Isolation of Human Oral Keratinocyte Progenitor/Stem Cells

Abstract

Progenitor/stem cell populations of epithelium are known to reside in the small-sized cell population. Our objective was to physically isolate and characterize an oral keratinocyte-enriched population of small-sized progenitor/stem cells. Primary human oral mucosal keratinocytes cultured in a chemically defined serum-free culture system, devoid of animal-derived feeder cells, were sorted by relative cell size and characterized by immunolabeling for beta1 integrin, nuclear transcription factor, peroxisome proliferator-activated receptor-gamma, and cell-cycle analysis. Sorted cells were distinguished as progenitor/stem cells by functional assays and their ability to regenerate an oral mucosal graft. Small-sized cells demonstrated the lowest expression of peroxisome proliferator-activated receptor-gamma, the highest colony-forming efficiency, a longer long-term proliferative potential, an enriched quiescent cell population, and the ability to regenerate an oral mucosal graft, implying that the small-sized cultured oral keratinocytes contained an enriched population of progenitor/stem cells.

Expression Profile of the Stem Cell Markers in Human Hertwig’s Epithelial Root Sheath/Epithelial Rests of Malassez Cells

“…These hierarchical models suggested the cellular and molecular mechanisms regulating the tissue regeneration and repair: the process of symmetrical or asymmetrical cell division leads to the hierarchy of ASCs. Additionally, it is known that epithelial stem cells are small in size and have little cytoplasmic complexity, and that the size and complexity of these cells increases in a differentiated state (De Paiva et al, 2006;Izumi et al, 2007;Romano et al, 2003). In this study, during the subculturing process, the size and complexity of the HERS/ERM cells were clearly increased, and the expression of stemnessrelated genes showed diverse patterns: the number of cells residing in increased cell size and complexity were highly increased but they showed the dramatically decreased expression of the EpSC and EmSC markers (data not shown).…”

Proteomics Characterization of Primary Human Oral Epithelial Cells Using a Novel Culture Technique for Use in Tissue Regeneration

“…Contamination of those large-sized cells cannot be ignored when we analyze the undifferentiated profiles. GACS was successfully used to enrich the population of small-sized undifferentiated cells [4]. In our results, proteins having higher expression in ePUKs small cells are involved in cellular movement, indicating they might have enhanced ability for wound healing and tissue regeneration.…”

“…ePUKs are floating, non-attached cells produced by large sized colonies in monolayer culture which are fed daily with 2-fold the usual amount of culture medium and the cell suspension containing ePUKs is poured into a new flask to form another monolayer without the use of enzymes to split the cultures [2]. Small-sized cultured keratinocytes are reported as a progenitor/stem-cell-enriched population since they have a high ability of colony formation and long term proliferative capacity [4,5]. However, when investigating the ePUKs undifferentiated profile, it is important to eliminate the contamination of the large sized dead or aged cells which are also floating in the media over laying a confluent monolayer, that lose their adherent ability.…”

Human exfoliated deciduous teeth and oral mucosa: promising applications in tissue regeneration

“…Izumi et al successfully isolated and characterized an oral keratinocyte-enriched population of progenitor-stem cells: able to in vitro form colonies, they ex vivo regenerate an oral mucosa equivalent 24 .…”

“…The demonstrated great potential of SHEDs and OMSCs 16,17,[24][25][26] and the relatively easy collecting and harvesting method, led us to suggest the opportunity to use the deciduous teeth, naturally lost in pediatric age, and/or the oral mucosa, for TR purposes of patients.…”