| Abstract |
Diabetes and congenital hyperinsulinism (CHI) are severe diseases
affecting the pancreas. Current models for testing drugs to treat these
diseases are in vivo in rodents or isolated rodent islets. Differences
between the human and rodent pancreas, and ethical issues, mean that in
vitro human models are needed. To develop a novel in vitro model for
pancreatic diseases, mesenchymal stem cells (MSCs) and induced pluripotent
stem cells (iPSCs) were derived from the pancreas of patients with CHI.
MSCs from three forms of CHI were phenotypically normal for MSCs, and
maintained the CHI-causing mutation. When compared to MSCs from bone
marrow, the CHI pancreatic MSCs expressed pancreas-specific gene ISL1 and
showed promoter hypomethylation of other pancreatic genes, including PDX1.
The CHI pMSCs could be differentiated to cells resembling immature beta-
cells, with some beta-cell gene expression (INS, PDX1), but no glucose
responsive insulin secretion. CHI associated hypersecretion of insulin was
not seen as the ATP-sensitive potassium KATP channels were not being
expressed. Addition of the Wnt inhibitor DKK1 markedly enhanced
differentiation via induction of neuronal genes. Alongside high insulin
secretion, CHI also features increased proliferation. CHI MSCs were also
hyperproliferative, and showed alterations to the cell cycle. These
changes were related to p27Kip1 localisation, a known affected protein in
CHI tissue, and CDK1, a novel regulator for CHI. iPSCs were also derived
from focal CHI MSCs and were also phenotypically normal, but did not
maintain the pancreatic hypomethylation present in MSCs. The CHI iPSCs
were efficiently differentiated to definitive endoderm and PDX1 positive
cells. Terminally differentiated iPSCs were endocrine, but were not mature
beta-cells. In conclusion, authentic MSCs and iPSCs were derived for the
first time from patients with CHI. These stem cells could be
differentiated towards beta-cells, but mature glucose responsive beta-
cells were not produced. MSC derived beta-like cells secreted insulin but
did not have KATP channels, whereas iPSC derived beta-like cells had KATP
channel gene expression but not INS. With further optimisation to resolve
these, CHI stem cell derived beta-cells may be used for in vitro modelling.
Further, the undifferentiated MSCs only show hyperproliferation associated
with p27Kip1 and CDK1 and so can be a useful resource for modelling
hyperproliferation seen in CHI.
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