIG-I Activation.

J Virol 2019 03 5;93(6). Epub 2019 Mar 5.

Vaccine and Infectious Disease Organization-International Vaccine Centre (VIDO-InterVac), University of Saskatchewan, Saskatoon, Saskatchewan, Canada

Pattern recognition receptors provide essential nonself immune surveillance within distinct cellular compartments. Retinoic acid-inducible gene I (RIG-I) is one of the primary cytosolic RNA sensors, with an emerging role in the nucleus. It is involved in the spatiotemporal sensing of influenza A virus (IAV) replication, leading to the induction of type I interferons (IFNs). Nonetheless, the physiological viral ligands activating RIG-I during IAV infection remain underexplored. Other than full-length viral genomes, cellular constraints that impede ongoing viral replication likely potentiate an erroneous viral polymerase generating aberrant viral RNA species with RIG-I-activating potential. Here, we investigate the origins of RIG-I-activating viral RNA under two such constraints. Using chemical inhibitors that inhibit continuous viral protein synthesis, we identify the incoming, but not -synthesized, viral defective interfering (DI) genomes contributing to RIG-I activation. In comparison, deprivation of viral nucleoprotein (NP), the key RNA chain elongation factor for the viral polymerase, leads to the production of aberrant viral RNA species activating RIG-I; however, their nature is likely to be distinct from that of DI RNA. Moreover, RIG-I activation in response to NP deprivation is not adversely affected by expression of the nuclear export protein (NEP), which diminishes the generation of a major subset of aberrant viral RNA but facilitates the accumulation of small viral RNA (svRNA). Overall, our results indicate the existence of fundamentally different mechanisms of RIG-I activation under cellular constraints that impede ongoing IAV replication. The induction of an IFN response by IAV is mainly mediated by the RNA sensor RIG-I. The physiological RIG-I ligands produced during IAV infection are not fully elucidated. Cellular constraints leading to the inhibition of ongoing viral replication likely potentiate an erroneous viral polymerase producing aberrant viral RNA species activating RIG-I. Here, we demonstrate that RIG-I activation during chemical inhibition of continuous viral protein synthesis is attributable to the incoming DI genomes. Erroneous viral replication driven by NP deprivation promotes the generation of RIG-I-activating aberrant viral RNA, but their nature is likely to be distinct from that of DI RNA. Our results thus reveal distinct mechanisms of RIG-I activation by IAV under cellular constraints impeding ongoing viral replication. A better understanding of RIG-I sensing of IAV infection provides insight into the development of novel interventions to combat influenza virus infection.
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http://dx.doi.org/10.1128/JVI.02066-18DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6401434PMC
March 2019

The NS1 Protein of Influenza A Virus Participates in Necroptosis by Interacting with MLKL and Increasing Its Oligomerization and Membrane Translocation.

J Virol 2019 01 4;93(2). Epub 2019 Jan 4.

Vaccine and Infectious Disease Organization-International Vaccine Centre (VIDO-InterVac), University of Saskatchewan, Saskatoon, Saskatchewan, Canada

Elimination of infected cells by programmed cell death is a well-recognized host defense mechanism to control the spread of infection. In addition to apoptosis, necroptosis is also one of the mechanisms of cell death that can be activated by viral infection. Activation of necroptosis leads to the phosphorylation of mixed-lineage kinase domain-like protein (MLKL) by receptor-interacting protein kinase 3 (RIPK3) and results in MLKL oligomerization and membrane translocation, leading to membrane disruption and a loss of cellular ion homeostasis. It has recently been reported that influenza A virus (IAV) infection induces necroptosis. However, the underlying mechanism of the IAV-mediated necroptosis process, particularly the roles of IAV proteins in necroptosis, remains unexplored. Here, we report that IAV infection induces necroptosis in macrophages and epithelial cells. We demonstrate that the NS1 protein of IAV interacts with MLKL. Coiled-coil domain 2 of MLKL has a predominant role in mediating the MLKL interaction with NS1. The interaction of NS1 with MLKL increases MLKL oligomerization and membrane translocation. Moreover, the MLKL-NS1 interaction enhances MLKL-mediated NLRP3 inflammasome activation, leading to increased interleukin-1β (IL-1β) processing and secretion. Necroptosis is a programmed cell death that is inflammatory in nature owing to the release of danger-associated molecular patterns from the ruptured cell membrane. However, necroptosis also constitutes an important arm of host immune responses. Thus, a balanced inflammatory response determines the disease outcome. We report that the NS1 protein of IAV participates in necroptosis by interacting with MLKL, resulting in increased MLKL oligomerization and membrane translocation. These results reveal a novel function of the NS1 protein and the mechanism by which IAV induces necroptosis. Moreover, we show that this interaction enhances NLRP3 inflammasome activation and IL-1β processing and secretion. This information may contribute to a better understanding of the role of necroptosis in IAV-induced inflammation.
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http://dx.doi.org/10.1128/JVI.01835-18DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6321931PMC
January 2019

