Pancreatic beta cell neogenesis: Debates and updates

Finding endogenous, renewable sources for insulin-producing beta cells in the adult pancreas is one of the holy grails of stem cell research and regenerative medicine. Through lineage tracing and scRNA-seq approaches, Gribben et al., 2021Gribben C. Lambert C. Messal H.A. Hubber E.L. Rackham C. Evans I. Heimberg H. Jones P. Sancho R. Behrens A. Ductal Ngn3-expressing progenitors contribute to adult β cell neogenesis in the pancreas.Cell Stem Cell. 2021; https://doi.org/10.1016/j.stem.2021.08.003Abstract Full Text Full Text PDF PubMed Scopus (12) Google Scholar have recently reported that Ngn3-expressing ductal cells could serve as progenitors for new beta cells in the adult pancreas. Finding endogenous, renewable sources for insulin-producing beta cells in the adult pancreas is one of the holy grails of stem cell research and regenerative medicine. Through lineage tracing and scRNA-seq approaches, Gribben et al., 2021Gribben C. Lambert C. Messal H.A. Hubber E.L. Rackham C. Evans I. Heimberg H. Jones P. Sancho R. Behrens A. Ductal Ngn3-expressing progenitors contribute to adult β cell neogenesis in the pancreas.Cell Stem Cell. 2021; https://doi.org/10.1016/j.stem.2021.08.003Abstract Full Text Full Text PDF PubMed Scopus (12) Google Scholar have recently reported that Ngn3-expressing ductal cells could serve as progenitors for new beta cells in the adult pancreas. The existence of facultative adult stem or progenitor cells that are capable of generating new functional beta cells in vivo in a process termed "neogenesis" has been controversial over the past two decades (Zhou and Melton, 2018Zhou Q. Melton D.A. Pancreas regeneration.Nature. 2018; 557: 351-358Crossref PubMed Scopus (183) Google Scholar). The abundance of conflicting data in this regard has led to an ongoing debate that warrants reflection on past efforts to validate adaptive beta cell neogenesis and, in particular, on caveats of technologies used in genetic lineage tracing of pancreatic cells that form the bedrock of support on both sides of the debate. To date, studies on adult beta cell regeneration have documented two major mechanisms: self-replication and differentiation from facultative progenitors. There is a broad consensus that self-replication occurs, but the existence and physiological significance of neogenesis remain controversial. In 2004, a milestone lineage tracing study revealed self-replication as the primary mechanism for the expansion and maintenance of beta cell mass (Dor et al., 2004Dor Y. Brown J. Martinez O.I. Melton D.A. Adult pancreatic beta-cells are formed by self-duplication rather than stem-cell differentiation.Nature. 2004; 429: 41-46Crossref PubMed Scopus (1900) Google Scholar). Teta et al. confirmed this independently using a novel DNA analog-based lineage tracing technique (Teta et al., 2007Teta M. Rankin M.M. Long S.Y. Stein G.M. Kushner J.A. Growth and regeneration of adult beta cells does not involve specialized progenitors.Dev. Cell. 2007; 12: 817-826Abstract Full Text Full Text PDF PubMed Scopus (463) Google Scholar). Since then, many studies have strengthened the view that pre-existing beta cells are the major cell source for new beta cells under most physiological conditions (Xiao et al., 2013Xiao X. Chen Z. Shiota C. Prasadan K. Guo P. El-Gohary Y. Paredes J. Welsh C. Wiersch J. Gittes G.K. No evidence for β cell neogenesis in murine adult pancreas.J. Clin. Invest. 2013; 123: 2207-2217Crossref PubMed Scopus (159) Google Scholar; Zhao et al., 2021Zhao H. Huang X. Liu Z. Pu W. Lv Z. He L. Li Y. Zhou Q. Lui K.O. Zhou B. Pre-existing beta cells but not progenitors contribute to new beta cells in the adult pancreas.Nat. Metab. 