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Influenza H5N1 and H1N1 virus replication and innate immune responses in bronchial epithelial cells are influenced by the state of differentiation.

Chan RW, Yuen KM, Yu WC, Ho CC, Nicholls JM, Peiris JS, Chan MC - PLoS ONE (2010)

Bottom Line: We found that the in vitro culture of the well differentiated NHBE cells acquired the physiological properties of normal human bronchi tissue which express high level of alpha2-6-linked sialic acid receptors and human airway trypsin-like (HAT) protease, in contrast to the low expression in the non-differentiated NHBE cells.The reduced virus replication efficiency partially accounts for the lower interferon-beta responses in influenza H5N1 virus infected well differentiated NHBE cells.Since influenza infection in the bronchial epithelium will lead to tissue damage and associate with the epithelium regeneration, the data generated from the undifferentiated NHBE cultures may also be relevant to disease pathogenesis.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Queen Mary Hospital, Pokfulam, Hong Kong SAR, People's Republic of China.

ABSTRACT
Influenza H5N1 virus continues to be enzootic in poultry and transmits zoonotically to humans. Although a swine-origin H1N1 virus has emerged to become pandemic, its virulence for humans remains modest in comparison to that seen in zoonotic H5N1 disease. As human respiratory epithelium is the primary target cells for influenza viruses, elucidating the viral tropism and host innate immune responses of influenza H5N1 virus in human bronchial epithelium may help to understand the pathogenesis. Here we established primary culture of undifferentiated and well differentiated normal human bronchial epithelial (NHBE) cells and infected with highly pathogenic influenza H5N1 virus (A/Vietnam/3046/2004) and a seasonal influenza H1N1 virus (A/Hong Kong/54/1998), the viral replication kinetics and cytokine and chemokine responses were compared by qPCR and ELISA. We found that the in vitro culture of the well differentiated NHBE cells acquired the physiological properties of normal human bronchi tissue which express high level of alpha2-6-linked sialic acid receptors and human airway trypsin-like (HAT) protease, in contrast to the low expression in the non-differentiated NHBE cells. When compared to H1N1 virus, the H5N1 virus replicated more efficiently and induced a stronger type I interferon response in the undifferentiated NHBE cells. In contrast, in well differentiated cultures, H5N1 virus replication was less efficient and elicited a lower interferon-beta response in comparison with H1N1 virus. Our data suggest that the differentiation of bronchial epithelial cells has a major influence in cells' permissiveness to human H1N1 and avian H5N1 viruses and the host innate immune responses. The reduced virus replication efficiency partially accounts for the lower interferon-beta responses in influenza H5N1 virus infected well differentiated NHBE cells. Since influenza infection in the bronchial epithelium will lead to tissue damage and associate with the epithelium regeneration, the data generated from the undifferentiated NHBE cultures may also be relevant to disease pathogenesis.

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Lectin binding: MAL-I binds to Sia-α2-3Galβ1-4GlcNAc (Panel A, D, G and J), MAL-II binds to Sia-α2-3Galβ1-3GalNAc (Panel B, E, H and K) and SNA binds to Sia-α2-6-linkage (Panel C, F, I and L) to determine the Sias distribution on the ud- and wd-NHBE cells.MAL-I, MAL-II and SNA bindings presented on the (A-C) en face staining of ud-NHBE, (D-F) cross-section staining of ud-NHBE, (G-I) cross-section staining of wd-NHBE cells in vitro cultures and (J-L) the human bronchial biopsy in reddish brown.
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pone-0008713-g002: Lectin binding: MAL-I binds to Sia-α2-3Galβ1-4GlcNAc (Panel A, D, G and J), MAL-II binds to Sia-α2-3Galβ1-3GalNAc (Panel B, E, H and K) and SNA binds to Sia-α2-6-linkage (Panel C, F, I and L) to determine the Sias distribution on the ud- and wd-NHBE cells.MAL-I, MAL-II and SNA bindings presented on the (A-C) en face staining of ud-NHBE, (D-F) cross-section staining of ud-NHBE, (G-I) cross-section staining of wd-NHBE cells in vitro cultures and (J-L) the human bronchial biopsy in reddish brown.

Mentions: Lectin histochemistry on the primary cultures of human ud- and wd-NHBE cells using SNA (which recognizes the human influenza receptor with Sia-α2-6 linkages) and MAL-I and MAL-II (which recognizes the avian influenza receptor with linkages Sia-α2-3Galβ1-4GlcNAc and Sia-α2-3Galβ1-3GalNAc, respectively). We found that both MAL-I (Figure 2A, en face and Figure 2D, cross-section) and MAL-II (Figure 2B, en face and Figure 2E, cross- section) bound strongly and the SNA (Figure 2C. en face and Figure 2F, cross-section) bound weakly to the ud-NHBE cells indicating that both Sia-α2-3Galβ1-3GalNAc and Sia-α2-6 linkages were present but with different abundance. Thus ud-NHBE cells expressed more avian influenza Sia-α2-3 linkage receptors. On the other hand, the wd-NHBE cells which had acquired the physiological properties of normal human bronchi tissue exhibited strong binding with MAL-I (Figure 2G and 2J) and SNA (Figure 2I and 2L) while MAL-II binding was sparse, focal and confined to ciliated cells (Figure 2H and 2K). This indicated that Sia-α2-3Galβ1-4GlcNAc and Sia-α2-6 linkages were abundant while Sia-α2-3Galβ1-3GalNAc was comparatively scarce in wd-NHBE cells and human bronchi tissue.


