- Review article
- Open Access
Respiratory Syncytial Virus Infection: From Biology to Therapy A Perspective
© World Allergy Organization; licensee BioMed Central Ltd. 2008
Received: 3 October 2007
Accepted: 13 December 2007
Published: 15 February 2008
Respiratory syncytial virus (RSV) is responsible for significant morbidity and mortality, particularly in infants younger than 18 months and in the elderly. To date, there are few effective treatment options available to prevent or treat RSV infections. Attractive therapeutic strategies include targeting host epithelial adhesion molecules required for RSV infection, enhancing localized cell-mediated immunity, interfering with RSV viral gene expression and developing a multigene DNA vaccine. The most recent data supporting the advantages and limitations of each of these approaches are discussed in detail. Several promising strategies offer hope for safe and effective prophylaxis and treatment of RSV infection.
Respiratory syncytial virus (RSV) is one of the most important respiratory pathogens targeting all age groups; however, infants (younger than 18 months) and the elderly experience the most severe aspects of the disease, which results in lower respiratory tract illnesses (ie, bronchiolitis and pneumonia) . Around 90% of infants are infected for the first time by the age of 2 years [1, 2]. Worldwide, about 5 million infants are hospitalized because of severe RSV infection. The first is usually the most severe, and previous findings indicate that infants with a history of premature birth, bronchopulmonary dysplasia, congenital heart disease, cystic fibrosis, or immunosuppression are more likely to develop the most severe clinical courses of bronchiolitis and pneumonia, which have the highest risk of death[1, 2].
However, an analysis of a comprehensive study done between 1979 and 1997 about RSV-associated deaths in US children suggests that most RSV-related deaths do not occur among children who are presumed to be at high risk for severe RSV lower respiratory tract illnesses . The leading cause in infant hospitalization is RSV bronchiolitis,  which imposes a severe burden upon health services. Costs related to emergency department visits between 1997 and 2000 amount to approximately 202 million US dollars . Complete immunity to RSV never develops, and reinfection throughout life is common. Although the major clinical manifestation of RSV in older children and adults is upper respiratory tract illness (rhinitis and acute otitis media), it may also cause up to 2.4% of community-acquired pneumonia in these population groups . In older adults, RSV was identified as responsible for 10% of winter hospital admissions and has a case-fatality rate that approaches 10%. In addition, 78% of RSV-associated deaths occur in individuals aged 65 years or older who have underlying cardiac and pulmonary pathology . In particular, RSV infection in adults with strong immunosuppression, for example, patients undergoing bone marrow transplantation is of great medical importance .
In the past 8 years, our research has identified both cellular and viral targets that may be useful for the prevention of RSV infection and its accompanying pathology. Differential microarray analysis was used to pinpoint gene expression changes in RSV-infected cells, and expression of candidate therapeutic genes was tested both in cultured lung epithelial cells in vitro and in animal models in vivo. Characterization of these gene expression changes includes immune modulation, signal transduction, and apoptosis. In this report, the biology of RSV and how these studies contribute to the basic mechanistic studies of RSV infection and have led to new targets to manage RSV infection will be discussed.
State of the Art in Treatment and Prophylaxis of RSV Infection
There is no treatment to protect against RSV infection, and the current treatment, Ribavirin, only produces modest short-term improvement in respiratory tract infection . Moreover, it is now restricted to a highly selected group of patients with T-cell immunodeficiency . Passive immunoprophylaxis, involving the administration of either a polyclonal antibody (Synagis) preparation or a humanized version of a monoclonal anti-RSV-F antibody (Palivizumab), is successful for protection of high-risk individuals against RSV infection. However, these approaches are only partially effective, expensive, and could generate resistant mutant RSV strains. Development of new and highly effective antibodies to modulate RSV infection remains a major medical and pharmaceutical goal.
To date, there is no licensed vaccine for the prevention of human RSV disease. Efforts have been made to develop active prophylaxis measures (vaccines), and both subunit and attenuated live vaccines are being pursed in clinical studies. Vaccine development has been limited after the testing of initial vaccines in the 1960s, which exacerbated the RSV disease [10, 11]. Some of the reasons for the lack of success in developing previous vaccines include the inadequate response to vaccination, the existence of 2 antigenically distinct RSV groups, and the history of disease enhancement after administration of a formalin-inactivated vaccine [12, 13].
Developing active or passive prophylaxis is important as they are expected to decrease the incidence of severe infections and thus may reduce or attenuate asthma pathogenesis. Recent advances in the vaccine area include research with plasmid-based DNA vaccines and small-interfering RNA (siRNA)-based approaches. To deliver these antiviral plasmids in the most effective way to target cells, a novel carrier system has been produced based on modified polysaccharide nanoparticles that protect the DNA and facilitate its introduction into the lungs. The advances in this field are reviewed in the following sections.
RSV Genome and Structure
Human RSV is in the genus Pneumovirus, subfamily Pneumovirinae, family Paramyxoviridae, order Mononegavirales, whose members consist of nonsegmented, negative-sense, single-stranded RNA viruses. In addition to human RSV, the genus Pneumovirus includes bovine RSV, ovine RSV, and pneumonia virus of mice. The RSV virions consist of a nucleocapsid contained within a lipid envelope of irregular spherical shape with sizes of 150 to 300 nm. Both infected cultures and viral preparations can also include filamentous forms of the virions that are 60 to 100 nm in diameter and up to 10 μm in length . The viral envelope is a lipid bilayer acquired from the host plasmatic membrane. The viral transmembrane glycoproteins--the fusion protein F, the attachment protein G, and the small hydrophobic protein SH--organize themselves to form spikes, which are visible under electron microscopy. Host lipid raft-derived proteins are also incorporated into the envelope of mature viral particles [15–17]. The envelope connects to the nucleocapsid through the viral matrix M protein. Using electron microscopy, the nucleocapsid is seen as an internal electrodense material with a diameter of 15 nm inside the round and filamentous forms of the virions . The nucleocapsids consist of the RNA genome and the associated nucleocapsid protein N, the phosphoprotein P, the large polymerase subunit L, and the antitermination factor M2-1. The viral RNA genome and the associated proteins in the nucleocapsid together form a very tight ribonucleoprotein complex, which is resistant to RNAse activity.
