Applied Biochemistry and Biotechnology

, Volume 170, Issue 4, pp 962–971

Extrinsic Factors Promoting In Vitro Differentiation of Insulin-Secreting Cells from Human Adipose Tissue-Derived Mesenchymal Stem Cells

Article

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 % CO2 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.

Keywords

Extrinsic factors Insulin-secreting cells Mesenchymal stem cells Transcriptional factors 

Abbreviations

ACT A

Activin A

α-MEM

Alpha modified Minimum Essential Media

B-hFGF

Basic-human fibroblast growth factor

DMEM

Dulbecco’s modified eagle’s medium

EB

Embryoid bodies

ESC

Embryonic stem cell

GLP

Glucagon-like peptide

H-AD-MSC

Human adipose tissue-derived mesenchymal stem cells

HGF

Hepatocyte growth factor

Isl-1

Islet 1 transcriptional factor

ICC

Islet-like cell clusters

IS-MSC

Insulin-secreting mesenchymal stem cells

NIC

Nicotinamide

Pax-6

Paired box genes-6

PBS

Phosphate-buffered saline

Pdx-1

Pancreatic and duodenal homobox 1

Png

Pentagastrin

RPM

Revolutions per minute

TGF-β

Transforming growth factor-β

Copyright information

© Springer Science+Business Media New York 2013

Authors and Affiliations

  1. 1.Department of Pathology, Laboratory Medicine, Transfusion Services and ImmunohematologyG. R. Doshi and K. M. Mehta Institute Of Kidney Diseases & Research Centre (IKDRC)- Dr. H.L. Trivedi Institute of Transplantation Sciences (ITS)AhmedabadIndia
  2. 2.Department of Nephrology and Transplantation MedicineG. R. Doshi and K. M. Mehta Institute of Kidney Diseases & Research Centre (IKDRC)–Dr. H.L. Trivedi Institute of Transplantation Sciences (ITS)AhmedabadIndia
  3. 3.Stem Cell Lab, Transplantation Biology Research Centre, Department of Pathology, Laboratory Medicine, Transfusion Services and ImmunohematologyG. R. Doshi and K. M. Mehta Institute of Kidney Diseases & Research Centre (IKDRC)- Dr. H.L. Trivedi Institute of Transplantation Sciences (ITS)AhmedabadIndia

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