Nuclear-resident RIG-I senses viral replication inducing antiviral immunity.

Nat Commun 2018 08 10;9(1):3199. Epub 2018 Aug 10.

Vaccine and Infectious Disease Organization-International Vaccine Centre (VIDO-InterVac), University of Saskatchewan, Saskatoon, SK, S7N 5E3, Canada.

The nucleus represents a cellular compartment where the discrimination of self from non-self nucleic acids is vital. While emerging evidence establishes a nuclear non-self DNA sensing paradigm, the nuclear sensing of non-self RNA, such as that from nuclear-replicating RNA viruses, remains unexplored. Here, we report the identification of nuclear-resident RIG-I actively involved in nuclear viral RNA sensing. The nuclear RIG-I, along with its cytoplasmic counterpart, senses influenza A virus (IAV) nuclear replication leading to a cooperative induction of type I interferon response. Its activation signals through the canonical signaling axis and establishes an effective antiviral state restricting IAV replication. The exclusive signaling specificity conferred by nuclear RIG-I is reinforced by its inability to sense cytoplasmic-replicating Sendai virus and appreciable sensing of hepatitis B virus pregenomic RNA in the nucleus. These results refine the RNA sensing paradigm for nuclear-replicating viruses and reveal a previously unrecognized subcellular milieu for RIG-I-like receptor sensing.
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http://dx.doi.org/10.1038/s41467-018-05745-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6086882PMC
August 2018

Swine Influenza Virus Induces RIPK1/DRP1-Mediated Interleukin-1 Beta Production.

Viruses 2018 08 9;10(8). Epub 2018 Aug 9.

Vaccine and Infectious Disease Organization-International Vaccine Centre (VIDO-InterVac), University of Saskatchewan, Saskatoon, SK S7N 5E3, Canada.

Nucleotide-binding domain and leucine-rich repeat-containing protein 3 (NLRP3) inflammasome plays a pivotal role in modulating lung inflammation in response to the influenza A virus infection. We previously showed that the swine influenza virus (SIV) infection induced NLRP3 inflammasome-mediated IL-1β production in primary porcine alveolar macrophages (PAMs), and we were interested in examining the upstream signaling events that are involved in this process. Here, we report that the SIV-infection led to dynamin-related protein 1 (DRP1) phosphorylation at serine 579 and mitochondrial fission in PAMs. IL-1β production was dependent on the reactive oxygen species (ROS) production, and DRP1 phosphorylation resulted in the upregulation of the NLRP3 inflammasome. Furthermore, the requirement of the kinase activity of receptor-interacting protein kinase 1 (RIPK1) for the IL-1β production and RIPK1-DRP1 association suggested that RIPK1 is an upstream kinase for DRP1 phosphorylation. Our results reveal a critical role of the RIPK1/DRP1 signaling axis, whose activation leads to mitochondrial fission and ROS release, in modulating porcine NLRP3 inflammasome-mediated IL-1β production in SIV-infected PAMs.
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http://dx.doi.org/10.3390/v10080419DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6116029PMC
August 2018

NS1 Protein of 2009 Pandemic Influenza A Virus Inhibits Porcine NLRP3 Inflammasome-Mediated Interleukin-1 Beta Production by Suppressing ASC Ubiquitination.

J Virol 2018 04 28;92(8). Epub 2018 Mar 28.

