scholarly journals Mechanism of Antiviral Activity of Nitazoxanide against the Influenza Virus: Effect of Tizoxanide on AdenosineTriphosphate in Influenza-virus Infected Madin Darby Canine Kidney Cells

2021 ◽  
Author(s):  
Jean-Francois Rossignol ◽  
Carl van Baalen ◽  
Aloys Tijsma
Keyword(s):  
Influenza Virus ◽  
Influenza A ◽  
Respiratory Infections ◽  
Mdck Cells ◽  
Dependent Manner ◽  
Madin Darby Canine Kidney ◽  
Atp Production ◽  
Viral Respiratory Infections ◽  
Darby Canine Kidney ◽  
Canine Kidney

Background: Nitazoxanide (NTZ) is a broad-spectrum antiviral undergoing clinical development for treating influenza and other viral respiratory infections such as those caused by rhinovirus/enterovirus and coronavirus including the emerging SARS-CoV-2. Methods: Nitazoxanide is a mild uncoupler of oxidative phosphorylation, which is modulating the ATP production in cells. ATP is an essential component of viral replication, and we have evaluated the effect of tizoxanide (TIZ), the active circulating metabolite of NTZ, on ATP in Madin-Darby canine kidney (MDCK) cells and in MDCK cells infected with influenza A and B viruses. Results: TIZ decreased cellular ATP in a dose-dependent manner in MDCK cells and in MDCK cells infected with influenza A and B viruses. Maximum inhibition of ATP in influenza infected or uninfected MDCK cells reached up to 45% after 6 and 24 hours of exposure to 100 micrometer TIZ. The decrease in cellular ATP did not affect cell viability and was reversible after eliminating TIZ from the culture. Conclusion: The concentrations of TIZ required to decrease cellular ATP levels were similar to those reported to inhibit replication of influenza A and B viruses in our laboratory. A decrease in ATP triggers activation of AMP-activated protein kinase, which is known to suppress the secretion of pro-inflammatory cytokines. Additional studies are warranted to evaluate the effect of TIZ on mitochondrial function.

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 Microtubule perturbation inhibits intracellular transport of an apical membrane glycoprotein in a substrate-dependent manner in polarized Madin-Darby canine kidney epithelial cells.

1990 ◽  
Vol 1 (12) ◽  
pp. 921-936 ◽  
Author(s):  
M J van Zeijl ◽  
K S Matlin
Keyword(s):  
Plasma Membrane ◽  
Epithelial Cells ◽  
Intracellular Transport ◽  
Mdck Cells ◽  
Sodium Dodecyl ◽  
Apical Plasma Membrane ◽  
Dependent Manner ◽  
Madin Darby Canine Kidney ◽  
Darby Canine Kidney ◽  
Canine Kidney

The effects of microtubule perturbation on the transport of two different viral glycoproteins were examined in infected Madin-Darby canine kidney (MDCK) cells grown on both permeable and solid substrata. Quantitative biochemical analysis showed that the microtubule-depolymerizing drug nocodazole inhibited arrival of influenza hemagglutinin on the apical plasma membrane in MDCK cells grown on both substrata. In contrast, the microtubule-stabilizing drug taxol inhibited apical appearance of hemagglutinin only when MDCK cells were grown on permeable substrata. On the basis of hemagglutinin mobility on sodium dodecyl sulfate gels and its sensitivity to endo H, it was evident that nocodazole and taxol arrested hemagglutinin at different intracellular sites. Neither drug caused a significant increase in the amount of hemagglutinin detected on the basolateral plasma membrane domain. In addition, neither drug had any noticeable effect on the transport of the vesicular stomatitis virus (VSV)-G protein to the basolateral surface. These results shed light on previous conflicting reports using this model system and support the hypothesis that microtubules play a role in the delivery of membrane glycoproteins to the apical, but not the basolateral, domain of epithelial cells.

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.