The genome for most of the virions is a negative-sense strand of RNA of 15,222 nucleotides in length. However, some virions are also found to have incorporated the positive-sense replicative intermediate (antigenomic RNA), which is synthesized during viral replication. Thus, this implies that during the viral assembly, there is no mechanism that allows discrimination in packaging. The viral genes are ordered from 3' to 5' in the following way: NS1-NS2-N-P-M-SH-G-F-M2-L. Glycoprotein G and F (and SH), respectively, mediate virus attachment and fusion to the host cell . In addition to fusion, protein F has also been postulated to participate in the attachment of the virus to the host cell membrane. Intercellular adhesion molecule 1 (ICAM-1), annexin-II, and Toll-like receptor 4 are receptors for protein F [19–21]. The matrix (M) protein forms a layer on the inner face of the viral envelope, and it plays an essential role in viral assembly through its interactions with the cell membrane, virus envelope, and virus nucleocapsid [22, 23]. The nucleocapsid-associated proteins N, P, M2-1, and L play essential roles at different stages for efficient viral transcription and replication. The nonstructural proteins NS1 and NS2 are thought to be antagonists of the interferon (IFN)-type I system. They seem to target the transcription factor IRF-3. Thus, the expression of these proteins helps the virus to reduce IFN-γ expression by infected cells [24, 25].
Prophylaxis and Treatment of RSV Infection
Summary of Studies Relating to RSV Infection
Ab, antisense RNA
F, G, SH, NS1, NS2, P, N
Development of Chitosan-Based Nanoparticles as a Platform for Gene and Drug Delivery
Numerous investigators, including those in our laboratory, have extensively studied chitosan, which we believe has the potential to be useful for the delivery of genes and drugs, as it has very low immunogenicity while having strong immunostimulatory properties . Moreover, as a carrier, it can most adequately provide heat stability to encapsulated or adsorbed vaccines. Chitosan, a natural biocompatible cationic polysaccharide extracted from crustacean shells, is capable of efficient drug and gene delivery [37–41]. Chitosan has many beneficial effects, including anticoagulant activity,  wound-healing properties,  and antimicrobial properties . In addition, chitosan is nontoxic, nonhemolytic, slowly biodegradable, and nuclease resistant, and it has been widely used in controlled drug delivery [37, 43–47]. Chitosan also increases transcellular and paracellular transport across the mucosal epithelium and, thus, may facilitate mucosal drug delivery and modulate immunity of the mucosal and bronchus-associated lymphoid tissues. Chitosan apparently binds to macrophages and myeloid cells via CD14.[49, 50]
The toxicity of mucosally administered chitosan has been studied in rodents. N-trimethyl chitosan and chitosan hydrochloride given intranasally do not alter the ciliary beat frequency of the rat nasal epithelium, and hence, both are considered to be nontoxic . In addition, the subacute oral toxicity of chitosan oligosaccharides was investigated in Sprague-Dawley rats of both sexes . The chitosan is metabolized and secreted through the viliary system. Thirty-six male and female rats were administered by gavage 500, 1000, and 2000 mg/kg per day of chitosan for 4 weeks (7 days per week), and their clinical signs, body weights, hematologic and biochemical parameters, and histopathology were examined. There were no significant differences in behavior, external appearance, body weight or food consumption between control and treated rats. In addition, no significant differences in urinalysis, hematology, blood biochemistry, relative organ weights, and histopathological findings were found in either control or treated rats. These results suggest that the acute toxicity of chitosan oligosaccharides is low and that the detection limit of toxicity is greater than 2000 mg/kg in rats. Furthermore, chlorophyllin-chitosan, an insoluble form of chlorophyllin, inhibits DNA adduct formation and mutagenesis by a heterocyclic food mutagen-carcinogen, 3-amino-1-methyl-5H-pyridoindole (Trp-P-2), in mice carrying the Escherichia coli rpsL gene as a mutagenesis reporter, this suggests that chlorophyllin-chitosan may be a candidate chemopreventive agent against the genotoxic action of Trp-P-2 and possibly other aromatic carcinogens in the diet.
The Environmental Protection Agency has ruled chitosan exempt from its tolerance guidelines because of its nontoxicity as evidenced by the: (1) literature search done for chitin, chitosan, N-acetyl-D-glucosamine, and D-glucosamine toxicity in humans using the databases PubMed, Hazardous Substances Data Bank, Integrated Risk Information System, Gene-Tox, Environmental Mutagen Information Center, Toxic Release Inventory, the Food and Drug Administration, the United States Department of Agriculture and ChemIDplus; (2) animal feeding studies, in which up to 5% of the diet is chitosan, that failed to show any adverse effects; and (3) the lack of reported complaints of toxicity against the database of 2700 complaints despite years of chitosan use in food and nutritional supplements.
A Nanoparticle Gene Expression Vaccine for RSV
The potential of vaccines has been intensely investigated since the discovery of the virus. All RSV proteins, except L, have been tested for immunogenicity and protective efficacy in rodents using recombinant vaccinia viruses [59–61]. A number of approaches, including recombinant live, attenuated, subunit vaccines, and DNA vaccines, are under intense investigation, [62–64] but none have crossed the clinical-phase hurdles and been licensed thus far. The development of RSV vaccines is complicated by the need to administer the vaccine at a very young age, between 6 weeks and 6 months, in the face of a premature immune system. In addition, because RSV is a mucosal pathogen, an effective vaccine must generate secreted mucosal antibodies, such as immunoglobulin A (IgA) and mucosal cytotoxic lymphocytes (CTLs) [65, 66]. The RSV-induced CTL response at mucosal sites is inadequate. Although evidence suggests the potential of a gene expression vaccine for RSV infection, the number of studies is limited. Previous reports using systemic injections of pDNA show variable results. The quantity of DNA used per unit body mass, as much as 10 mg/kg, and the route of administration chosen are inconvenient for infants and are suboptimal for inducing mucosal immunity against a pulmonary infection.
