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Trend of telomerase activity change during human iPSC self-renewal and differentiation revealed by a quartz crystal microbalance based assay.

Zhou Y, Zhou P, Xin Y, Wang J, Zhu Z, Hu J, Wei S, Ma H - Sci Rep (2014)

Bottom Line: Telomerase plays an important role in governing the life span of cells for its capacity to extend telomeres.The pluripotency of hiPSCs during differentiation could be estimated through monitoring telomerase activity and compared with the expression levels of markers of pluripotency gene via quantitative real time PCR.Regular assessment for factors associated with pluripotency or stemness was expensive and requires excessive sample consuming, thus TREAQ could be a promising alternative technology for routine monitoring of telomerase activity and estimate the pluripotency of stem cells.

View Article: PubMed Central - PubMed

Affiliation: Division of Nanobiomedicine, Suzhou Institute of Nano-Tech and Nano-Bionics, Chinese Academy of Sciences, Suzhou 215123, P. R. China.

ABSTRACT
Telomerase plays an important role in governing the life span of cells for its capacity to extend telomeres. As high activity of telomerase has been found in stem cells and cancer cells specifically, various methods have been developed for the evaluation of telomerase activity. To overcome the time-consuming procedures and complicated manipulations of existing methods, we developed a novel method named Telomeric Repeat Elongation Assay based on Quartz crystal microbalance (TREAQ) to monitor telomerase activity during the self-renewal and differentiation of human induced pluripotent stem cells (hiPSCs). TREAQ results indicated hiPSCs possess invariable telomerase activity for 11 passages on Matrigel and a steady decline of telomerase activity when differentiated for different periods, which is confirmed with existing golden standard method. The pluripotency of hiPSCs during differentiation could be estimated through monitoring telomerase activity and compared with the expression levels of markers of pluripotency gene via quantitative real time PCR. Regular assessment for factors associated with pluripotency or stemness was expensive and requires excessive sample consuming, thus TREAQ could be a promising alternative technology for routine monitoring of telomerase activity and estimate the pluripotency of stem cells.

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FACS analysis of hNF1-4C11 hiPSCs after 1 (A), 3 (B), 5 (C), 7 (D), 9 (E), 11(F) passages on Matrigel™ for Oct-4.The percentage of positive cells is listed in parentheses, which indicate the stable pluripotency of hiPSCs.
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f8: FACS analysis of hNF1-4C11 hiPSCs after 1 (A), 3 (B), 5 (C), 7 (D), 9 (E), 11(F) passages on Matrigel™ for Oct-4.The percentage of positive cells is listed in parentheses, which indicate the stable pluripotency of hiPSCs.

Mentions: Our quantitative RT-PCR data quantified the Oct4, Nanog, Sox2 and Rex-1 transcripts relative to the ACTB housekeeping gene in differentiated hiPSCs. As shown in Figure 7, Oct-4 and Sox2 expression significantly decreased during differentiation. On the other hand, flow cytometric results indicated invariable Oct4 expression during the self-renewal of hiPSCs (Fig. 8).


Trend of telomerase activity change during human iPSC self-renewal and differentiation revealed by a quartz crystal microbalance based assay.

Zhou Y, Zhou P, Xin Y, Wang J, Zhu Z, Hu J, Wei S, Ma H - Sci Rep (2014)

FACS analysis of hNF1-4C11 hiPSCs after 1 (A), 3 (B), 5 (C), 7 (D), 9 (E), 11(F) passages on Matrigel™ for Oct-4.The percentage of positive cells is listed in parentheses, which indicate the stable pluripotency of hiPSCs.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4225532&req=5

f8: FACS analysis of hNF1-4C11 hiPSCs after 1 (A), 3 (B), 5 (C), 7 (D), 9 (E), 11(F) passages on Matrigel™ for Oct-4.The percentage of positive cells is listed in parentheses, which indicate the stable pluripotency of hiPSCs.
Mentions: Our quantitative RT-PCR data quantified the Oct4, Nanog, Sox2 and Rex-1 transcripts relative to the ACTB housekeeping gene in differentiated hiPSCs. As shown in Figure 7, Oct-4 and Sox2 expression significantly decreased during differentiation. On the other hand, flow cytometric results indicated invariable Oct4 expression during the self-renewal of hiPSCs (Fig. 8).

Bottom Line: Telomerase plays an important role in governing the life span of cells for its capacity to extend telomeres.The pluripotency of hiPSCs during differentiation could be estimated through monitoring telomerase activity and compared with the expression levels of markers of pluripotency gene via quantitative real time PCR.Regular assessment for factors associated with pluripotency or stemness was expensive and requires excessive sample consuming, thus TREAQ could be a promising alternative technology for routine monitoring of telomerase activity and estimate the pluripotency of stem cells.

View Article: PubMed Central - PubMed

Affiliation: Division of Nanobiomedicine, Suzhou Institute of Nano-Tech and Nano-Bionics, Chinese Academy of Sciences, Suzhou 215123, P. R. China.

ABSTRACT
Telomerase plays an important role in governing the life span of cells for its capacity to extend telomeres. As high activity of telomerase has been found in stem cells and cancer cells specifically, various methods have been developed for the evaluation of telomerase activity. To overcome the time-consuming procedures and complicated manipulations of existing methods, we developed a novel method named Telomeric Repeat Elongation Assay based on Quartz crystal microbalance (TREAQ) to monitor telomerase activity during the self-renewal and differentiation of human induced pluripotent stem cells (hiPSCs). TREAQ results indicated hiPSCs possess invariable telomerase activity for 11 passages on Matrigel and a steady decline of telomerase activity when differentiated for different periods, which is confirmed with existing golden standard method. The pluripotency of hiPSCs during differentiation could be estimated through monitoring telomerase activity and compared with the expression levels of markers of pluripotency gene via quantitative real time PCR. Regular assessment for factors associated with pluripotency or stemness was expensive and requires excessive sample consuming, thus TREAQ could be a promising alternative technology for routine monitoring of telomerase activity and estimate the pluripotency of stem cells.

Show MeSH
Related in: MedlinePlus