Influenza H5N1 and H1N1 virus replication and innate immune responses in bronchial epithelial cells are influenced by the state of differentiation.

Chan RW, Yuen KM, Yu WC, Ho CC, Nicholls JM, Peiris JS, Chan MC - PLoS ONE (2010)

Lectin binding: MAL-I binds to Sia-α2-3Galβ1-4GlcNAc (Panel A, D, G and J), MAL-II binds to Sia-α2-3Galβ1-3GalNAc (Panel B, E, H and K) and SNA binds to Sia-α2-6-linkage (Panel C, F, I and L) to determine the Sias distribution on the ud- and wd-NHBE cells.MAL-I, MAL-II and SNA bindings presented on the (A-C) en face staining of ud-NHBE, (D-F) cross-section staining of ud-NHBE, (G-I) cross-section staining of wd-NHBE cells in vitro cultures and (J-L) the human bronchial biopsy in reddish brown.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2806912&req=5

pone-0008713-g002: Lectin binding: MAL-I binds to Sia-α2-3Galβ1-4GlcNAc (Panel A, D, G and J), MAL-II binds to Sia-α2-3Galβ1-3GalNAc (Panel B, E, H and K) and SNA binds to Sia-α2-6-linkage (Panel C, F, I and L) to determine the Sias distribution on the ud- and wd-NHBE cells.MAL-I, MAL-II and SNA bindings presented on the (A-C) en face staining of ud-NHBE, (D-F) cross-section staining of ud-NHBE, (G-I) cross-section staining of wd-NHBE cells in vitro cultures and (J-L) the human bronchial biopsy in reddish brown.
Mentions: Lectin histochemistry on the primary cultures of human ud- and wd-NHBE cells using SNA (which recognizes the human influenza receptor with Sia-α2-6 linkages) and MAL-I and MAL-II (which recognizes the avian influenza receptor with linkages Sia-α2-3Galβ1-4GlcNAc and Sia-α2-3Galβ1-3GalNAc, respectively). We found that both MAL-I (Figure 2A, en face and Figure 2D, cross-section) and MAL-II (Figure 2B, en face and Figure 2E, cross- section) bound strongly and the SNA (Figure 2C. en face and Figure 2F, cross-section) bound weakly to the ud-NHBE cells indicating that both Sia-α2-3Galβ1-3GalNAc and Sia-α2-6 linkages were present but with different abundance. Thus ud-NHBE cells expressed more avian influenza Sia-α2-3 linkage receptors. On the other hand, the wd-NHBE cells which had acquired the physiological properties of normal human bronchi tissue exhibited strong binding with MAL-I (Figure 2G and 2J) and SNA (Figure 2I and 2L) while MAL-II binding was sparse, focal and confined to ciliated cells (Figure 2H and 2K). This indicated that Sia-α2-3Galβ1-4GlcNAc and Sia-α2-6 linkages were abundant while Sia-α2-3Galβ1-3GalNAc was comparatively scarce in wd-NHBE cells and human bronchi tissue.

Bottom Line: We found that the in vitro culture of the well differentiated NHBE cells acquired the physiological properties of normal human bronchi tissue which express high level of alpha2-6-linked sialic acid receptors and human airway trypsin-like (HAT) protease, in contrast to the low expression in the non-differentiated NHBE cells.The reduced virus replication efficiency partially accounts for the lower interferon-beta responses in influenza H5N1 virus infected well differentiated NHBE cells.Since influenza infection in the bronchial epithelium will lead to tissue damage and associate with the epithelium regeneration, the data generated from the undifferentiated NHBE cultures may also be relevant to disease pathogenesis.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Queen Mary Hospital, Pokfulam, Hong Kong SAR, People's Republic of China.

ABSTRACT
Influenza H5N1 virus continues to be enzootic in poultry and transmits zoonotically to humans. Although a swine-origin H1N1 virus has emerged to become pandemic, its virulence for humans remains modest in comparison to that seen in zoonotic H5N1 disease. As human respiratory epithelium is the primary target cells for influenza viruses, elucidating the viral tropism and host innate immune responses of influenza H5N1 virus in human bronchial epithelium may help to understand the pathogenesis. Here we established primary culture of undifferentiated and well differentiated normal human bronchial epithelial (NHBE) cells and infected with highly pathogenic influenza H5N1 virus (A/Vietnam/3046/2004) and a seasonal influenza H1N1 virus (A/Hong Kong/54/1998), the viral replication kinetics and cytokine and chemokine responses were compared by qPCR and ELISA. We found that the in vitro culture of the well differentiated NHBE cells acquired the physiological properties of normal human bronchi tissue which express high level of alpha2-6-linked sialic acid receptors and human airway trypsin-like (HAT) protease, in contrast to the low expression in the non-differentiated NHBE cells. When compared to H1N1 virus, the H5N1 virus replicated more efficiently and induced a stronger type I interferon response in the undifferentiated NHBE cells. In contrast, in well differentiated cultures, H5N1 virus replication was less efficient and elicited a lower interferon-beta response in comparison with H1N1 virus. Our data suggest that the differentiation of bronchial epithelial cells has a major influence in cells' permissiveness to human H1N1 and avian H5N1 viruses and the host innate immune responses. The reduced virus replication efficiency partially accounts for the lower interferon-beta responses in influenza H5N1 virus infected well differentiated NHBE cells. Since influenza infection in the bronchial epithelium will lead to tissue damage and associate with the epithelium regeneration, the data generated from the undifferentiated NHBE cultures may also be relevant to disease pathogenesis.

Show MeSH
Related in: MedlinePlus