scholarly journals Transcriptomic Characterization Reveals Attributes of High Influenza Virus Productivity in MDCK Cells

Viruses ◽  
2021 ◽  
Vol 13 (11) ◽  
pp. 2200
Author(s):  
Qian Ye ◽  
Thu Phan ◽  
Wei-Shou Hu ◽  
Xuping Liu ◽  
Li Fan ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Cell Line ◽  
Inflammatory Responses ◽  
Mdck Cells ◽  
Virus Production ◽  
Viral Mrnas ◽  
Factors Affecting ◽  
High Productivity ◽  
Antiviral Responses ◽  
Darby Canine Kidney

The Madin–Darby Canine Kidney (MDCK) cell line is among the most commonly used cell lines for the production of influenza virus vaccines. As cell culture-based manufacturing is poised to replace egg-based processes, increasing virus production is of paramount importance. To shed light on factors affecting virus productivity, we isolated a subline, H1, which had twice the influenza virus A (IAV) productivity of the parent (P) through cell cloning, and characterized H1 and P in detail on both physical and molecular levels. Transcriptome analysis revealed that within a few hours after IAV infection, viral mRNAs constituted over one fifth of total mRNA, with several viral genes more highly expressed in H1 than P. Functional analysis of the transcriptome dynamics showed that H1 and P responded similarly to IAV infection, and were both subjected to host shutoff and inflammatory responses. Importantly, H1 was more active in translation and RNA processing intrinsically and after infection. Furthermore, H1 had more subdued inflammatory and antiviral responses. Taken together, we postulate that the high productivity of IAV hinges on the balance between suppression of host functions to divert cellular resources and the sustaining of sufficient activities for virus replication. Mechanistic insights into virus productivity can facilitate the process optimization and cell line engineering for advancing influenza vaccine manufacturing.

Electrophysiological Measurements of a Toad Renal Epithelial Cell Line (A6) as an Assay for Predicting Ocular Eye Irritancy

1992 ◽  
Vol 20 (2) ◽  
pp. 218-221
Author(s):  
Henning F. Bjerregaard
Keyword(s):  
Epithelial Cell ◽  
Cell Line ◽  
South African ◽  
Mdck Cells ◽  
Epithelial Cell Line ◽  
Renal Epithelial Cell ◽  
Electrophysiological Measurements ◽  
Renal Epithelial Cell Line ◽  
Darby Canine Kidney

An established epithelial cell line (A6) from a South African clawed toad (Xenopus laevis) kidney was used as a model for the corneal epithelium of the eye in order to determine ocular irritancy. When grown on Millipore filter inserts, A6 cells form a monolayer epithelium of high electrical resistance and generate a trans-epithelial potential difference. These two easily-measured electrophysiological endpoints showed a dose-related decrease after exposure for 24 hours to seven selected chemicals of different ocular irritancy potential. It was demonstrated that both trans-epithelial resistance and potential ranked closely with in vivo eye irritancy data and correlated well (r = 0.96) with loss of trans-epithelial impermeability of Madin-Darby canine kidney (MDCK) cells, detected by use of a fluorescein leakage assay.

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.

CAP, a new human suspension cell line for influenza virus production

2012 ◽  
Vol 97 (1) ◽  
pp. 111-122 ◽  
Author(s):  
Yvonne Genzel ◽  
Ilona Behrendt ◽  
Jana Rödig ◽  
Erdmann Rapp ◽  
Claudia Kueppers ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Cell Line ◽  
Virus Production ◽  
Suspension Cell ◽  
Suspension Cell Line

Wave microcarrier cultivation of MDCK cells for influenza virus production in serum containing and serum-free media

Vaccine ◽  
2006 ◽  
Vol 24 (35-36) ◽  
pp. 6074-6087 ◽  
Author(s):  
Y. Genzel ◽  
R.M. Olmer ◽  
B. Schäfer ◽  
U. Reichl
Keyword(s):  
Influenza Virus ◽  
Mdck Cells ◽  
Virus Production ◽  
Serum Free ◽  
Free Media

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 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.

scholarly journals An Intact Dilysine-like Motif in the Carboxyl Terminus of MAL Is Required for Normal Apical Transport of the Influenza Virus Hemagglutinin Cargo Protein in Epithelial Madin-Darby Canine Kidney Cells

2001 ◽  
Vol 12 (6) ◽  
pp. 1869-1883 ◽  
Author(s):  
Rosa Puertollano ◽  
José Angel Martı́nez-Menárguez ◽  
Alicia Batista ◽  
José Ballesta ◽  
Miguel Angel Alonso
Keyword(s):  
Influenza Virus ◽  
Mdck Cells ◽  
Protein Sorting ◽  
Apical Surface ◽  
Carboxy Terminus ◽  
Madin Darby Canine Kidney ◽  
Influenza Virus Hemagglutinin ◽  
Carboxyl Terminal ◽  
Darby Canine Kidney ◽  
Canine Kidney

The MAL proteolipid, a component of the integral protein sorting machinery, has been demonstrated as being necessary for normal apical transport of the influenza virus hemagglutinin (HA) and the overall apical membrane proteins in Madin-Darby canine kidney (MDCK) cells. The MAL carboxy terminus ends with the sequence Arg-Trp-Lys-Ser-Ser (RWKSS), which resembles dilysine-based motifs involved in protein sorting. To investigate whether the RWKSS pentapeptide plays a role in modulating the distribution of MAL and/or its function in apical transport, we have expressed MAL proteins with distinct carboxy terminus in MDCK cells whose apical transport was impaired by depletion of endogenous MAL. Apical transport of HA was restored to normal levels by expression of MAL with an intact but not with modified carboxyl terminal sequences bearing mutations that impair the functioning of dilysine-based sorting signals, although all the MAL proteins analyzed incorporated efficiently into lipid rafts. Ultrastructural analysis indicated that compared with MAL bearing an intact RWKSS sequence, a mutant with lysine −3 substituted by serine showed a twofold increased presence in clathrin-coated cytoplasmic structures and a reduced expression on the plasma membrane. These results indicate that the carboxyl-terminal RWKSS sequence modulates the distribution of MAL in clathrin-coated elements and is necessary for HA transport to the apical surface.

