Characterization of ALDHhi/CD133+ Cells in the Human Fetal Pancreas.

Introduction: Understanding how endocrine progenitors commit to a β-cell lineage during human fetal development is necessary for the design of in vitro β-cell differentiation protocols. Previous research has determined that high aldehyde dehydrogenase (ALDH) activity is necessary for the development and survival of β-cells. The purpose of the current study is to assess the endocrine lineage commitment of ALDH^hi/CD133⁺ human fetal pancreatic cells and determine their potential for forming a pool of endocrine cell precursors. Methods: Human fetal pancreata (18-22 weeks) were dissociated to a single cell suspension, labeled for ALDH and CD133, and sorted into double positive (ALDH^hi/CD133⁺) and double negative (ALDH^lo/CD133^-) populations using fluorescence-activated cell sorting (FACS). Sorted populations were then cultured for an extended period to assess their phenotypic stability and then characterized using immunofluorescence staining and qRT-PCR to quantify various transcription factors and cellular markers indicative of endocrine cell commitment. Results: Fluorescence-activated cell sorting yielded a larger population of double positive cells than double negative cells. Expression of ALDH and endocrine-specific transcription factors was not detected in either cell population after extended cell culture. Both of the expanded populations were positive for vimentin, Ki-67, CK19, SOX9 and β-catenin, with no significant differences in expression levels. Conclusions: Our preliminary results indicate that neither population is capable of maintaining expression of ALDH or endocrine-specific markers after extended culture. Further studies should assess each population's capacity for forming islet-like cell clusters to further characterize any underlying differences in endocrine lineage commitment. [ABSTRACT FROM AUTHOR]