Identification of telomerase activity of human adipose-derived mesenchymal stem cells from various donors and at varied passages / 中国组织工程研究

ABSTRACT

BACKGROUND:It is not certain that whether human adipose-derived mesanchymal stem cells express telomerase reverse transcriptase (TERT).OBJECTIVE:To investigate whether human adipose-derived mesenchymal stem cells express telomerase reverse transcriptase or not.DESIGN,TIME AND SETTING:The cytology in vitro experiments were performed at TEDA Life and Technology Research Center,Institute of Hematology and Blood Diseases Hospital,Chinese Academy of Medical Sciences and Peking Union Medical College in August 2009.MATERIALSAdult adipose tissue was obtained from 5 healthy donors undergoing liposuction at the Tianjin Yili Medical Cosmetology Plastics Outpatient.Hela cells were supplied by the Cell Center of Basic Medical Sciences,Institute of Basic Medical Sciences,Chinese Academy of Medical Sciences.METHODS:Human adipose-derived mesenchymal stem cells were isolated from 20 mL human edipose tissue of each healthy donor by collagenase digestion.Cells were digested by trypsin when 80% confluency.Hela cells served as controls,and treated with 1640 medium containing 10% fetal bovine serum,and then digested using trypsin when 80% confluency.MAIN OUTCOME MEASURES:Morphology,phenotype and differentiation of human adipose-derived mesenchymal stem cells.TERT of human adipose-derived mesenchymal stem calls were detected by reverse transcription-polymerase chain reaction (RT-PCR).Telomerase activity was measured by telomeric repeat amplification protocol assay ELISA (TRAP-ELISA).RESULTS:Human adipose-derived mesenchymal stem cells adhered to the flask,presented spindle shape,and whirlpool-shape when cell density was large.Human adipose-derived mesenchymal stem cells were labeled positively for CD73,CD90 and CD105,but negatively for CD19,CD34,CD11b,CD45,and HLA-DR.Following adipogenic induction,oil red O staining showed significant red oil drop.Following ostesgenic induction,Von Kossa staining showed black particles,which indicated calcium deposition.Human adipose-derived mesenchymal stem cells at P1,P4 and P7 did not express telomerase reverse transcriptase gane.Human adipose-derived mesenchymal stem cells from 5 different donors did not expression telomerase reverse transcriptase gane.Hela cells had telomerase activation,but human adipose-derived mesanchymal stem cells did not express telomerase activation.CONCLUSION:Human adipose-derived mesanchymal stem cells do not express mRNA of hTERT,nor the activity of telomerase reverse transcriptase.