Influenza A virus infection of human middle ear cells in vitro

C. A. Buchman, Nevis L. Fregien

Research output: Contribution to journalArticle

10 Citations (Scopus)

Abstract

Hypothesis: Human-derived normal middle ear mucosal cells can be harvested and cultured and will support influenza A virus (INF A) infection. Study Design: Protocols for the collection and in vitro culture of middle ear mucosal cells were developed and used to investigate the effects of INF A infection as it relates to the pathogenesis of otitis media. Materials and Methods: Middle ear mucosa was harvested during surgeries that opened the normal middle ear. Middle ear mucosal cells were plated and grown in collagen-coated dishes. Cells were characterized before and after INF A exposure using phase-contrast and immunofluorescence microscopy as well as reverse transcriptase-polymerase chain reaction (RT-PCR) for cytokeratin 18 gene expression and INF A. Results: Primary cultures of human middle ear epithelial cells were established. Prolonged growth of middle ear cells yielded a second cell type that failed to stain for cytokeratin on immunofluorescence but continued to produce positive RT-PCR results on cytokeratin 18 analysis. After INF A exposure, cytological changes and immunofluorescence staining showed cellular infection. RT-PCR analysis using INF A-specific primers showed positive results for up to 72 hours after viral exposure. Conclusions: Primary cultures of human middle ear mucosal cells have been established. Two distinctly different cell culture systems have been developed: 1) middle ear epithelial cells and 2) either dedifferentiated epithelial cells or fibroblasts. Exposure of both cell types to INF A demonstrates that each can support cellular infection and viral replication. These models should be useful for studies of the pathogenesis of virus-mediated otitis media.

Original languageEnglish
Pages (from-to)1739-1744
Number of pages6
JournalLaryngoscope
Volume110
Issue number10 I
StatePublished - Jan 1 2000

Fingerprint

Influenza A virus
Middle Ear
Virus Diseases
Reverse Transcriptase Polymerase Chain Reaction
Keratin-18
Epithelial Cells
Otitis Media
Fluorescent Antibody Technique
In Vitro Techniques
Phase-Contrast Microscopy
Keratins
Fluorescence Microscopy
Mucous Membrane
Coloring Agents
Collagen
Cell Culture Techniques
Fibroblasts
Staining and Labeling
Viruses
Gene Expression

Keywords

  • Cell culture
  • Influenza virus
  • Middle ear epithelium
  • Otitis media

ASJC Scopus subject areas

  • Otorhinolaryngology

Cite this

Influenza A virus infection of human middle ear cells in vitro. / Buchman, C. A.; Fregien, Nevis L.

In: Laryngoscope, Vol. 110, No. 10 I, 01.01.2000, p. 1739-1744.

Research output: Contribution to journalArticle

Buchman, C. A. ; Fregien, Nevis L. / Influenza A virus infection of human middle ear cells in vitro. In: Laryngoscope. 2000 ; Vol. 110, No. 10 I. pp. 1739-1744.
@article{a0f4db74deb64f90a8668d995e000048,
title = "Influenza A virus infection of human middle ear cells in vitro",
abstract = "Hypothesis: Human-derived normal middle ear mucosal cells can be harvested and cultured and will support influenza A virus (INF A) infection. Study Design: Protocols for the collection and in vitro culture of middle ear mucosal cells were developed and used to investigate the effects of INF A infection as it relates to the pathogenesis of otitis media. Materials and Methods: Middle ear mucosa was harvested during surgeries that opened the normal middle ear. Middle ear mucosal cells were plated and grown in collagen-coated dishes. Cells were characterized before and after INF A exposure using phase-contrast and immunofluorescence microscopy as well as reverse transcriptase-polymerase chain reaction (RT-PCR) for cytokeratin 18 gene expression and INF A. Results: Primary cultures of human middle ear epithelial cells were established. Prolonged growth of middle ear cells yielded a second cell type that failed to stain for cytokeratin on immunofluorescence but continued to produce positive RT-PCR results on cytokeratin 18 analysis. After INF A exposure, cytological changes and immunofluorescence staining showed cellular infection. RT-PCR analysis using INF A-specific primers showed positive results for up to 72 hours after viral exposure. Conclusions: Primary cultures of human middle ear mucosal cells have been established. Two distinctly different cell culture systems have been developed: 1) middle ear epithelial cells and 2) either dedifferentiated epithelial cells or fibroblasts. Exposure of both cell types to INF A demonstrates that each can support cellular infection and viral replication. These models should be useful for studies of the pathogenesis of virus-mediated otitis media.",
keywords = "Cell culture, Influenza virus, Middle ear epithelium, Otitis media",
author = "Buchman, {C. A.} and Fregien, {Nevis L.}",
year = "2000",
month = "1",
day = "1",
language = "English",
volume = "110",
pages = "1739--1744",
journal = "Laryngoscope",
issn = "0023-852X",
publisher = "Wiley-Blackwell",
number = "10 I",

