In vitro susceptibility to rhinovirus infection is greater for bronchial than for nasal airway epithelial cells in human subjects Nilceia Lopez-Souza,

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In vitro susceptibility to rhinovirus infection is greater for bronchial than for nasal airway epithelial cells in human subjects Nilceia Lopez-Souza, MD, PhD, Silvio Favoreto, DDS, PhD, Hofer Wong, BS, Theresa Ward, RN, Shigeo Yagi, PhD, David Schnurr, MD, Walter E. Finkbeiner, MD, PhD, Gregory M. Dolganov, PhD, Jonathan H. Widdicombe, PhD, Homer A. Boushey, MD, Pedro C. Avila, MD Journal of Allergy and Clinical Immunology Volume 123, Issue 6, Pages 1384-1390.e2 (June 2009) DOI: 10.1016/j.jaci.2009.03.010 Copyright © 2009 American Academy of Allergy, Asthma & Immunology Terms and Conditions

Fig 1 Light microscopy of epithelial cultures. Cross-sections of passage 1 HNE (A and B) and HBE (C and D) cell cultures from control (Fig 1, A and C) and asthmatic (Fig 1, B and D) subjects grown at an air-liquid interface for 21 days and stained with hematoxylin and eosin. Cultures obtained from all groups showed similar morphology, forming multilayered sheets containing variable numbers of ciliated cells and scattered intraepithelial vacuoles. Scale bar = 25 μm. Journal of Allergy and Clinical Immunology 2009 123, 1384-1390.e2DOI: (10.1016/j.jaci.2009.03.010) Copyright © 2009 American Academy of Allergy, Asthma & Immunology Terms and Conditions

Fig 2 HRV-16 replication on HNE and HBE cells measured by means of real-time PCR 48 hours after viral inoculation. Intracellular HRV RNA load was higher in all infected (RV) compared with noninfected control (C) cultures (∗P < .002, generalized estimating equations) from asthmatic (n = 6) and healthy (n = 5) subjects. HRV replicated to a greater extent in HBE cell cultures than in HNE cell cultures in both groups. There were no differences between groups for nasal or bronchial cells. Bars represent the median, and whiskers represent the interquartile range. Journal of Allergy and Clinical Immunology 2009 123, 1384-1390.e2DOI: (10.1016/j.jaci.2009.03.010) Copyright © 2009 American Academy of Allergy, Asthma & Immunology Terms and Conditions

Fig 3 Immunocytochemistry for rhinovirus RNA (HRV-16). HRV-16 virus capsid protein 2 was stained with Cyanine 3 (red), and nucleic acids were stained with YO-PRO-1 iodide (green). A, Micrograph of noninfected control HBE cells showing no HRV-16. B, Micrograph of infected HBE cells showing a high number of HRV-16–infected cells, some with condensed chromatin/nuclei (bright green). Journal of Allergy and Clinical Immunology 2009 123, 1384-1390.e2DOI: (10.1016/j.jaci.2009.03.010) Copyright © 2009 American Academy of Allergy, Asthma & Immunology Terms and Conditions

Fig 4 Levels of IL-6, IL-1α, interferon-inducible protein 10 (IP-10), and RANTES in supernatants from HBE and HNE cell cultures collected 48 hours after HRV-16 (RV) or sham (C, control) inoculation. Cultures from asthmatic (n = 6) and healthy (n = 5) subjects displayed similar results. HBE cell cultures, but not HNE cell cultures, produced greater amounts of mediators after HRV-16 infection. Bars represent the median, and whiskers represent the interquartile range. Data analysis was performed with generalized estimating equations. Journal of Allergy and Clinical Immunology 2009 123, 1384-1390.e2DOI: (10.1016/j.jaci.2009.03.010) Copyright © 2009 American Academy of Allergy, Asthma & Immunology Terms and Conditions