Expression of IL-6, IL-8, and RANTES on human bronchial epithelial cells, NCI-H292, induced by influenza virus A Satoshi Matsukura, MD, Fumio Kokubu,

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Presentation transcript:

Expression of IL-6, IL-8, and RANTES on human bronchial epithelial cells, NCI-H292, induced by influenza virus A Satoshi Matsukura, MD, Fumio Kokubu, MD, Hiromichi Noda, MD, Hisahiro Tokunaga, MD, Mitsuru Adachi, MD Journal of Allergy and Clinical Immunology Volume 98, Issue 6, Pages 1080-1087 (December 1996) DOI: 10.1016/S0091-6749(96)80195-3 Copyright © 1996 Mosby, Inc. Terms and Conditions

FIG. 1 Detection of viral RNA by RT-PCR. One hundred microliters of viral stock (1 × 106 PFU/ml influenza virus A, H3N2) was added to culture plates of NCI-H292 cells. At 10 minutes after addition of viral stock or at 1 hour after rocking of plates every 10 minutes, supernatant of the plates was removed and 2 ml of fresh RPMI-1640 medium was added. At 10 minutes or at 24 hours after viral inoculation, cells were harvested. Hemagglutinin 3 gene of influenza virus A was detected from collected cells by RT-PCR. M, Size marker; 1, cells collected at 10 minutes after virus inoculation; 2, cells collected at 24 hours after virus inoculation. Representative of three separate experiments. Journal of Allergy and Clinical Immunology 1996 98, 1080-1087DOI: (10.1016/S0091-6749(96)80195-3) Copyright © 1996 Mosby, Inc. Terms and Conditions

FIG. 2 Concentration of IL-6 and IL-8 in culture medium. Bronchial epithelial cells, NCI-H292, were incubated with 10 ng/ml IL-1β or TNF-α. NCI-H292 cells were also infected with influenza virus A. Culture medium was collected at 6, 24, 48, and 72 hours after stimulation or infection. Concentrations of IL-6 and IL-8 in collected medium were determined by ELISA. A, IL-6 concentration. B, IL-8 concentration. *p < 0.05 compared with control. Five separate experiments were performed. Journal of Allergy and Clinical Immunology 1996 98, 1080-1087DOI: (10.1016/S0091-6749(96)80195-3) Copyright © 1996 Mosby, Inc. Terms and Conditions

FIG. 2 Concentration of IL-6 and IL-8 in culture medium. Bronchial epithelial cells, NCI-H292, were incubated with 10 ng/ml IL-1β or TNF-α. NCI-H292 cells were also infected with influenza virus A. Culture medium was collected at 6, 24, 48, and 72 hours after stimulation or infection. Concentrations of IL-6 and IL-8 in collected medium were determined by ELISA. A, IL-6 concentration. B, IL-8 concentration. *p < 0.05 compared with control. Five separate experiments were performed. Journal of Allergy and Clinical Immunology 1996 98, 1080-1087DOI: (10.1016/S0091-6749(96)80195-3) Copyright © 1996 Mosby, Inc. Terms and Conditions

FIG. 3 Concentrations of RANTES in culture medium. Bronchial epithelial cells, NCI-H292, were infected with influenza virus A. Medium was collected 6, 24, 48, and 72 hours later. Concentrations of RANTES in medium of control cells and cells infected with influenza virus were determined by ELISA. *p < 0.05 compared with control. Five separate experiments were performed. Journal of Allergy and Clinical Immunology 1996 98, 1080-1087DOI: (10.1016/S0091-6749(96)80195-3) Copyright © 1996 Mosby, Inc. Terms and Conditions

FIG. 4 Effects of antiserum on cytokine production. Viral stock was incubated with antiserum and then added to NCI-H292 cells. After incubation for 10 minutes, supernatant was removed and fresh RPMI-1640 medium was added. Forty-eight hours later, culture medium was collected, and concentration of each cytokine was determined by ELISA. A, Concentration of IL-6. B, Concentration of IL-8. C, Concentration of RANTES. Three separate experiments were performed. Journal of Allergy and Clinical Immunology 1996 98, 1080-1087DOI: (10.1016/S0091-6749(96)80195-3) Copyright © 1996 Mosby, Inc. Terms and Conditions

FIG. 4 Effects of antiserum on cytokine production. Viral stock was incubated with antiserum and then added to NCI-H292 cells. After incubation for 10 minutes, supernatant was removed and fresh RPMI-1640 medium was added. Forty-eight hours later, culture medium was collected, and concentration of each cytokine was determined by ELISA. A, Concentration of IL-6. B, Concentration of IL-8. C, Concentration of RANTES. Three separate experiments were performed. Journal of Allergy and Clinical Immunology 1996 98, 1080-1087DOI: (10.1016/S0091-6749(96)80195-3) Copyright © 1996 Mosby, Inc. Terms and Conditions

FIG. 4 Effects of antiserum on cytokine production. Viral stock was incubated with antiserum and then added to NCI-H292 cells. After incubation for 10 minutes, supernatant was removed and fresh RPMI-1640 medium was added. Forty-eight hours later, culture medium was collected, and concentration of each cytokine was determined by ELISA. A, Concentration of IL-6. B, Concentration of IL-8. C, Concentration of RANTES. Three separate experiments were performed. Journal of Allergy and Clinical Immunology 1996 98, 1080-1087DOI: (10.1016/S0091-6749(96)80195-3) Copyright © 1996 Mosby, Inc. Terms and Conditions

FIG. 5 RANTES mRNA detected by RT-PCR. NCI-H 292 cells were stimulated with IL-1β, TNF-α, and IL-6 plus IL-8 (10 ng/ml each). NCI-H292 cells were also infected with influenza virus A. After 24 hours, epithelial cells were harvested and mRNA of RANTES (A) and β-actin (B) were assayed by RT-PCR. M, Size marker; 1, control cells; 2, cells infected by influenza virus A; 3, cells stimulated with IL-1β; 4, cells stimulated with TNF-α; 5, cells stimulated with IL-6 plus IL-8. Representative of five separate experiments. Journal of Allergy and Clinical Immunology 1996 98, 1080-1087DOI: (10.1016/S0091-6749(96)80195-3) Copyright © 1996 Mosby, Inc. Terms and Conditions

FIG. 5 RANTES mRNA detected by RT-PCR. NCI-H 292 cells were stimulated with IL-1β, TNF-α, and IL-6 plus IL-8 (10 ng/ml each). NCI-H292 cells were also infected with influenza virus A. After 24 hours, epithelial cells were harvested and mRNA of RANTES (A) and β-actin (B) were assayed by RT-PCR. M, Size marker; 1, control cells; 2, cells infected by influenza virus A; 3, cells stimulated with IL-1β; 4, cells stimulated with TNF-α; 5, cells stimulated with IL-6 plus IL-8. Representative of five separate experiments. Journal of Allergy and Clinical Immunology 1996 98, 1080-1087DOI: (10.1016/S0091-6749(96)80195-3) Copyright © 1996 Mosby, Inc. Terms and Conditions