Generation of human islets through expansion and differentiation of non-islet pancreatic cells discarded (pancreatic discard) after islet isolation.
OBJECTIVE
Islet transplantation is hampered by the shortage of donor tissues. Our objective was to generate islet-like cell clusters (ICCs) from cultures of non-islet pancreatic cells.


METHODS
The starting cultured cells came from the non-islet fractions of human pancreases after enzymatic digestion and purification for the purpose of islet isolation. Initially, these cells expanded in monolayer cultures and became confluent on collagen-coated flasks. After trypsination and suspension of these cells in a defined islet differentiation medium, the cells aggregated to form ICCs.


RESULTS
The initial cell population consisted of less than 1% of insulin-positive cells, 44% amylase-positive cells, and 41% cytokeratin (CK) 7-positive, or CK19 cells, but PDX-1 cells were absent. Cells from later stages of the monolayer cultures showed signs of dedifferentiation/transdifferentiation. At the time of harvesting, more than 90% of the cells were positive for CK 7/19 and PDX-1, but less than 1% of the cells were insulin-positive. After aggregation, the ICCs appeared redifferentiated, and contained glucose-responsive, insulin-secreting cells with an insulin content measuring 20% of that found in freshly isolated islets isolated from the same pancreas. ICCs transplanted into athymic mice and removed after 4 months did acquire the morphology of mature islets, indicating further maturation of the ICCs in vivo after transplantation. Human C-peptide was detected in recipient animal sera.


CONCLUSIONS
Using the specified culture methods, non-islet pancreas cells can generate cell clusters resembling islets. These ICCs, obtained from fractions of the pancreas that are otherwise discarded, continue to differentiate after transplantation to become mature islets.
Southern California Islet Cell Resources Center, Department of Diabetes, Endocrinology and Metabolism, City of Hope National Medical Center/Beckman Research Institute, Duarte, CA 91010, USA. itodorov@coh.org
Ivan
I
Todorov
Keiko
K
Omori
Michael
M
Pascual
Jeffery
J
Rawson
Indu
I
Nair
Luis
L
Valiente
Tommy
T
Vuong
Takeru
T
Matsuda
Chris
C
Orr
Kevin
K
Ferreri
Craig V
CV
Smith
Fouad
F
Kandeel
Yoko
Y
Mullen
D002102:Cadaver; D018929:Cell Culture Techniques / Q000379:methods; D002454:Cell Differentiation; D002455:Cell Division; D002469:Cell Separation; D006801:Humans; D007328:Insulin / Q000378:metabolism; D000078790:Insulin Secretion; D007515:Islets of Langerhans / Q000166:cytology* / Q000378:metabolism; D016381:Islets of Langerhans Transplantation / Q000706:statistics & numerical data; D010179:Pancreas / Q000166:cytology*; D014019:Tissue Donors
D007328:Insulin