β-Cell replacement therapy is a promising field of research that is currently evaluating new sources of cells for clinical use. Pancreatic epithelial cells are potent candidates for β-cell engineering, but their large-scale expansion has not been evidenced yet. Here we describe the efficient expansion and β-cell differentiation of purified human pancreatic duct cells (DCs). When cultured in endothelial growth-promoting media, purified CA19-9(+) cells proliferated extensively and achieved up to 22 population doublings over nine passages. While proliferating, human pancreatic duct-derived cells (HDDCs) downregulated most DC markers, but they retained low CK19 and SOX9 gene expression. HDDCs acquired mesenchymal features but differed from fibroblasts or pancreatic stromal cells. Coexpression of duct and mesenchymal markers suggested that HDDCs were derived from DCs via a partial epithelial-to-mesenchymal transition (EMT). This was supported by the blockade of HDDC appearance in CA19-9(+) cell cultures after incubation with the EMT inhibitor A83-01. After a differentiation protocol mimicking pancreatic development, HDDC populations contained about 2% of immature insulin-producing cells and showed glucose-unresponsive insulin secretion. Downregulation of the mesenchymal phenotype improved β-cell gene expression profile of differentiated HDDCs without affecting insulin protein expression and secretion. We show that pancreatic ducts represent a new source for engineering large amounts of β-like-cells with potential for treating diabetes.

Recent results indicate that the reduction of β-adrenergic signaling impairs angiogenesis under ischemic conditions. Because angiogenesis may occur in the absence of ischemia, it remains to be determined whether and how β-adrenergic signaling regulates angiogenesis, which develops under normoxic conditions. The effect of β-adrenergic ligands on angiogenesis was investigated using 3-dimensional cultures of mouse aortic rings embedded in collagen type I, in which luminized microvessels develop in response to vascular endothelial growth factor (VEGF). Under normoxic conditions, both isoproterenol, a β-adrenergic receptor (β-AR) agonist, and forskolin, an adenylate cyclase activator, were unable to influence aortic microvessel sprouting. On the contrary, treatment with propranolol, a β-AR antagonist, caused an approximately 70% increase in VEGF-mediated microvessel sprouting. This effect was abolished in rings from both double β-AR and β1-AR knockout mice, but not in rings from β2-AR knockout mice. Significant increases in microvessel sprouting were also observed when mouse aortic rings from C57BL/6 mice were treated with the β1-AR-selective antagonists metoprolol and bisoprolol or with the β2-AR-selective antagonist ICI 118,551. Conversely, carvedilol, a nonselective β-AR antagonist, was unable to affect aortic sprouting. These findings suggest that some β-blockers display proangiogenic activity through a mechanism that is independent of their ability to antagonize catecholamine action. The present results also identify a new function for β-AR signaling as a facilitator for VEGF-mediated angiogenesis and have implications for understanding the mechanisms that regulate angiogenic responses under normoxic conditions.

Since insulin discovery, islet transplantation was the first protocol to show the possibility to cure patients with type 1 diabetes using low-risk procedures. The scarcity of pancreas donors triggered a burst of studies focused on the production of new β cells in vitro. These were rapidly dominated by pluripotent stem cells (PSCs) demonstrating diabetes-reversal potential in diabetic mice. Subsequent enthusiasm fostered a clinical trial with immunoisolated embryonic-derived pancreatic progenitors. Yet safety is the Achilles' heel of PSCs, and a whole branch of β cell engineering medicine focuses on transdifferentiation of adult pancreatic cells. New data showed the possibility to chemically stimulate acinar or α cells to undergo β cell neogenesis and provide opportunities to intervene in situ without the need for a transplant, at least after weighing benefits against systemic adverse effects. The current studies suggested the pancreas as a reservoir of facultative progenitors (e.g., in the duct lining) could be exploited ex vivo for expansion and β cell differentiation in timely fashion and without the hurdles of PSC use. Diabetes cell therapy is thus a growing field not only with great potential but also with many pitfalls to overcome for becoming fully envisioned as a competitor to the current treatment standards.

β-Cell replacement therapy represents the most promising approach to restore glucose homeostasis in patients with type 1 diabetes. This study shows an innovative and robust in vitro system for large-scale production of β-like cells from human pancreatic duct-derived cells (HDDCs) using a nonintegrative RNA-based reprogramming technique. V-Maf musculoaponeurotic fibrosarcoma oncogene homolog A overexpression was efficient and sufficient to induce β-cell differentiation and insulin secretion from HDDCs in response to glucose stimulation, allowing the cells to mitigate hyperglycemia in diabetic SCID-beige mice. The data describe a new, reliable, and fast procedure in adult human pancreatic cells to generate clinically relevant amounts of new β cells with the potential to reverse diabetes.

Thorough research on the capacity of human islet transplantation to cure type 1 diabetes led to the achievement of 3-to 5-year-long insulin independence in nearly half of transplanted patients. Yet, translation of this technique to clinical routine is limited by organ shortage and the need for long-term immunosuppression, restricting its use to adults with unstable disease. The production of new bona fide β cells in vitro was thus investigated and finally achieved with human pluripotent stem cells (PSCs). Besides ethical concerns about the use of human embryos, studies are now evaluating the possibility of circumventing the spontaneous tumor formation associated with transplantation of PSCs. These issues fueled the search for cell candidates for β-cell engineering with safe profiles for clinical translation. In vivo studies revealed the regeneration capacity of the exocrine pancreas after injury that depends at least partially on facultative progenitors in the ductal compartment. These stimulated subpopulations of pancreatic ductal cells (PDCs) underwent β-cell transdifferentiation through reactivation of embryonic signaling pathways. In vitro models for expansion and differentiation of purified PDCs toward insulin-producing cells were described using cocktails of growth factors, extracellular-matrix proteins and transcription factor overexpression. In this review, we will describe the latest findings in pancreatic β-cell mass regeneration due to adult ductal progenitor cells. We will further describe recent advances in human PDC transdifferentiation to insulin-producing cells with potential for clinical translational studies.