Overexpression of hTERT extends replicative capacity of human nucleus pulposus cells, and protects against serum starvation-induced apoptosis and cell cycle arrest.
The nucleus pulposus (NP) cells are chondrocyte-like cells that are required for the resistance of compressive loads through the synthesis of collagen fibrils and proteoglycan aggrecans, and the generation of a hydrostatic swelling pressure, and thus play an important role in the intervertebral disc. Here, we report the production and characterization of an immortalized human NP cell line from normal NP cells using stable transfection of recombinant human telomerase reverse transcriptase (hTERT) gene. The hTERT-transfected NP cells exhibited morphological characteristics typical of native cells. When compared with the first generation of normal NP cells, the hTERT-transfected NP cells grew faster and had an increased level of IGF-1 and TGF-β gene expression. They were successfully passaged over 20 generations without significant change in the levels of type II collagen and proteoglycan aggrecan expression. In addition, they showed resistance to serum starvation-induced apoptosis, G1 cell cycle arrest, and gene expression of p53, CCNE1, Fas, and Caspase 3. Moreover, histology revealed that no tumorigenicity of NP cells over expressing hTERT was observed after they were implanted in nude mice. Taken together, an immortalized human NP cell line was established, which has an extended lifespan, retains phenotypic features similar to primary parent NP cells, and should provide a suitable model for studying the biology of NP cells.
Guangzhou Institute of Traumatic Surgery, the Fourth Affiliated Hospital of Medical College, Jinan University, Guangzhou 510220, China. liangweiguo@tom.com
Weiguo
W
Liang
Dongping
D
Ye
Libing
L
Dai
Yan
Y
Shen
Jiake
J
Xu
D000818:Animals; D017209:Apoptosis; D053148:Caspase 3 / Q000096:biosynthesis; D018929:Cell Culture Techniques; D059447:Cell Cycle Checkpoints; D002460:Cell Line; D049109:Cell Proliferation; D019902:Chondrocytes; D003094:Collagen / Q000096:biosynthesis; D019927:Cyclin E / Q000096:biosynthesis; D006801:Humans; D006874:Hydrostatic Pressure; D007334:Insulin-Like Growth Factor I / Q000096:biosynthesis; D007403:Intervertebral Disc / Q000166:cytology* / Q000378:metabolism / Q000502:physiology; D051379:Mice; D008819:Mice, Nude; D015513:Oncogene Proteins / Q000096:biosynthesis; D011509:Proteoglycans / Q000096:biosynthesis; D019098:Telomerase / Q000096:biosynthesis / Q000235:genetics*; D014162:Transfection; D016212:Transforming Growth Factor beta / Q000096:biosynthesis; D016159:Tumor Suppressor Protein p53 / Q000096:biosynthesis; D019014:fas Receptor / Q000096:biosynthesis
C475820:CCNE1 protein, human; D019927:Cyclin E; C491901:FAS protein, human; D015513:Oncogene Proteins; D011509:Proteoglycans; D016212:Transforming Growth Factor beta; D016159:Tumor Suppressor Protein p53; D019014:fas Receptor; D007334:Insulin-Like Growth Factor I; D003094:Collagen; C509186:TERT protein, human; D019098:Telomerase; D053148:Caspase 3