2021; 3: 352-365Crossref PubMed Scopus (16) Google Scholar), with the exception of extreme beta cell loss resulting in transdifferentiation of beta cells from delta and alpha cells. Conversely, evidence supporting facultative stem cell-driven beta cell neogenesis in vivo has also been accumulating. In 2008, neurogenin3 (Ngn3)-expressing ductal cells were demonstrated to differentiate into beta cells in a murine pancreatic duct ligation (PDL) injury model (Xu et al., 2008Xu X. D'Hoker J. Stangé G. Bonné S. De Leu N. Xiao X. Van de Casteele M. Mellitzer G. Ling Z. Pipeleers D. et al.Beta cells can be generated from endogenous progenitors in injured adult mouse pancreas.Cell. 2008; 132: 197-207Abstract Full Text Full Text PDF PubMed Scopus (809) Google Scholar). Others also reported stem cell activities in adult pancreatic cells expressing a variety of markers including CAII, Ptf1a, CD133, ALDH1, Pdx1, and Sox9. In spite of these studies, more evidence is needed to convincingly support the contribution of stem cells to new beta cells in the adult pancreas. In their new study in Cell Stem Cell, Gribben and colleagues reported that the ductal Ngn3+ endocrine progenitors contribute to beta cells in homeostasis and during diabetes (Gribben et al., 2021Gribben C. Lambert C. Messal H.A. Hubber E.L. Rackham C. Evans I. Heimberg H. Jones P. Sancho R. Behrens A. Ductal Ngn3-expressing progenitors contribute to adult β cell neogenesis in the pancreas.Cell Stem Cell. 2021; https://doi.org/10.1016/j.stem.2021.08.003Abstract Full Text Full Text PDF PubMed Scopus (12) Google Scholar). The authors used both genetic lineage tracing and single-cell RNA sequencing (scRNA-seq) approaches to prove the existence of the facultative Ngn3+ beta cell progenitors and suggested that beta cell neogenesis maintains the adult beta cell mass in addition to pre-existing beta cell replication. In proof-of-principle experiments, they performed pulse-chase experiments utilizing Ins1-CreER and found dilution of labeling during homeostasis from 78% to 66% at 1 and 6 months post-tamoxifen treatment. The dilution rate of labeled beta cells was about 0.68% per week, suggesting infiltration of new beta cells from non-labeled progenitors. Meanwhile, they used Hnf1b-CreER to label pancreatic ductal cells and observed an increased number of traced beta cells by 0.66% per week, indicating the contribution of ductal cells to beta cell mass over time. Gribben et al. also used Ngn3-CreER to trace the cell fate of a subset of ductal cells that expressed Ngn3 by a tamoxifen-mediated pulse-chase strategy (Figure 1A). Their results showed that a small population of Ngn3+ ductal cells expressing the endocrine marker somatostatin were capable of leaving the ducts, contributing to beta cells in homeostasis (Figure 1A). Interestingly, the number of beta cells derived from Ngn3+ ductal cells increased in diabetic mice. The differentiation of Ngn3+ ductal cells to the adult beta cells was also supported by single-cell transcriptomic analysis of pancreatic islet cells. In summary, this study highlights the existence of an Ngn3+ progenitor cell population in ductal epithelium and their contribution to adult beta cell neogenesis, providing new insights into potential treatment of diabetes. This work by Gribben and colleagues is interesting and important to strengthen our understanding of beta cell neogenesis. It may provide new hope that we may one day activate the endogenous resident progenitors to generate new beta cells for treating diabetes. Nevertheless, several questions remain to be addressed. For example, the authors reported the dilution of labeled beta cells by Ins1-CreER lineage tracing (Gribben et al., 2021Gribben C. Lambert C. Messal H.A. Hubber E.L. Rackham C. Evans I. Heimberg H. Jones P. Sancho R. Behrens A. Ductal Ngn3-expressing progenitors contribute to adult β cell neogenesis in the pancreas.Cell Stem Cell. 2021; https://doi.org/10.1016/j.stem.2021.08.003Abstract Full Text Full Text PDF PubMed Scopus (12) Google Scholar), suggesting new beta cell formation from progenitors. However, this result is in sharp contrast to previous studies utilizing a similar strategy (Dor et al., 2004Dor Y. Brown J. Martinez O.I. Melton D.A. Adult pancreatic beta-cells are formed by self-duplication rather than stem-cell differentiation.Nature. 2004; 429: 41-46Crossref PubMed Scopus (1900) Google Scholar; Zhao et al., 2021Zhao H. Huang X. Liu Z. Pu W. Lv Z. He L. Li Y. Zhou Q. Lui K.O. Zhou B. Pre-existing beta cells but not progenitors contribute to new beta cells in the adult pancreas.Nat. Metab. 