Proteome of H-411E (liver) cells exposed to insulin and tumor necrosis factor-α: Analysis of proteins involved in insulin resistance

Solomon S. Solomon, Nicholas Buss, James Shull, Susanne Monnier, Gipsy Majumdar, Jian Wu, Ivan Gerling

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Abstract

Insulin resistance may be modeled in H-411E liver cells in tissue culture with the use of the cytokine tumor necrosis factor-α (TNF-α) and insulin. This tissue-culture model nicely mimics IR in human type 2 diabetes mellitus. After incubation of liver cells in tissue culture with INS alone, TNF-α alone, and TNF-α plus insulin, as well as a control sample, liver-cell extracts were separated on 2D polyacrylamide-gel electrophoresis on the basis of isoelectric point and molecular weight. We analyzed the gel images with the use of PD Quest software (Bio-Rad Laboratories, Hercules, Calif) to identify differentially expressed protein spots (ie, up or down with insulin vs down or up with TNF-α plus insulin). In separate experiments, phosphorus-32 incorporation/autoradiography and phosphoprotein staining were used to characterize treatment-induced phosphorylations. Affected protein spots were identified with the use of peptide fingerprinting and matrix-assisted laser desorption ionization time of flight mass spectrometry. The first series of experiments identified 6 differentially expressed proteins: eukaryotic translation initiation factor-3, subunit 2, regulator of G-protein signaling-5, superoxide dismutase, protein disulfide isomerase A6, proteasome subunit-α type 3, and regucalcin. In addition, we observed changes in the phosphorylation of protein disulfide isomerase A6. A second series of experiments identified 7 additional proteins with significantly altered differential expression: cell-division protein kinase-4, kinogen heavy chain, carbonic anhydrase-7, E 3 ubiquitin protein ligase, URE-B1; Rab GDP dissociation inhibitor-β, Rab GDP dissociation inhibitor-β2, and MAWDBP. It can be seen that differentially expressed proteins, affected by treatment with insulin or with TNF-α plus insulin, include regulators of translation, protein degradation, cellular Ca++, G-proteins, and free-radical production. Although one cannot detail the mechanism or mechanisms of TNF-α induced IR from this data alone, it is easy to relate all of these proteins to a role in insulin signal transduction and, hence, insulin resistance.

Original languageEnglish (US)
Pages (from-to)275-283
Number of pages9
JournalJournal of Laboratory and Clinical Medicine
Volume145
Issue number5
DOIs
StatePublished - May 1 2005

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Proteome
Liver
Insulin Resistance
Tumor Necrosis Factor-alpha
Insulin
Guanine Nucleotide Dissociation Inhibitors
Tissue culture
Proteins
Protein Disulfide-Isomerases
Phosphorylation
Prokaryotic Initiation Factor-3
Eukaryotic Initiation Factor-3
Translational Peptide Chain Initiation
GTP-Binding Protein Regulators
Cyclin-Dependent Kinase 4
Eukaryotic Initiation Factors
Liver Extracts
Peptide Mapping
Ubiquitin-Protein Ligases
Carbonic Anhydrases

All Science Journal Classification (ASJC) codes

  • Pathology and Forensic Medicine

Cite this

Proteome of H-411E (liver) cells exposed to insulin and tumor necrosis factor-α : Analysis of proteins involved in insulin resistance. / Solomon, Solomon S.; Buss, Nicholas; Shull, James; Monnier, Susanne; Majumdar, Gipsy; Wu, Jian; Gerling, Ivan.

In: Journal of Laboratory and Clinical Medicine, Vol. 145, No. 5, 01.05.2005, p. 275-283.

Research output: Contribution to journalArticle

Solomon, Solomon S. ; Buss, Nicholas ; Shull, James ; Monnier, Susanne ; Majumdar, Gipsy ; Wu, Jian ; Gerling, Ivan. / Proteome of H-411E (liver) cells exposed to insulin and tumor necrosis factor-α : Analysis of proteins involved in insulin resistance. In: Journal of Laboratory and Clinical Medicine. 2005 ; Vol. 145, No. 5. pp. 275-283.
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