DAS181, a sialidase fusion protein, protects human airway epithelium against influenza virus infection: an in vitro pharmacodynamic analysis

AbstractInfluenza A virus (IAV) causes seasonal epidemics and periodic pandemics, resulting in significant morbidity and mortality in the human population. Tethered mucin 1 (MUC1) is highly expressed in airway epithelium, the primary site of IAV replication, and also by other cell types that influence IAV infection, including macrophages. MUC1 has the potential to influence infection dynamics through physical interactions and/or signaling activity, and recent work suggests MUC1 acts as a releasable decoy receptor and anti-inflammatory molecule during IAV infection. Still, the modulation of MUC1 and its impact during viral pathogenesis remains unclear. Thus, we sought to further investigate the interplay between MUC1 and IAV in an in vitro model of primary human airway epithelium (HAE). Our data indicate that a recombinant IAV hemagglutinin (H3) and H3N2 virus can bind endogenous HAE MUC1. We find that infection of HAE cultures with H1N1 or H3N2 IAV strains does not trigger enhanced MUC1 shedding, but instead stimulates an increase in cell-associated MUC1 protein. We observed a similar increase after stimulation with either type I or type III interferon (IFN); however, inhibition of IFN signaling during H1N1 infection only partially abrogated this increase, indicating multiple soluble factors contribute to MUC1 upregulation during the antiviral response. We expanded these findings and demonstrate that in addition to HAE, primary human monocyte-derived macrophages also upregulate MUC1 protein in response to both IFN treatment and conditioned media from IAV-infected HAE cultures. We then developed HAE genetically depleted for MUC1 to determine its impact on IAV pathogenesis, finding that MUC1 knock-out cultures exhibited enhanced viral growth compared to control cultures. Together, our data support a model whereby MUC1 antagonizes productive uptake of IAV in HAE. Infection then stimulates MUC1 expression on multiple cell types through IFN-dependent and -independent mechanisms that may further impact infection dynamics.Author SummaryThe mucosal surface of the respiratory epithelium is an important site of first contact for viral respiratory pathogens. Large and heavily glycosylated molecules known as tethered mucins extend from the cell surface and may physically restrict access to underlying cells. Recently, one of these tethered mucins, MUC1, has also been shown to influence cell signaling and inflammation. Still, despite its abundance in the airway and multifunctional capability, the role of MUC1 during influenza virus infection in the human respiratory tract remains unclear. Here, we demonstrate that influenza virus directly interacts with MUC1 in a physiologically-relevant model of human airway epithelium and find that MUC1 protein expression is elevated throughout the epithelium and in primary human monocyte-derived macrophages in response to important antiviral signals produced during infection. Using genetically-modified human airway cultures lacking MUC1, we then provide evidence of more efficient influenza virus infection in the absence of this mucin. Our data suggest that MUC1 not only physically restricts influenza virus uptake, but also represents a dynamic component of the host response that acts to further stem viral spread.

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Neuraminidase Is Important for the Initiation of Influenza Virus Infection in Human Airway Epithelium

Journal of Virology ◽

10.1128/jvi.78.22.12665-12667.2004 ◽

2004 ◽

Vol 78 (22) ◽

pp. 12665-12667 ◽

Cited By ~ 371

Author(s):

Mikhail N. Matrosovich ◽

Tatyana Y. Matrosovich ◽

Thomas Gray ◽

Noel A. Roberts ◽

Hans-Dieter Klenk

Keyword(s):

Influenza Virus ◽

Airway Epithelium ◽

Influenza Virus Infection ◽

Influenza Viruses ◽

Avian Influenza Viruses ◽

Oseltamivir Carboxylate ◽

Human Airway ◽

Human Airway Epithelium ◽

Prophylactic Use ◽

Influenza Virus Neuraminidase

ABSTRACT Influenza virus neuraminidase (NA) plays an essential role in release and spread of progeny virions, following the intracellular viral replication cycle. To test whether NA could also facilitate virus entry into cell, we infected cultures of human airway epithelium with human and avian influenza viruses in the presence of the NA inhibitor oseltamivir carboxylate. Twenty- to 500-fold less cells became infected in drug-treated versus nontreated cultures (P < 0.0001) 7 h after virus application, indicating that the drug suppressed the initiation of infection. These data demonstrate that viral NA plays a role early in infection, and they provide further rationale for the prophylactic use of NA inhibitors.