Vaccine and Infectious Disease Organization-International Vaccine Centre (VIDO-InterVac), University of Saskatchewan, Saskatoon, SK, Canada

The inflammasome represents a molecular platform for innate immune regulation and controls proinflammatory cytokine production. The NLRP3 inflammasome is comprised of NLRP3, ASC, and procaspase-1. When the NLRP3 inflammasome is activated, it causes ASC speck formation and caspase-1 activation, resulting in the maturation of interleukin-1β (IL-1β). The NLRP3 inflammasome is regulated at multiple levels, with one level being posttranslational modification. Interestingly, ubiquitination of ASC has been reported to be indispensable for the activation of the NLRP3 inflammasome. Influenza A virus (IAV) infection induces NLRP3 inflammasome-dependent IL-1β secretion, which contributes to the host antiviral defense. However, IAVs have evolved multiple antagonizing mechanisms, one of which is executed by viral NS1 protein to suppress the NLRP3 inflammasome. In this study, we compared IL-1β production in porcine alveolar macrophages in response to IAV infection and found that the 2009 pandemic H1N1 induced less IL-1β than swine influenza viruses (SIVs). Further study revealed that the NS1 C terminus of pandemic H1N1 but not that of SIV was able to significantly inhibit NLRP3 inflammasome-mediated IL-1β production. This inhibitory function was attributed to impaired ASC speck formation and suppression of ASC ubiquitination. Moreover, we identified two target lysine residues, K110 and K140, which are essential for both porcine ASC ubiquitination and NLRP3 inflammasome-mediated IL-1β production. These results revealed a novel mechanism by which the NS1 protein of the 2009 pandemic H1N1 suppresses NLRP3 inflammasome activation. Influenza A virus (IAV) infection activates the NLRP3 inflammasome, resulting in the production of IL-1β, which contributes to the host innate immune response. ASC, an adaptor protein of NLRP3, forms specks that are critical for inflammasome activation. Here, we report that the NS1 C terminus of the 2009 pandemic H1N1 has functions to suppress porcine IL-1β production by inhibiting ASC speck formation and ASC ubiquitination. Furthermore, the ubiquitination sites on porcine ASC were identified. The information gained here may contribute to an in-depth understanding of porcine inflammasome activation and regulation in response to different IAVs, helping to further enhance our knowledge of innate immune responses to influenza virus infection in pigs.
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http://dx.doi.org/10.1128/JVI.00022-18DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5874408PMC
April 2018

[A validation study of national early warning score in evaluation of death risk in elderly patients with critical illness].

Zhonghua Wei Zhong Bing Ji Jiu Yi Xue 2016 May;28(5):387-90

Objective: To verify the validity and feasibility of national early warning score (NEWS) in evaluation of death risk in elderly patients with critical illness,in order to find out which scoring method is more suitable for elderly critical illness patients.

Methods: A prospective case-control study was conducted. The critical illness patients aged over 60 years old with the length of hospital stay over 24 hours, and admitted to Department of Emergency of Qingdao Municipal Hospital from January to December 2015 were enrolled. The clinical data including in emergency and the actual outcome of patients were collected, and the patients were divided into death group and survival group according to 30-day outcome. Patients in the two groups were assessed by using NEWS and risk classification according to the first results of vital signs monitoring. Multivariate logistic regression model was used to analyze the relationship between the NEWS classification and the risk of death in elderly critical ill patients.

Results: 1 950 emergency elderly patients with critical illness were enrolled, with 78 cases (4.0%) dead within 30 days and 1 872 survived (96.0%).Compared with the survival group, patients in death group were older (years:79.8 ± 10.8 vs.75.3 ± 8.9,t =4.335,P <0.001),and had higher acute physiology and chronic health evaluation Ⅱ (APACHE Ⅱ) score (22.9± 4.6 vs.18.2 ± 4.8,t =8.487,P < 0.001),lower Glasgow coma scale (GCS) score (12.2 ± 4.5 vs.13.4 ± 5.2,t =-2.007,P =0.045),higher incidence of respiratory system diseases (29.5% vs.17.9%,x 2 =12.742,P =0.013),higher NEWS score (11.2 ± 5.5 vs.3.9 ± 2.7,t =22.063,P < 0.001),as well as higher proportion of patients with NEWS classification of high risk and very high risk (65.4% vs.15.8%,x 2 =263.125,P < 0.001).With the increase of NEWS risk classification, mortality rate was also increased, and the mortality rate in the patients with low, medium, high and very high risk were 0.81% (9/1 108),3.63% (18/496),5.83% (13/223),30.89% (38/123),respectively, with statistically significant difference (x 2 =179.741,P < 0.001).It was showed by logistic regression analysis that the NEWS score of elderly patients with critical illness were positively correlated with 30-day death. The 30-day death risk of patients with middle risk, high risk and very high risk was 4.600,9.052 and 54.598 folds of the patients with low risk respectively.

