Generierung von insulinproduzierenden »Neo-Inselzellen« aus humanen Monozyten — Analyse des Differenzierungsstatus

Abstract

Allogeneic islet cell transplantation is currently the only curative treatment of diabetes mellitus. However due to organ shortage islet transplantation is limited and immunsupressive protocols are hazardous and complicated side effects. The incidence of diabetes is constantly growing and a treatment with autologous insulin producing cells would hold great promise for the therapy of diabetes. We herein describe the generation of insulin producing »Neo-islet cells« from human peripheral blood monocytes, which are able to secret insulin in a glucose dependent fashion and normalize blood glucose levels after transplantation into diabetic mice for a period of ten days. In comparison with pancreatic islet cells »Neo-islet cells« exhibit a gene expression pattern which is comparable to an endocrine precursor cell as known from embryonic pancreas developmental studies.

Human monocytes were treated with MCSF and IL-3 in order to undergo a step of dedifferentiation. Further culture in EGF, HGF and nicotinamide for 4 to 8 days resulted in the formation of cell aggregates expressing insulin, somatostatin and glucagon. Insulin and C-peptide secretion was shown to be glucose dependent after incubation with 22mM glucose whereas 3mM glucose did not induce insulin secretion. MRNA was isolated from »Neo-islet cells« at different times and analyzed for expression of insulin, somatostatin, glucagon, PDX-1, Ngn3, carboxypeptidase, Nkx6.1, NeuroD, Glut2, Ngn3, MAFa and Pax4. Stable expression of insulin, carboxypeptidase, glucagon, Maffa, Nkn6.1, Ngn3 and Neuro D was found. The expression of somatostatin, Isl-1, pro-hormone-convertase, Glut2, Pax 4 and PDX 1 was not reproducible in all experiments and so far not stable. Although »Neoislet cells« are not fully differentiated they were able to normalize blood glucose in diabetic mice transiently. We hope to characterize these cells in more detail in order to optimize the differentiation protocol as a perspective for autologous cell therapy of diabetes.