title: An ex vivo model of coxsackievirus infection using multilayered human conjunctival epithelial cells.
abstract: BACKGROUND
  The purpose of this study was to establish an ex vivo model of coxsackievirus infection since there seems to be no suitable disease model currently.
  
  
  METHODS
  Human conjunctival epithelial cells (HCECs) were cultured for 2 weeks in a serum-free air-liquid interface system to produce a multilayered structure. The cells were infected with coxsackievirus A24 (CVA24). Histological changes were investigated by staining the cells with H&E and DAPI, and apoptosis was evaluated using the TUNEL technique. Virus replication was measured in HeLa cells infected with viral progeny from multilayered HCECs, after 1 and 3 days, using the TCID(50) method.
  
  
  RESULTS
  Cultured HCECs formed multiple layers. The cells showed characteristics of conjunctival epithelial cells and goblet cells, being immunohistochemically positive for CK19 and MUC5AC, respectively. CVA24 replicated readily in cultured multilayered HCECs. A mild cytopathic effect was noted 1 day after viral inoculation. Cell damage was extensive at 3 days. TUNEL imaging confirmed that the cytopathology was attributable to apoptosis. The TCID(50) data from HeLa cells indicated that the virus was actively replicating at both 1 and 3 days after inoculation.
  
  
  CONCLUSIONS
  This novel infection model may be useful in investigating the pathogenesis of acute hemorrhagic conjunctivitis and the effectiveness of antiviral treatments.
authors.affiliation: Department of Ophthalmology, University of Ulsan College of Medicine, Asan Medical Center, 388-1 Pungnap-2dong, Songpa-gu, Seoul 138-736, Korea.
authors.forename: Jooeun
authors.initials: J
authors.lastname: Lee
authors.forename: Eun Jung
authors.initials: EJ
authors.lastname: Jun
authors.forename: Jung Hae
authors.initials: JH
authors.lastname: Sunwoo
authors.forename: Eun Soon
authors.initials: ES
authors.lastname: Kim
authors.forename: Jae-hyung
authors.initials: JH
authors.lastname: Kim
authors.forename: Jae Yong
authors.initials: JY
authors.lastname: Kim
authors.forename: Myoung Joon
authors.initials: MJ
authors.lastname: Kim
authors.forename: Yoo Kyum
authors.initials: YK
authors.lastname: Kim
authors.forename: Heuiran
authors.initials: H
authors.lastname: Lee
authors.forename: Hungwon
authors.initials: H
authors.lastname: Tchah
mesh_terms: D000208:Acute Disease; D017209:Apoptosis; D018929:Cell Culture Techniques; D003228:Conjunctiva / Q000166:cytology*; D003236:Conjunctivitis, Viral / Q000473:pathology* / Q000821:virology; D003384:Coxsackievirus Infections / Q000473:pathology*; D030015:Enterovirus C, Human / Q000254:growth & development; D004847:Epithelial Cells / Q000166:cytology / Q000821:virology*; D006367:HeLa Cells; D006470:Hemorrhage / Q000473:pathology / Q000821:virology; D006801:Humans; D020287:In Situ Nick-End Labeling; D014779:Virus Replication