Our laboratory developed a nanoparticle multigene vaccination strategy against RSV infection using a complementary DNA cocktail produced by cloning 9 RSV antigens (NS1, NS2, M, SH, F, M2, N, G, or P in a pVAX plasmid) complexed with chitosan nanoparticles, referred to as nanoparticle gene expression vaccine (NGXV). The NGXV was administered to mice by the intranasal route. The rationale for developing this vaccine is based on the following reports. All of the RSV proteins, except L, have been tested individually (and in some cases, in combination) for immunogenicity and protective efficacy in rodents using recombinant vaccinia viruses [59–63]. The F and G proteins are the antigens that induce most of the the neutralizing antibodies against RSV [68–70]. The CTL repertoire in humans revealed that the N, SH, F, M, M2, and NS2 proteins were strong target antigens. In BALB/c mice, the F, N, and M2 proteins are major target antigens [61, 71–73]. Protection against and recovery from RSV infection are mediated largely by the immune system, with the specific direct effectors being secretory antibodies, serum antibodies, and major histocompatibility complex class I-restricted CTLs.
The results demonstrate that a single vaccination of about 1 mg/kg body weight of NGXV decreases viral titers by 2 orders of magnitude (100-fold) upon primary infection. In addition, NGXV significantly decreases pulmonary inflammation and does not alter airway hyperresponsiveness, thus making it a potentially safe vaccine. This may represent a major breakthrough in RSV vaccine development.
Host Gene Expression
Prophylactic IFN-γ gene transfer in BALB/c mice decreases viral replication and induces a Th1-like (increased production of IFN-γ and interleukin-12), instead of a Th2-like (decreased interleukin-5) immune response against RSV infection [74–76]. Viral infections induce IFN-γ, which in turn facilitates the resolution of viral infection . Levels of IFN-γ have been compared in bronchoalveolar lavage fluids after infection with RSV in control and pIFN-γ-treated mice. A 3-to 6-fold increase in IFN-γ production was found in RSV-infected mice compared with uninfected mice. Such increases are considered to be relatively low compared with other viral infections [74–76]. The finding that a natural live virus infection is cleared by elevated IFN-γ production, a response similar to that seen after live viral infection in mice, suggests that the results from this animal model may be applicable to human RSV disease.
A new prophylactic approach consists of taking advantage of the RNA interference mechanism initially discovered in plant cells and that is present in all species including mammals. RNA interference is triggered by double-stranded RNA that is cleaved by an RNAse III-like enzyme, Dicer, into 21-25-nucleotide fragments (siRNAs) with characteristic 5' and 3' termini [77, 78]. These siRNAs act as guides for a multiprotein complex, including a PAZ/PIWI domain, containing the protein Argonaute2, which cleaves the target messenger RNA (mRNA) . These gene-silencing mechanisms are highly specific and induce inhibition of gene expression throughout an organism. RNA interference is a known phenomenon that has been proven effective in silencing a number of genes of different viruses [80–82]. The siRNA to viral P or NS-1 mRNAs prevents RSV infection in cellular and animal model studies [83, 84]. Prophylactic intranasal administration of an siRNA formulation specific for RSV-P mRNA is able to significantly reduce the viral load and the disease parameters in RSV-infected mice . A carrier in the formulation is not required. In addition, a very low dose is effective in showing a protective effect. Moreover, siRNA-resistant virus did not appear after using this formulation . Although intranasal administration of naked siRNA to humans was found to be safe in a phase I study, other studies have shown toxicity.
Because the synthesis of RNA oligonucleotide-based siRNA is expensive, our laboratory engineered DNAvector-based approaches to introduce siNS1 into RSV-infected human cells and animal models. This is based on the principle of the intracellular transcription of small RNA molecules that are synthesized from a DNA template under the control of RNA polymerase III promoters, such as U6 . NS-1 was selected as the target because the NS1 protein interferes with type-1 IFN-mediated host antiviral responses [24, 86]. Silencing of the NS-1 gene attenuated RSV replication and boosted the immune response through an increase in IFN-γ production . The prophylactic intranasal administration of this formulation, combined with chitosan, significantly reduced the viral load and ameliorated the pulmonary pathology in RSV-infected mice . In addition, mice treated with this formulation develop protection against reinfection.
Moreover, this formulation also drives human dendritic cells to promote a Th1-like profile . Overall, siRNA-mediated silencing of the NS1 gene up-regulates host-antiviral genes and suppresses RSV replication compared with control groups. Studies confirm the role of siNS1 in a rat model of RSV infection. A phase I study is currently under development using the nanoparticle-incorporated siNS1, and it may represent a novel prophylaxis/therapy that can be used in a global population.
Summary and Conclusion
The RSV is the major pathogen responsible for serious upper and lower respiratory tract infections, primarily in infants, but also in the elderly worldwide. The precise molecular and cellular mechanisms are unclear, and satisfactory prophylaxis or treatment strategies are yet to emerge. This research has resulted in the understanding of the pathology and complexity of signaling pathways involved in successful infection; the role of host defense molecules such as ICAM-1, IFN-γ, and related pathways; and how they can be exploited to develop less costly prophylaxis and treatments for RSV infection. Finally, the potential to develop safe and effective prophylaxis and/or treatment by targeting important RSV genes is under investigation.
S. S. Mohapatra is supported by grants from the Veterans' Affairs Merit Review and VA Career Scientist Awards, and by US National Institutes of Health grant no. 5RO1HL71101-01A2. The support from the Joy McCann Culverhouse and Mabel and Ellsworth Simmons Endowments to the Division of Allergy and Immunology, Department of Internal Medicine, College of Medicine, and the Veterans' Affairs Hospital is also gratefully acknowledged.
