Cell-specific expression of RANTES, MCP-1, and MIP-1alpha by lower airway epithelial cells and eosinophils infected with respiratory syncytial virus - PubMed (original) (raw)

Cell-specific expression of RANTES, MCP-1, and MIP-1alpha by lower airway epithelial cells and eosinophils infected with respiratory syncytial virus

B Olszewska-Pazdrak et al. J Virol. 1998 Jun.

Abstract

Respiratory syncytial virus (RSV) is the major cause of acute bronchiolitis in infancy, a syndrome characterized by wheezing, respiratory distress, and the pathologic findings of peribronchial mononuclear cell infiltration and release of inflammatory mediators by basophil and eosinophil leukocytes. Composition and activation of this cellular response are thought to rely on the discrete target cell selectivity of C-C chemokines. We demonstrate that infection in vitro of human epithelial cells of the lower respiratory tract by RSV induced dose- and time-dependent increases in mRNA and protein secretion for RANTES (regulated upon activation, normal T-cell expressed and presumably secreted), monocyte chemotactic protein-1 (MCP-1), and macrophage inflammatory protein-1alpha (MIP-1alpha). Production of MCP-1 and MIP-1alpha was selectively localized only in epithelial cells of the small airways and lung. Exposure of epithelial cells to gamma interferon (IFN-gamma), in combination with RSV infection, induced a significant increase in RANTES production that was synergistic with respect to that obtained by RSV infection or IFN-gamma treatment alone. Epithelial cell-derived chemokines exhibited a strong chemotactic activity for normal human blood eosinophils. Furthermore, eosinophils were susceptible to RSV and released RANTES and MIP-1alpha as a result of infection. Therefore, the inflammatory process in RSV-induced bronchiolitis appears to be triggered by the infection of epithelial cells and further amplified via mechanisms driven by IFN-gamma and by the secretion of eosinophil chemokines.

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Figures

FIG. 1

FIG. 1

C-C chemokine secretion by RSV-infected epithelial cells. NHBE, SAE, and A549 cells were infected with RSV at an MOI of 1 (RSV) or cultured in control medium (Ctrl) for 48 h. RANTES, MCP-1, and MIP-1α concentrations were determined in the supernatant by specific ELISA. The results are expressed as mean ± SD of five experiments. ∗, P < 0.05 compared with uninfected control cells; †, P < 0.02 for A549 cells compared with SAE and NHBE cells; ‡, P < 0.01 for SAE cells compared with NHBE cells.

FIG. 2

FIG. 2

Effect of RSV infectious dose and UV-inactivated RSV on RANTES production by airway epithelial cells. NHBE cells were infected with RSV at MOIs of 5, 1, 0.5, and 0.2 for 48 h and A549 cells at MOIs of 1, 0.5, and 0.2 for 24 h. In experiments designed to determine the requirement of replicating virus for RANTES induction, NHBE and A549 cells were exposed to UV-inactivated RSV (UV). Data are expressed as mean ± SD of three experiments. ∗, P < 0.05 for each infectious dose compared with control or with UV-inactivated RSV.

FIG. 3

FIG. 3

Kinetics of RANTES and MIP-1α accumulation in supernatant from RSV-infected A549, SAE, and NHBE cells. Epithelial cells were infected with RSV at an MOI of 1 (RSV) or cultured in control medium (Ctrl). After indicated time of incubation, supernatants were collected for RANTES and MIP-1α determination by ELISA. The results are expressed as mean ± SD of four experiments. ∗, P < 0.05 compared with control at each time point.

FIG. 4

FIG. 4

Time-dependent expression of chemokine mRNA by RSV-infected NHBE and A549 cells. Total RNA was extracted from control (Ctrl) and RSV-infected (RSV) cells at the indicated time. RT-PCR was performed with specific primers for human RANTES, MIP-1α, MCP-1, and β-actin. PCR products were visualized with ethidium bromide staining on agarose gel. Expression of the housekeeping gene encoding β-actin is shown for comparison to demonstrate relative equal loading of the RT-PCR mixtures for all samples. A representative experiment from three performed on separate cell culture samples is shown.