2021; 3: 352-365Crossref PubMed Scopus (16) Google Scholar). Whether difference in mouse strains, husbandry conditions, immunostaining technique, or quantification methods may influence the lineage tracing results remains unclear. Of note, the major conclusions of this study were drawn from experiments using Cre-loxP genetic lineage tracing. The fidelity of genetic lineage tracing hinges entirely on the specificity of Cre or CreER, the linchpin of the Cre-loxP-based lineage tracing (He et al., 2017He L. Li Y. Li Y. Pu W. Huang X. Tian X. Wang Y. Zhang H. Liu Q. Zhang L. et al.Enhancing the precision of genetic lineage tracing using dual recombinases.Nat. Med. 2017; 23: 1488-1498Crossref PubMed Scopus (129) Google Scholar). To unambiguously establish that ductal Ngn3+ progenitor cells contribute to beta cell mass in homeostasis or after injury, Ngn3 itself cannot be expressed in beta cells. However, Ngn3 could be expressed by some pre-existing beta cells, in addition to ductal cells, based on genetic tracing models with Ngn3-tTA, Ngn3-EGFP, and Ngn3-CreER (Wang et al., 2009Wang S. Jensen J.N. Seymour P.A. Hsu W. Dor Y. Sander M. Magnuson M.A. Serup P. Gu G. Sustained Neurog3 expression in hormone-expressing islet cells is required for endocrine maturation and function.Proc. Natl. Acad. Sci. USA. 2009; 106: 9715-9720Crossref PubMed Scopus (136) Google Scholar). Similarly, scRNA-seq profiling tracking human beta cell differentiation from pluripotent stem cells also reveals the Ngn3+Ins+ beta cell population particularly in the immature stage (Li et al., 2020Li X. Yang K.Y. Chan V.W. Leung K.T. Zhang X.B. Wong A.S. Chong C.C.N. Wang C.C. Ku M. Lui K.O. Single-cell RNA-seq reveals that CD9 is a negative marker of glucose-responsive pancreatic β-like cells derived from human pluripotent stem cells.Stem Cell Reports. 2020; 15: 1111-1126Abstract Full Text Full Text PDF PubMed Scopus (19) Google Scholar). Given the potential issues involved in ectopic lineage tracing of beta cells in addition to Ngn3+ ductal cells (Figure 1B), more work is needed to independently re-assess the contribution of Ngn3+ stem cells to beta cells. To this end, more advanced technology, such as the dual orthogonal recombinase-based lineage tracing strategy that has been developed recently, may offer a system to simultaneously label a putative stem cell marker while excluding beta cell labeling (He et al., 2017He L. Li Y. Li Y. Pu W. Huang X. Tian X. Wang Y. Zhang H. Liu Q. Zhang L. et al.Enhancing the precision of genetic lineage tracing using dual recombinases.Nat. Med. 2017; 23: 1488-1498Crossref PubMed Scopus (129) Google Scholar). Taken together, this study provides insight into an endogenous source of progenitors for new beta cells in homeostasis and during diabetes. Unraveling the underlying mechanisms that drive the activation and expansion of these potential beta cell progenitors would advance the field and provide significant therapeutic implications for targeting diabetes and other insulin-dependent disorders. Meanwhile, the existence of beta cell neogenesis remains controversial and new strategies with more state-of-the-art technologies are required to fully address the potential existence of adult stem or progenitor cells in beta cell neogenesis in the future. We thank Dr. Qiao Zhou (Cornell University, USA) for valuable advice. This work was supported by the National Science Foundation of China (31730112, 82088101, 32050087, 91849202, and 31625019). The authors declare no competing interests. Ductal Ngn3-expressing progenitors contribute to adult β cell neogenesis in the pancreasGribben et al.Cell Stem CellSeptember 2, 2021In BriefA better understanding of adult beta cell neogenesis would open new approaches for diabetes treatment. Gribben et al. used lineage tracing and scRNA-seq to characterize a duct-resident somatostatin-positive endocrine progenitor cell population that is a source of beta cells in homeostasis and during diabetes. Full-Text PDF Open Access