scholarly journals Expression of Factor X in BHK-21 Cells Promotes Low Pathogenic Influenza Viruses Replication

2015 ◽  
Vol 2015 ◽  
pp. 1-6 ◽  
Author(s):  
Shahla Shahsavandi ◽  
Mohammad Majid Ebrahimi ◽  
Shahin Masoudi ◽  
Hasan Izadi
Keyword(s):  
Influenza Virus ◽  
Mdck Cells ◽  
High Titer ◽  
Influenza Viruses ◽  
Factor X ◽  
Proteolytic Activation ◽  
Influenza Virus Hemagglutinin ◽  
Pathogenic Avian Influenza Virus ◽  
Replication Procedure ◽  
Cell Expression

A cDNA clone for factor 10 (FX) isolated from chicken embryo inserted into the mammalian cell expression vector pCDNA3.1 was transfected into the baby hamster kidney (BHK-21) cell line. The generated BHK-21 cells with inducible expression of FX were used to investigate the efficacy of the serine transmembrane protease to proteolytic activation of influenza virus hemagglutinin (HA) with monobasic cleavage site. Data showed that the BHK-21/FX stably expressed FX after ten serial passages. The cells could proteolytically cleave the HA of low pathogenic avian influenza virus at multiplicity of infection 0.01. Growth kinetics of the virus on BHK-21/FX, BHK-21, and MDCK cells were evaluated by titrations of virus particles in each culture supernatant. Efficient multicycle viral replication was markedly detected in the cell at subsequent passages. Virus titration demonstrated that BHK-21/FX cell supported high-titer growth of the virus in which the viral titer is comparable to the virus grown in BHK-21 or MDCK cells with TPCK-trypsin. The results indicate potential application for the BHK-21/FX in influenza virus replication procedure and related studies.

scholarly journals TMPRSS2 and TMPRSS4 Facilitate Trypsin-Independent Spread of Influenza Virus in Caco-2 Cells

2010 ◽  
Vol 84 (19) ◽  
pp. 10016-10025 ◽  
Author(s):  
Stephanie Bertram ◽  
Ilona Glowacka ◽  
Paulina Blazejewska ◽  
Elizabeth Soilleux ◽  
Paul Allen ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Serine Proteases ◽  
Alveolar Epithelium ◽  
Influenza Viruses ◽  
Target Cells ◽  
Human Respiratory Tract ◽  
Proteolytic Activation ◽  
Viral Spread ◽  
Endogenous Expression ◽  
Influenza Virus Hemagglutinin

ABSTRACT Proteolysis of influenza virus hemagglutinin by host cell proteases is essential for viral infectivity, but the proteases responsible are not well defined. Recently, we showed that engineered expression of the type II transmembrane serine proteases (TTSPs) TMPRSS2 and TMPRSS4 allows hemagglutinin (HA) cleavage. Here we analyzed whether TMPRSS2 and TMPRSS4 are expressed in influenza virus target cells and support viral spread in the absence of exogenously added protease (trypsin). We found that transient expression of TMPRSS2 and TMPRSS4 resulted in HA cleavage and trypsin-independent viral spread. Endogenous expression of TMPRSS2 and TMPRSS4 in cell lines correlated with the ability to support the spread of influenza virus in the absence of trypsin, indicating that these proteases might activate influenza virus in naturally permissive cells. Indeed, RNA interference (RNAi)-mediated knockdown of both TMPRSS2 and TMPRSS4 in Caco-2 cells, which released fully infectious virus without trypsin treatment, markedly reduced the spread of influenza virus, demonstrating that these proteases were responsible for efficient proteolytic activation of HA in this cell line. Finally, TMPRSS2 was found to be coexpressed with the major receptor determinant of human influenza viruses, 2,6-linked sialic acids, in human alveolar epithelium, indicating that viral target cells in the human respiratory tract express TMPRSS2. Collectively, our results point toward an important role for TMPRSS2 and possibly TMPRSS4 in influenza virus replication and highlight the former protease as a potential therapeutic target.

scholarly journals The MAL Proteolipid Is Necessary for Normal Apical Transport and Accurate Sorting of the Influenza Virus Hemagglutinin in Madin-Darby Canine Kidney Cells

1999 ◽  
Vol 145 (1) ◽  
pp. 141-151 ◽  
Author(s):  
Rosa Puertollano ◽  
Fernando Martín-Belmonte ◽  
Jaime Millán ◽  
María del Carmen de Marco ◽  
Juan P. Albar ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Mdck Cells ◽  
Basolateral Membrane ◽  
Ectopic Expression ◽  
Apical Surface ◽  
Madin Darby Canine Kidney ◽  
Transport Pathway ◽  
Influenza Virus Hemagglutinin ◽  
Darby Canine Kidney ◽  
Canine Kidney

