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(1/561) Interferon gamma expressed by a recombinant respiratory syncytial virus attenuates virus replication in mice without compromising immunogenicity.

Interferon gamma (IFN-gamma) has pleiotropic biological effects, including intrinsic antiviral activity as well as stimulation and regulation of immune responses. An infectious recombinant human respiratory syncytial virus (rRSV/mIFN-gamma) was constructed that encodes murine (m) IFN-gamma as a separate gene inserted into the G-F intergenic region. Cultured cells infected with rRSV/mIFN-gamma secreted 22 microg mIFN-gamma per 10(6) cells. The replication of rRSV/mIFN-gamma, but not that of a control chimeric rRSV containing the chloramphenicol acetyl transferase (CAT) gene as an additional gene, was 63- and 20-fold lower than that of wild-type (wt) RSV in the upper and lower respiratory tract, respectively, of mice. Thus, the attenuation of rRSV/mIFN-gamma in vivo could be attributed to the activity of mIFN-gamma and not to the presence of the additional gene per se. The mice were completely resistant to subsequent challenge with wt RSV. Despite its growth restriction, infection of mice with rRSV/mIFN-gamma induced a level of RSV-specific antibodies that, on day 56, was comparable to or greater than that induced by infection with wt RSV. Mice infected with rRSV/mIFN-gamma developed a high level of IFN-gamma mRNA and an increased amount of interleukin 12 p40 mRNA in their lungs, whereas other cytokine mRNAs tested were unchanged compared with those induced by wt RSV. Because attenuation of RSV typically is accompanied by a reduction in immunogenicity, expression of IFN-gamma by an rRSV represents a method of attenuation in which immunogenicity can be maintained rather than be reduced.  (+info)

(2/561) Two neutralizing human anti-RSV antibodies: cloning, expression, and characterization.

BACKGROUND: Respiratory syncytial virus (RSV) infection is a major problem in the newborn and aging populations. Fully human monoclonal antibodies with the ability to neutralize RSV could have a major impact on the immunotherapy of the disease. The generation of human antibodies has been difficult because there exists no general way to activate B cells against an antigen of choice in vitro. MATERIALS AND METHODS: Human spleen cells from individuals exposed to RSV were used to repopulate SCID mice. Hu-SCID mice were boosted with RSV fusion (F)-protein and subsequently developed B cell tumors. The tumors were removed and cultured and subcloned in vitro, using a feeder layer of CD154-expressing T cells. Two of these tumors produced the antibodies designated RF-1 and RF-2. VL genes were isolated by standard PCR techniques, however, it was necessary to use high-temperature reverse transcriptase to clone the VH genes. RESULTS: RF-1 and RF-2 VH genes were both found to be closely related members of the VH2 family. Vk genes originated from the VK III family. RF-1 and RF-2 recombinant antibodies expressed in CHO cells (cRF-1 and cRF-2) were found to have affinities for RSV F-protein of 0.1 nM and 0.07 nM, respectively, and both were able to neutralize several A and B subtypes of RSV. CONCLUSION: The technique of immortalizing human B lymphocytes, by passage in SCID mice and expression as recombinant antibodies in CHO cells, provides a method by which high-affinity human antibodies can be developed for immunotherapy of viral diseases.  (+info)

(3/561) Molecular epidemiology of respiratory syncytial virus in The Gambia.

Respiratory syncytial virus (RSV) infection in The Gambia occurs seasonally in association with the rainy season. This study examined the genetic variability of RSV isolates from four consecutive epidemics from 1993-6. Each epidemic was made up of a number of variants which were replaced in subsequent epidemics. Analysis of attachment (G) protein gene sequences showed that isolates were closely related to those observed in the rest of the world. However, many isolates from 1993 and 1994 were unlike other isolates observed in the developed world during this period and were more similar to isolates from 1984 in Europe. In addition, the most commonly observed genotype in the UK in the 1990s was not detected in The Gambia during this period.  (+info)

(4/561) Respiratory syncytial virus infection: immune response, immunopathogenesis, and treatment.

Respiratory syncytial virus (RSV) is the single most important cause of lower respiratory tract infection during infancy and early childhood. Once RSV infection is established, the host immune response includes the production of virus-neutralizing antibodies and T-cell-specific immunity. The humoral immune response normally results in the development of anti-RSV neutralizing-antibody titers, but these are often suboptimal during an infant's initial infection. Even when the production of RSV neutralizing antibody following RSV infection is robust, humoral immunity wanes over time. Reinfection during subsequent seasons is common. The cellular immune response to RSV infection is also important for the clearance of virus. This immune response, vital for host defense against RSV, is also implicated in the immunopathogenesis of severe lower respiratory tract RSV bronchiolitis. Many details of the immunology and immunopathologic mechanisms of RSV disease known at present have been learned from rodent models of RSV disease and are discussed in some detail. In addition, the roles of immunoglobulin E, histamine, and eosinophils in the immunopathogenesis of RSV disease are considered. Although the treatment of RSV bronchiolitis is primarily supportive, the role of ribavirin is briefly discussed. Novel approaches to the development of new antiviral drugs with promising anti-RSV activity in vitro are also described.  (+info)

(5/561) Incomplete regulation of NF-kappaB by IkappaBalpha during respiratory syncytial virus infection in A549 cells.