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Linear polysialoside outperforms dendritic analogs for inhibition of influenza virus infection in vitro and in vivo

Biomaterials ◽

10.1016/j.biomaterials.2017.05.028 ◽

2017 ◽

Vol 138 ◽

pp. 22-34 ◽

Cited By ~ 43

Author(s):

Sumati Bhatia ◽

Daniel Lauster ◽

Markus Bardua ◽

Kai Ludwig ◽

Stefano Angioletti-Uberti ◽

...

Keyword(s):

Influenza Virus ◽

Virus Infection ◽

Influenza Virus Infection

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An approach to testing undiluted e-cigarette aerosol in vitro using 3D reconstituted human airway epithelium

Toxicology in Vitro ◽

10.1016/j.tiv.2018.01.010 ◽

2019 ◽

Vol 54 ◽

pp. 391-401 ◽

Cited By ~ 15

Author(s):

E. Bishop ◽

L. Haswell ◽

J. Adamson ◽

S. Costigan ◽

D. Thorne ◽

...

Keyword(s):

Airway Epithelium ◽

Human Airway ◽

Human Airway Epithelium

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Incorporation of biodegradable nanoparticles into human airway epithelium cells—in vitro study of the suitability as a vehicle for drug or gene delivery in pulmonary diseases

Biochemical and Biophysical Research Communications ◽

10.1016/j.bbrc.2004.04.067 ◽

2004 ◽

Vol 318 (2) ◽

pp. 562-570 ◽

Cited By ~ 80

Author(s):

M Brzoska ◽

K Langer ◽

C Coester ◽

S Loitsch ◽

T.O.F Wagner ◽

...

Keyword(s):

Gene Delivery ◽

Airway Epithelium ◽

In Vitro Study ◽

Pulmonary Diseases ◽

Biodegradable Nanoparticles ◽

Human Airway ◽

Human Airway Epithelium ◽

Or Gene ◽

Epithelium Cells

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Geniposide demonstrates anti-inflammatory and antiviral activity against pandemic A/Jiangsu/1/2009 (H1N1) influenza virus infection in vitro and in vivo

Antiviral Therapy ◽

10.3851/imp3152 ◽

2017 ◽

Vol 22 (7) ◽

pp. 599-611 ◽

Cited By ~ 15

Author(s):

Zhang Yunshi ◽

Yao Jing ◽

Qi Xian ◽

Liu Xing ◽

Lu Xieqin ◽

...

Keyword(s):

Influenza Virus ◽

Virus Infection ◽

Antiviral Activity ◽

Influenza Virus Infection ◽

H1n1 Influenza ◽

H1n1 Influenza Virus ◽

2009 H1n1 ◽

Anti Inflammatory

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Tropism and Infectivity of a Seasonal A(H1N1) and a Highly Pathogenic Avian A(H5N1) Influenza Virus in Primary Differentiated Ferret Nasal Epithelial Cell Cultures

Journal of Virology ◽

10.1128/jvi.00080-19 ◽

2019 ◽

Vol 93 (10) ◽

Cited By ~ 6

Author(s):

Hui Zeng ◽

Cynthia S. Goldsmith ◽

Amrita Kumar ◽

Jessica A. Belser ◽

Xiangjie Sun ◽

...

Keyword(s):

