Figure 1
Reaggregation of human islet cells to pseudoislets specifically improves GSIS. (A) Experimental design of pseudoislet formation. Isolated human islets were cultured overnight. Non-dispersed islets were partly used for insulin secretion. The others (1000–2000 islets) were digested into single cells using trypsin. After digestion, a defined number of single cells were reaggregated in hanging drops of 20 µl over 3 days. After 2 days or longer culture in non-adhesive multi-well plates, the pseudoislets were used in the experiments. (B) DNA content of single islets and pseudoislets (formed from 2000 cells and cultured for 2 days) from the same donors (n = 2) were measured as described under Materials and Methods. (C,D) Comparison of GSIS of islets and pseudoislets reaggregated from 1000, 2000 and 4000 cells as indicated expressed as (C) fold-stimulation (12 mM glucose over 2.8 mM glucose). (D) % of insulin content (white bars, 2.8 mM glucose and grey bars, 12 mM glucose). Results are expressed as means + s.e.m. of n = 5 independent preparations. (E–N) GSIS of individual preparations of islets (black signs) and pseudoislets (red signs) expressed as (E–I) fold-stimulation and (J–N) % of insulin content. *p < 0.05, ***p < 0.0002 denotes significant GSIS; #p < 0.05 denotes significant difference between secretion at 12 mM glucose of islets and the respective pseudoislet. §p < 0.05 denotes significance to secretion of islets at 2.8 mM glucose. €p < 0.05 significance between secretion at 12 mM glucose of 1000-cells and 4000-cells pseudoislets.
