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. 1989 Feb;75(2):324–328.

Flow cytometric analysis of pulmonary lymphocytes from mice infected with respiratory syncytial virus.

P J Openshaw 1
PMCID: PMC1542109  PMID: 2784741

Abstract

BALB/c mice were infected intra-nasally with respiratory syncytial virus (RSV) and cells recovered from the lungs by single or repeated bronchoalveolar lavage (BAL). Single BAL gave enough cells (1 - 2 x 10(5) cells/mouse) for morphological study of Giemsa stained cytospin preparations, and allowed the same mouse lungs to be assayed for virus titre or processed for histology. In normal or sham-infected mice the majority of recovered cells were macrophages, with less than 5% lymphocytes. Following infection the proportion of lymphocytes increased to about 20% between days 10 and 16 and decreased thereafter. By contrast, histological changes in the lung peaked at day 7 and resolved by day 9 or 10. Repeated BAL yielded a higher proportion of lymphocytes and provided enough cells for flow cytometric study of cell surface markers. The results of two-colour stains with antibodies to L3T4 (CD4), Lyt2 (CD8), mouse CD3, Thy 1.2 and surface immunoglobulin (SIg) were studied by flow cytometry. Analysis of the small non-granular cells (lymphocytes) showed that fewer than 5% of recovered lymphocytes were SIg+ (B cells), while most bore T cell surface markers. Early during infection (day 3-6), 30-60% of lymphocytes were Thy 1.2-, T3-, CD4-/CD8- and SIg- CD8+ cells outnumbered CD4+ cells 2:1 or 4:1 from day 6 of infection. In conclusion, CD8+ T cells constitute the major subpopulation of lymphocytes recovered from the lungs of mice recovering from RSV infection. BAL provides a useful quantitative method of assessing the cellular immune response in the lung following RSV infection.