Conclusion: NEWS score can be used to assess the risk of death in emergency elderly patients with critical illness. NEWS risk classification can quantify and classify the risk of death in the elderly patients with critical illness.
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May 2016

DDX3 Interacts with Influenza A Virus NS1 and NP Proteins and Exerts Antiviral Function through Regulation of Stress Granule Formation.

J Virol 2016 Jan 20;90(7):3661-75. Epub 2016 Jan 20.

Vaccine and Infectious Disease Organization-International Vaccine Center, University of Saskatchewan, Saskatoon, Saskatchewan, Canada Vaccinology & Immunotherapeutics Program, School of Public Health, University of Saskatchewan, Saskatoon, Saskatchewan, Canada

Unlabelled: DDX3 belongs to the DEAD box RNA helicase family and is a multifunctional protein affecting the life cycle of a variety of viruses. However, its role in influenza virus infection is unknown. In this study, we explored the potential role of DDX3 in influenza virus life cycle and discovered that DDX3 is an antiviral protein. Since many host proteins affect virus life cycle by interacting with certain components of the viral machinery, we first verified whether DDX3 has any viral interaction partners. Immunoprecipitation studies revealed NS1 and NP as direct interaction partners of DDX3. Stress granules (SGs) are known to be antiviral and do form in influenza virus-infected cells expressing defective NS1 protein. Additionally, a recent study showed that DDX3 is an important SG-nucleating factor. We thus explored whether DDX3 plays a role in influenza virus infection through regulation of SGs. Our results showed that SGs were formed in infected cells upon infection with a mutant influenza virus lacking functional NS1 (del NS1) protein, and DDX3 colocalized with NP in SGs. We further determined that the DDX3 helicase domain did not interact with NS1 and NP; however, it was essential for DDX3 localization in virus-induced SGs. Knockdown of DDX3 resulted in impaired SG formation and led to increased virus titers. Taken together, our results identified DDX3 as an antiviral protein with a role in virus-induced SG formation.

Importance: DDX3 is a multifunctional RNA helicase and has been reported to be involved in regulating various virus life cycles. However, its function during influenza A virus infection remains unknown. In this study, we demonstrated that DDX3 is capable of interacting with influenza virus NS1 and NP proteins; DDX3 and NP colocalize in the del NS1 virus-induced SGs. Furthermore, knockdown of DDX3 impaired SG formation and led to a decreased virus titer. Thus, we provided evidence that DDX3 is an antiviral protein during influenza virus infection and its antiviral activity is through regulation of SG formation. Our findings provide knowledge about the function of DDX3 in the influenza virus life cycle and information for future work on manipulating the SG pathway and its components to fight influenza virus infection.
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http://dx.doi.org/10.1128/JVI.03010-15DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4794679PMC
January 2016

RNA interference of influenza A virus replication by microRNA-adapted lentiviral loop short hairpin RNA.

J Gen Virol 2015 Oct 17;96(10):2971-2981. Epub 2015 Jul 17.

Vaccine and Infectious Disease Organization - International Vaccine Center (VIDO-InterVac), University of Saskatchewan, Saskatoon, Saskatchewan, Canada.

