Pancreatic beta-cell purification by altering FAD and NAD(P)H metabolism

M J Smelt, M M Faas, B J de Haan, P de Vos

Research output: Contribution to journalArticleAcademicpeer-review

Abstract

Isolation of primary beta cells from other cells within in the pancreatic islets is of importance for many fields of islet research. However, up to now, no satisfactory method has been developed that gained high numbers of viable beta cells, without considerable alpha-cell contamination. In this study, we investigated whether rat beta cells can be isolated from nonbeta endocrine cells by manipulating the flavin adenine dinucleotide (FAD) and nicotinamide-adenine dinucleotide phosphate (NAD(P)H) autofluorescence. Beta cells were isolated from dispersed islets by flow cytometry, based on their high FAD and NAD(P)H fluorescence. To improve beta cell yield and purity, the cellular FAD and NAD(P)H contents were altered by preincubation in culture media containing varying amounts of D-glucose and amino acids. Manipulation of the cellular FAD and NAD(P)H fluorescence improves beta cell yield and purity after sorting. This method is also a fast and reliable method to measure beta cell functional viability. A conceivable application is assessing beta cell viability before transplantation.

Original languageEnglish
Pages (from-to)1-11
JournalExperimental diabetes research
Volume2008
DOIs
Publication statusPublished - 2008
Externally publishedYes

Keywords

  • Animals
  • Cell Culture Techniques
  • Cell Separation/methods
  • Culture Media
  • Flavin-Adenine Dinucleotide/metabolism
  • Flow Cytometry/methods
  • Glucagon-Secreting Cells/cytology
  • Immunohistochemistry
  • Insulin-Secreting Cells/cytology
  • Islets of Langerhans/cytology
  • Male
  • NAD/metabolism
  • NADP/metabolism
  • Rats
  • Rats, Inbred Lew

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