- Simoes EA: Respiratory syncytial virus infection. Lancet. 1999, 354: 847-852.View ArticlePubMedGoogle Scholar
- Respiratory syncytial virus activity-United States, 1999-2000 season. MMWR Morb Mortal Wkly Rep. 2000, 49: 1091-1093.Google Scholar
- Shay DK, Holman RC, Roosevelt GE, Clarke MJ, Anderson LJ: Bronchiolitis-associated mortality and estimates of respiratory syncytial virus-associated deaths among US children, 1979-1997. J Infect Dis. 2001, 183: 16-22. 10.1086/317655.View ArticlePubMedGoogle Scholar
- Leader S, Kohlhase K: Recent trends in severe respiratory syncytial virus (RSV) among US infants, 1997 to 2000. J Pediatr. 2003, 143 (suppl): S127-S132.View ArticlePubMedGoogle Scholar
- Dowell SF, Anderson LJ, Gary HE, Erdman DD, Plouffe JF, et al: Respiratory syncytial virus is an important cause of community-acquired lower respiratory infection among hospitalized adults. J Infect Dis. 1996, 174: 456-462. 10.1093/infdis/174.3.456.View ArticlePubMedGoogle Scholar
- Thompson WW, Shay DK, Weintraub E, Brammer L, Cox N, Anderson LJ, Fukuda K: Mortality associated with influenza and respiratory syncytial virus in the United States. JAMA. 2003, 289: 179-186. 10.1001/jama.289.2.179.View ArticlePubMedGoogle Scholar
- Thomas NJ, Hollenbeak CS, Ceneviva GD, Geskey JM, Young MJ: Palivizumab prophylaxis to prevent respiratory syncytial virus mortality after pediatric bone marrow transplantation: a decision analysis model. J Pediatr Hematol Oncol. 2007, 29: 227-232. 10.1097/MPH.0b013e3180437ded.View ArticlePubMedGoogle Scholar
- Falsey AR, Cunningham CK, Barker WH, Kouides RW, Yuen JB, et al: Respiratory syncytial virus and influenza A infections in the hospitalized elderly. J Infect Dis. 1995, 172: 389-394. 10.1093/infdis/172.2.389.View ArticlePubMedGoogle Scholar
- Kimpen JL: Prevention and treatment of respiratory syncytial virus bronchiolitis and postbronchiolitic wheezing. Respir Res. 2002, 3 (suppl): S40-S45. 10.1186/rr183.PubMedPubMed CentralGoogle Scholar
- Kalina WV, Woolums AR, Berghaus RD, Gershwin LJ: Formalin-inactivated bovine RSV vaccine enhances a Th2 mediated immune response in infected cattle. Vaccine. 2004, 22: 1465-1474. 10.1016/j.vaccine.2003.10.024.View ArticlePubMedGoogle Scholar
- Castilow EM, Olson MR, Varga SM: Understanding respiratory syncytial virus (RSV) vaccine-enhanced disease. Immunol Res. 2007, 39: 225-239. 10.1007/s12026-007-0071-6.View ArticlePubMedGoogle Scholar
- Piedra PA: Clinical experience with respiratory syncytial virus vaccines. Pediatr Infect Dis J. 2003, 22 (suppl): S94-S99. 10.1097/01.inf.0000053893.15894.ff.PubMedGoogle Scholar
- Dudas RA, Karron RA: Respiratory syncytial virus vaccines. Clin Microbiol Rev. 1998, 11: 430-439.PubMedPubMed CentralGoogle Scholar
- Bachi T, Howe C: Morphogenesis and ultrastructure of respiratory syncytial virus. J Virol. 1973, 12: 1173-1180.PubMedPubMed CentralGoogle Scholar
- Brown G, Jeffree CE, McDonald T, Rixon HW, Aitken JD, Sugrue RJ: Analysis of the interaction between respiratory syncytial virus and lipid-rafts in Hep2 cells during infection. Virology. 2004, 327: 175-185. 10.1016/j.virol.2004.06.038.View ArticlePubMedGoogle Scholar
- Brown G, Rixon HW, Sugrue RJ: Respiratory syncytial virus assembly occurs in GM1-rich regions of the host-cell membrane and alters the cellular distribution of tyrosine phosphorylated caveolin-1. J Gen Virol. 2002, 83: 1841-1850.View ArticlePubMedGoogle Scholar
- Brown G, Aitken J, Rixon HW, Sugrue RJ: Caveolin-1 is incorporated into mature respiratory syncytial virus particles during virus assembly on the surface of virus-infected cells. J Gen Virol. 2002, 83: 611-621.View ArticlePubMedGoogle Scholar
- Bourgeois C, Bour JB, Lidholt K, Gauthray C, Pothier P: Heparin-like structures on respiratory syncytial virus are involved in its infectivity in vitro. J Virol. 1998, 72: 7221-7227.PubMedPubMed CentralGoogle Scholar
- Malhotra R, Ward M, Bright H, et al: Isolation and characterisation of potential respiratory syncytial virus receptor(s) on epithelial cells. Microbes Infect. 2003, 5: 123-133. 10.1016/S1286-4579(02)00079-5.View ArticlePubMedGoogle Scholar
- Kurt-Jones EA, Popova L, Kwinn L, et al: Pattern recognition receptors TLR4 and CD14 mediate response to respiratory syncytial virus. Nat Immunol. 2000, 1: 398-401. 10.1038/80833.View ArticlePubMedGoogle Scholar
- Behera AK, Matsuse H, Kumar M, Kong X, Lockey RF, Mohapatra SS: Blocking intercellular adhesion molecule-1 on human epithelial cells decreases respiratory syncytial virus infection. Biochem Biophys Res Commun. 2001, 280: 188-195. 10.1006/bbrc.2000.4093.View ArticlePubMedGoogle Scholar
