Background: Long term exposure of INS-1 beta cells to elevated glucose concentrations has been observed to impair insulin expression, a phenomenon termed as glucose toxicity. Despite its potential relevance in the progression of diabetes, the underlying mechanisms is not yet fully understood.
Aim: Target analysis of single, or a sub-group of metabolites has until now been the method of choice in mammalian cells to investigate the pathological state in terms of small molecule composition. We therefore aimed to show that metabolite profiling is useful to detect multitude changes in metabolism in INS-1 cells after chronic high glucose exposure. We further suggest that mitogen-activated protein kinases (MAPK) participate in the regulation of insulin gene expression.
Methods: INS-1 cells were incubated with medium containing 3 mM and 16 mM glucose±MAPK inhibitor for 48 h. Insulin mRNA was measured using quantitative real time RT-PCR. Intracellular metabolites were extracted with a chloroform:methanol:water mixture and analysed by GC-tofMS.
Results: Insulin gene expression is significantly reduced after exposure of INS-1 cells to chronic high glucose levels. The application of INS-1 extracts to GC-tofMS allowed the relative quantification of several hundred biological peaks. The use of principal component analysis demonstrates a clear differentiation of cells treated with low or high glucose. Further in-depth analysis revealed details of glucose-derived changes in single metabolites. The inhibition of MAPK results in a restoration of insulin gene expression at high glucose levels that may be explained due to improvement of specific metabolite levels.
Conclusion: Metabolite profiling allowed the identification of core metabolites which were present at significantly different levels in low and high glucose treated INS-1 cells and which may participate in the regulation of insulin gene expression.
03 - 07 May 2008
European Society of Endocrinology