Objectives: Several studies have demonstrated that chronic exposure of pancreatic beta-cells or
islets to supraphysiological glucose concentrations results in loss of the glucose
responsiveness of the insulin promoter and reduced insulin expression. We aimed to
investigate the signalling pathways involved in this process in more detail using
the insulinoma cell line INS-1.
Methods: INS-1 cells were treated with 16mM glucose for 48 hours. Various signalling pathways
were investigated by inhibition with specific pharmacological inhibitors. Expression
of insulin and various differentiation markers of beta-cells were determined by semi-quantitative
real-time RT-PCR.
Results: High glucose levels resulted in down-regulation of insulin expression and specific
regulation of genes characteristic for beta cells (for example Glut2, IAPP) and also
activators and repressors of insulin gene transcription (for example PDX-1, BETA2,
MafA, C/EBPbeta). Inhibition of fatty acid synthesis by C75 and activation of AMP-activated
protein kinase by AICAR resulted in a similar increase of insulin gene expression
under basal and high glucose concentrations. However, the glucotoxic effect itself
was not neutralised and we thus determined the implication of the mitogen-activated
protein kinases (MAPK) that are known to be activated in response to cellular stress.
Our data indicate that blockage of ERK1/2 and p38 activity using the specific pathway
inhibitors PD098059 and SB202190 respectively, resulted in neutralisation of hyperglycaemia-induced
reduction of insulin gene expression.
Conclusion: Taken together, reducing intracellular fatty acid accumulation by inhibiting biosynthesis
and concurrently stimulating beta-oxidation resulted in elevated insulin gene expression
under basal and glucotoxic conditions. However, our data suggest that the glucotoxic
effects depend on MAPK ERK1/2 and p38.