FIG. 5

FIG. 5

Synergistic effect of RSV and IFN-γ on RANTES protein release by NHBE cells. Epithelial cells were infected with RSV at an MOI of 0.2 and treated with IFN-γ (100 U/ml) alone or in combination with RSV. RANTES was detected in supernatants by ELISA after 48 h of incubation. The results are expressed as mean ± SD of three experiments. ∗, P < 0.01 compared with control, RSV, and IFN-γ alone.

FIG. 6

FIG. 6

Biological activity of RANTES released by RSV-infected NHBE and A549 cells. Eosinophil chemotaxis was determined in the presence of supernatant from RSV-infected cells (RSV-CM) or from uninfected cells (Ctrl). Neutralizing experiments were performed in the presence of specific anti-RANTES MAb or mouse IgG isotype control. PAF (10−7 M) was used as a positive control. Results are expressed as mean ± SD of four determinations for each set of experiments. ∗, P < 0.01 for RSV-CM, RSV-CM plus mouse IgG, and PAF compared with control; ∗∗, P < 0.01 for RSV-CM plus anti-RANTES MAb compared with RSV-CM.

FIG. 7

FIG. 7

Eosinophil infection by RSV. Eosinophils were cultured with control medium (A) or infected with RSV for 2 h (B) or 16 h (C). Cytospin preparations of eosinophils were stained with anti-RSV Fgp MAb followed by a fluorescein isothiocyanate-conjugated anti-mouse F(ab′)2 IgG antibody. In the preparations of eosinophils that were exposed to RSV for 2 h, the Fgp staining was concentrated in a pericytoplasmic halo. Upon RSV infection for 16 h, typical intracytoplasmic granular fluorescence immunoreactivity was observed.

FIG. 8

FIG. 8

RANTES (A) and MIP-1α (B) production by eosinophils cultured with purified RSV. Eosinophils were infected with RSV at an MOI of 10 (E+RSV) or incubated with culture medium (E). After 16 h, the supernatants were collected for RANTES and MIP-1α determination by ELISA. For the measurement of total content of RANTES and MIP-1α, eosinophils were lysed by Triton X-100 (E max). The results are expressed as mean ± SD of five experiments using eosinophil preparations from different donors. ∗, P < 0.01 compared with eosinophils cultured with medium or lysed by Triton X-100.

References

    1. Aherne W, Bird T, Court S D, Gardner P S, McQuillin J. Pathological changes in virus infections of the lower respiratory tract in children. J Clin Pathol. 1970;23:7–18. - PMC - PubMed
    1. Alam R, Let-Brown M A, Forsythe P A, Anderson-Walters D J, Kenamore C, Kormos C, Grant J A. Monocyte chemotactic and activating factor is a potent histamine-releasing factor for basophils. J Clin Invest. 1992;89:723–728. - PMC - PubMed
    1. Alam R, Stafford S, Forsythe P, Harrison R, Faubion D, Lett-Brown M A, Grant J A. RANTES is a chemotactic and activating factor for human eosinophils. J Immunol. 1993;150:3442–3447. - PubMed
    1. Alam, R., J. York, M. Boyers, S. Stafford, J. A. Grant, J. Lee, P. Forsythe, T. Sim, and N. Ida. Increased MCP-1, RANTES, and MIP-1α in bronchoalveolar lavage fluid of allergic asthmatic patients. 1996. Am. J. Respir. Crit. Care Med. 153:1398–1404. - PubMed
    1. Baggiolini M, Dewald B, Moser B. Interleukin-8 and related chemotactic, cytokines—CXC and CC chemokines. Adv Immunol. 1994;55:97–179. - PubMed

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