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)

H(+)-peptide cotransport in Madin-Darby canine kidney cells: expression and calmodulin-dependent regulation

1995 ◽  
Vol 268 (3) ◽  
pp. F391-F397 ◽  
Author(s):  
M. Brandsch ◽  
V. Ganapathy ◽  
F. H. Leibach
Keyword(s):  
Mdck Cells ◽  
Peptide Transport ◽  
Maximal Velocity ◽  
Dependent Manner ◽  
Beta Lactam ◽  
Madin Darby Canine Kidney ◽  
Carbonyl Cyanide ◽  
Lactam Antibiotic ◽  
Darby Canine Kidney ◽  
Canine Kidney

The transport of the dipeptide glycylsarcosine was studied in the kidney cell lines OK, LLC-PK1, and Madin-Darby canine kidney (MDCK), grown as confluent monolayers on impermeable plastic supports. Uptake of the dipeptide in OK and LLC-PK1 cells was slow, was not inhibited by other peptides, and was not influenced by an inwardly directed H+ gradient, indicating lack of expression of the H(+)-peptide cotransport system in these cells under our conditions. In contrast, uptake of the dipeptide in MDCK cells was rapid and was found to be stimulated by an inwardly directed H+ gradient. This stimulation was markedly reduced by the protonophore carbonyl cyanide p-trifluoromethoxy-phenylhydrazone. The H+ gradient-dependent uptake of glycylsarcosine was inhibited by dipeptides and tripeptides and by the beta-lactam antibiotic cephalexin but not by the amino acids glycine and leucine. The uptake was saturable and apparently occurred via a single transport system. The Michaelis-Menten constant for the system was 1.3 +/- 0.1 mM, and the maximal velocity was 13.3 +/- 0.7 nmol.30.min-1.mg protein-1. Treatment of MDCK cells with the calmodulin antagonists N-(6-aminohexyl)-5-chloro-1-napthalenesulfonamide (W-7), CGS-9343B, or calmidazolium inhibited the glycylsarcosine uptake by 40-50% in a time- and dose-dependent manner. In contrast, the uptake of alanine, leucine, glucose, and taurine was found to be stimulated by treatment with W-7. Kinetic analysis revealed that the inhibition of the peptide transport activity was mainly associated with a decrease of the maximal velocity of the system.(ABSTRACT TRUNCATED AT 250 WORDS)

Arginine vasopressin stimulates H+-ATPase in MDCK cells via V1 (cell Ca2+) and V2 (cAMP) receptors

2004 ◽  
Vol 286 (2) ◽  
pp. F402-F408 ◽  
Author(s):  
Maria Oliveira-Souza ◽  
Raif Musa-Aziz ◽  
Gerhard Malnic ◽  
Margarida de Mello Aires
Keyword(s):  
Arginine Vasopressin ◽  
Mdck Cells ◽  
Specific Inhibitor ◽  
Cytosolic Calcium ◽  
Dependent Manner ◽  
Madin Darby Canine Kidney ◽  
Dose Dependent ◽  
Darby Canine Kidney ◽  
Canine Kidney ◽  
Stimulation Of

The effect of arginine vasopressin (AVP) and/or atrial natriuretic peptide (ANP) on the regulation of intracellular pH (pHi) via H+-ATPase and of cytosolic calcium ([Ca2+]i) was investigated in Madin-Darby canine kidney (MDCK) cells by the fluorescent probes BCECF-AM and fluo-4-AM, respectively. The pHi recovery rate was examined after intracellular acidification following an NH4Cl pulse, in the presence of zero Na+ plus Schering 28080 (a specific inhibitor of H+-K+-ATPase). AVP (10-12-10-6 M) increased the rate of pHi recovery and [Ca2+]i in a dose-dependent manner. V1- or V2-receptor antagonists impaired the effect of AVP on both processes, and DDAVP (10-12-10-6 M; a V2-selective agonist) caused a dose-dependent stimulation of them. [Ca2+]i or cAMP (as increased by 10-5 M thapsigargin or 8-BrcAMP, respectively) alone had no effect on H+-ATPase, but their synergic action was necessary to stimulate H+-ATPase. In agreement with these findings, ANP (10-6 M) or dimethyl-BAPTA-AM (5 × 10-5 M), impairing the increase of [Ca2+]i in response to AVP, blocks the stimulatory effect of AVP on H+-ATPase.