- Marty A, Meanger J, Mills J, Shields B, Ghildyal R: Association of matrix protein of respiratory syncytial virus with the host cell membrane of infected cells. Arch Virol. 2004, 149: 199-210.View ArticlePubMedGoogle Scholar
- Rodriguez L, Cuesta I, Asenjo A, Villanueva N: Human respiratory syncytial virus matrix protein is an RNA-binding protein: binding properties, location and identity of the RNA contact residues. J Gen Virol. 2004, 85: 709-719. 10.1099/vir.0.19707-0.View ArticlePubMedGoogle Scholar
- Spann KM, Tran KC, Chi B, Rabin RL, Collins PL: Suppression of the induction of alpha, beta, and lambda interferons by the NS1 and NS2 proteins of human respiratory syncytial virus in human epithelial cells and macrophages. J Virol. 2004, 78: 4363-4369. 10.1128/JVI.78.8.4363-4369.2004.View ArticlePubMedPubMed CentralGoogle Scholar
- Bossert B, Conzelmann KK: Respiratory syncytial virus (RSV) nonstructural (NS) proteins as host range determinants: a chimeric bovine RSV with NS genes from human RSV is attenuated in interferon-competent bovine cells. J Virol. 2002, 76: 4287-4293. 10.1128/JVI.76.9.4287-4293.2002.View ArticlePubMedPubMed CentralGoogle Scholar
- van Schaik SM, Enhorning G, Vargas I, Welliver RC: Respiratory syncytial virus affects pulmonary function in BALB/c mice. J Infect Dis. 1998, 177: 269-276. 10.1086/514208.View ArticlePubMedGoogle Scholar
- Graham BS, Perkins MD, Wright PF, Karzon DT: Primary Respiratory Syncytial Virus-Infection in Mice. J Med Virol. 1988, 26: 153-162. 10.1002/jmv.1890260207.View ArticlePubMedGoogle Scholar
- Graham BS, Henderson GS, Tang YW, Lu XT, Neuzil KM, Colley DG: Priming Immunization Determines T-Helper Cytokine Messenger-Rna Expression Patterns in Lungs of Mice Challenged with Respiratory Syncytial Virus. J Immunol. 1993, 151: 2032-2040.PubMedGoogle Scholar
- Hashimoto K, Peebles RS, Sheller JR, et al: Suppression of airway hyperresponsiveness induced by ovalbumin sensitisation and RSV infection with Y-27632, a Rho kinase inhibitor. Thorax. 2002, 57: 524-527. 10.1136/thorax.57.6.524.View ArticlePubMedPubMed CentralGoogle Scholar
- Peebles RS, Sheller JR, Johnson JE, Mitchell DB, Graham BS: Respiratory syncytial virus infection prolongs methacholine-induced airway hyperresponsiveness in ovalbumin-sensitized mice. J Med Virol. 1999, 57: 186-192. 10.1002/(SICI)1096-9071(199902)57:2<186::AID-JMV17>3.0.CO;2-Q.View ArticlePubMedGoogle Scholar
- Peebles RS, Sheller JR, Collins RD, Jarzecka K, Mitchell DB, Graham BS: Respiratory syncytial virus (RSV)-induced airway hyperresponsiveness in allergically sensitized mice is inhibited by live RSV and exacerbated by formalin-inactivated RSV. J Infect Dis. 2000, 3: 671-677.View ArticleGoogle Scholar
- Peebles RS, Hashimoto K, Graham BS: The complex relationship between respiratory syncytial virus and allergy in lung disease. Viral Immunol. 2003, 16: 25-34. 10.1089/088282403763635429.View ArticlePubMedGoogle Scholar
- Tang YW, Graham BS: T cell source of type 1 cytokines determines illness patterns in respiratory syncytial virus-infected mice. J Clin Invest. 1997, 99: 2183-2191. 10.1172/JCI119391.View ArticlePubMedPubMed CentralGoogle Scholar
- Hussell T, Khan U, Openshaw P: IL-12 treatment attenuates T helper cell type 2 and Bcell responses but does not improve vaccine-enhanced lung illness. J Immunol. 1997, 159: 328-334.PubMedGoogle Scholar
- Walzl G, Matthews S, Kendall S, Gutierrez-Ramos JC, Coyle AJ, Openshaw PJM, Hussell T: Inhibition of T1/ST2 during respiratory syncytial virus infection prevents T helper cell type 2 (Th2)- but not Th1-driven immunopathology. J Exp Med. 2001, 193: 785-792. 10.1084/jem.193.7.785.View ArticlePubMedPubMed CentralGoogle Scholar
- Kneuer C, Sameti M, Bakowsky U, Schiestel T, Schirra H, Schmidt H, Lehr CM: A nonviral DNA delivery system based on surface modified silica-nanoparticles can efficiently transfect cells in vitro. Bioconjug Chem. 2000, 11: 926-932. 10.1021/bc0000637.View ArticlePubMedGoogle Scholar
- Wrobel I, Collins D: Fusion of cationic liposomes with mammalian cells occurs after endocytosis. Biochim Biophys Acta. 1995, 1235: 296-304. 10.1016/0005-2736(95)80017-A.View ArticlePubMedGoogle Scholar
- Koltover I, Salditt T, Radler JO, Safinya CR: An inverted hexagonal phase of cationic liposome-DNA complexes related to DNA release and delivery. Science. 1998, 281: 78-81.View ArticlePubMedGoogle Scholar
- Hughes RC: The complex carbohydrates of mammalian cell surfaces and their biological roles. Essays Biochem. 1975, 11: 1-36.PubMedGoogle Scholar
- Kabanov AV: Taking polycation gene delivery systems from in vitro to in vivo. 1999, 2: 365-372.Google Scholar
- Nettelbeck DM, Jerome V, Muller R: Gene therapy: designer promoters for tumour targeting. Trends Genet. 2000, 16: 174-181. 10.1016/S0168-9525(99)01950-2.View ArticlePubMedGoogle Scholar
