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Stem Cell Res. 2009 Jan;2(1):29-40. doi: 10.1016/j.scr.2008.07.004. Epub 2008 Jul 31.

Derivation of functional insulin-producing cell lines from primary mouse embryo culture.

Author information

  • 1Cardiovascular Research Institute, National University Medical Institutes, Yong Loo Lin School of Medicine, National University of Singapore, Singapore. nmiligd@nus.edu.sg

Abstract

We have previously described the derivation of insulin-producing cell lines from mouse embryonic stem cells (mESCs) by differentiation of an intermediate lineage-restricted E-RoSH cell line through nutrient depletion in the presence of nicotinamide followed by limiting dilution. Here we investigated whether insulin-producing cell lines could be similarly derived directly from mouse embryo cells or tissues. Using a similar approach, we generated the RoSH2.K and MEPI-1 to -14 insulin-producing cell lines from the 5.5-dpc embryo-derived E-RoSH-analogous RoSH2 cell line and a 6.0-dpc mouse embryo culture, respectively. Insulin content was approximately 8 microg/10(6) MEPI-1 cells and 0.5 microg/10(6) RoSH2.K cells. Like insulin-producing mESC-derived ERoSHK cell lines, both MEPI and RoSH2.K lines were amenable to repeated cycles of freeze and thaw, replicated for months with a doubling time of 3-4 days, and exhibited genomic, structural, biochemical, and pharmacological properties of pancreatic beta-cells, including storage and release of insulin and C-peptide in an equimolar ratio. Transplantation of these cells also reversed hyperglycemia in streptozotocin-treated SCID mice and did not induce teratoma. Like ERoSHK cells, both RoSH2.K and MEPI-1 cells also induced hypoglycemia in the mice. Therefore, our protocol is robust and could reproducibly generate insulin-producing cell lines from different embryonic cell sources.

PMID:
19383407
[PubMed - indexed for MEDLINE]
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