Effect of Riluzole on Ca2+ Movement and Cytotoxicity in Madin-Darby Canine Kidney Cells

2006 ◽  
Vol 25 (8) ◽  
pp. 461-469 ◽  
Author(s):  
W-C Chen ◽  
H-H Cheng ◽  
C-J Huang ◽  
C-T Chou ◽  
S-I Liu ◽  
...  
Keyword(s):  
Mdck Cells ◽  
Dependent Manner ◽  
Madin Darby Canine Kidney ◽  
Concentration Dependent Manner ◽  
Intracellular Ca ◽  
Pre Treatment ◽  
Lateral Sclerosis ◽  
Darby Canine Kidney ◽  
Canine Kidney

Riluzole is a drug used in the treatment of amyotrophic lateral sclerosis; however, its in vitro action is unclear. In this study, the effect of riluzole on intracellular Ca2- concentration ([Ca2-]i) in Madin-Darby canine kidney (MDCK) cells was investigated using the Ca2--sensitive fluorescent dye, fura-2. Riluzole (100 -500 mM) caused a rapid and sustained increase of [Ca2-]i in a concentration-dependent manner (EC50 = 150 mM). Some 40 and 50% of this [Ca2-]i increase was prevented by the removal of extracellular Ca2- and the addition of La3-, respectively, but was unchanged by dihydropyridines, verapamil and diltiazem. In Ca2--free medium, thapsigargin -an inhibitor of the endoplasmic reticulum (ER) Ca2--ATPase -caused a monophasic [Ca2-]i increase, after which the increasing effect of riluzole on [Ca2-]iwas attenuated by 70%; in addition, pre-treatment with riluzole abolished thapsigargin-induced [Ca2-]i increases. U73122, an inhibitor of phospholipase C (PLC), abolished ATP (but not riluzole)-induced [Ca2-]i increases. At concentrations of 250 and 500 mM, riluzole killed 40 and 95% cells, respectively. The cytotoxic effect of riluzole (250 mM) was unaltered by pre-chelating cytosolic Ca2- with BAPTA. Collectively, in MDCK cells, riluzole rapidly increased [Ca2-]i by stimulating extra-cellular Ca2- influx via an La3--sensitive pathway and intracellular Ca2- release from the ER via, as yet, unidentified mechanisms. Furthermore, riluzole caused Ca2--unrelated cytotoxicity in a concentration-depen-dent manner.

scholarly journals Comparative Susceptibility of Madin–Darby Canine Kidney (MDCK) Derived Cell Lines for Isolation of Swine Origin Influenza A Viruses from Different Clinical Specimens

Viruses ◽  
2021 ◽  
Vol 13 (12) ◽  
pp. 2346
Author(s):  
Matthew Suderman ◽  
Mariko Moniwa ◽  
Tamiru N. Alkie ◽  
Davor Ojkic ◽  
Andre Broes ◽  
...  
Keyword(s):  
Cell Lines ◽  
Influenza A ◽  
Mdck Cells ◽  
Influenza A Viruses ◽  
Madin Darby Canine Kidney ◽  
Clinical Specimens ◽  
Oral Fluids ◽  
Nasal Swabs ◽  
Darby Canine Kidney ◽  
Canine Kidney

Madin–Darby canine kidney (MDCK) cells are commonly used for the isolation of mammalian influenza A viruses. The goal of this study was to compare the sensitivity and suitability of the original MDCK cell line in comparison with MDCK-derived cell lines, MDCK.2, MDCK SIAT-1 and MDCK-London for isolation of swine-origin influenza A viruses (IAV-S) from clinical specimens. One-hundred thirty clinical specimens collected from pigs in the form of nasal swabs, lung tissue and oral fluids that were positive by PCR for the presence of IAV-S RNA were inoculated in the cell cultures listed above. MDCK-SIAT1 cells yielded the highest proportion of positive IAV-S isolations from all specimen types. For nasal swabs, 58.62% of the specimens were IAV-S positive in MDCK-SIAT1 cells, followed by MDCK-London (36.21%), and conventional MDCK and MDCK.2 cells (27.5%). For lung specimens, 59.38% were IAV-S positive in MDCK-SIAT1 cells, followed by MDCK-London (40.63%), and conventional MDCK and MDCK.2 cells (18.75–31.25%). Oral fluids yielded the lowest number of positive virus isolation results, but MDCK-SIAT1 cells were still had the highest rate (35%) of IAV-S isolation, whereas the isolation rate in other cells ranged from 5–7.5%. Samples with lower IAV-S PCR cycle threshold (Ct) values were more suitable for culturing and isolation. The isolated IAV-S represented H1N1-β, H1N2-α, H1N1pdm and H3N2 cluster IV and cluster IVB viruses. The result of the current study demonstrated the importance of using the most appropriate MDCK cells when isolating IAV-S from clinical samples.

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