Author:

Keywords:

Reprogramming, Endodermal progenitors

Abstract:

Large-scale application of islet transplantation is hampered by immunological challenges as well as by donor shortage. An alternative source for human cadaveric islets is generating insulin-producing β-cells from stem cells. A challenge in this area is to find adequate stem or progenitor cells and to identify the mechanisms by which functional insulin-producing beta cells can be produced from such stem/progenitor cells. The development of techniques to reprogram somatic cells to stem cells, specifically the description of induced pluripotent stem cells, has provided the basis for our hypothesis that, using defined transcription factors (TFs), it should be possible to reprogram human bone marrow (BM) Multipotent Adult Progenitors (MAPC) into endoderm progenitors. To test this hypothesis we selected 16 transcription factors, known to play a key role during (pancreatic) endoderm development combined with transcription factors expressed in pluripotent stem cells. Here we demonstrated that endodermal progenitors (termed iEndo) can be generated from human BM derived MAPC cells by transduction of 16TFs and culture in endoderm conditions. Following transduction and culture in endoderm conditions, hMAPC underwent mesenchyme to epithelium transition (MET), forming cluster of epithelioid cells that express endodermal marker genes as demonstrated by qRT-PCR and FACS. By selectively removing 1 or more TFs from the combination of 16 TFs, we have shown that at least 1 TFs are essential while the other 15 may not be essential, for induction of MET, or induction of endogenous endodermal marker expression. Based on these studies, we had transduced MAPCs with a cocktail of 7 or 6 TFs to induce iEndo cells that are not yet committed to more mature endodermal cells. We will then differentiate iEndo cells to pancreatic beta cells, those function will be tested in vitro and in vivo. A preliminary study from 14TFs iEndo cells to pancreatic differentiation shows significant induction of pancreatic and endocrine markers expression at mRNA level. Our results provide a novel approach for generating insulin producing β-cells for cell-based therapy and disease modelling.