Hepatic proteomic analysis revealed altered metabolic pathways in insulin resistant Akt1(+/-)/Akt2(-/-) mice

Abstract

Objective: The aim of this study was to identify liver proteome changes in a mouse model of severe insulin resistance and markedly decreased leptin levels.

Methods: Two-dimensional differential gel electrophoresis was utilized to identify liver proteome changes in AKT1(+/-)/AKT2(-/-) mice. Proteins with altered levels were identified with tandem mass spectrometry. Ingenuity Pathway Analysis was performed for the interpretation of the biological significance of the observed proteomic changes.

Results: 11 proteins were identified from 2 biological replicates to be differentially expressed by a ratio of at least 1.3 between age-matched insulin resistant (Akt1(+/-)/Akt2(-/-)) and wild type mice. Albumin and mitochondrial ornithine aminotransferase were detected from multiple spots, which suggest post-translational modifications. Enzymes of the urea cycle were common members of top regulated pathways.

Conclusion: Our results help to unveil the regulation of the liver proteome underlying altered metabolism in an animal model of severe insulin resistance.

Keywords: AKT; Insulin resistance; Liver; Mouse; Proteomics.

Figure 1. Analyses of body weights, random serum glucose levels, OGTT and liver histology

(A) Adult body weights of wild type and Akt1^+/-/Akt2^-/- mutant mice. (B) Random blood glucose levels of wild type and Akt1^+/-/Akt2^-/- mutant mice. (C) Blood glucose levels during OGTT of 20-week- old of wild type and Akt1^+/-/Akt2^-/- mutant mice. (D) Plasma insulin levels during OGTT of 20-week- old of wild type and Akt1^+/-/Akt2^-/- mutant mice. (E) Representative photomicrographs (×200) of liver sections stained with H&E from 20-week-old wild type and Akt1^+/-/Akt2^-/- mutant mice. All results are mean +/- SD. The markings indicate statistical significance in comparison to wild type mice at the same time point (*p<0.001, ^‡p<0.005).

Figure 2. Representative 2D-DIGE gel of liver proteins

The gel depicts proteins with an pI 3-7 further separated by molecular weight in a 12.5% polyacrylamide gel. Statistically significant, differentially expressed proteins from both replicates are circled on the above gel. Red circles indicate increased and blue circles indicate decreased normalized protein levels in AKT1^+/-/AKT2^-/- mice in comparison to wild type mice. Information on their identities and differential expression is in Table 1.

Figure 3. Classification of identified proteins and biological pathway analysis

(A) Cellular component localization of the 11 differentially expressed proteins identified as per UniProt. For each cellular component, the number of proteins in each class is given. (B) Enriched biological process networks by IPA software for the 11 differentially expressed proteins. The –log(p-value) for the enriched networks is as is per the height of the bars along the y-axis on the left. Upregulated pathways are colored yellow and downregulated pathways are colored blue. The ratio of the number of enriched proteins from our dataset in comparison to the total number of proteins within a pathway is indicated by the orange line and as per the y-axis on the right.