Respiratory syncytial virus (RSV) infection of airway epithelial cells results in persistent NF-kappaB activation and NF-kappaB-mediated interleukin-8 production. Previous studies in airway epithelial cells demonstrated that tumor necrosis factor alpha (TNF-alpha)-induced NF-kappaB activation is transient due to regulation by IkappaBalpha. However, during RSV infection, IkappaBalpha has only a partial inhibitory effect on NF-kappaB activation. Studies presented here demonstrate that neither increased IkappaBalpha production which occurs as a result of RSV-induced NF-kappaB activation nor inhibition of proteasome-mediated IkappaBalpha degradation results in a reversal of RSV-induced NF-kappaB activation. Thus, while manipulation of IkappaBalpha results in reversal of TNF-alpha-induced NF-kappaB activation, manipulation of IkappaBalpha does not result in a reversal of RSV-induced NF-kappaB activation.  (+info)

(6/561) CD8 T cells are essential in the development of respiratory syncytial virus-induced lung eosinophilia and airway hyperresponsiveness.

Viral respiratory infections can cause bronchial hyperresponsiveness and exacerbate asthma. In mice, respiratory syncytial virus (RSV) infection results in airway hyperresponsiveness (AHR) and eosinophil influx into the airways. The immune cell requirements for these responses to RSV infection are not well defined. To delineate the role of CD8 T cells in the development of RSV-induced AHR and lung eosinophilia, we tested the ability of mice depleted of CD8 T cells to develop these symptoms of RSV infection. BALB/c mice were depleted of CD8 T cells using anti-CD8 Ab treatment before intranasal administration of infectious RSV. Six days postinfection, airway responsiveness to inhaled methacholine was assessed by barometric body plethysmography, and numbers of lung eosinophils and levels of IFN-gamma, IL-4, and IL-5 in bronchoalveolar lavage fluid were monitored. RSV infection resulted in airway eosinophilia and AHR in control mice, but not in CD8-depleted animals. Further, whereas RSV-infected mice secreted increased amounts of IL-5 into the airways as compared with noninfected controls, no IL-5 was detectable in both bronchoalveolar lavage fluid and culture supernatants from CD8-depleted animals. Treatment of CD8-depleted mice with IL-5 fully restored both lung eosinophilia and AHR. We conclude that CD8 T cells are essential for the influx of eosinophils into the lung and the development of AHR in response to RSV infection.  (+info)

(7/561) Absence of lung immunopathology following respiratory syncytial virus (RSV) challenge in mice immunized with a recombinant RSV G protein fragment.

The relative immunopathogenic potential of a recombinant fusion protein incorporating residues 130-230 of respiratory syncytial virus (RSV-A) G protein (BBG2Na), formalin-inactivated RSV-A (FI-RSV), and phosphate-buffered saline (PBS) was investigated in mice after immunization and RSV challenge. FI-RSV priming resulted in massive infiltration of B cells and activated CD4(+) and CD8(+) T lymphocytes in mediastinal lymph nodes (MLN) and lungs, where eosinophilia and elevated IFN-gamma, IL-2, -4, -5, -10, and -13 mRNA transcripts were also detected. PBS-primed mice showed only elevated pulmonary IL-2 and IFN-gamma mRNAs, while an activated CD8(+) T cell peak was detected in MLN and lungs. Cell infiltration also occurred in MLN of BBG2Na-immunized mice. However, there was no evidence of T cell, B cell, or granulocyte infiltration or activation in lungs, while transient transcription of Th1-type cytokine genes was evident. The absence of pulmonary infiltration is unlikely due to insufficient viral antigen. Thus, this recombinant fusion RSV G fragment does not prime for adverse pulmonary immunopathologic responses.  (+info)

(8/561) Identification of multiple protective epitopes (protectopes) in the central conserved domain of a prototype human respiratory syncytial virus G protein.

A recombinant fusion protein (BBG2Na) comprising the central conserved domain of the respiratory syncytial virus subgroup A (RSV-A) (Long) G protein (residues 130 to 230) and an albumin binding domain of streptococcal protein G was shown previously to protect mouse upper (URT) and lower (LRT) respiratory tracts against intranasal RSV challenge (U. F. Power, H. Plotnicky-Gilquin, T. Huss, A. Robert, M. Trudel, S. Stahl, M. Uhlen, T. N. Nguyen, and H. Binz, Virology 230:155-166, 1997). Panels of monoclonal antibodies (MAbs) and synthetic peptides were generated to facilitate dissection of the structural elements of this domain implicated in protective efficacy. All MAbs recognized native RSV-A antigens, and five linear B-cell epitopes were identified; these mapped to residues 152 to 163, 165 to 172, 171 to 187 (two overlapping epitopes), and 196 to 204, thereby covering the highly conserved cysteine noose domain. Antibody passive-transfer and peptide immunization studies revealed that all epitopes were implicated in protection of the LRT, but not likely the URT, against RSV-A challenge. Pepscan analyses of anti-RSV-A and anti-BBG2Na murine polyclonal sera revealed lower-level epitope usage within the central conserved region in the former, suggesting diminished immunogenicity of the implicated epitopes in the context of the whole virus. However, Pepscan analyses of RSV-seropositive human sera revealed that all of the murine B-cell protective epitopes (protectopes) that mapped to the central conserved domain were recognized in man. Should these murine protectopes also be implicated in human LRT protection, their clustering around the highly conserved cysteine noose region will have important implications for the development of RSV vaccines.  (+info)