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Selected References

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  1. Cannon M. J. Microplaque immunoperoxidase detection of infectious respiratory syncytial virus in the lungs of infected mice. J Virol Methods. 1987 Jul;16(4):293–301. doi: 10.1016/0166-0934(87)90014-0. [DOI] [PubMed] [Google Scholar]
  2. Dialynas D. P., Wilde D. B., Marrack P., Pierres A., Wall K. A., Havran W., Otten G., Loken M. R., Pierres M., Kappler J. Characterization of the murine antigenic determinant, designated L3T4a, recognized by monoclonal antibody GK1.5: expression of L3T4a by functional T cell clones appears to correlate primarily with class II MHC antigen-reactivity. Immunol Rev. 1983;74:29–56. doi: 10.1111/j.1600-065x.1983.tb01083.x. [DOI] [PubMed] [Google Scholar]
  3. Dickstein R. A., Kiremidjian-Schumacher L., Stotzky G. Effect of lidocaine on production of migration inhibitory factor and on macrophage motility: in vitro exposure of guinea pig lymphocytes and macrophages. J Leukoc Biol. 1984 Nov;36(5):621–632. doi: 10.1002/jlb.36.5.621. [DOI] [PubMed] [Google Scholar]
  4. Ennis F. A., Wells M. A., Butchko G. M., Albrecht P. Evidence that cytotoxic T cells are part of the host's response to influenza pneumonia. J Exp Med. 1978 Nov 1;148(5):1241–1250. doi: 10.1084/jem.148.5.1241. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Fulginiti V. A., Eller J. J., Sieber O. F., Joyner J. W., Minamitani M., Meiklejohn G. Respiratory virus immunization. I. A field trial of two inactivated respiratory virus vaccines; an aqueous trivalent parainfluenza virus vaccine and an alum-precipitated respiratory syncytial virus vaccine. Am J Epidemiol. 1969 Apr;89(4):435–448. doi: 10.1093/oxfordjournals.aje.a120956. [DOI] [PubMed] [Google Scholar]
  6. Haslam P. L., Turton C. W., Heard B., Lukoszek A., Collins J. V., Salsbury A. J., Turner-Warwick M. Bronchoalveolar lavage in pulmonary fibrosis: comparison of cells obtained with lung biopsy and clinical features. Thorax. 1980 Jan;35(1):9–18. doi: 10.1136/thx.35.1.9. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Hijlstra A., Van Dorp W., Daha M. R., Leslie G. Q. The effect of lidocaine on the processing of soluble immune aggregates and immune complexes by peritoneal macrophages. Immunology. 1980 Sep;41(1):237–244. [PMC free article] [PubMed] [Google Scholar]
  8. Holt P. G. Alveolar macrophages. I. A simple technique for the preparation of high numbers of viable alveolar macrophages from small laboratory animals. J Immunol Methods. 1979;27(2):189–198. doi: 10.1016/0022-1759(79)90264-3. [DOI] [PubMed] [Google Scholar]
  9. Hunninghake G. W., Kawanami O., Ferrans V. J., Young R. C., Jr, Roberts W. C., Crystal R. G. Characterization of the inflammatory and immune effector cells in the lung parenchyma of patients with interstitial lung disease. Am Rev Respir Dis. 1981 Apr;123(4 Pt 1):407–412. doi: 10.1164/arrd.1981.123.4.407. [DOI] [PubMed] [Google Scholar]
  10. Kim H. W., Canchola J. G., Brandt C. D., Pyles G., Chanock R. M., Jensen K., Parrott R. H. Respiratory syncytial virus disease in infants despite prior administration of antigenic inactivated vaccine. Am J Epidemiol. 1969 Apr;89(4):422–434. doi: 10.1093/oxfordjournals.aje.a120955. [DOI] [PubMed] [Google Scholar]
  11. Leo O., Foo M., Sachs D. H., Samelson L. E., Bluestone J. A. Identification of a monoclonal antibody specific for a murine T3 polypeptide. Proc Natl Acad Sci U S A. 1987 Mar;84(5):1374–1378. doi: 10.1073/pnas.84.5.1374. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Marshak-Rothstein A., Fink P., Gridley T., Raulet D. H., Bevan M. J., Gefter M. L. Properties and applications of monoclonal antibodies directed against determinants of the Thy-1 locus. J Immunol. 1979 Jun;122(6):2491–2497. [PubMed] [Google Scholar]
  13. Ortaldo J. R., Herberman R. B. Heterogeneity of natural killer cells. Annu Rev Immunol. 1984;2:359–394. doi: 10.1146/annurev.iy.02.040184.002043. [DOI] [PubMed] [Google Scholar]
  14. Prince G. A., Horswood R. L., Berndt J., Suffin S. C., Chanock R. M. Respiratory syncytial virus infection in inbred mice. Infect Immun. 1979 Nov;26(2):764–766. doi: 10.1128/iai.26.2.764-766.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Prince G. A., Jenson A. B., Hemming V. G., Murphy B. R., Walsh E. E., Horswood R. L., Chanock R. M. Enhancement of respiratory syncytial virus pulmonary pathology in cotton rats by prior intramuscular inoculation of formalin-inactiva ted virus. J Virol. 1986 Mar;57(3):721–728. doi: 10.1128/jvi.57.3.721-728.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Prince G. A., Jenson A. B., Horswood R. L., Camargo E., Chanock R. M. The pathogenesis of respiratory syncytial virus infection in cotton rats. Am J Pathol. 1978 Dec;93(3):771–791. [PMC free article] [PubMed] [Google Scholar]
  17. Rabinovitch M., DeStefano M. J. Use of the local anesthetic lidocaine for cell harvesting and subcultivation. In Vitro. 1975 Nov-Dec;11(6):379–381. doi: 10.1007/BF02616374. [DOI] [PubMed] [Google Scholar]
  18. Reynolds H. Y. Bronchoalveolar lavage. Am Rev Respir Dis. 1987 Jan;135(1):250–263. doi: 10.1164/arrd.1987.135.1.250. [DOI] [PubMed] [Google Scholar]
  19. Rodgers B. C., Mims C. A. Role of macrophage activation and interferon in the resistance of alveolar macrophages from infected mice to influenza virus. Infect Immun. 1982 Jun;36(3):1154–1159. doi: 10.1128/iai.36.3.1154-1159.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Sarmiento M., Glasebrook A. L., Fitch F. W. IgG or IgM monoclonal antibodies reactive with different determinants on the molecular complex bearing Lyt 2 antigen block T cell-mediated cytolysis in the absence of complement. J Immunol. 1980 Dec;125(6):2665–2672. [PubMed] [Google Scholar]
  21. Sims D. G., Downham M. A., McQuillin J., Gardner P. S. Respiratory syncytial virus infection in north-east England. Br Med J. 1976 Nov 6;2(6044):1095–1098. doi: 10.1136/bmj.2.6044.1095. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Stott E. J., Taylor G., Ball L. A., Anderson K., Young K. K., King A. M., Wertz G. W. Immune and histopathological responses in animals vaccinated with recombinant vaccinia viruses that express individual genes of human respiratory syncytial virus. J Virol. 1987 Dec;61(12):3855–3861. doi: 10.1128/jvi.61.12.3855-3861.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Stott E. J., Taylor G. Respiratory syncytial virus. Brief review. Arch Virol. 1985;84(1-2):1–52. doi: 10.1007/BF01310552. [DOI] [PubMed] [Google Scholar]
  24. Taylor G., Stott E. J., Hayle A. J. Cytotoxic lymphocytes in the lungs of mice infected with respiratory syncytial virus. J Gen Virol. 1985 Dec;66(Pt 12):2533–2538. doi: 10.1099/0022-1317-66-12-2533. [DOI] [PubMed] [Google Scholar]
  25. Taylor G., Stott E. J., Hughes M., Collins A. P. Respiratory syncytial virus infection in mice. Infect Immun. 1984 Feb;43(2):649–655. doi: 10.1128/iai.43.2.649-655.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Wyde P. R., Peavy D. L., Cate T. R. Morphological and cytochemical characterization of cells infiltrating mouse lungs after influenza infection. Infect Immun. 1978 Jul;21(1):140–146. doi: 10.1128/iai.21.1.140-146.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]

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