Changes in cardiac mitochondrial proteome of lean Ossabaw minipigs are associated with reduced mitochondrial function prior the manifestation of metabolic syndrome – compared with Göttingen minipigs

Background: Ossabaw minipigs differ from Göttingen minipigs in their unique genetic predisposition to develop a full metabolic syndrome and their non-responsiveness to cardioprotective interventions, even before they develop their diseased phenotype. Previous DNA sequencing data revealed differences in a cluster of mitochondrial genes between the two pig strains. Alterations in mitochondrial protein composition affect their function: Krebs cycle, oxidative phosphorylation, mitochondrial biogenesis. All of them play a crucial role in the development of metabolic syndrome as well as in myocardial ischemia-reperfusion injury and cardioprotection.

Aim: Comparison of the cardiac mitochondrial proteome and mitochondrial function of adult lean Ossabaw minipigs with that of adult lean Göttingen minipigs.

Methods and Results: Mitochondria were isolated by differential centrifugation from cardiac left ventricular tissue samples of both pig strains (n=4, each). The mitochondrial proteome was analyzed by LC-MS/MS. Proteins were classified corresponding to their mitochondrial localization and intra-mitochondrial function according to MitoCarta3.0 and Beyond MitoCarta. A total of 810 proteins were identified in at least three of four samples, 654 proteins were classified as mitochondrial proteins. Among them, 22 proteins were differentially expressed between both pig strains (see Figure). Most of them were classified as related to mitochondrial metabolism (9 higher/7 lower expressed in Ossabaw minipigs), four (two higher/two lower expressed) as related to mitochondrial transcription and translation, and three as related to small molecule transport (one higher/two lower expressed), two as related to oxidative phosphorylation (one higher/one lower expressed), and one as related to dynamics and surveillance (lower expressed; see Figure). Since these differences are directly and indirectly related to mitochondrial oxidative phosphorylation and ATP production, we compared these parameters between the two pig strains (n=26 Ossabaw minipigs, n=13 Göttingen minipigs). The respiratory control ratio (state 3/state 4 respiration) was reduced in Ossabaw minipigs compared to Göttingen minipigs (11.6±3.3 vs. 14.5±5.3, p<0.05), which was reflected in a decreased ATP production (38±27 vs. 75±17 µmol ATP/mg protein, p<0.001).

Conclusion: The mitochondrial proteome is largely comparable between Ossabaw and Göttingen minipigs. Proteins that are differentially expressed are related to mitochondrial metabolism, transcription and translation, small molecule transport, oxidative phosphorylation and dynamics and surveillance. Indeed, these changes are associated with a reduced respiratory control ratio and ATP production in Ossabaw minipig mitochondria compared to Göttingen minipig mitochondria. In a next step, the results of the proteomic approach need to be confirmed by Western blotting and their causal role in mitochondrial function needs to be established by the use of antagonists.