ARTICLESIn vivo reprogramming of adult pancreat icexocrine cells to b-cellsQiao Zhou1, Juliana Brown2, Andrew Kanarek1, Jayaraj Rajagopal1& Douglas A. Melton1One goal of regenerative medicine is to instructively convert adult cells into other cell types for tissue repair andregeneration. Although isolated examples of adult cell reprogramming are known, there is no general understanding of howto turn one cell type into another in a controlled manner. Here, using a strategy of re-expressing key developmentalregulators in vivo, we identify a specific combination of three transcription factors (Ngn3 (also known as Neurog3) Pdx1 andMafa) that reprograms differentiated pancreatic exocrine cells in adult mice into cells that closely resemble b-cells. Theinduced b-cells are indistinguishable from endogenous islet b-cells in size, shape and ultrastructure. They express genesessential for b-cell function and can ameliorate hyperglycaemia by remodelling local vasculature and secreting insulin. Thisstudy provides an example of cellular reprogramming using defined factors in an adult organ and suggests a generalparadigm for directing cell reprogramming without reversion to a pluripotent stem cell state.Cells of adult organisms arise from sequential differentiation steps thatare generally thought to be irreversible1. Biologists often describe thisprocess of development as proceeding from an undifferentiated(embryonic) cell to a terminally differentiated cell that forms part ofan adult tissue or organ. There are rare examples, however, in whichcells of one type can be converted to another type in a process calledcellular reprogramming or lineage reprogramming2,3. Various forms ofcellular reprogramming are referred to in the literature as transdiffe-rentiation, dedifferentiation or transdetermination4. For example, cel-lular reprogramming occurs in amphibian limb regeneration and flyimaginal disc identity switches5,6, and it may be central to certain typesof pathological metaplasia4. There is long-standing interestand fascina-tion in reprogramming studies, in part because of the promise of har-nessing this phenomenon for regenerativemedicine whereby abundantadult cells that can be easily collected would be converted to othermedically important cell types to repair diseased or damaged tissues.Somatic cell nuclear transfer (SCNT), developed in the 1960s,demonstrated that nuclei from differentiated adult cells could be repro-grammed to a totipotent state after injection into enucleated eggs2,7.More recently, it was shown that a small number of transcriptionfactors can reprogram cultured adult skin cells to induced pluripotentstem (iPS) cells8–13. These studies point to the possibility of regeneratingmammalian tissues by first reverting skin or other adult cells to pluri-potent stem cells and then redifferentiating theseinto various cell types.Alternatively, it should be possible to convert one cell type into anotherdirectly, without the need to first revert the cell to an undifferentiatedpluripotent state. Indeed, there are examples in the literature that sug-gest that this approach is feasible. For example, studies with embryoniccells have shown that dermal fibroblasts and retinal epithelial cells canbe converted into muscle-like cells14, and pancreatic tissue to liver15. Inadult animals, mature B lymphocytes have been reprogrammed intomacrophages16or pro-B cells17. Today, well documented examples ofcellular reprogramming, especially in adult animals, remain rare andhave generally been restricted to cases in which a single inducing factoris involved. The recent work on iPS formation suggests that a specificcombination of multiple factors, instead of a single one, might be themost effective way to reprogram adult cells8–13.We developed a strategy to identify adult cell reprogrammingfactors by re-expressing multiple embryonic genes in living adultanimals. Our focus on embryonic genes is based in part on regenera-tion studies in newts, frogs and fish, wherein it has been shown thatdedifferentiation of adult cells to progenitors, a form of cellularreprogramming, is accompanied by reactivation of embryonic regu-lators5,18,19. These studies suggest that re-expression of appropriateembryonic genes may reprogram differentiated cells.To search for factors that could reprogram adult cells into b-cells,we focused on transcription factors, a class of genes enriched forfactors that regulate cell fates during embryogenesis. An in situhybridization screen of more than 1,100 transcription factors iden-tified groups of transcription factors with cell-type-specific expres-sions in the embryonic pancreas20. There are at least 20 transcriptionfactors expressed in mature b-cells and their immediate precursors,the endocrine progenitors (Supplementary Table 1). Of these, 9 genesexhibited b-cell developmental phenotypes when mutated21,22, andthese were selected for initial reprogramming experiments.We chose mature exocrine cells of the adult pancreas as target cellsfor reprogramming. Exocrine cells derive from pancreatic endoderm,as do b-cells23, and exocrine cells can turn on endocrineprograms whendissociated and cultured in vitro24,25. We carried out our experiments invivo so that any induced b-cells would reside in their native envir-onment, which might promote their survival and/or maturation. Inaddition, this approach allows for a direct comparison of endogenousand induced b-cells. The transcription factors were delivered into thepancreas in adenoviral vectors. It has been shown that adenoviruspreferentially infects pancreatic exocrine cells, but not islet cells26,and, because most endogenous b-cells reside within islets (Fig. 1b),any newly formed (induced) b-cells could be easily detected as extra-islet insulin1cells.Induction of insulin1cells in adult miceAdenovirus that co-expresses each transcription factor together withnuclear GFP (nGFP) was purified. All nine viruses were pooled andinjected as a mixture (referred to as M9, for mixture of nine) into thepancreata of 2-month-old adult mice (Fig. 1a). The immune-deficient1Department of Stem Cell and Regenerative Biology, Howard Hughes Medical Institute, Harvard Stem Cell Institute, Harvard University, 7 Divinity Avenue, Cambridge, Massachusetts02138, USA.2Department of Pathology, Children’s Hospital, Boston, Harvard Medical School, Harvard Stem Cell Institute, 300 Longwood Avenue, Boston, Massachusetts 02115-5724, USA.doi:10.1038/nature073141