Influenza Virus ◽

Virus Infection ◽

Avian Influenza ◽

H5n1 Virus ◽

H1n1 Virus ◽

Influenza Virus Infection ◽

Influenza Viruses ◽

Host Interactions ◽

Culture Model

ABSTRACTFerrets represent an invaluable animal model to study influenza virus pathogenesis and transmission. To further characterize this model, we developed a differentiated primary ferret nasal epithelial cell (FNEC) culture model for investigation of influenza A virus infection and virus-host interactions. This well-differentiated culture consists of various cell types, a mucociliary clearance system, and tight junctions, representing the nasal ciliated pseudostratified respiratory epithelium. Both α2,6-linked and α2,3-linked sialic acid (SA) receptors, which preferentially bind the hemagglutinin (HA) of human and avian influenza viruses, respectively, were detected on the apical surface of the culture with different cellular tropisms. In accordance with the distribution of SA receptors, we observed that a pre-2009 seasonal A(H1N1) virus infected both ciliated and nonciliated cells, whereas a highly pathogenic avian influenza (HPAI) A(H5N1) virus primarily infected nonciliated cells. Transmission electron microscopy revealed that virions were released from or associated with the apical membranes of ciliated, nonciliated, and mucin-secretory goblet cells. Upon infection, the HPAI A(H5N1) virus replicated to titers higher than those of the human A(H1N1) virus at 37°C; however, replication of the A(H5N1) virus was significantly attenuated at 33°C. Furthermore, we found that infection with the A(H5N1) virus induced higher expression levels of immune mediator genes and resulted in more cell damage/loss than with the human A(H1N1) virus. This primary differentiated FNEC culture model, recapitulating the structure of the nasal epithelium, provides a useful model to bridgein vivoandin vitrostudies of cellular tropism, infectivity, and pathogenesis of influenza viruses during the initial stages of infection.IMPORTANCEAlthough ferrets serve as an important model of influenza virus infection, much remains unknown about virus-host interactions in this species at the cellular level. The development of differentiated primary cultures of ferret nasal epithelial cells is an important step toward understanding cellular tropism and the mechanisms of influenza virus infection and replication in the airway milieu of this model. Using lectin staining and microscopy techniques, we characterized the sialic acid receptor distribution and the cellular composition of the culture model. We then evaluated the replication of and immune response to human and avian influenza viruses at relevant physiological temperatures. Our findings offer significant insight into this first line of defense against influenza virus infection and provide a model for the evaluation of emerging influenza viruses in a well-controlledin vitroenvironmental setting.

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Dendritic Cells Targeting Lactobacillus plantarum Strain NC8 with a Surface-Displayed Single-Chain Variable Fragment of CD11c Induce an Antigen-Specific Protective Cellular Immune Response

Infection and Immunity ◽

10.1128/iai.00759-19 ◽

2019 ◽

Vol 88 (2) ◽

Author(s):

Jing Liu ◽

Guilian Yang ◽

Haibin Huang ◽

Chunwei Shi ◽

Xing Gao ◽

...

Keyword(s):

T Cells ◽

Dendritic Cells ◽

Influenza Virus ◽

Virus Infection ◽

Lactobacillus Plantarum ◽

Influenza Virus Infection ◽

Cell Culture Supernatant ◽

Single Chain ◽

Ifn Γ

ABSTRACT Influenza A virus (H1N1) is an acute, highly contagious respiratory virus. The use of lactic acid bacteria (LAB) to deliver mucosal vaccines against influenza virus infection is a research hot spot. In this study, two recombinant Lactobacillus plantarum strains expressing hemagglutinin (HA) alone or coexpressing aCD11c-HA to target HA protein to dendritic cells (DCs) by fusion to an anti-CD11c single-chain antibody (aCD11c) were constructed. The activation of bone marrow dendritic cells (BMDCs) by recombinant strains and the interaction of activated BMDCs and sorted CD4+ or CD8+ T cells were evaluated through flow cytometry in vitro, and cellular supernatants were assessed by using an enzyme-linked immunosorbent assay kit. The results demonstrated that, compared to the HA strain, the aCD11c-HA strain significantly increased the activation of BMDCs and increased the production of CD4+ gamma interferon-positive (IFN-γ+) T cells, CD8+ IFN-γ+ T cells, and IFN-γ in the cell culture supernatant in vitro. Consistent with these results, the aCD11c-HA strain clearly increased the activation and maturation of DCs, the HA-specific responses of CD4+ IFN-γ+ T cells, CD8+ IFN-γ+ T cells, and CD8+ CD107a+ T cells, and the proliferation of T cells in the spleen, finally increasing the levels of specific antibodies and neutralizing antibodies in mice. In addition, the protection of immunized mice was observed after viral infection, as evidenced by improved weight loss, survival, and lung pathology. The adoptive transfer of CD8+ T cells from the aCD11c-HA mice to NOD/Lt-SCID mice resulted in a certain level of protection after influenza virus infection, highlighting the efficacy of the aCD11c targeting strategy.

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