The MAL (MAL/VIP17) proteolipid is a nonglycosylated integral membrane protein expressed in a restricted pattern of cell types, including T lymphocytes, myelin-forming cells, and polarized epithelial cells. Transport of the influenza virus hemagglutinin (HA) to the apical surface of epithelial Madin-Darby canine kidney (MDCK) cells appears to be mediated by a pathway involving glycolipid- and cholesterol- enriched membranes (GEMs). In MDCK cells, MAL has been proposed previously as being an element of the protein machinery for the GEM-dependent apical transport pathway. Using an antisense oligonucleotide-based strategy and a newly generated monoclonal antibody to canine MAL, herein we have approached the effect of MAL depletion on HA transport in MDCK cells. We have found that MAL depletion diminishes the presence of HA in GEMs, reduces the rate of HA transport to the cell surface, inhibits the delivery of HA to the apical surface, and produces partial missorting of HA to the basolateral membrane. These effects were corrected by ectopic expression of MAL in MDCK cells whose endogenous MAL protein was depleted. Our results indicate that MAL is necessary for both normal apical transport and accurate sorting of HA.

scholarly journals A review of H5Nx avian influenza viruses

2019 ◽  
Vol 7 ◽  
pp. 251513551882162 ◽  
Author(s):  
Ivette A. Nuñez ◽  
Ted M. Ross
Keyword(s):  
Influenza Virus ◽  
Avian Influenza ◽  
Avian Influenza Virus ◽  
Highly Pathogenic Avian Influenza ◽  
Influenza Viruses ◽  
Avian Influenza Viruses ◽  
Highly Pathogenic ◽  
Bird Populations ◽  
Pathogenic Avian Influenza ◽  
Pathogenic Avian Influenza Virus

Highly pathogenic avian influenza viruses (HPAIVs), originating from the A/goose/Guangdong/1/1996 H5 subtype, naturally circulate in wild-bird populations, particularly waterfowl, and often spill over to infect domestic poultry. Occasionally, humans are infected with HPAVI H5N1 resulting in high mortality, but no sustained human-to-human transmission. In this review, the replication cycle, pathogenicity, evolution, spread, and transmission of HPAIVs of H5Nx subtypes, along with the host immune responses to Highly Pathogenic Avian Influenza Virus (HPAIV) infection and potential vaccination, are discussed. In addition, the potential mechanisms for Highly Pathogenic Avian Influenza Virus (HPAIV) H5 Reassorted Viruses H5N1, H5N2, H5N6, H5N8 (H5Nx) viruses to transmit, infect, and adapt to the human host are reviewed.

scholarly journals Proteolytic Activation of Influenza Viruses by Serine Proteases TMPRSS2 and HAT from Human Airway Epithelium

2006 ◽  
Vol 80 (19) ◽  
pp. 9896-9898 ◽  
Author(s):  
Eva Böttcher ◽  
Tatyana Matrosovich ◽  
Michaela Beyerle ◽  
Hans-Dieter Klenk ◽  
Wolfgang Garten ◽  
...  
Keyword(s):  
Serine Proteases ◽  
Mammalian Cells ◽  
Mdck Cells ◽  
Influenza Viruses ◽  
Human Respiratory Tract ◽  
Proteolytic Activation ◽  
Mammalian Expression ◽  
Human Airway Epithelium

ABSTRACT Host cell proteases that cleave the hemagglutinin (HA) of influenza viruses in the human respiratory tract are still not identified. Here we cloned two human type II transmembrane serine proteases with known airway localization, TMPRSS2 and HAT, into mammalian expression vector. Cotransfection of mammalian cells with plasmids encoding HA and either protease resulted in HA cleavage in situ. Transient expression of either protease in MDCK cells enabled multicycle replication of influenza viruses in these cells in the absence of exogenous trypsin. These data suggest that TMPRSS2 and HAT are candidates for proteolytic activation of influenza viruses in vivo.

scholarly journals A single amino acid change in the cytoplasmic domain alters the polarized delivery of influenza virus hemagglutinin.

