Figure 8. In-vitro and in-vivo functional characterization of ICAs.

Static stimulation assay for day14 ICAs. ICAs were sequentially treated with low glucose (5.5 mM) and high glucose (22 mM) concentration in triplicate for one hour. Cell supernatants were collected and analyzed for c-peptide release by EIA assay (p values≤0.05) (n = 3) (A). Total intracellular c-peptide content of day 14 ICAs (p≤0.05) was measure in acid-ethanol and compared equal number of undifferentiated h-ASCs (p≤0.05), (n = 3) (B). In-vivo characterization of h-ASCs derived ICAs. Transplantation of day-14 ICAs in STZ induced diabetic Swiss albino mice (n = 3+3) showed hypoglycemic effect within two weeks. Day-14 ICAs (1000–1200) were encapsulated in calcium-alginate and packed into biocompatible capsules (made of PU-PVP-IPN) and transplanted intraperitoneally to diabetic mice. Control group of animals included sham control of diabetic mice transplanted with empty capsule (n = 3) and diabetic mice transplanted with encapsulated undifferentiated h-ASCs (n = 3+3). Overnight fasting blood glucose levels of mice in each experimental group for three week time period is represented (C). Quantification of human C-peptide in blood serum of mice transplanted with ICAs and diabetic control mice (D). Serum samples of experimental mice were collected by retro orbital bleeding on 28^th day of transplantation, for the detection of human c-peptide/mouse c-peptide in blood with ultrasensitive human c-peptide/mouse c-peptide kit were used. Abbreviations: ICAs, Islet like cell aggregates; STZ, streptozotocin; IP-GTT, intra-peritoneal glucose tolerance test; DAPI, (4′, 6-diamidoino-2-phenylindole).