Abstract:
PURPOSE:To characterize a new nontransfected, spontaneously immortalized epithelial cell line from normal human conjunctiva (IOBA-NHC), both morphologically and functionally, to determine whether the differentiated phenotype of conjunctival epithelial cells is preserved. METHODS:Outgrowing cells from explanted conjunctival tissue were successively passaged and preliminarily characterized at passage 3 to assess epithelial origin. The cells were further characterized at passages 15 to 20, 40, 60, and 100 by analyzing (1) proliferation and in vitro behavior (viability, plating efficiency, colony forming efficiency and colony size, and Ki-67 protein expression), (2) karyotype and G-banding, (3) epithelial marker expression (cytokeratins, desmoplakins, EGF receptor), (4) absence of contaminating cell types, (5) expression of conjunctival differentiation markers (mucin gene expression), and (6) functional capability in response to proinflammatory stimuli. IOBA-NHC cells were analyzed by light and electron (transmission and scanning) microscopy, immunohistochemistry, electrophoresis and Western blot analysis, flow cytometry, and reverse transcription-polymerase chain reaction (RT-PCR). RESULTS:IOBA-NHC cells showed high proliferative ability in vitro and typical epithelial morphology. Cytokeratins and GalNAc, GluNAc, mannose, and sialic acid residues were immunodetected in these cells. No contaminating cell types were found. MUC1, -2, and -4, but not -5AC or -7 mucin genes were expressed in every cell passage tested. Exposure of cells to inflammatory mediators (IFNgamma and/or TNFalpha) resulted in increased expression of intercellular adhesion molecule (ICAM)-1 and HLA-DR. CONCLUSIONS:Morphologic and functional characterization of the nontransfected, spontaneously immortalized IOBA-NHC cell line shows that this new cell line may be a useful experimental tool in the field of ocular surface cell biology.
journal_name
Invest Ophthalmol Vis Scijournal_title
Investigative ophthalmology & visual scienceauthors
Diebold Y,Calonge M,Enríquez de Salamanca A,Callejo S,Corrales RM,Sáez V,Siemasko KF,Stern MEdoi
10.1167/iovs.03-0560keywords:
subject
Has Abstractpub_date
2003-10-01 00:00:00pages
4263-74issue
10eissn
0146-0404issn
1552-5783journal_volume
44pub_type
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