Abstract
The search for new antiviral strategies to treat influenza A virus (IAV) infections is one major international health care activity. Hereby, the IAV-caused misuse of cellular nuclear factor kappa B (NF-κB) signaling pathways in infected cells represents one target for antiviral therapy. In the present study, pyrrolidine dithiocarbamate (PDTC), which is known as an antioxidant and as an inhibitor of IAV-induced NF-κB activation, was studied in vivo. After the antiviral activity of PDTC was confirmed in MDCK cells, mice—infected with the mouse-adapted strain of IAV A/PR/8/34 (H1N1)—were treated intraperitoneally simultaneously with PDTC (75, 150, 200 mg/kg body weight). The influence of PDTC administrations was evaluated on viral replication and inflammatory reactions in lung tissue up to 14 days postinfection (p. i.). This therapy increased survival up to 80% and reduced IAV-caused weight loss and viral replication in lung tissue in a dose-dependent manner. Protective effects were less pronounced, if the therapy started later on during an ongoing IAV infection. In addition, simultaneous PDTC treatment also limited IAV-caused infiltration of immune cells as well as local interferon-γ expression in lung tissue. These results imply that PDTC decreases IAV-caused disease in mice significantly. Therefore, the development of drugs like PDTC that interfere with NF-κB signaling may represent a modern focus of anti-IAV therapy.
Similar content being viewed by others
References
Taubenberger JK, Kash JC (2010) Influenza virus evolution, host adaptation, and pandemic formation. Cell Host Microbe 7(6):440–451
Taubenberger JK, Morens DM (2010) Influenza: the once and future pandemic. Public Health Rep 125(Suppl 3):16–26
Krug RM, Aramini JM (2009) Emerging antiviral targets for influenza A virus. Trends Pharmacol Sci 30(6):269–277
Lin C, Zimmer SG, Lu Z, Holland RE Jr, Dong Q, Chambers TM (2001) The involvement of a stress-activated pathway in equine influenza virus-mediated apoptosis. Virology 287(1):202–213
Morris SJ, Price GE, Barnett JM, Hiscox SA, Smith H, Sweet C (1999) Role of neuraminidase in influenza virus-induced apoptosis. J Gen Virol 80:137–146
Takizawa T, Matsukawa S, Higuchi Y, Nakamura S, Nakanishi Y, Fukuda R (1993) Induction of programmed cell death (apoptosis) by influenza virus infection in tissue culture cells. J Gen Virol 74:2347–2355
Uchide N, Ohyama K, Bessho T, Yuan B, Yamakawa T (2002) Effect of antioxidants on apoptosis induced by influenza virus infection: inhibition of viral gene replication and transcription with pyrrolidine dithiocarbamate. Antiviral Res 56(3):207–217
Miyamoto D, Kusagaya Y, Endo N, Sometani A, Takeo S, Suzuki T, Arima Y, Nakajima K, Suzuki Y (1998) Thujaplicin-copper chelates inhibit replication of human influenza viruses. Antiviral Res 39(2):89–100
Uchide N, Ohyama K, Bessho T, Yuan B, Yamakawa T (2002) Apoptosis in cultured human fetal membrane cells infected with influenza virus. Biol Pharm Bull 25(1):109–114
Shi XL, Shi ZH, Huang H, Zhu HG, Zhou P, Ju D (2010) Therapeutic effect of recombinant human catalase on H1N1 influenza-induced pneumonia in mice. Inflammation 33(3):166–172
Knobil K, Choi AM, Weigand GW, Jacoby DB (1998) Role of oxidants in influenza virus-induced gene expression. Am J Physiol 274:134–142
