TY - JOUR
T1 - hTERT extends proliferative lifespan and prevents oxidative stress-induced apoptosis in human lens epithelial cells
AU - Huang, Xiao Qin
AU - Wang, Juan
AU - Liu, Jin Ping
AU - Feng, Hao
AU - Liu, Wen Bin
AU - Yan, Qin
AU - Liu, Yan
AU - Sun, Shu Ming
AU - Deng, Mi
AU - Gong, Lili
AU - Liu, Yun
AU - Li, David Wan Cheng
PY - 2005
Y1 - 2005
N2 - PURPOSE. Telomerase is a specialized polymerase that catalyzes synthesis of telomeres in most eukaryotes. When introduced into somatic cells, it extends the proliferative lifespan and prevents replicative senescence. Whether it has similar functions in lens epithelial cells, especially in human lens epithelial cells (HLECs) remains to be determined. In this study, the human telomerase reverse transcriptase (hTERT) catalytic subunit was introduced into HLECs. A stable cell line expressing hTERT was established and the functions of hTERT were studied. METHODS. The telomeric repeat amplification protocol (TRAP) assay was used to analyze the telomerase activity. Western blot analysis was used to examine hTERT expression. Southern blot analysis was used to detect telomere length. HLECs isolated from intact lenses were cultured in DMEM and transfected with hTERT cDNA. The expression of the exogenous hTERT was examined with RT-PCR, Western blot analysis, and TRAP assay. The functions of hTERT were examined with various techniques. RESULTS. Among the human, bovine, and rabbit lenses examined, only the central epithelium from the 6-month rabbit lens displayed telomerase activity. In both transparent and cataractous human lenses, hTERT activity and expression were not detected. However, the template RNA was present in both types of human lenses. The telomeres in transparent lenses were approximately 1 kb longer than those in cataractous lenses. The primary cultures and later passages of HLECs also displayed no detectable telomerase activity. Introduction of hTERT cDNA into HLECs followed by G418 selection yielded a stable line of HLECs expressing hTERT. In this line, hTERT has supported normal growth after 48 population doublings (PDs) to date and also enhanced antiapoptotic activity against oxidative stress. CONCLUSIONS. Telomere lengths may be associated with cataractogenesis. hTERT introduced into HLECs prevents replicative senescence through telomere synthesis. Furthermore, hTERT displays functions beyond telomere synthesis in normal HLECs.
AB - PURPOSE. Telomerase is a specialized polymerase that catalyzes synthesis of telomeres in most eukaryotes. When introduced into somatic cells, it extends the proliferative lifespan and prevents replicative senescence. Whether it has similar functions in lens epithelial cells, especially in human lens epithelial cells (HLECs) remains to be determined. In this study, the human telomerase reverse transcriptase (hTERT) catalytic subunit was introduced into HLECs. A stable cell line expressing hTERT was established and the functions of hTERT were studied. METHODS. The telomeric repeat amplification protocol (TRAP) assay was used to analyze the telomerase activity. Western blot analysis was used to examine hTERT expression. Southern blot analysis was used to detect telomere length. HLECs isolated from intact lenses were cultured in DMEM and transfected with hTERT cDNA. The expression of the exogenous hTERT was examined with RT-PCR, Western blot analysis, and TRAP assay. The functions of hTERT were examined with various techniques. RESULTS. Among the human, bovine, and rabbit lenses examined, only the central epithelium from the 6-month rabbit lens displayed telomerase activity. In both transparent and cataractous human lenses, hTERT activity and expression were not detected. However, the template RNA was present in both types of human lenses. The telomeres in transparent lenses were approximately 1 kb longer than those in cataractous lenses. The primary cultures and later passages of HLECs also displayed no detectable telomerase activity. Introduction of hTERT cDNA into HLECs followed by G418 selection yielded a stable line of HLECs expressing hTERT. In this line, hTERT has supported normal growth after 48 population doublings (PDs) to date and also enhanced antiapoptotic activity against oxidative stress. CONCLUSIONS. Telomere lengths may be associated with cataractogenesis. hTERT introduced into HLECs prevents replicative senescence through telomere synthesis. Furthermore, hTERT displays functions beyond telomere synthesis in normal HLECs.
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U2 - 10.1167/iovs.05-0154
DO - 10.1167/iovs.05-0154
M3 - Article
C2 - 15980242
AN - SCOPUS:20744441769
SN - 0146-0404
VL - 46
SP - 2503
EP - 2513
JO - Investigative Ophthalmology and Visual Science
JF - Investigative Ophthalmology and Visual Science
IS - 7
ER -