Metabolomic and proteomic analysis of a clonal insulin-producing beta-cell line (INS-1 832/13).

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Metabolomic and proteomic analysis of a clonal insulin-producing beta-cell line (INS-1 832/13). / Fernandez, Celine; Krus, Ulrika; Hallgard, Elna; Spégel, Peter; Holm, Cecilia; Krogh, Morten; Wårell, Kristofer; James, Peter; Mulder, Hindrik.

I: Journal of Proteome Research, Vol. 7, Nr. 1, 2008, s. 400-411.

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T1 - Metabolomic and proteomic analysis of a clonal insulin-producing beta-cell line (INS-1 832/13).

AU - Fernandez, Celine

AU - Krus, Ulrika

AU - Hallgard, Elna

AU - Spégel, Peter

AU - Holm, Cecilia

AU - Krogh, Morten

AU - Wårell, Kristofer

AU - James, Peter

AU - Mulder, Hindrik

PY - 2008

Y1 - 2008

N2 - Metabolites generated from fuel metabolism in pancreatic beta-cells control exocytosis of insulin, a process which fails in type 2 diabetes. To identify and quantify these metabolites, global and unbiased analysis of cellular metabolism is required. To this end, polar metabolites, extracted from the clonal 832/13 beta-cell line cultured at 2.8 and 16.7 mM glucose for 48 h, were derivatized followed by identification and quantification, using gas chromatography (GC) and mass spectrometry (MS). After culture at 16.7 mM glucose for 48 h, 832/13 beta-cells exhibited a phenotype reminiscent of glucotoxicity with decreased content and secretion of insulin. The metabolomic analysis revealed alterations in the levels of 7 metabolites derived from glycolysis, the TCA cycle and pentose phosphate shunt, and 4 amino acids. Principal component analysis of the metabolite data showed two clusters, corresponding to the cells cultured at 2.8 and 16.7 mM glucose, respectively. Concurrent changes in protein expression were analyzed by 2-D gel electrophoresis followed by LC-MS/MS. The identities of 86 spots corresponding to 75 unique proteins that were significantly different in 832/13 beta-cells cultured at 16.7 mM glucose were established. Only 5 of these were found to be metabolic enzymes that could be involved in the metabolomic alterations observed. Anticipated changes in metabolite levels in cells exposed to increased glucose were observed, while changes in enzyme levels were much less profound. This suggests that substrate availability, allosteric regulation, and/or post-translational modifications are more important determinants of metabolite levels than enzyme expression at the protein level.

AB - Metabolites generated from fuel metabolism in pancreatic beta-cells control exocytosis of insulin, a process which fails in type 2 diabetes. To identify and quantify these metabolites, global and unbiased analysis of cellular metabolism is required. To this end, polar metabolites, extracted from the clonal 832/13 beta-cell line cultured at 2.8 and 16.7 mM glucose for 48 h, were derivatized followed by identification and quantification, using gas chromatography (GC) and mass spectrometry (MS). After culture at 16.7 mM glucose for 48 h, 832/13 beta-cells exhibited a phenotype reminiscent of glucotoxicity with decreased content and secretion of insulin. The metabolomic analysis revealed alterations in the levels of 7 metabolites derived from glycolysis, the TCA cycle and pentose phosphate shunt, and 4 amino acids. Principal component analysis of the metabolite data showed two clusters, corresponding to the cells cultured at 2.8 and 16.7 mM glucose, respectively. Concurrent changes in protein expression were analyzed by 2-D gel electrophoresis followed by LC-MS/MS. The identities of 86 spots corresponding to 75 unique proteins that were significantly different in 832/13 beta-cells cultured at 16.7 mM glucose were established. Only 5 of these were found to be metabolic enzymes that could be involved in the metabolomic alterations observed. Anticipated changes in metabolite levels in cells exposed to increased glucose were observed, while changes in enzyme levels were much less profound. This suggests that substrate availability, allosteric regulation, and/or post-translational modifications are more important determinants of metabolite levels than enzyme expression at the protein level.

U2 - 10.1021/pr070547d

DO - 10.1021/pr070547d

M3 - Article

C2 - 18062666

VL - 7

SP - 400

EP - 411

JO - Journal of Proteome Research

JF - Journal of Proteome Research

SN - 1535-3893

IS - 1

ER -