1. Volume 141, Issue 2, Pages 731-741.e4 (August 2011)
Lineage Tracing Evidence for Transdifferentiation of Acinar to Duct Cells and Plasticity of Human Pancreas 
Isabelle Houbracken, Evelien de Waele, Jessy Lardon, Zhidong Ling, Harry Heimberg, Ilse Rooman, Luc Bouwens 
Gastroenterology 
Volume 141, Issue 2, Pages e4 (August 2011)
DOI: /j.gastro
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2. Figure 1
Immunofluorescence characterization of the human exocrine cells during culture.
(A–C) Expression of the ductal marker CK19 (red) and the acinar marker chymotrypsin (green) in the freshly isolated exocrine fraction (A), at day 4 (B), and at day 7 in culture (C). Arrows and inset indicate cells coexpressing CK19 and chymotrypsin at day 4.
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3. Figure 2
Expression of duct cell markers at the start of the culture. (A–E) The different ductal markers are coexpressed in the freshly isolated exocrine cells: (A) CK19 (red)/HNF1B (green), (B) CK19 (red)/SOX9 (green), (C) CD133 (red)/SOX9 (green), (D) CAII (red)/SOX9 (green). (E) CFTR (green) is expressed in chymotrypsin (red)-negative cells. Blue represents nuclear staining with 4′,6-diamidino-2-phenylindole (DAPI).
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4. Figure 3
Nongenetic lineage tracing of acinar cells with UEA-1 lectin. (A and B) Immunofluorescent staining of FITC-conjugated UEA-1–labeled cells with (A) chymotrypsin and (B) CK19 immediately after the labeling period of 3 hours. Blue represents nuclear staining with DAPI. (C and D) Fraction of UEA-1–labeled chymotrypsin-positive and CK19-positive cells. In red is the fraction of (C) UEA-1–labeled chymotrypsin-positive (n = 4) or (D) UEA-1–labeled CK19-positive (n = 5) cells indicated for each time point in culture, and in green is the fraction of (C) UEA-1–labeled chymotrypsin-positive or (D) UEA-1–labeled CK19-positive cells. During culture, the percentage of UEA-1–labeled chymotrypsin-positive cells decreases, whereas the fraction of UEA-1–labeled CK19-positive cells increases.
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5. Figure 4
Expression of duct cell markers at the end of culture in UEA-1–labeled cells. (A–F) After a culture period of 7 days, the FITC-conjugated UEA-1–labeled cells (green) express (A) CK19, (B) CAII, (C) CD133, (D) SOX9, (E) HNF1B, and (F) CFTR. These ductal markers are stained in red. Blue represents nuclear staining with DAPI.
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6. Figure 5
Genetic lineage tracing of human acinar cells. (A) Schematic of the 2 adenoviral vectors. (B and C) Human exocrine cells on day 2, after transduction with both Ad-CMV-LSL-EGFP and Ad-Amy-Cre. The immunocytochemical staining for (B) chymotrypsin (red) or (C) CK19 (red) and EGFP (green) indicate stringent control of the EGFP expression in acinar cells. (D and E) Monolayer culture of human exocrine cells on day 7, after transduction with both Ad-CMV-LSL-EGFP and Ad-Amy-Cre. The immunocytochemical staining for (D) chymotrypsin (red) or (E) CK19 (red) and EGFP (green) indicate acinoductal transdifferentiation. Blue represents nuclear staining with DAPI.
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7. Figure 6
Analysis of the process by which acinar cells give rise to duct cells. (A) Quantitative reverse-transcription polymerase chain reaction analysis of the expression of a ductal marker (CK19) and acinar digestive enzymes (amylase, lipase, and chymotrypsin) in response to treatment with UO126, compound E, or EGF. Results are normalized to cycloA and to the control condition on day 7 (untreated, ctrl). Note the extremely high expression level of acinar enzymes on day 1 compared with day 7. (B) Quantification of the percentage of UEA-1–labeled CK19-positive cells on CK19-positive cells and UEA-1–labeled chymotrypsin-positive cells on total cells on day 7 in cultures treated with UO126, compound E, epidermal growth factor, or untreated (ctrl). (C) Quantitative reverse-transcription polymerase chain reaction analysis of the expression of pancreatic progenitor markers NKX6.1, PDX1, PTF1A, NOTCH1, and HES1 during culture (*P < .05).
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8. Supplementary Figure 1
Validation and optimization of the genetic tracing system. (A–C) AR42J cells were transfected with (A) pCMV-EGFP, (B) pShuttle-CMV-Lox-STOP-Lox-EGFP (CMV-LSL-EGFP) only, or (C) CMV-LSL-EGFP and pShuttle-Amylase-nlsCre (Amy-Cre) to evaluate the functionality of the adenoviral tracing system. Cells transfected with both Cre-lox constructs expressed EGFP (green), contrary to cells transfected with CMV-LSL-EGFP only. (D) Percentage of EGFP-positive cells on total cells on day 7 of monolayer culture of human exocrine fraction, transduced with Ad-CMV-EGFP with increasing MOIs on day 1.
Gastroenterology  , e4DOI: ( /j.gastro )
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