Metabolome Response to Glucose in the β-Cell Line INS-1 832/13

Charles F Burant,Matthew A Lorenz,Mahmoud A El Azzouny,Robert T Kennedy

doi:10.1074/jbc.m112.414961

Charles F Burant, Matthew A Lorenz + Show 2 more

Open Access

https://doi.org/10.1074/jbc.m112.414961

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Abstract

Glucose-stimulated insulin secretion (GSIS) from pancreatic β-cells is triggered by metabolism of the sugar to increase ATP/ADP ratio that blocks the KATP channel leading to membrane depolarization and insulin exocytosis. Other metabolic pathways believed to augment insulin secretion have yet to be fully elucidated. To study metabolic changes during GSIS, liquid chromatography with mass spectrometry was used to determine levels of 87 metabolites temporally following a change in glucose from 3 to 10 mM glucose and in response to increasing concentrations of glucose in the INS-1 832/13 β-cell line. U-[(13)C]Glucose was used to probe flux in specific metabolic pathways. Results include a rapid increase in ATP/ADP, anaplerotic tricarboxylic acid cycle flux, and increases in the malonyl CoA pathway, support prevailing theories of GSIS. Novel findings include that aspartate used for anaplerosis does not derive from the glucose fuel added to stimulate insulin secretion, glucose flux into glycerol-3-phosphate, and esterification of long chain CoAs resulting in rapid consumption of long chain CoAs and de novo generation of phosphatidic acid and diacylglycerol. Further, novel metabolites with potential roles in GSIS such as 5-aminoimidazole-4-carboxamide ribotide (ZMP), GDP-mannose, and farnesyl pyrophosphate were found to be rapidly altered following glucose exposure.

Highlights

The biochemical pathways underlying glucose-stimulated insulin secretion have not been fully elucidated
Insulin secretion rate following a step change from 0.5 to 10 mM glucose showed a relatively sharp peak at ϳ4 min (28 ng/mg protein/min) and a smaller broad peak with maxima at ϳ25 min corresponding to 1st phase and 2nd phase of insulin secretion (Fig. 1B, see supplemental Fig. S2 for calculation of secretion rate), consistent with previous reports of Glucose-stimulated insulin secretion (GSIS) in islets [14] and INS-1 832/13 cells [15]
GDP-fucose, a product of GDPmannose metabolism [57], was unchanged. While determination of these metabolites in ␤-cells have not been previously explored, we have previously described that activation of the insulin-like growth factor II (IGF-II)/mannose-6-phosphate (M-6-P) receptor by IGF-II results in augmentation of insulin secretion, even at low concentrations of glucose [58]

Summary

Background

The biochemical pathways underlying glucose-stimulated insulin secretion have not been fully elucidated. Results: Mass spectrometry analysis revealed rapid and substantial metabolic reprogramming evoked by glucose in INS-1 cells. Glucose-stimulated insulin secretion (GSIS) from pancreatic ␤-cells is triggered by metabolism of the sugar to increase ATP/ ADP ratio that blocks the KATP channel leading to membrane depolarization and insulin exocytosis. To study metabolic changes during GSIS, liquid chromatography with mass spectrometry was used to determine levels of 87 metabolites temporally following a change in glucose from 3 to 10 mM glucose and in response to increasing concentrations of glucose in the INS-1 832/13 ␤-cell line. Results include a rapid increase in ATP/ADP, anaplerotic tricarboxylic acid cycle flux, and increases in the malonyl CoA pathway, support prevailing theories of GSIS. The metabolic pathways that facilitate 1st and 2nd phase of glucose-stimulated insulin secretion (GSIS) by ␤-cells are not fully understood [1]. ␤-Cell Metabolomics play a role in the dynamics of insulin secretion following glucose exposure

EXPERIMENTAL PROCEDURES

RESULTS AND DISCUSSION

CONCLUSIONS