An In Vitro Model of the Blood-Brain Barrier: The Response of Madin-Darby Canine Kidney Cells to Triethyl Tin

1996 ◽  
Vol 24 (3) ◽  
pp. 349-357
Author(s):  
Bellina Veronesi ◽  
Kent Carlsón ◽  
Marion Ehrich
Keyword(s):  
Cell Line ◽  
Blood Brain Barrier ◽  
Electrical Resistance ◽  
Mdck Cells ◽  
Brain Barrier ◽  
Madin Darby Canine Kidney ◽  
Blood Brain ◽  
Darby Canine Kidney ◽  
Canine Kidney

The development of a cell culture model which simulates the properties of the blood–brain barrier (BBB) is necessary for the detection of neurotoxic chemicals that can disrupt the barrier, and to provide a more “risk relevant” in vitro screening battery. The present study evaluates the Madin-Darby canine kidney (MDCK) epithelial cell line for this purpose. Changes in electrical resistance and enzyme activities were correlated in confluent MDCK cells exposed to the neurotoxic metal, triethyl tin (TET). Concentrations of TET (0.001–10μM) were established that produced depression in electrical resistance of the MDCK cells after exposure for 8 hours or caused fluorescein leakage after exposure for 72 hours. Confluent cultures of MDCK cells were then exposed to these concentrations of TET and assayed after exposure for 24 hours and 72 hours for changes in those enzymes common to both epithelial and cerebral endothelial cells. The results indicated that increased alkaline phosphatase (APP), γ-glutamyl transpeptidase (GGTP) and superoxide dismutase (SOD) characterised the loss of electrical resistance and permeability disruption in TET-exposed MDCK confluent cultures. Relative increases in APP and decreases in GGTP activities preceded cytotoxicity, which was associated with a high SOD activity. Such enzyme changes may be predictive endpoints of barrier cell disruption by neurotoxic metals in this cell line and support the additional evaluation of the MDCK cell line as an in vitro “screen” for chemicals that disrupt the BBB.

Madin Darby canine kidney cell-lines used in influenza virus production has resulted in canine 28s being integrated in the influenza virus genome - evidence from Influenza A patients in Hong Kong

2020 ◽  
Author(s):  
Sandeep Chakraborty
Keyword(s):  
Influenza Virus ◽  
Hong Kong ◽  
Influenza Vaccine ◽  
Mdck Cell ◽  
Influenza A ◽  
Mdck Cells ◽  
Virus Genome ◽  
Kidney Cells ◽  
Darby Canine Kidney ◽  
Canine Kidney

In 1989, 54 nucleotides from chicken 18s were seen to be inserted into the haemagglutinin gene of an influenza virus increasing viral pathogenicity [1]. Previously, I have reported human 18s sequences (from sequence vectors) appended to the influenza virus genome in Covid19 patients from Wuhan and Hong Kong [2]. These human ribosomal sequences are supposed to increase the transcription of the virus in the human cell, and thus will be more pathogenic. Here, I report the circulation of Influenza A genomes with 28s from canine integrated, showing that the escape of lab-made viruses is quite prevalent.Canine 28s sequence appended to flu genomesA recent (2020,Accid:PRJNA605947) study from the University of Hong Kong that did Nanopore sequencing to find novel targets for detection and surveillance of Influenza A viruses [3] shows the integration of canine 28s (Fig 1) sequence to the flu genome. The full read (SRR11067307.3179,SI:fullread.fa) splits into the flu virus (1-1605, SI:flu.baltimore.fa,(Baltimore/R0197/2017(H1N1)) nucleocapsid protein (NP) gene) and canine 28 (1606-1973, SI:canine.28s.fa Accid:XR 004817748.1, Canis lupus dingo 28S)). There is no reason to find canine 28s in clinical samples, barring the fact that canine kidney cells are used to manufacture the virus for several applications, including vaccines.Madin Darby canine kidney cells (MDCK) - why MDCK?The advantages of using MDCK influenza production is well known [4]. MDCK is better in the replication of live attenuated influenza viruses than most other cell lines (like Vero), thus yielding more virus in large-scale production of influenza virus [5]. The virus replicates rapidly in MDCK to ‘produce high titers in MDCK cells in as few as 3 to 10 passages, i.e., in 10–30 days’ [4]. Also, MDCK cells are also good for the production of certain influenza B virus vaccines, and MDCK cell-derived components are not allergenic [6]. Vaccines made using MDCK cellsInfluvac, a split virus vaccine produced in adherent MDCK cells, in the Netherlands in 1999 [7]. The use of MDCK cells (MedImmune) for production of live attenuated influenza vaccine in both serum containing and serum-free media was found to be more efficient [8]. Another trivalent MDCK cell culture-derived influenza vaccine is Optaflu [9]. ”Flucelvax Quadrivalent is the only cell-based inactivated flu vaccine that has been licensed by the FDA for use in the United States.” (https://www.cdc.gov/flu/prevent/cell-based.htm)

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