Lowy RJ, Dimitrov DS (1997) Characterization of influenza virus-induced death of J774.1 macrophages. Exp Cell Res 234(2):249–258
Olsen CW, Dybdahl-Sissoko N, Hinshaw VS (1996) The influence of calcium and reactive oxygen species on influenza virus-induced apoptosis. Cell Death Differ 3(2):191–197
Meyer M, Schreck R, Baeuerle PA (1993) H2O2 and antioxidants have opposite effects on activation of NF-kappa B and AP-1 in intact cells: AP-1 as secondary antioxidant-responsive factor. EMBO J 12(5):2005–2015
Schreck R, Meier B, Mannel DN, Droge W, Baeuerle PA (1992) Dithiocarbamates as potent inhibitors of nuclear factor kappa B activation in intact cells. J Exp Med 175(5):1181–1194
Hayakawa M, Miyashita H, Sakamoto I, Kitagawa M, Tanaka H, Yasuda H, Karin M, Kikugawa K (2003) Evidence that reactive oxygen species do not mediate NF-kappa B activation. EMBO J 22(13):3356–3366
Kim I, Kim CH, Kim JH, Lee J, Choi JJ, Chen ZA, Lee MG, Chung KC, Hsu CY, Ahn YS (2004) Pyrrolidine dithiocarbamate and zinc inhibit proteasome-dependent proteolysis. Exp Cell Res 298(1):229–238
Borrello S, Demple B (1997) NF kappa B-independent transcriptional induction of the human manganous superoxide dismutase gene. Arch Biochem Biophys 348(2):289–294
Hartsfield CL, Alam J, Choi AM (1998) Transcriptional regulation of the heme oxygenase 1 gene by pyrrolidine dithiocarbamate. FASEB J 12(15):1675–1682
Wild AC, Mulcahy RT (1999) Pyrrolidine dithiocarbamate up-regulates the expression of the genes encoding the catalytic and regulatory subunits of gamma-glutamylcysteine synthetase and increases intracellular glutathione levels. Biochem J 338:659–665
Uchide N, Ohyama K, Bessho T, Toyoda H (2005) Inhibition of influenza-virus-induced apoptosis in chorion cells of human fetal membranes by nordihydroguaiaretic Acid. Intervirology 48(5):336–340
Uchide N, Ohyama K (2003) Antiviral function of pyrrolidine dithiocarbamate against influenza virus: the inhibition of viral gene replication and transcription. J Antimicrob Chemother 52(1):8–10
Flory E, Kunz M, Scheller C, Jassoy C, Stauber R, Rapp UR, Ludwig S (2000) Influenza virus-induced NF-kappa B-dependent gene expression is mediated by overexpression of viral proteins and involves oxidative radicals and activation of Ikappa B kinase. J Biol Chem 275(12):8307–8314
Nimmerjahn F, Dudziak D, Dirmeier U, Hobom G, Riedel A, Schlee M, Staudt LM, Rosenwald A, Behrends U, Bornkamm GW, Mautner J (2004) Active NF-kappa B signalling is a prerequisite for influenza virus infection. J Gen Virol 85:2347–2356
Wurzer WJ, Ehrhardt C, Pleschka S, Berberich-Siebelt F, Wolff T, Walczak H, Planz O, Ludwig S (2004) NF-kappa B-dependent induction of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) and Fas/FasL is crucial for efficient influenza virus propagation. J Biol Chem 279(30):30931–30937
Schmolke M, Viemann D, Roth J, Ludwig S (2009) Essential impact of NF-kappaB signaling on the H5N1 influenza A virus-induced transcriptome. J Immunol 183(8):5180–5189
Wei L, Sandbulte MR, Thomas PG, Webby RJ, Homayouni R, Pfeffer LM (2006) NFkappa B negatively regulates interferon-induced gene expression and anti-influenza activity. J Biol Chem 281(17):11678–11684
Uchide N, Toyoda H (2008) Future target molecules for influenza treatment. Mini Rev Med Chem 8(5):491–495
