The transcription factor Neurogenin3 (Ngn3) is required for islet-cell type specification. Here, we show that hepatic gene transfer of Ngn3 transiently induces insulin in terminally differentiated hepatocytes but fails to transdifferentiate them, i.e., switch their lineage into islet cells. However, Ngn3 leads to long-term diabetes reversal in mice due to the emergence of periportal islet-like cell clusters. These neo-islets display glycemia-regulated insulin, beta-cell-specific transcripts, and an islet-specific transcription cascade, and they produce all four major islet hormones. They appear to arise from hepatic progenitor cells, most likely endoderm-derived oval cells. Thus, transfer of a single lineage-defining transcription factor, Ngn3, is sufficient to induce cell-lineage switching from a hepatic to an islet lineage in these progenitor cells, a process consistent with transdetermination, i.e, lineage switching in lineage-determined, but not terminally differentiated, cells. This paradigm of induced transdetermination of receptive progenitor cells in vivo may be generally applicable to therapeutic organogenesis for multiple diseases, including diabetes.

Test whether Neurog3 is sufficient to induce islet organogenesis in the mouse liver.

Diabetes was induced by IP streptozotocin on two consecutive days. STZ-induced diabetic mice were transfected with Ngn3-Btc, Neurog3, Btc, and empty vector using helper-dependent adenoviral vectors (HDAds) by intravenous injection via tail vein after 1 week of stable diabetes. Total RNA extracted from the liver of transfected mice and the liver and the pancreas of normal mice was subjected to quantitative Real-Time RT-PCR using specific primers and SYBR Green mix and normalized to the expression of two housekeeping genes (Gapdh and Eef1g). The primers were designed to probe mRNAs of a set of islet hormones and beta-cell-specific proteins (Ins1, Ins2, Gcg, Ppy, Sst, Abcc8, Kcnj11, Pcsk1, Pcsk2, Thy1) and a set of transcription factors (Pdx1, Neurog3, Neurod1, Isl1, Pax4, Pax6, Nkx2-2, Nkx6-1). Ngn3-Btc treated mice were sacrificed at two time points: 3-6 weeks and 12-16 weeks after HDAd treatment.

By quantitative Real-Time RT-PCR, transcripts of a set of islet hormones and beta-cell-specific proteins (Ins1, Ins2, Gcg, PP, Sst, Abcc8, Kcnj11, Pcsk1, Pcsk2, Thy1) and a set of transcription factors (Pdx1, Neurog3, Neurod1, Isl1, Pax4, Pax6, Nkx2-2, Nkx6-1) were detected in the liver of Ngn3-Btc treated mice (at two time points) and Ngn3-only treated mice. RNA abundance was detected at a higher level in Ngn3-Btc treated mice.

Neurog3 gene transfer, with or without Btc, was sufficient to induce the expression of genes encoding a set of major islet hormones and a set of transcription factors in the mouse liver.

• Ngn3: 1110, 1470, 2520, 2800, 3021, 3100, 3425, 3733, 3795, 4195, 4374,
• transdifferentiation: 3795, 4397, 4495,