- Ezekowitz RA, Williams DJ, Koziel H, Armstrong MY, Warner A, Richards FF, Rose RM: Uptake of Pneumocystis carinii mediated by the macrophage mannose receptor. Nature. 1991, 351: 155-158. 10.1038/351155a0.View ArticlePubMedGoogle Scholar
- Wileman T, Boshans R, Stahl P: Uptake and transport of mannosylated ligands by alveolar macrophages. Studies on ATP-dependent receptor-ligand dissociation. J Biol Chem. 1985, 260: 7387-7393.PubMedGoogle Scholar
- Ferkol T, Perales JC, Mularo F, Hanson RW: Receptor-mediated gene transfer into macrophages. Proc Natl Acad Sci USA. 1996, 93: 101-105. 10.1073/pnas.93.1.101.View ArticlePubMedPubMed CentralGoogle Scholar
- Ferkol T, Mularo F, Hilliard J, Lodish S, Perales JC, Ziady A, Konstan M: Transfer of the human Alpha1-antitrypsin gene into pulmonary macrophages in vivo. Am J Respir Cell Mol Biol. 1998, 18: 591-601. 10.1165/ajrcmb.18.5.2874.View ArticlePubMedGoogle Scholar
- Rojanasakul Y, Wang LY, Malanga CJ, Ma JK, Liaw J: Targeted gene delivery to alveolar macrophages via Fc receptor-mediated endocytosis. Pharm Res. 1994, 11: 1731-1736. 10.1023/A:1018959231951.View ArticlePubMedGoogle Scholar
- Behera AK, Kumar M, Lockey RF, Mohapatra SS: Adenovirus-mediated interferon gamma gene therapy for allergic asthma: involvement of interleukin 12 and STAT4 signaling. Hum Gene Ther. 2002, 13: 1697-1709. 10.1089/104303402760293547.View ArticlePubMedGoogle Scholar
- Kumar M, Behera AK, Matsuse H, Lockey RF, Mohapatra SS: Intranasal IFN-gamma gene transfer protects BALB/c mice against respiratory syncytial virus infection. Vaccine. 1999, 18: 558-567. 10.1016/S0264-410X(99)00185-1.View ArticlePubMedGoogle Scholar
- Otterlei M, Varum KM, Ryan L, Espevik T: Characterization of binding and TNF-alpha-inducing ability of chitosans on monocytes: the involvement of CD14. Vaccine. 1994, 12: 825-832. 10.1016/0264-410X(94)90292-5.View ArticlePubMedGoogle Scholar
- Bianco ID, Balsinde J, Beltramo DM, Castagna LF, Landa CA, Dennis EA: Chitosan-induced phospholipase A2 activation and arachidonic acid mobilization in P388D1 macrophages. FEBS Lett. 2000, 466: 292-294. 10.1016/S0014-5793(00)01089-9.View ArticlePubMedGoogle Scholar
- Haffejee N, Du Plessis J, Muller DG, Schultz C, Kotze AF, Goosen C: Intranasal toxicityof selected absorption enhancers. Pharmazie. 2001, 56: 882-888.PubMedGoogle Scholar
- Kim SK, Park PJ, Yang HP, Han SS: Subacute toxicity of chitosan oligosaccharide in Sprague-Dawley rats. Arzneimittelforschung. 2001, 51: 769-774.PubMedGoogle Scholar
- Anzai N, Taniyama T, Nakandakari N, Sugiyama C, Negishi T, Hayatsu H, Negishi K: Inhibition of DNA adduct formation and mutagenic action of 3-amino-1-methyl-5h-pyrido[4,3-b]indole by chlorophyllin-chitosan in rpsL transgenic mice. Jpn J Cancer Res. 2001, 92: 848-853. 10.1111/j.1349-7006.2001.tb01171.x.View ArticlePubMedGoogle Scholar
- Bokura H, Kobayashi S: Chitosan decreases total cholesterol in women: a randomized, double-blind, placebo-controlled trial. Eur J Clin Nutr. 2003, 57: 721-725. 10.1038/sj.ejcn.1601603.View ArticlePubMedGoogle Scholar
- Metso S, Ylitalo R, Nikkila M, Wuolijoki E, Ylitalo P, Lehtimaki T: The effect of long-term microcrystalline chitosan therapy on plasma lipids and glucose concentrations in subjects with increased plasma total cholesterol: a randomised placebo-controlled double-blind crossover trial in healthy men and women. Eur J Clin Pharmacol. 2003, 59: 741-746. 10.1007/s00228-003-0691-2.View ArticlePubMedGoogle Scholar
- Wuolijoki E, Hirvela T, Ylitalo P: Decrease in serum LDL cholesterol with microcrystalline chitosan. Methods Find Exp Clin Pharmacol. 1999, 21: 357-361. 10.1358/mf.19220.127.116.113477.View ArticlePubMedGoogle Scholar
- Barroso Aranda J, Contreras F, Bagchi D, Preuss HG: Efficacy of a novel chitosan formulation on fecal fat excretion: a double-blind, crossover, placebo-controlled study. J Med. 2002, 33: 209-225.PubMedGoogle Scholar
- Pittler MH, Abbot NC, Harkness EF, Ernst E: Randomized, double-blind trial of chitosan for body weight reduction. Eur J Clin Nutr. 1999, 53: 379-381. 10.1038/sj.ejcn.1600733.View ArticlePubMedGoogle Scholar
- Brandenburg AH, Neijens HJ, Osterhaus A: Pathogenesis of RSV lower respiratory tract infection: implications for vaccine development. Vaccine. 2001, 19: 2769-2782. 10.1016/S0264-410X(00)00536-3.View ArticlePubMedGoogle Scholar
- Cherrie AH, Anderson K, Wertz GW, Openshaw PJM: Human Cytotoxic T-Cells Stimulated by Antigen on Dendritic Cells Recognize the N, Sh, F, M, 22k, and 1b Proteins of Respiratory Syncytial Virus. J Virol. 1992, 66: 2102-2110.PubMedPubMed CentralGoogle Scholar
