(A) Schematic diagram of experimental strategy. Adenoviruses co-expressing reprogramming factor (R.F.) and mCherry (cherry) were used to directly induce conversion of acinar cells in adult pancreas. … see more
(A). Diagrams of the constructs used. CMV: cytomegaloviral promoter. Dark gray bar: 2A peptide that mediates polycistronic protein expression. Cherry: monomeric cherry fluorescent protein. (B). … see more
(A–L) Mafa alone, Pdx1 alone, Pdx1.Mafa (polycistronic coexpression), Ngn3.Pdx1 (polycistronic coexpression) do not induce the three principle hormones of pancreatic islets. Sst, somatostatin; Gcg, … see more
We performed qPCR analyses at four different time points after viral infection (day 2, 10, 30, and 60) in Ngn3cherry mediated delta cell induction (A) or Ngn3cherry+Mafacherry mediated alpha cell … see more
(A–C) Induced δ-cells co-express somatostatin (SST) and cholecystokinin receptor B (Cckbr) (A and A′). They also co-express the endocrine markers Pax6 (B and B′) and Synaptophysin (Syn, C and C′). … see more
The induced δ-cells are detectable in adult pancreas 2 month after induction and they continue to express Pax6. Ecad: E-cadherin.
Representative images of endogenous α-, δ-, and β-cells are presented in A, C, E, whereas that of induced α- and δ-cells are presented in B, D. Images in A′–E′ are magnified views of the boxed areas … see more
The genomic region around the transcriptional start site (TSS) is shown (1 kb upstream and 1 kb downstream). CpG are represented as triangles. CpG analyzed in this study are shown as solid … see more
Endogenous δ- and α-cells were purified by staining wide-type islet cells and intracellular FACS (A), yield 10.1% glucagon+ cells and 5.5% somatostatin+ cells from islets. Induced δ-cells were … see more
(A) The genomic region around the transcriptional start site (TSS) of somatostatin gene is shown (1 kb upstream and 1 kb downstream). CpG are represented as triangles. CpG analyzed in this study are … see more
Combined bisulfite restriction analysis (COBRA) confirmed that the Amylase 2 promoter is lightly methylated in acinar cells (middle lane), but heavily methylated in both islet δ-cells and induced … see more
(A) Co-infection of two separate viruses carrying Ngn3 and Mafa led to the induction of Glucagon (Gcg+) cells. Somatostatin (Sst+) cells were also induced as a separate population (A′). (B–D) … see more
The induced α-cells are detectable in adult pancreas 2 month after induction. They are Pax6+ and Ecadherin+.
Induced α-cells were isolated by harvesting the acinar fraction of infected pancreatic samples 20 days after infection (co-infection with Ngn3cherry and Mafacherry), following by intracellular FACS. … see more
A small number of induced Gcg+ α-cells express the acinar factor Amylase 30 days after induction. White arrows indicate properly converted Gcg+Amylase− cells. Yellow arrows indicate partially … see more
(A–B) Genetic lineage tracing of induced δ- and α-like cells. Tamoxifen induction of bigenic Ptf1aCreER::RosaYFP animals led to specific and indelible labeling of approximately 20% of adult … see more
Immunohistochemistry showed that 4 days after expression of the three reprogramming factors individually in the pancreas, Ngn3 and Mafa, but not Pdx1, strongly suppressed the expression of the … see more
Compared with the robust induction of Pax6 and Sst by Ngn3 alone (A, D, G), co-expression of Nr5a2 and Ngn3 (by co-infection of two separate viruses) strongly inhibited the activation of both … see more
In acinar to δ-cell conversion induced by Ngn3, strong suppression of the acinar factor Mist1 was observed in the Cherry+-infected cells at day 2 (A). The pan-endocrine factor Pax6 was also induced … see more
In acinar to β-cell conversion induced by M3 (Ngn3+Mafa+Pdx1), near complete suppression of the acinar factor Mist1 was observed in the Cherry+-infected cells at day 2 (A). The pan-endocrine factor … see more
Compared with robust induction of Sst+ cells by Ngn3 alone (A), polycistronic co-expression of Mafa and Ngn3 led to strong reduction of Sst+ cells (B). Polycistronic co-expression of Pdx1 and Ngn3 … see more
(A) Summary of acinar to islet endocrine conversion with defined factors. (B) Our studies suggest that there are two main processes in pancreatic acinar to endocrine reprogramming. Ngn3 plays a … see more
PCR primers for DNA methylation assays