We have described multipotent progenitor-like cells within the major pancreatic ducts (MPDs) of the human pancreas. They express PDX1, its surrogate surface marker P2RY1, and the bone morphogenetic protein (BMP) receptor 1A (BMPR1A)/activin-like kinase 3 (ALK3), but not carbonic anhydrase II (CAII). Here we report the single-cell RNA sequencing (scRNA-seq) of ALK3 ^bright+ -sorted ductal cells, a fraction that harbors BMP-responsive progenitor-like cells. Our analysis unveiled the existence of multiple subpopulations along two major axes, one that encompasses a gradient of ductal cell differentiation stages, and another featuring cells with transitional phenotypes toward acinar tissue. A third potential ducto-endocrine axis is revealed upon integration of the ALK3 ^bright+ dataset with a single-cell whole-pancreas transcriptome. When transplanted into immunodeficient mice, P2RY1 ⁺ /ALK3 ^bright+ populations (enriched in PDX1 ⁺ /ALK3 ⁺ /CAII ⁻ cells) differentiate into all pancreatic lineages, including functional β-cells. This process is accelerated when hosts are treated systemically with an ALK3 agonist. We found PDX1 ⁺ /ALK3 ⁺ /CAII ⁻ progenitor-like cells in the MPDs of types 1 and 2 diabetes donors, regardless of the duration of the disease. Our findings open the door to the pharmacological activation of progenitor cells in situ.

Regenerative medicine is transitioning into clinical programs using stem/progenitor cell therapies for repair of damaged organs. We summarize those for liver and pancreas, organs that share endodermal stem cell populations, biliary tree stem cells (hBTSCs), located in peribiliary glands. They are precursors to hepatic stem/progenitors in canals of Hering and to committed progenitors in pancreatic duct glands. They give rise to maturational lineages along a radial axis within bile duct walls and a proximal‐to‐distal axis starting at the duodenum and ending with mature cells in the liver or pancreas. Clinical trials have been ongoing for years assessing effects of determined stem cells (fetal‐liver‐derived hepatic stem/progenitors) transplanted into the hepatic artery of patients with various liver diseases. Immunosuppression was not required. Control subjects, those given standard of care for a given condition, all died within a year or deteriorated in their liver functions. Subjects transplanted with 100‐150 million hepatic stem/progenitor cells had improved liver functions and survival extending for several years. Full evaluations of safety and efficacy of transplants are still in progress. Determined stem cell therapies for diabetes using hBTSCs remain to be explored but are likely to occur following ongoing preclinical studies. In addition, mesenchymal stem cells (MSCs) and hematopoietic stem cells (HSCs) are being used for patients with chronic liver conditions or with diabetes. MSCs have demonstrated significant effects through paracrine signaling of trophic and immunomodulatory factors, and there is limited evidence for inefficient lineage restriction into mature parenchymal or islet cells. HSCs' effects are primarily via modulation of immune mechanisms. Stem Cells 2013;31:2047–2060

In type 1 diabetes (T1D), autoimmune destruction of pancreatic β cells leads to insulin deficiency and loss of glycemic control. However, knowledge about human pancreas pathophysiology in T1D remains incomplete. To address this limitation, we established a pancreas tissue slice platform of donor organs with and without diabetes, facilitating the first live cell studies of human pancreas in T1D pathogenesis to our knowledge. We show that pancreas tissue slices from organ donors allow thorough assessment of processes critical for disease development, including insulin secretion, β cell physiology, endocrine cell morphology, and immune infiltration within the same donor organ. Using this approach, we compared detailed pathophysiological profiles for 4 pancreata from donors with T1D with 19 nondiabetic control donors. We demonstrate that β cell loss, β cell dysfunction, alterations of β cell physiology, and islet infiltration contributed differently to individual cases of T1D, allowing insight into pathophysiology and heterogeneity of T1D pathogenesis. Thus, our study demonstrates that organ donor pancreas tissue slices represent a promising and potentially novel approach in the search for successful prevention and reversal strategies of T1D.

The exocrine pancreas can give rise to endocrine insulin-producing cells upon ectopic expression of key transcription factors. However, the need for genetic manipulation remains a translational hurdle for diabetes therapy. Here we report the conversion of adult human nonendocrine pancreatic tissue into endocrine cell types by exposure to bone morphogenetic protein 7. The use of this U.S. Food and Drug Administration–approved agent, without any genetic manipulation, results in the neogenesis of clusters that exhibit high insulin content and glucose responsiveness both in vitro and in vivo. In vitro lineage tracing confirmed that BMP-7–induced insulin-expressing cells arise mainly from extrainsular PDX-1+, carbonic anhydrase II− (mature ductal), elastase 3a (acinar)−, and insulin− subpopulations. The nongenetic conversion of human pancreatic exocrine cells to endocrine cells is novel and represents a safer and simpler alternative to genetic reprogramming.

Treatment of human pancreatic non-endocrine tissue with Bone Morphogenetic Protein 7 (BMP-7) leads to the formation of glucose-responsive β-like cells. Here, we show that BMP-7 acts on extrainsular cells expressing PDX1 and the BMP receptor activin-like kinase 3 (ALK3/BMPR1A). In vitro lineage tracing indicates that ALK3+ cell populations are multipotent. PDX1+/ALK3+ cells are absent from islets but prominently represented in the major pancreatic ducts and pancreatic duct glands. We identified the purinergic receptor P2Y1 (P2RY1) as a surrogate surface marker for PDX1. Sorted P2RY1+/ALK3bright+ cells form BMP-7-expandable colonies characterized by NKX6.1 and PDX1 expression. Unlike the negative fraction controls, these colonies can be differentiated into multiple pancreatic lineages upon BMP-7 withdrawal. RNA-seq further corroborates the progenitor-like nature of P2RY1+/ALK3bright+ cells and their multilineage differentiation potential. Our studies confirm the existence of progenitor cells in the adult human pancreas and suggest a specific anatomical location within the ductal and glandular networks.

Despite progress in our knowledge about pancreatic islet specification, most attempts at differentiating stem/progenitor cells into functional, transplantable β cells have met only with moderate success thus far. A major challenge is the intrinsic simplicity of in vitro culture systems, which cannot approximate the physiological complexity of in vivo microenvironments. Oxygenation is a critical limitation of standard culture methods, and one of special relevance for the development of β cells, known for their high O2 requirements. Based on our understanding of islet physiology, we have tested the hypothesis that enhanced O2 delivery (as provided by novel perfluorocarbon-based culture devices) may result in higher levels of β-cell differentiation from progenitor cells in vitro. Using a mouse model of pancreatic development, we demonstrate that a physiological-like mode of O2 delivery results in a very significant upregulation of endocrine differentiation markers (up to 30-fold for insulin one and 2), comparable to relevant in vivo controls. This effect was not observed by merely increasing environmental O2 concentrations in conventional settings. Our findings indicate that O2 plays an important role in the differentiation of β cells from their progenitors and may open the door to more efficient islet differentiation protocols from embryonic and/or adult stem cells. Disclosure of potential conflicts of interest is found at the end of this article.