HNF6 in human pancreatic duct development

Background Recently, we identified the novel diabetes gene HNF6 regulating the transcriptional and epigenetic machinery critical for proper pancreatic endocrine development. In addition to a diabetic phenotype, patients lacking functional HNF6 also suffered from exocrine pancreatic insufficiency, absence of gallbladder and of extrahepatic biliary duct. Moreover, HNF6 was shown to play an important role during duct morphogenesis in mice supporting a functional role during human pancreatic duct development. Therefore, we aim to investigate the underlying molecular basis by which loss of HNF6 causes different pancreatic phenotypes in patients.

Methods We used genome-edited human embryonic stem cells to dissect the functional consequences of defective HNF6 in pancreatic ductal development. Our novel and unique differentiation protocol yields virtually pure pancreatic duct-like cells resembling key features of adult human pancreatic ducts. Finally, various stages of development were characterized to assess the morphology, differentiation, and proliferation of HNF6-depleted ductal cells.

Results During in vitro differentiation, loss of HNF6 results in diminished formation of pancreatic progenitor (PP) cells. To efficiently initiate ductal differentiation, we purified PP cells using the stage-specific surface marker GP2. HNF6-depleted ductal organoids showed initially an increased cyst size similar to the mouse model but failed to maintain organoids in consecutive passages supported by loss of proliferation marker Ki67. Moreover, ductal organoids expressed markers such as CK19, CK8 and CFTR but showed increases in maturation marker CK7. This might correlate with an early decrease in NKX6.1 leading to premature formation of ducts and loss of proliferation capacity. Also, organoids lacking HNF6 tend to lose primary cilia during maturation in accordance with the mouse model.

Conclusion Our in vitro approach to direct human HNF6-depleted PSCs towards pancreatic duct-like cells recapitulates aspects of patient phenotypes and therefore, provides a valuable methodology to characterize pancreatic diseases. However, further time-resolved RNA- and ATAC-seq analysis is necessary to reveal exact transcriptional and molecular mechanisms during ductal development.

Publication History

Article published online:
08 September 2020