- Connors M, Collins PL, Firestone CY, Murphy BR: Respiratory Syncytial Virus (Rsv) F-Protein, G-Protein, M2-Protein (22k), and N-Proteins Each Induce Resistance to Rsv Challenge, but Resistance Induced by M2-Proteins and N-Proteins Is Relatively Short-Lived. J Virol. 1991, 65: 1634-1637.PubMedPubMed CentralGoogle Scholar
- Schmidt AC, Wenzke DR, McAuliffe JM, St Claire M, Elkins WR, Murphy BR, Collins PL: Mucosal immunization of rhesus monkeys against respiratory syncytial virus subgroups A and B and human parainfluenza virus type 3 by using a live cDNA-derived vaccine based on a host range-attenuated bovine parainfluenza virus type 3 vector backbone. J Virol. 2002, 76: 1089-1099. 10.1128/JVI.76.3.1089-1099.2002.View ArticlePubMedPubMed CentralGoogle Scholar
- Teng MN, Collins PL: Altered growth characteristics of recombinant respiratory syncytial viruses which do not produce NS2 protein. J Virol. 1999, 73: 466-473.PubMedPubMed CentralGoogle Scholar
- Teng MN, Whitehead SS, Bermingham A, St Claire M, Elkins WR, Murphy BR, Collins PL: Recombinant respiratory syncytial virus that does not express the NS1 or M2-2 protein is highly attenuated and immunogenic in chimpanzees. Genetic vaccination strategies for enhanced cellular, humoral and mucosal immunity. J Virol. 2000, 74: 9317-9321. 10.1128/JVI.74.19.9317-9321.2000.View ArticlePubMedPubMed CentralGoogle Scholar
- Ramsay AJ, Kent SJ, Strugnell RA, Suhrbier A, Thomson SA, Ramshaw IA: Immunogenicity and safety of respiratory syncytial virus subunit vaccine in seropositive children 18-36 months old. Immunol Rev. 1999, 171: 27-44. 10.1111/j.1600-065X.1999.tb01341.x.View ArticlePubMedGoogle Scholar
- Tristram DA, Welliver RC, Mohar CK, Hogerman DA, Hildreth SW, Paradiso P: J Infect Dis. 1993, 167: 191-195. 10.1093/infdis/167.1.191.View ArticlePubMedGoogle Scholar
- Huang DB, Wu JJ, Tyring SK: A review of licensed viral vaccines, some of their safety concerns, and the advances in the development of investigational viral vaccines. J Infect. 2004, 49: 179-209. 10.1016/j.jinf.2004.05.018.View ArticlePubMedGoogle Scholar
- Walsh EE, Schlesinger JJ, Brandriss MW: Purification and characterization of GP90, one of the envelope glycoproteins of respiratory syncytial virus. J Gen Virol. 1984, 65: 761-767. 10.1099/0022-1317-65-4-761.View ArticlePubMedGoogle Scholar
- Walsh EE, Cote PJ, Fernie BF, Schlesinger JJ, Brandriss MW: Analysis of the respiratory syncytial virus fusion protein using monoclonal and polyclonal antibodies. J Gen Virol. 1986, 67: 505-513. 10.1099/0022-1317-67-3-505.View ArticlePubMedGoogle Scholar
- Walsh EE, Hall CB, Briselli M, Brandriss MW, Schlesinger JJ: Immunization with glycoprotein subunits of respiratory syncytial virus to protect cotton rats against viral infection. J Infect Dis. 1987, 155: 1198-1204. 10.1093/infdis/155.6.1198.View ArticlePubMedGoogle Scholar
- Wyatt LS, Whitehead SS, Venanzi KA, Murphy BR, Moss B: Priming and boosting immunity to respiratory syncytial virus by recombinant replication-defective vaccinia virus MVA. Vaccine. 1999, 18: 392-397. 10.1016/S0264-410X(99)00257-1.View ArticlePubMedGoogle Scholar
- Nicholas JA, Rubino KL, Levely ME, Adams EG, Collins PL: Cytolytic T-lymphocyte responses to respiratory syncytial virus: effector cell phenotype and target proteins. J Virol. 1990, 64: 4232-4241.PubMedPubMed CentralGoogle Scholar
- Domachowske JB, Rosenberg RF: Respiratory syncytial virus infection: immune response, immunopathogenesis, and treatment. Clin Microbiol Rev. 1999, 12: 298-309.PubMedPubMed CentralGoogle Scholar
- Durbin JE, Johnson TR, Durbin RK, Mertz SE, Morotti RA, Peebles RS, Graham BS: The role of IFN in respiratory syncytial virus pathogenesis. J Immunol. 2002, 168: 2944-2952.View ArticlePubMedGoogle Scholar
- Kumar M, Behera AK, Matsuse H, Lockey RF, Mohapatra SS: Intranasal IFN-gamma gene transfer protects BALB/c mice against respiratory syncytial virus infection. Vaccine. 1999, 18: 558-567. 10.1016/S0264-410X(99)00185-1.View ArticlePubMedGoogle Scholar
- Bukreyev A, Whitehead SS, Bukreyeva N, Murphy BR, Collins PL: Interferon gamma expressed by a recombinant respiratory syncytial virus attenuates virus replication in mice without compromising immunogenicity. Proc Natl Acad Sci USA. 1999, 96: 2367-2372. 10.1073/pnas.96.5.2367.View ArticlePubMedPubMed CentralGoogle Scholar
- Provost P, Silverstein RA, Dishart D, et al: Dicer is required for chromosome segregation and gene silencing in fission yeast cells. Proc Natl Acad Sci USA. 2002, 99: 16648-16653. 10.1073/pnas.212633199.View ArticlePubMedPubMed CentralGoogle Scholar
- Provost P, Dishart D, Doucet J, Frendewey D, Samuelsson B, Radmark O: Ribonuclease activity and RNA binding of recombinant human Dicer. EMBO J. 2002, 21: 5864-5874. 10.1093/emboj/cdf578.View ArticlePubMedPubMed CentralGoogle Scholar
