Novel Mitochondrial and Nuclear Targets of Acute PKCε Activation in the Aged Female Rat Heart

Open Access
Jefferson, Sarah Jane
Area of Honors:
Bachelor of Science
Document Type:
Thesis Supervisors:
  • Donna Hope Korzick, Thesis Supervisor
  • Stephen Jacob Piazza, Honors Advisor
  • Craig Eugene Cameron, Faculty Reader
  • mitochondria
  • PKC
  • heart
  • menopause
Mortality due to myocardial infarction (MI) increases as women experience menopause, implicating a protective role for estradiol (E2). However, studies have produced conflicting results on the effects of hormone replacement therapy (HRT) on occurrence of MI. This points to the need for development of therapeutic methods for reducing ischemia/reperfusion (I/R) injury in the aged, E2-deficient heart. This study focused on the cardioprotective effects of PKCε and its mechanism of action in the mitochondria as well as its effects on gene expression. Using a high-throughput proteomics approach, we aimed to characterize altered expression levels of mitochondrial proteins contributing to both reduced ischemic tolerance in the aged, E2-deficient female rat heart and cardioprotection in PKCε-activated hearts. We also aimed to characterize changes in mRNA level of two proteins involved in PKCε-mediated cardioprotection. A Langendorff-perfused heart model was utilized to measure functional recovery in adult (5 mo) and aged (23 mo) female F344 ovary-intact or ovariectomized (OVX) rats administered a PKCε-activator (ψεRACK) prior to 47 min ischemia and 60 min reperfusion. mRNA was isolated from the LV’s of these hearts and subjected to RT-PCR. Proteomic analysis was conducted on the LV mitochondrial fractions of hearts undergoing control perfusions with or without ψεRACK, utilizing isobaric tags for relative and absolute quantitation (iTRAQ) 8-plex labeling and tandem mass spectrometry. The aged OVX hearts showed reduced functional recovery following I/R when compared with the adult ovary-intact hearts. Administration of ψεRACK prior to I/R increased functional recovery, measured by recovery of LVDP, in both adult intact and aged OVX groups. Proteomic analysis revealed three possible targets of PKCε cardioprotective signaling, the antioxidant enzymes glutathione peroxidase and superoxide dismutase 2, as well as the heat shock protein Hsp 10. Finally, decreased levels of Cx43 and RACK2 mRNA were seen with age, while administration of ψεRACK prior to I/R at least partially abrogated this effect. These results indicate a protective role for PKCε in the aged, E2-deficient heart and identify novel nuclear and mitochondrial targets of acute PKCε activation.