Title:Characterization of Induced Pluripotent Stem Cells from Human Epidermal Melanocytes by Transduction with Two Combinations of Transcription Factors
Volume: 19
Issue: 6
关键词:
表皮，黑素细胞，多能，重编程，iPSCS，转录。
摘要:
Objective: In order to generate induced Pluripotent Stem Cells (iPSCs) more efficiently, it
is crucial to identify somatic cells that are easily accessible and possibly require fewer factors for conversion
into iPSCs.
Methods: Human epidermal melanocytes were transduced with lentiviral vectors carrying 3 transcription
factors (OCT-4, KLF-4 and c-MYC, 3F) or 4 transcription factors (OCT-4, KLF-4, c-MYC and
SOX-2, 4F). Once the clones had formed, assays related to stem cell pluripotency, including alkaline
phosphatase staining, DNA methylation levels, expression of stem cell markers and ultrastructure
analysis were carried out. The iPSCs obtained were then induced to differentiate into the cells representing
the three embryonic layers in vitro.
Results: Seven days after the transduction of epidermal melanocytes with 3F or 4F, clones were
formed that were positive for alkaline phosphatase staining. Fluorescent staining with antibodies
against OCT-4 and SOX-2 was strongly positive, and the cells showed a high nucleus-cytoplasm ratio
and active karyokinesis. No melanosomes were found in the cytoplasm by ultrastructural analysis.
There were obvious differences in DNA methylation levels between the cloned cells and their parental
cells. However, there was not a significant difference between 3F or 4F transfected clonal cells.
Meanwhile, the iPSCs successfully differentiated into the three germ layer cells in vitro.
Conclusion: Human epidermal melanocytes do not require ectopic SOX-2 expression for conversion
into iPSCs, and may serve as an alternative source for deriving patient-specific iPSCs with fewer genetic
elements.