- Hammond SM, Boettcher S, Caudy AA, Kobayashi R, Hannon GJ: Argonaute2, a link between genetic and biochemical analyses of RNAi. Science. 2001, 293: 1146-1150. 10.1126/science.1064023.View ArticlePubMedGoogle Scholar
- Ahlquist P: RNA-dependent RNA polymerases, viruses, and RNA silencing. Science. 2002, 296: 1270-1273. 10.1126/science.1069132.View ArticlePubMedGoogle Scholar
- Tabara H, Sarkissian M, Kelly WG, et al: The rde-1 gene, RNA interference, and transposon silencing in C-elegans. Cell. 1999, 99: 123-132. 10.1016/S0092-8674(00)81644-X.View ArticlePubMedGoogle Scholar
- Fire A: RNA-triggered gene silencing. Trends Genet. 1999, 15: 358-363. 10.1016/S0168-9525(99)01818-1.View ArticlePubMedGoogle Scholar
- Bitko V, Musiyenko A, Shulyayeva O, Barik S: Inhibition of respiratory viruses by nasally Inhibition of respiratory syncytial virus infection with intranasal siRNA nanoparticles targeting the viral NS1 gene. Nat Med. 2005, 11: 50-55. 10.1038/nm1164.View ArticlePubMedGoogle Scholar
- Zhang WD, Yang H, Kong XY, Mohapatra S, San Juan-Vergara H, et al: Inhibition of respiratory syncytial virus infection with intranasal siRNA nanoparticles targeting the viral NS1 gene. Nat Med. 2005, 11: 56-62. 10.1038/nm1174.View ArticlePubMedGoogle Scholar
- Oshima K, Kawasaki H, Soda Y, Tani K, Asano S, Taira K: Maxizymes and small hairpin-type RNAs that are driven by a tRNA promoter specifically cleave a chimeric gene associated with leukemia in vitro and in vivo. Cancer Res. 2003, 63: 6809-6814.PubMedGoogle Scholar
- Bossert B, Marozin S, Conzelmann KK: Nonstructural proteins NS1 and NS2 of bovine respiratory syncytial virus block activation of interferon regulatory factor 3. J Virol. 2003, 77: 8661-8668. 10.1128/JVI.77.16.8661-8668.2003.View ArticlePubMedPubMed CentralGoogle Scholar
- Matsuse H, Behera AK, Kumar M, Lockey RF, Mohapatra SS: Differential cytokine mRNA expression in Dermatophagoides farinae allergen-sensitized and respiratory syncytial virus-infected mice. Microbes Infect. 2000, 2: 753-759. 10.1016/S1286-4579(00)00500-1.View ArticlePubMedGoogle Scholar
- Matsuse H, Behera AK, Kumar M, Rabb H, Lockey RF, Mohapatra SS: Recurrent respiratory syncytial virus infections in allergen-sensitized mice lead to persistent airway inflammation and hyperresponsiveness. J Immunol. 2000, 164: 6583-6592.View ArticlePubMedGoogle Scholar
- Matsuse H, Kong X, Hu J, Wolf SF, Lockey RF, Mohapatra SS: Intranasal IL-12 produces discreet pulmonary and systemic effects on allergic inflammation and airway reactivity. Int Immunopharmacol. 2003, 3: 457-468. 10.1016/S1567-5769(02)00250-3.View ArticlePubMedGoogle Scholar
- Kumar M, Kong X, Behera A, Hellermann G, Lockey RF, Mohapatra SS: Chitosan IFN-?-pDNA Nanoparticle (CIN) Therapy for Allergic Asthma. Genet Vaccines Ther. 2003, 1: 3-10.1186/1479-0556-1-3.View ArticlePubMedPubMed CentralGoogle Scholar
- Kumar M, Behera AK, Hu J, Lockey RF, Mohapatra SS: IFN-gamma and IL-12 plasmid DNAs as vaccine adjuvant in a murine model of grass allergy. J Allergy Clin Immunol. 2001, 108: 402-408. 10.1067/mai.2001.117261.View ArticlePubMedGoogle Scholar
- Behera AK, Kumar M, Lockey RF, Mohapatra SS: 2'-5' Oligoadenylate synthetase plays a critical role in interferon-gamma inhibition of respiratory syncytial virus infection of human epithelial cells. J Biol Chem. 2002, 277: 25601-25608. 10.1074/jbc.M200211200.View ArticlePubMedGoogle Scholar
- Kong X, San Juan H, Behera A, Peeples ME, Wu J, Lockey RF, Mohapatra SS: ERK-1/2 activity is required for efficient RSV infection. FEBS Lett. 2004, 559: 33-38. 10.1016/S0014-5793(04)00002-X.View ArticlePubMedGoogle Scholar
- Kong X, San Juan H, Kumar M, Behera AK, Mohapatra SS, et al: Respiratory syncytial virus infection activates STAT signaling in human epithelial cells. Biochem Biophys Res Commun. 2003, 306: 616-622. 10.1016/S0006-291X(03)01008-8.View ArticlePubMedGoogle Scholar
- San-Juan-Vergara H, Peeples ME, Lockey RF, Mohapatra SS: Protein kinase C-alpha activity is required for respiratory syncytial virus fusion to human bronchial epithelial cells. J Virol. 2004, 78: 13717-13726. 10.1128/JVI.78.24.13717-13726.2004.View ArticlePubMedPubMed CentralGoogle Scholar
- Zhang W, Yang H, Kong X, Mohapatra S, San Juan-Vergara H, et al: Chitosan IFN-gamma-pDNA Nanoparticle (CIN) Therapy for Allergic Asthma. Nat Med. 2005, 11: 56-62. 10.1038/nm1174.View ArticlePubMedGoogle Scholar
- Mohapatra SS: Mucosal gene expression vaccine: a novel vaccine strategy for respiratory syncytial virus. Infect Dis J. 2003, 22 (suppl): S100-S103. 10.1097/01.inf.0000053894.31944.26.Google Scholar
This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.