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 α2-6-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.
Highlights
Pathogenic avian influenza (HPAI) H5N1 virus continues to be enzootic in poultry in parts of Asia and Africa and transmits zoonotically to humans
We evaluated the mRNA expression level of human airway trypsin-like (HAT) proteases in wd-normal human bronchial epithelial (NHBE) cell cultures and found a 700-fold increase in gene expression in Air liquid interface (ALI)-NHBE cell cultures differentiated for 21 days when compared to the ud-NHBE cells (Figure 1H)
The fact that a low-pathogenic H1N1 virus replicates efficiently in these cells without exogenous trypsin and that addition of trypsin did not further enhance viral replication suggests that the wd-NHBE cells make sufficient HAT to support low pathogenic influenza viral replication, as occurs in vivo
Summary
Pathogenic avian influenza (HPAI) H5N1 virus continues to be enzootic in poultry in parts of Asia and Africa and transmits zoonotically to humans. From 2003 to November 2009, influenza H5N1 virus has caused 444 confirmed human cases and 262 of them were fatal. Human H5N1 cases were found in 15 countries; the three most affected countries being Vietnam, China and Indonesia, where the fatality rates ranged from 42– 82% [1]. A swine origin influenza H1N1 virus (H1N1pdm) has recently emerged to become pandemic, its virulence for humans so far remains modest in comparison with that seen in zoonotic H5N1 disease [2]. Elucidating the pathogenesis underlying the unusual virulence of H5N1 virus may help understand the pathogenesis of acute respiratory distress syndrome in severe viral pneumonia, including that seen occasionally in pandemic H1N1 [3]
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