Chabicovsky M, Prieschl-Grassauer E, Seipelt J, Muster T, Szolar OH, Hebar A, Doblhoff-Dier O (2010) Pre-clinical safety evaluation of pyrrolidine dithiocarbamate. Basic Clin Pharmacol Toxicol 107(3):758–767
Reed LJ, Muench H (1938) A simple method of estimating fifty percent endpoints. Am J Hyg 27:493–497
Chieux V, Chehadeh W, Harvey J, Haller O, Wattre P, Hober D (2001) Inhibition of coxsackievirus B4 replication in stably transfected cells expressing human MxA protein. Virology 283(1):84–92
Chomczynski P, Sacchi N (1987) Single-step method of RNA isolation by acid guanidinium thiocyanate- phenol-chloroform extraction. Anal Biochem 162(1):156–159
Pfaffl MW (2001) A new mathematical model for relative quantification in real-time RT-PCR. Nucleic Acids Res 29(9):e45
Henke A, Launhardt H, Klement K, Stelzner A, Zell R, Munder T (2000) Apoptosis in coxsackievirus B3-caused diseases: interaction between the capsid protein VP2 and the proapoptotic protein siva. J Virol 74(9):4284–4290
Kim CH, Kim JH, Hsu CY, Ahn YS (1999) Zinc is required in pyrrolidine dithiocarbamate inhibition of NF-kappa B activation. FEBS Lett 449(1):28–32
Ulmer JB, Deck RR, DeWitt CM, Friedman A, Donnelly JJ, Liu MA (1994) Protective immunity by intramuscular injection of low doses of influenza virus DNA vaccines. Vaccine 12(16):1541–1544
Henke A (2002) DNA immunization—a new chance in vaccine research? Med Microbiol Immunol 191(3–4):187–190
Schmidt RE (2002) Drug under test: influenza—Relenza in daily practice. Experience during the influenza season 1999/2000. Med Microbiol Immunol 191(3–4):175–179
Harrod ME, Emery S, Dwyer DE (2006) Antivirals in the management of an influenza pandemic. Med J Aust 185(Suppl 10):S58–S61
Monto AS, McKimm-Breschkin JL, Macken C, Hampson AW, Hay A, Klimov A, Tashiro M, Webster RG, Aymard M, Hayden FG, Zambon M (2006) Detection of influenza viruses resistant to neuraminidase inhibitors in global surveillance during the first 3 years of their use. Antimicrob Agents Chemother 50(7):2395–2402
Sheu TG, Deyde VM, Okomo-Adhiambo M, Garten RJ, Xu X, Bright RA, Butler EN, Wallis TR, Klimov AI, Gubareva LV (2008) Surveillance for neuraminidase inhibitor resistance among human influenza A, B viruses circulating worldwide from 2004 to 2008. Antimicrob Agents Chemother 52(9):3284–3292
Mazur I, Wurzer WJ, Ehrhardt C, Pleschka S, Puthavathana P, Silberzahn T, Wolff T, Planz O, Ludwig S (2007) Acetylsalicylic acid (ASA) blocks influenza virus propagation via its NF-kappa B-inhibiting activity. Cell Microbiol 9(7):1683–1694
Gaudernak E, Seipelt J, Triendl A, Grassauer A, Kuechler E (2002) Antiviral effects of pyrrolidine dithiocarbamate on human rhinoviruses. J Virol 76(12):6004–6015
Kang MS, Choi EK, Choi DH, Ryu SY, Lee HH, Kang HC, Koh JT, Kim OS, Hwang YC, Yoon SJ, Kim SM, Yang KH, Kang IC (2008) Antibacterial activity of pyrrolidine dithiocarbamate. FEMS Microbiol Lett 280(2):250–254
Krenn BM, Holzer B, Gaudernak E, Triendl A, van Kuppeveld FJ, Seipelt J (2005) Inhibition of polyprotein processing and RNA replication of human rhinovirus by pyrrolidine dithiocarbamate involves metal ions. J Virol 79(22):13892–13899
