Analysis of telomere lengths in human corneal endothelial cells from donors of different ages

Catherine A. Egan, Isabelle Savre-Train, Jerry W. Shay, Steven E. Wilson, William M. Bourne

Research output: Contribution to journalArticle

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Abstract

PURPOSE. To investigate the telomere hypothesis of cellular aging as the mechanism for cell cycle arrest in normal human corneal endothelium. METHODS. The corneal endothelium and epithelium from 21 human corneas from 13 donors 5 weeks to 84 years of age were dissected and frozen at -70°C. Purified DNA, digested with the restriction enzyme, HinfI, was run on 0.7% agarose gels, probed with radiolabeled (AATCCC)4, and exposed to a phosphor screen. The length of the terminal restriction fragment (TRF) was determined by densitometry. RESULTS. The cells of the corneal endothelium had TRF lengths ranging from 11.0 to 14.0 kbp (mean, 12.2 ± 0.9). Corneal epithelial specimens showed TRF lengths that were always less than (mean, 10.4 ± 1.0; range 9.0-12.0) the corresponding endothelial TRF lengths. Human corneal endothelial cells, transformed with human papilloma-virus type 16 oncogenes E6 and E7, showed decreasing TRF lengths from 11 kbp at population doubling level (PDL) 15 to 9.5 kbp at PDL 73. Neither the endothelial and epithelial cells from human donors nor the transformed pre-immortalized human endothelial cells showed evidence of telomerase activity. CONCLUSIONS. Human corneal endothelial cells have long telomeres throughout life. Their limited replicative ability does not appear to result from critically short telomere lengths.

Original languageEnglish (US)
Pages (from-to)648-653
Number of pages6
JournalInvestigative Ophthalmology and Visual Science
Volume39
Issue number3
StatePublished - Mar 1998

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Telomere
Endothelial Cells
Corneal Endothelium
Papillomaviridae
Corneal Epithelium
Densitometry
Cell Aging
Telomerase
Cell Cycle Checkpoints
Oncogenes
Sepharose
Cornea
Population
Gels
Epithelial Cells
DNA
Enzymes

ASJC Scopus subject areas

  • Ophthalmology

Cite this

Analysis of telomere lengths in human corneal endothelial cells from donors of different ages. / Egan, Catherine A.; Savre-Train, Isabelle; Shay, Jerry W.; Wilson, Steven E.; Bourne, William M.

In: Investigative Ophthalmology and Visual Science, Vol. 39, No. 3, 03.1998, p. 648-653.

Research output: Contribution to journalArticle

Egan, Catherine A. ; Savre-Train, Isabelle ; Shay, Jerry W. ; Wilson, Steven E. ; Bourne, William M. / Analysis of telomere lengths in human corneal endothelial cells from donors of different ages. In: Investigative Ophthalmology and Visual Science. 1998 ; Vol. 39, No. 3. pp. 648-653.
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N2 - PURPOSE. To investigate the telomere hypothesis of cellular aging as the mechanism for cell cycle arrest in normal human corneal endothelium. METHODS. The corneal endothelium and epithelium from 21 human corneas from 13 donors 5 weeks to 84 years of age were dissected and frozen at -70°C. Purified DNA, digested with the restriction enzyme, HinfI, was run on 0.7% agarose gels, probed with radiolabeled (AATCCC)4, and exposed to a phosphor screen. The length of the terminal restriction fragment (TRF) was determined by densitometry. RESULTS. The cells of the corneal endothelium had TRF lengths ranging from 11.0 to 14.0 kbp (mean, 12.2 ± 0.9). Corneal epithelial specimens showed TRF lengths that were always less than (mean, 10.4 ± 1.0; range 9.0-12.0) the corresponding endothelial TRF lengths. Human corneal endothelial cells, transformed with human papilloma-virus type 16 oncogenes E6 and E7, showed decreasing TRF lengths from 11 kbp at population doubling level (PDL) 15 to 9.5 kbp at PDL 73. Neither the endothelial and epithelial cells from human donors nor the transformed pre-immortalized human endothelial cells showed evidence of telomerase activity. CONCLUSIONS. Human corneal endothelial cells have long telomeres throughout life. Their limited replicative ability does not appear to result from critically short telomere lengths.

AB - PURPOSE. To investigate the telomere hypothesis of cellular aging as the mechanism for cell cycle arrest in normal human corneal endothelium. METHODS. The corneal endothelium and epithelium from 21 human corneas from 13 donors 5 weeks to 84 years of age were dissected and frozen at -70°C. Purified DNA, digested with the restriction enzyme, HinfI, was run on 0.7% agarose gels, probed with radiolabeled (AATCCC)4, and exposed to a phosphor screen. The length of the terminal restriction fragment (TRF) was determined by densitometry. RESULTS. The cells of the corneal endothelium had TRF lengths ranging from 11.0 to 14.0 kbp (mean, 12.2 ± 0.9). Corneal epithelial specimens showed TRF lengths that were always less than (mean, 10.4 ± 1.0; range 9.0-12.0) the corresponding endothelial TRF lengths. Human corneal endothelial cells, transformed with human papilloma-virus type 16 oncogenes E6 and E7, showed decreasing TRF lengths from 11 kbp at population doubling level (PDL) 15 to 9.5 kbp at PDL 73. Neither the endothelial and epithelial cells from human donors nor the transformed pre-immortalized human endothelial cells showed evidence of telomerase activity. CONCLUSIONS. Human corneal endothelial cells have long telomeres throughout life. Their limited replicative ability does not appear to result from critically short telomere lengths.

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