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M. Orea, H. Martinez–Osorio, C. García–Vazquez, M. Calonge, Y. Diebold; Telomerase Expression in the Conjunctival Cell Line IOBA–NHC . Invest. Ophthalmol. Vis. Sci. 2006;47(13):4947.
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© ARVO (1962-2015); The Authors (2016-present)
The spontaneously immortalized cell line IOBA–NHC, derived from human conjunctiva epithelium, is highly demanded for ocular surface research since its publication in 2003. This interest has prompted us to try to elucidate the mechanism by which this cell line became immortalized. Constitutive expression of telomerase prevents telomeres shortening and cell senescence, and it is associated with cellular immortality. Therefore, the purpose of this study was to determine whether telomerase activity is present in the IOBA–NHC cell line.
Telomerase activity was measured by a telomerase repeat amplification protocol (TRAP) using a TRAPeze enzyme–linked assay kit in normal human conjunctival biopsy samples (n=6), IOBA–NHC cells (passages 67 and 103), conjunctival epithelial cells obtained by deep conjunctival impression cytology (CIC) (n=10), and human conjunctival fibroblasts (HCF). Telomerase activity was calculated by comparing the ratio of telomerase products to an internal standard for each cell lysate (optimum lysate is equivalent to 1000 cells). A catalytic subunit telomerase reverse transcriptase (hTERT)–immortalized epithelial cell line from human cornea (HCEC) was used as positive control. Additionally, to determine whether a process associated with aging occurred, senescence was evaluated using a ß–galactosidase staining kit and HCF cells as positive control.
IOBA–NHC cells expressed increased telomerase activity whereas human conjunctiva lysates and HCF cells expressed low or undetectable levels of this activity. Telomerase was also detected in deep CIC cells, although in lower levels than in IOBA–NHC cells. ß–galactosidase activity was undetectable in the immortalized IOBA–NHC cell line.
These results suggest that an enhanced telomerase activity in IOBA–NHC cell line is associated with its spontaneous immortalization process, providing this cell line with an increased replicative potential. These results offer further support to its use as an in vitro system.
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