1991 ◽  
Vol 114 (3) ◽  
pp. 413-421 ◽  
Author(s):  
C B Brewer ◽  
M G Roth
Keyword(s):  
Influenza Virus ◽  
Amino Acid ◽  
Amino Acid Change ◽  
Mdck Cells ◽  
Single Amino Acid ◽  
Apical Surface ◽  
Membrane Glycoproteins ◽  
Coated Pits ◽  
Influenza Virus Hemagglutinin ◽  
Single Amino Acid Change

In the polarized kidney cell line MDCK, the influenza virus hemagglutinin (HA) has been well characterized as a model for apically sorted membrane glycoproteins. Previous work from our laboratory has shown that a single amino acid change in the cytoplasmic sequence of HA converts it from a protein that is excluded from coated pits to one that is efficiently internalized. Using trypsin or antibodies to mark protein on the surface, we have shown in MDCK cells that HA containing this mutation is no longer transported to the apical surface but instead is delivered directly to the basolateral plasma membrane. We propose that a cytoplasmic feature similar to an endocytosis signal can cause exclusive basolateral delivery.

scholarly journals Characterization of a Porcine Lung Epithelial Cell Line Suitable for Influenza Virus Studies

2001 ◽  
Vol 75 (19) ◽  
pp. 9517-9525 ◽  
Author(s):  
Sang Heui Seo ◽  
Olga Goloubeva ◽  
Richard Webby ◽  
Robert G. Webster
Keyword(s):  
Influenza Virus ◽  
Epithelial Cell ◽  
Cell Line ◽  
Mdck Cells ◽  
Influenza Viruses ◽  
Lung Epithelial Cell ◽  
Clinical Samples ◽  
Epithelial Cell Line ◽  
Porcine Endogenous Retrovirus ◽  
Lung Epithelial

ABSTRACT We established a porcine lung epithelial cell line designated St. Jude porcine lung cells (SJPL) and demonstrated that all tested influenza A and B viruses replicated in this cell line. The infectivity titers of most viruses in SJPL cells were comparable to or better than those in MDCK cells. The propagation of influenza viruses from clinical samples in SJPL cells did not lead to antigenic changes in the hemagglutinin molecule. The numbers of both Sia2-3Gal and Sia2-6Gal receptors on SJPL cells were greater than those on MDCK cells. Influenza virus infection of SJPL cells did not lead to apoptosis, as did infection of MDCK cells. No porcine endogenous retrovirus was detected in SJPL cells, and in contrast to MDCK cells, SJPL cells did not cause tumors in nude mice.

scholarly journals A Prevalent Focused Human Antibody Response to the Influenza Virus Hemagglutinin Head Interface

mBio ◽  
2021 ◽  
Author(s):  
Kevin R. McCarthy ◽  
Jiwon Lee ◽  
Akiko Watanabe ◽  
Masayuki Kuraoka ◽  
Lindsey R. Robinson-McCarthy ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Herd Immunity ◽  
Influenza Viruses ◽  
Influenza Vaccines ◽  
Human Antibody ◽  
Human Influenza ◽  
Human Antibodies ◽  
Influenza Virus Hemagglutinin ◽  
Selection For ◽  
Rapid Appearance

The rapid appearance of mutations in circulating human influenza viruses and selection for escape from herd immunity require prediction of likely variants for an annual updating of influenza vaccines. The identification of human antibodies that recognize conserved surfaces on the influenza virus hemagglutinin (HA) has prompted efforts to design immunogens that might selectively elicit such antibodies.

scholarly journals APOPTOSIS STUDY OF INDONESIAN AVIAN INFLUENZA VIRUS SUBTYPE H5N1 IN MADIN-DARBY CANINE KIDNEY CELLS

2017 ◽  
Vol 11 (1) ◽  
pp. 39-44
Author(s):  
NLP Indi Dharmayanti ◽  
Dwi Rillah Ukhti ◽  
Farida Syamsiah ◽  
Risza Hartawan
Keyword(s):  
Influenza Virus ◽  
Avian Influenza ◽  
Avian Influenza Virus ◽  
Mdck Cells ◽  
Madin Darby Canine Kidney ◽  
Dna Ladder ◽  
Pathogenic Avian Influenza Virus ◽  
Virus Subtype ◽  
Darby Canine Kidney ◽  
Canine Kidney

This study aimed to determine the ability of highly pathogenic avian influenza virus (HPAI) virus subtype H5N1 originated from Indonesia to induce apoptosis in Madin-Darby Canine Kidney (MDCK) cells. Three HPAI virus subtype H5N1 isolates with different genetic characteristic namely A/Bird/Bali1/2011, A/Chicken/East Java/BwiI2/2010 and A/Chicken/West Java/1074/2003, were cultured in MDCK cells. Apoptosis was identified by deoxyribonucleic acid (DNA) fragmentation of infected MDCK cells using Apoptotic DNA Ladder Kit. The results showed that all three HPAI virus isolates used in this study did not able to induce apoptosis in the MDCK cells within 5 to 72 hours post infection.

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