Extrinsic factors promoting in vitro differentiation of insulin-secreting cells from human adipose tissue-derived mesenchymal stem cells

Abstract

Understanding of β cell regeneration is needed to develop new treatment modalities in diabetes mellitus. We present our experience of glucose-sensitive insulin-secreting mesenchymal stem cells (IS-MSC) generated and differentiated from human adipose tissue (h-AD) with application of specific differentiation media, sans xenogenic material. h-AD from donor abdominal wall was collected in proliferation medium composed of α-Minimum Essential Media, albumin, fibroblast-growth factor and antibiotics, minced, incubated in collagenase I at 37 °C with shaker and centrifuged. Supernatant and pellets were separately cultured in proliferation medium on cell + plates at 37 °C with 5 % CO(2) for 10 days. Cells were harvested, checked for viability, sterility, quantification, flow-cytometry (CD45(-)/90(+)/73(+)), and differentiated into insulin-expressing cells using medium composed of Dulbecco's modified eagle's medium, gene expressing upregulators and antibiotics for 3 days. They were studied for transcriptional factors paired box genes-6(Pax-6), islet 1 transcriptional factor (Isl-1), pancreatic and duodenal homobox-1(Pdx-1). C-peptide and insulin were measured by chemiluminescence. IS-MSC showed presence of all three transcriptional factors and showed rise in insulin and c-peptide level in presence of glucose stimuli. It can be concluded that the specific extrinsic factors used in the defined differentiation media effectively and safely promote differentiation of glucose-sensitive insulin-secreting cells from human adipose tissue, without any genetic modulation.