Lanke K, Krenn BM, Melchers WJ, Seipelt J, van Kuppeveld FJ (2007) PDTC inhibits picornavirus polyprotein processing and RNA replication by transporting zinc ions into cells. J Gen Virol 88(Pt 4):1206–1217
Wurzer WJ, Planz O, Ehrhardt C, Giner M, Silberzahn T, Pleschka S, Ludwig S (2003) Caspase 3 activation is essential for efficient influenza virus propagation. EMBO J 22(11):2717–2728
Erl W, Weber C, Hansson GK (2000) Pyrrolidine dithiocarbamate-induced apoptosis depends on cell type, density, and the presence of Cu2+ and Zn2+. Am J Physiol Cell Physiol 278(6):C1116–C1125
Zabel U, Schreck R, Baeuerle PA (1991) DNA binding of purified transcription factor NF-kappa B. Affinity, specificity, Zn2+ dependence, and differential half-site recognition. J Biol Chem 266(1):252–260
Shumilla JA, Wetterhahn KE, Barchowsky A (1998) Inhibition of NF-kappa B binding to DNA by chromium, cadmium, mercury, zinc, and arsenite in vitro: evidence of a thiol mechanism. Arch Biochem Biophys 349(2):356–362
Oxford JS, Perrin DD (1974) Inhibition of the particle-associated RNA-dependent RNA polymerase activity of influenza viruses by chelating agents. J Gen Virol 23(1):59–71
Michaelis M, Geiler J, Naczk P, Sithisarn P, Ogbomo H, Altenbrandt B, Leutz A, Doerr HW, Cinatl J Jr (2010) Glycyrrhizin inhibits highly pathogenic H5N1 influenza A virus-induced pro-inflammatory cytokine and chemokine expression in human macrophages. Med Microbiol Immunol 199:291–297
Geiler J, Michaelis M, Sithisarn P, Cinatl J Jr (2010) Comparison of pro-inflammatory cytokine expression and cellular signal transduction in human macrophages infected with different influenza A viruses. Med Microbiol Immunol. doi:10.1007/s00430-010-0173-y
Julkunen I, Sareneva T, Pirhonen J, Ronni T, Melen K, Matikainen S (2001) Molecular pathogenesis of influenza A virus infection and virus-induced regulation of cytokine gene expression. Cytokine Growth Factor Rev 12(2–3):171–180
Geiler J, Michaelis M, Naczk P, Leutz A, Langer K, Doerr HW, Cinatl J Jr (2009) N-acetyl-L-cysteine (NAC) inhibits virus replication and expression of pro-inflammatory molecules in A549 cells infected with highly pathogenic H5N1 influenza A virus. Biochem Pharmacol 79(3):413–420
Cheung CY, Poon LL, Lau AS, Luk W, Lau YL, Shortridge KF, Gordon S, Guan Y, Peiris JS (2002) Induction of proinflammatory cytokines in human macrophages by influenza A (H5N1) viruses: a mechanism for the unusual severity of human disease? Lancet 360(9348):1831–1837
Starko KM (2009) Salicylates and pandemic influenza mortality, 1918–1919 pharmacology, pathology, and historic evidence. Clin Infect Dis 49(9):1405–1410
Eyers S, Weatherall M, Shirtcliffe P, Perrin K, Beasley R (2010) The effect on mortality of antipyretics in the treatment of influenza infection: systematic review and meta-analysis. J R Soc Med 103(10):403–411
Ethical standards
All experiments comply with the current laws of Germany.
Conflict of interest
The authors declare that they have no conflict of interest.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Wiesener, N., Zimmer, C., Jarasch-Althof, N. et al. Therapy of experimental influenza virus infection with pyrrolidine dithiocarbamate. Med Microbiol Immunol 200, 115–126 (2011). https://doi.org/10.1007/s00430-010-0182-x
Received:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s00430-010-0182-x