N-acetylcysteine lacks universal inhibitory activity against influenza A viruses
© Garigliany and Desmecht; licensee BioMed Central Ltd. 2011
Received: 29 November 2010
Accepted: 9 May 2011
Published: 9 May 2011
N-acetylcysteine (NAC) has been recently proposed as an adjuvant therapeutic drug for influenza pneumonia in humans. This proposal is based on its ability to restrict influenza virus replication in vitro and to attenuate the severity of the disease in mouse models. Although available studies were made with different viruses (human and avian), published information related to the anti-influenza spectrum of NAC is scarce. In this study, we show that NAC is unable to alter the course of a fatal influenza pneumonia caused by inoculation of a murinized swine H1N1 influenza virus. NAC was indeed able to inhibit the swine virus in vitro but far less than reported for other strains. Therefore, susceptibility of influenza viruses to NAC appears to be strain-dependent, suggesting that it cannot be considered as a universal treatment for influenza pneumonia.
About 10 percent of the human population is affected by influenza annually and several pandemic episodes have occurred throughout recorded history . This context explains why continued efforts are made to identify new therapeutic molecules. Among these, N-acetylcysteine (NAC), which is commonly used for its mucolytic activity in humans, was shown to inhibit influenza virus both in mouse models, alone or in combination, with the A/PR/8 strain [2, 3], and in vitro, with H5N1 strains . Recently, NAC treatment was reported to reduce symptoms of influenza-like illness in humans  and administration of the dose of 100 mg/kg supposedly contributed to the success of the treatment of a patient infected with the 2009 pandemic H1N1 virus .
In order to determine whether these successful results can be extrapolated to other viral strains than A/PR/8 and H5N1 strains, the effect of NAC on the clinical course and outcome of experimental influenza was assessed in mice inoculated with a lethal dose of our murinized swine H1N1 influenza strain . In spite of a significant but very partial anti-influenza effect in vitro, neither percent survival nor body weight loss were altered by NAC treatment in vivo, suggesting that NAC-susceptibility of influenza A viruses is strain-dependent.
In vivo study
Two groups of ten 8-wk old female CD-1 mice were intranasally inoculated with 10 MLD50 of murinized A/swine/Iowa/4/1976 (H1N1) virus . The first group received 100 mg/kg NAC (Sigma) daily by gavage, from day 1 to day 7 post-infection (pi), while the second received the vehicle only. Clinical status, body weight (BW) and mortality were recorded daily up to day 14 pi. Challenge studies were approved by the Belgian Council for Laboratory Animal Science, under the guidance of the Institutional Animal Care and Use Committees of the University of Liège.
In vitro study
Near confluent Vero cells (ATCC CCL-81) were infected at a multiplicity of infection of 0.01 with a Vero cell-adapted variant of the A/swine/Iowa/4/1976 (H1N1) virus in 6-well plates. One hour after infection, fresh DMEM supplemented with 0.2% bovine serum albumin (Invitrogen), 2 μg/ml TPCK-treated trypsin (Sigma), and either 0, 0.5, 1.5 or 2.5 mg/ml of NAC (Sigma) was added onto the cells. Culture supernatants were harvested 48 h after infection and titrated by standard plaque assays.
Results and discussion
Overall, the in vitro and in vivo results gathered here show that susceptibility of influenza viruses to NAC is clearly strain-dependent, which suggests that NAC cannot be considered to be a universal treatment for influenza pneumonia. A systematic testing of anti-influenza activity of NAC should be implemented whenever a new strain emerges.
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- Simonsen L, Fukuda K, Schonberger LB, Cox NJ: The impact of influenza epidemics on hospitalizations. J Infect Dis. 2000, 181: 831-837. 10.1086/315320.View ArticlePubMedGoogle Scholar
- Ungheri D, Pisani C, Sanson G, Bertani A, Schioppacassi G, Delgado R, Sironi M, Ghezzi P: Protective effect of n-acetylcysteine in a model of influenza infection in mice. Int J Immunopathol Pharmacol. 2000, 13: 123-128.PubMedGoogle Scholar
- Ghezzi P, Ungheri D: Synergistic combination of N-acetylcysteine and ribavirin to protect from lethal influenza viral infection in a mouse model. Int J Immunopathol Pharmacol. 2004, 17: 99-102.PubMedGoogle Scholar
- Geiler J, Michaelis M, Naczk P, Leutz A, Langer K, Doerr HW, Cinatl J: 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. 2010, 79: 413-420. 10.1016/j.bcp.2009.08.025.View ArticlePubMedGoogle Scholar
- De Flora S, Grassi C, Carati L: Attenuation of influenza-like symptomatology and improvement of cell-mediated immunity with long-term N-acetylcysteine treatment. Eur Respir J. 1997, 10: 1535-1541. 10.1183/09031936.97.10071535.View ArticlePubMedGoogle Scholar
- Lai KY, Ng WY, Osburga Chan PK, Wong KF, Cheng F: High-dose N-acetylcysteine therapy for novel H1N1 influenza pneumonia. Ann Intern Med. 2010, 152: 687-688.View ArticlePubMedGoogle Scholar
- Garigliany MM, Habyarimana A, Lambrecht B, Van de Paar E, Cornet A, van den Berg T, Desmecht D: Influenza A strain-dependent pathogenesis in fatal H1N1 and H5N1 subtype infections of mice. Emerg Infect Dis. 2010, 16: 595-603.View ArticlePubMedPubMed CentralGoogle Scholar
- Garozzo A, Tempera G, Ungheri D, Timpanaro R, Castro A: N-acetylcysteine synergizes with oseltamivir in protecting mice from lethal influenza infection. Int J Immunopathol Pharmacol. 2007, 20: 349-354.PubMedGoogle Scholar
- American International Chemical, Acetylcysteine Material Safety Data Sheet. http://www.aicma.com/msds/Acetylcysteine%20USP.pdf
- Gowdy KM, Krantz QT, King C, Boykin E, Jaspers I, Linak WP, Gilmour MI: Role of oxidative stress on diesel-enhanced influenza infection in mice. Part Fibre Toxicol. 2010, 7: 34-10.1186/1743-8977-7-34.View ArticlePubMedPubMed CentralGoogle Scholar
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