Journal: Cells

Article Title: Efficient Generation of Pancreatic Progenitor Cells from Induced Pluripotent Stem Cells Derived from a Non-Invasive and Accessible Tissue Source—The Plucked Hair Follicle

doi: 10.3390/cells13121010

Figure Lengend Snippet: Cryopreserved hair follicle derived primary keratinocytes can be reprogrammed using a feeder-free system across multiple demographics. Expanded primary keratinocytes (5 × 10 5 cells) from 11 participants were taken for reprogramming at passages 2–3 ( A ). The cells were electroporated and reprogrammed using the Epi5 vector system and gradually placed into N2B27 media. At day 9, the media were switched to TeSR E7, after which the development of Embryonic Stem Cell (ESC)-like colonies was observed until day 21. The non-reprogrammed keratinocytes flattened and ceased to proliferate in culture. Scale bar = 100 µm and 200 µm for d33 ( B ). All 11 participants were able to generate at least one colony, which could be further expanded by day 21 ( C ). In total, 4 participant cell lines from the older (>60) and younger demographic (<40) were further expanded, and episome-free lines were generated. All the cell lines exhibited ESC morphology. Scale bar = ( D ). The reprogrammed participants’ ages were correlated to their reprogramming efficiency (# of colonies formed on day 21). There was no significant difference observed ( E ). There was also no significant difference between reprogramming efficiency and sex, with both sexes having similar colony-forming units. (ns: non-significant.) ( F ). There was a significant positive correlation when comparing reprogramming efficiency and % outgrowth success (# of follicles with outgrowth/10) ( G ).

Article Snippet: In total, 1 μL of Epi5 Reprogramming Vectors (Oct4, Sox2, Klf4, L-Myc, and Lin28) and 1 μL of Epi5 p53 and EBNA vectors (Life Technologies, Carlsbad, CA, USA) were added to each 100 μL solution.

Techniques: Derivative Assay, Plasmid Preparation, Generated

Journal: Cells

Article Title: Efficient Generation of Pancreatic Progenitor Cells from Induced Pluripotent Stem Cells Derived from a Non-Invasive and Accessible Tissue Source—The Plucked Hair Follicle

doi: 10.3390/cells13121010

Figure Lengend Snippet: Induced pluripotent stem cells derived from cryopreserved hair follicles maintain pluripotency in vitro and are genetically stable. The episome-free lines were evaluated for pluripotency marker expression using immunofluorescence and confocal imaging. All the cell lines were positive for OCT4 (Green = FITC488]), Nanog (Green = FITC488), and SOX2 (Red = CY5647), as represented by cell line N3. Scale Bars = 200 μm ( A ). The cells were then expanded at various passages and cell population doublings (CPD), and were analyzed using G-banding analysis. Routine G-banding analysis was carried out, and 15 metaphases per cell line were examined. All four cell lines had no major chromosomal abnormalities, and the lines exhibited their respective XY: XX chromosome patterning ( B ).

Article Snippet: In total, 1 μL of Epi5 Reprogramming Vectors (Oct4, Sox2, Klf4, L-Myc, and Lin28) and 1 μL of Epi5 p53 and EBNA vectors (Life Technologies, Carlsbad, CA, USA) were added to each 100 μL solution.

Techniques: Derivative Assay, In Vitro, Marker, Expressing, Immunofluorescence, Imaging