S. Andrikopoulos et al., Two novel immortal pancreatic beta-cell lines expressing and secreting human islet amyloid polypeptide do not spontaneously develop islet amyloid, DIABETES, 48(10), 1999, pp. 1962-1970
Type 2 diabetes is characterized by islet amyloid deposits, which are prima
rily composed of the amyloidogenic human form of islet amyloid polypeptide
(IAPP, amylin). The mechanism of islet amyloido-genesis is not known, but o
ther products (e.g., apolipoprotein E and perlecan) contained within Islet
amyloid may be necessary: Because rodent IAPP does not form islet amyloid,
the currently available beta-cell lines are not useful for studying process
es involved in amyloid formation. To develop a suitable in vivo cell system
for the study of islet amyloid formation, we generated two new beta-cell l
ines that express the amyloidogenic human IAPP. We did this by crossbreedin
g human IAPP transgenic mice with RIP-Tag mice that develop islet tumors an
d then culturing one of these islet tumors from two separate offspring of t
his cross. The resultant 2350-2C0 and 2511 cell Lines produce human as well
as mouse IAPP-like immunoreactivity (IAPP-LI) and immunoreactive insulin (
IRI). Incubation of both these cell Lines with 16.7 mmol/l glucose resulted
in a two- to fourfold increase in human IAPP-LI, mouse I;IPP-LI, and IRI s
ecretion compared with 1.67 mmol/l glucose and the combination of 16.7 mmol
/l glucose and 10 mmol/l arginine, 0.1 mmol/l 3-isobutyl-1-methylxanthine (
IBMX), and 5 mu mol/l carbachol induced a >50-fold increase in the release
of these peptides. The omission of calcium from the above secretagogue cock
tail reduced secretion of all three peptides to only two- to sixfold higher
than the 16.7 mmol/l glucose condition. Perifusion with 16.7 mmol/l glucos
e plus 0.1 mmol/l IBMX caused a biphasic secretion of human IAPP-LI and mou
se IAPP-LI, as well as IRI, in both cell lines, with the peak of the first
phase being five- to sixfold higher than the prestimulated 1.67 mmol/l gluc
ose condition. Immunoelectron microscopic inspection of both 2350-2C0 and 2
511 cells after 7 days of culture did not reveal the presence of amyloid fi
brils, suggesting the need for other critical components. We conclude that
we have established two novel beta-cell lines that produce and secrete huma
n IAPP in a regulated manner. These cell lines will be a useful tool to inv
estigate the secretion of human LAPP as well as the necessity of other comp
onents for islet amyloid formation.