Telomerase Reverses Pluripotency and Blocks Spontaneous Differentiation

Objectives:

Cellular senescence and spontaneous differentiation are critical phenomena that impair long-term study of cells in vitro. The same problems are similarly encountered in the investigation of human mesenchymal stem cells (hMSCs), despite its high proliferation rate and multipotency comparing to those of differentiated cells derived from different tissues. The present study was designed to overcome these problems and also explored to drive hMSCs toward stemness.

Methods: and Results:

Ectopic expression of telomerase in a previously developed hMSC line was found to enhance pluripotency and block spontaneous differentiation of the cells. Surprisingly, the telomerase-transfected hMSCs (3A6) had a differentiation potential far beyond the normal hMSCs. They expressed trophoectoderm and germline specific markers in vitro, similar to those of embryonic stem cells upon perturbations with BMP4 and retinoic acid, respectively. Furthermore, the telomerase-transfected hMSCs (3A6) displayed higher osteogenic and neural differentiation efficiency than their parental cells while there was a decrease in DNA methylation level as proved by a global CpG island methylation profile analysis. Possible underlying mechanisms were assayed by DNA methylation and its regulation enzymes. Notably, the demethylated CpG islands were found to be highly associated with development and differentiation associated genes. Principal component analysis further pointed out the expression profile of the cells converged toward embryonic stem cells.

Conclusions:

These data demonstrate the reversion of pluripotency and blockage of spontaneous differentiation by ectopic expression of telomerase.

Funding

These studies were supported in part by grants from National Scientific Council (NSC-95-2314-B-075-047-MY3, NSC-95-2627-B-010-009- , NSC-96-2627-B-010-009-) and grants from National Yang-Ming University, Ministry of Education, Aim for the Top University Plan, and Veterans General Hospital-Taipei (V95E1-002 & V95E1-003).