No evidence for β cell neogenesis in murine adult pancreas
Xiangwei Xiao, … , John Wiersch, George K. Gittes
Xiangwei Xiao, … , John Wiersch, George K. Gittes
Published April 24, 2013
Citation Information: J Clin Invest. 2013;123(5):2207-2217. https://doi.org/10.1172/JCI66323.
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Research Article

No evidence for β cell neogenesis in murine adult pancreas

Abstract

Whether facultative β cell progenitors exist in the adult pancreas is a major unsolved question. To date, lineage-tracing studies have provided conflicting results. To track β cell neogenesis in vivo, we generated transgenic mice that transiently coexpress mTomato and GFP in a time-sensitive, nonconditional Cre-mediated manner, so that insulin-producing cells express GFP under control of the insulin promoter, while all other cells express mTomato (INSCremTmG mice). Newly differentiated β cells were detected by flow cytometry and fluorescence microscopy, taking advantage of their transient coexpression of GFP and mTomato fluorescent proteins. We found that β cell neogenesis predominantly occurs during embryogenesis, decreases dramatically shortly after birth, and is completely absent in adults across various models of β cell loss, β cell growth and regeneration, and inflammation. Moreover, we demonstrated upregulation of neurogenin 3 (NGN3) in both proliferating ducts and preexisting β cells in the ligated pancreatic tail after pancreatic ductal ligation. These results are consistent with some recent reports, but argue against the widely held belief that NGN3 marks cells undergoing endocrine neogenesis in the pancreas. Our data suggest that β cell neogenesis in the adult pancreas occurs rarely, if ever, under either normal or pathological conditions.

Authors

Xiangwei Xiao, Zean Chen, Chiyo Shiota, Krishna Prasadan, Ping Guo, Yousef El-Gohary, Jose Paredes, Carey Welsh, John Wiersch, George K. Gittes

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Figure 7

An inflammatory milieu can upregulate NGN3 expression in β cells.

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An inflammatory milieu can upregulate NGN3 expression in β cells. We hyp...
We hypothesized that the release of acinar cell contents and the inflammatory milieu in the ligated PDL pancreas may induce preexisting β cells to upregulate their NGN3 expression. To examine this hypothesis (schematic in A), we isolated purified β cells from MIP-GFP mice by FACS. Reaggregated β cells were cultured overnight before exposure to nucleotide-free normal exocrine extract (from GFP– cells in MIP-GFP pancreas), or PDL tail extracts, or control medium. After 2 hours’ culture, the cells were harvested for RNA extraction and qPCR (B) or cytospun for immunostaining (C). (B) Both nucleotide-free exocrine extract and nucleotide-free PDL tail extracts were used as templates to amplify some genes. The absence of PCR products from these extracts confirmed the absence of nucleotide contamination. Amylase, Ck19, Cd31, and vimentin transcripts were also undetected in the β cells, confirming the purity of the MIP-GFP β cells by FACS. Ngn3 transcripts were low in β cells treated with control medium, but were 17.3 ± 0.7-fold upregulated when they were exposed to the exocrine extract for 2 hours, and 68.5 ± 2.1-fold upregulated in β cells when they were exposed to PDL tail extract for 2 hours. (C) Immunostaining of cytospun cells confirmed the qPCR results. NGN3 in red, insulin in green, and Hoechst (HO) in blue. Scale bar: 20 μm. **P < 0.01.