Development of a conditionally immortalized human pancreatic β cell line

Abstract

Diabetic patients exhibit a reduction in β cells, which secrete insulin to help regulate glucose homeostasis; however, little is known about the factors that regulate proliferation of these cells in human pancreas. Access to primary human β cells is limited and a challenge for both functional studies and drug discovery progress. We previously reported the generation of a human β cell line (EndoC-βH1) that was generated from human fetal pancreas by targeted oncogenesis followed by in vivo cell differentiation in mice. EndoC-βH1 cells display many functional properties of adult β cells, including expression of β cell markers and insulin secretion following glucose stimulation; however, unlike primary β cells, EndoC-βH1 cells continuously proliferate. Here, we devised a strategy to generate conditionally immortalized human β cell lines based on Cre-mediated excision of the immortalizing transgenes. The resulting cell line (EndoC-βH2) could be massively amplified in vitro. After expansion, transgenes were efficiently excised upon Cre expression, leading to an arrest of cell proliferation and pronounced enhancement of β cell-specific features such as insulin expression, content, and secretion. Our data indicate that excised EndoC-βH2 cells are highly representative of human β cells and should be a valuable tool for further analysis of human β cells.

Human pancreas development

“…By contrast, although the early PDX1 + pancreatic progenitor cells were proliferative, once cells had undergone β-cell differentiation replication rates were greatly reduced (Castaing et al, 2005). More recently, Scharfmann and colleagues have used the same murine in vivo incubation technique of human fetal progenitor cells as part of a protocol to achieve the exciting development of a glucose-sensitive human β-cell line (Ravassard et al, 2011;Scharfmann et al, 2014).…”

The quest to make fully functional human pancreatic beta cells from embryonic stem cells: climbing a mountain in the clouds

“…It remains to be shown whether the insulin levels in the explanted devices are quantitatively equivalent to those produced by human beta cells and whether the cells respond to relevant secretagogues. It should be noted that if these beta-like cells are proliferating a rate higher than 1% then it is likely that they are not fully differentiated [46,67]. Nevertheless, this progress has precipitated a major consolidation in the field, with the merger of the Janssen subsidiary BetaLogics with ViaCyte (which was also supported by Janssen prior to the merger).…”

“…At this moment, stem cell derived beta-like cells are probably the functional equivalent of the best-characterised human and rodent beta cell lines [46]. In my opinion, clear evidence has not yet been presented that fully functional beta cells can be found in stem cell derived cultures.…”

Non-coding genome functions in diabetes

“…Furthermore, these cells secrete insulin in a glucose-responsive manner, and their transplantation reverses chemically induced diabetes in mice (Ravassard et al 2011). Subsequent work from the same team allowed fine-tuning of this methodology, resulting in the generation of the EndoC-bH2 line, which allows excision of the transgenes that confer cell immortalization and, hence, a better approximation to the physiological features of true b cells (Scharfmann et al 2014).…”