}

TY - JOUR

T1 - Influenza A virus infection of human middle ear cells in vitro

AU - Buchman, C. A.

AU - Fregien, Nevis L.

PY - 2000/1/1

Y1 - 2000/1/1

N2 - Hypothesis: Human-derived normal middle ear mucosal cells can be harvested and cultured and will support influenza A virus (INF A) infection. Study Design: Protocols for the collection and in vitro culture of middle ear mucosal cells were developed and used to investigate the effects of INF A infection as it relates to the pathogenesis of otitis media. Materials and Methods: Middle ear mucosa was harvested during surgeries that opened the normal middle ear. Middle ear mucosal cells were plated and grown in collagen-coated dishes. Cells were characterized before and after INF A exposure using phase-contrast and immunofluorescence microscopy as well as reverse transcriptase-polymerase chain reaction (RT-PCR) for cytokeratin 18 gene expression and INF A. Results: Primary cultures of human middle ear epithelial cells were established. Prolonged growth of middle ear cells yielded a second cell type that failed to stain for cytokeratin on immunofluorescence but continued to produce positive RT-PCR results on cytokeratin 18 analysis. After INF A exposure, cytological changes and immunofluorescence staining showed cellular infection. RT-PCR analysis using INF A-specific primers showed positive results for up to 72 hours after viral exposure. Conclusions: Primary cultures of human middle ear mucosal cells have been established. Two distinctly different cell culture systems have been developed: 1) middle ear epithelial cells and 2) either dedifferentiated epithelial cells or fibroblasts. Exposure of both cell types to INF A demonstrates that each can support cellular infection and viral replication. These models should be useful for studies of the pathogenesis of virus-mediated otitis media.

AB - Hypothesis: Human-derived normal middle ear mucosal cells can be harvested and cultured and will support influenza A virus (INF A) infection. Study Design: Protocols for the collection and in vitro culture of middle ear mucosal cells were developed and used to investigate the effects of INF A infection as it relates to the pathogenesis of otitis media. Materials and Methods: Middle ear mucosa was harvested during surgeries that opened the normal middle ear. Middle ear mucosal cells were plated and grown in collagen-coated dishes. Cells were characterized before and after INF A exposure using phase-contrast and immunofluorescence microscopy as well as reverse transcriptase-polymerase chain reaction (RT-PCR) for cytokeratin 18 gene expression and INF A. Results: Primary cultures of human middle ear epithelial cells were established. Prolonged growth of middle ear cells yielded a second cell type that failed to stain for cytokeratin on immunofluorescence but continued to produce positive RT-PCR results on cytokeratin 18 analysis. After INF A exposure, cytological changes and immunofluorescence staining showed cellular infection. RT-PCR analysis using INF A-specific primers showed positive results for up to 72 hours after viral exposure. Conclusions: Primary cultures of human middle ear mucosal cells have been established. Two distinctly different cell culture systems have been developed: 1) middle ear epithelial cells and 2) either dedifferentiated epithelial cells or fibroblasts. Exposure of both cell types to INF A demonstrates that each can support cellular infection and viral replication. These models should be useful for studies of the pathogenesis of virus-mediated otitis media.

KW - Cell culture

KW - Influenza virus

KW - Middle ear epithelium

KW - Otitis media

UR - http://www.scopus.com/inward/record.url?scp=0033780384&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0033780384&partnerID=8YFLogxK

M3 - Article

VL - 110

SP - 1739

EP - 1744

JO - Laryngoscope

JF - Laryngoscope

SN - 0023-852X

IS - 10 I

ER -