Limitations of the current vaccines and antivirals against influenza A virus (IAV) pandemic underscore the urgent need for developing novel anti-influenza strategies. RNA interference (RNAi) induced by small interfering RNA (siRNA) has become a powerful new means to inhibit viral infection in a gene-specific manner. However, the efficacy of the siRNA delivery platform and the relatively high cost of administration have hindered widespread application of siRNA. In this study, we developed a microRNA (miRNA)-30-based lentivirus delivery system by embedding a synthetic short hairpin RNA (shRNA) stem into the context of endogenous precursor of miRNA-30 (shRNAmir) to express a silencer of the influenza gene. We showed that the miRNA-based lentivirus vector was able to express and process a single nucleoprotein (NP)-targeting shRNAmir, which could potently inhibit IAV replication. We further showed that miRNA-based lentivirus vector carrying tandemly linked NP and polymerase PB1 shRNAmirs could express and process double shRNAmirs. Despite the relatively low levels of NP and PB1 miRNAs produced in the stably transduced cells, the combination of two miRNAs exerted a great degree of inhibition on influenza infection. Given the advantage of combinatorial RNAi in preventing emergence of mutant virus, miRNA-based lentiviral vectors are valuable tools for anitiviral activities. To the best of our knowledge, this is the first study demonstrating that a miRNA-based RNAi strategy can be applied for better control of influenza virus infection.
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http://dx.doi.org/10.1099/jgv.0.000247DOI Listing
October 2015

Influenza A Virus Panhandle Structure Is Directly Involved in RIG-I Activation and Interferon Induction.

J Virol 2015 Jun 25;89(11):6067-79. Epub 2015 Mar 25.

Vaccine and Infectious Disease Organization-International Vaccine Center, University of Saskatchewan, Saskatoon, Saskatchewan, Canada Vaccinology & Immunotherapeutics Program, School of Public Health, University of Saskatchewan, Saskatoon, Saskatchewan, Canada Department of Veterinary Microbiology, Western College of Veterinary Medicine, University of Saskatchewan, Saskatoon, Saskatchewan, Canada

Unlabelled: Retinoic acid-inducible gene I (RIG-I) is an important innate immune sensor that recognizes viral RNA in the cytoplasm. Its nonself recognition largely depends on the unique RNA structures imposed by viral RNA. The panhandle structure residing in the influenza A virus (IAV) genome, whose primary function is to serve as the viral promoter for transcription and replication, has been proposed to be a RIG-I agonist. However, this has never been proved experimentally. Here, we employed multiple approaches to determine if the IAV panhandle structure is directly involved in RIG-I activation and type I interferon (IFN) induction. First, in porcine alveolar macrophages, we demonstrated that the viral genomic coding region is dispensable for RIG-I-dependent IFN induction. Second, using in vitro-synthesized hairpin RNA, we showed that the IAV panhandle structure could directly bind to RIG-I and stimulate IFN production. Furthermore, we investigated the contributions of the wobble base pairs, mismatch, and unpaired nucleotides within the wild-type panhandle structure to RIG-I activation. Elimination of these destabilizing elements within the panhandle structure promoted RIG-I activation and IFN induction. Given the function of the panhandle structure as the viral promoter, we further monitored the promoter activity of these panhandle variants and found that viral replication was moderately affected, whereas viral transcription was impaired dramatically. In all, our results indicate that the IAV panhandle promoter region adopts a nucleotide composition that is optimal for balanced viral RNA synthesis and suboptimal for RIG-I activation.

Importance: The IAV genomic panhandle structure has been proposed to be an RIG-I agonist due to its partial complementarity; however, this has not been experimentally confirmed. Here, we provide direct evidence that the IAV panhandle structure is competent in, and sufficient for, RIG-I activation and IFN induction. By constructing panhandle variants with increased complementarity, we demonstrated that the wild-type panhandle structure could be modified to enhance RIG-I activation and IFN induction. These panhandle variants posed moderate influence on viral replication but dramatic impairment of viral transcription. These results indicate that the IAV panhandle promoter region adopts a nucleotide composition to achieve optimal balance of viral RNA synthesis and suboptimal RIG-I activation. Our results highlight the multifunctional role of the IAV panhandle promoter region in the virus life cycle and offer novel insights into the development of antiviral agents aiming to boost RIG-I signaling or virus attenuation by manipulating this conserved region.
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http://dx.doi.org/10.1128/JVI.00232-15DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4442436PMC
June 2015

Suppression of ERK activation in urethral epithelial cells infected with Neisseria gonorrhoeae and its isogenic minD mutant contributes to anti-apoptosis.

Microbes Infect 2015 Apr 29;17(4):317-22. Epub 2014 Dec 29.

Department of Biology, College of Arts and Science, University of Saskatchewan, W.P. Thompson Building 112 Science Place, Saskatoon, Saskatchewan, S7N5E2, Canada; Vaccine and Infectious Disease Organization-International Vaccine Center (VIDO-InterVac), University of Saskatchewan, 120 Veterinary Road, Saskatoon, Saskatchewan, S7N5E3, Canada; Department of Microbiology and Immunology, College of Medicine, University of Saskatchewan, Saskatoon, Saskatchewan, S7N5E5, Canada. Electronic address:

In gonococci-infected transduced human urethral epithelial cells (THUEC), the role of ERK, a mitogen-activated protein kinase (MAPK), in apoptosis is unknown. We observed lowering of ERK activation in THUEC following infection with anti-apoptosis-inducing Neisseria gonorrhoeae strain CH811. An isogenic cell division mutant of this strain, Ng CJSD1 (minD deficient), which is large and abnormally shaped, reduced ERK phosphorylation levels even more than its parental strain in THUEC. This led to higher anti-apoptosis in mutant-infected cells as compared to the parental strain-infected cells. Our results suggest that N. gonorrhoeae infection reduces ERK activation in THUEC contributing to anti-apoptosis.
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http://dx.doi.org/10.1016/j.micinf.2014.12.012DOI Listing
April 2015

Estimating the submarine groundwater and nutrients discharge of Yellow River delta with cross-section method.

Water Sci Technol 2010 ;62(2):394-402

Key Lab of Marine Environmental Science and Ecology, Ministry of Education, College of Environmental Science and Engineering, Ocean University of China, Qingdao 266100, China.

In this paper, cross-section method was used to estimate the groundwater and nutrients discharge fluxing to the Bohai Sea from the Yellow River Delta. The flux of shallow phreatic groundwater (within 10 m) in the Yellow River Farm discharging into sea was 2.9x10(-5) m3/m d in 2004 and 3.1x10(-5) m3/m d in 2005. Time distribution monthly mean flux is consistent with the Yellow River's runoff but taking on lag effect. And the volume of the phreatic water discharging from the whole delta is 3.71-3.77x10(3) m3, which is 2x10(-5)% of the Yellow River's annual runoff. The transport amount of shallow confined water (buried depth 15-20 m) from 2004 to 2005 was 5.7-6.2x10(-3) m3/m d in the Yellow River delta, 0.0037-0.004% of the runoff of Yellow River. There is low concentration of NO3-, NH4+ and PO4(3-) but high dissolved SiO2 in the shallow confined aquifer. Despite the high concentration of phreatic nitrate, it weakly influences the seawater because of the little flux of discharge into sea.
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http://dx.doi.org/10.2166/wst.2010.279DOI Listing
October 2010

A case study to detect the leakage of underground pressureless cement sewage water pipe using GPR, electrical, and chemical data.

Environ Sci Technol 2002 Mar;36(5):1077-85

Department of Environmental Engineering, College of Geoscience, Ocean University of Qingdao, Shandong, PR China.

The exploration and determination of leakage of underground pressureless nonmetallic pipes is difficult to deal with. A comprehensive method combining Ground Penetrating Rader (GPR), electric potential survey and geochemical survey is introduced in the leakage detection of an underground pressureless nonmetallic sewage pipe in this paper. Theoretically, in the influencing scope of a leakage spot, the obvious changes of the electromagnetic properties and the physical-chemical properties of the underground media will be reflected as anomalies in GPR and electrical survey plots. The advantages of GPR and electrical survey are fast and accurate in detection of anomaly scope. In-situ analysis of the geophysical surveys can guide the geochemical survey. Then water and soil sampling and analyzing can be the evidence for judging the anomaly is caused by pipe leakage or not. On the basis of previous tests and practical surveys, the GPR waveforms, electric potential curves, contour maps, and chemical survey results are all classified into three types according to the extent or indexes of anomalies in orderto find out the leakage spots. When three survey methods all show their anomalies as type I in an anomalous spot, this spot is suspected as the most possible leakage location. Otherwise, it will be down grade suspected point. The suspect leakage spots should be confirmed by referring the site conditions because some anomalies are caused other factors. The excavation afterward proved that the method for determining the suspected location by anomaly type is effective and economic. Comprehensive method of GRP, electric potential survey, and geochemical survey is one of the effective methods in the leakage detection of underground nonmetallic pressureless pipe with its advantages of being fast and accurate.
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http://dx.doi.org/10.1021/es001954sDOI Listing
March 2002
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