Mitochondria as Central Mediators of Cardiac Cell Death with Ischemia and Reperfusion Injury in Aged Female Rats
Open Access
- Author:
- Garvin, Alexandra M
- Graduate Program:
- Physiology
- Degree:
- Doctor of Philosophy
- Document Type:
- Dissertation
- Date of Defense:
- October 17, 2016
- Committee Members:
- Donna Hope Korzick, Dissertation Advisor/Co-Advisor
Donna Hope Korzick, Committee Chair/Co-Chair
Connie Jo Rogers, Committee Member
Joshua D Lambert, Committee Member
Gregory C Shearer, Outside Member
Chee Lim, Special Member - Keywords:
- heart
ischemia/reperfusion
mitochondria
aging
female
cell death - Abstract:
- Acute myocardial infarction (AMI) is exacerbated with both advancing age and estrogen (E2) loss in the female heart. However, the mechanistic underpinnings of age-related differences in cell death following ischemia/reperfusion (I/R) injury in females remains largely unexplored. This line of investigative research is critical if therapeutic targets to restore and preserve cardiac function in aged females are to be realized. Here, it was hypothesized that programmed necrosis (PN), in concert with altered mitochondrial respiration, morphology, and quality control, conspire to produce mitochondrial dysfunction and increased susceptibility to cell death upon an ischemic stimulus in the aged female heart. Given known reductions in cardioprotective reserve capacity with senescence, the role of PN inhibition as a pathway to protection was also employed and a novel model to simulate post-menopausal E2 deficiency in rodents established. The first study (Chapter 3) aimed to determine the relationship between mitochondrial ultrastructure and subpopulation function as a result of advancing age and I/R injury in the female heart. Fischer 344 female rats were ovariectomized (OVX) at 15 mo and studied at 22 mo (MO OVX) vs adult (6 mo). Hearts were subjected to in vivo coronary artery ligation (CAL) with 31or 55 min I and variable times of R (10 min, 6 hr, and 24 hr) to establish PN. Respiration rates of isolated mitochondria were reduced selectively by age and I/R in subsarcolemmal (SSM) and interfibrillar (IFM) mitochondria, respectively, leading to a significant reduction in left ventricular function (p<0.05). Mitochondrial morphology assessed by transmission electron microscopy (TEM) suggests an age-dependent alteration in the time course of autophagic flux. Toward a mechanism of protection and therapeutic intervention in the E2-deficient female heart, the aim of the second study (Chapter 4) was to determine the efficacy of PN inhibition on infarct size reduction and preservation of LV function following I/R injury. Necrostatin-1 (nec-1; 3.5 mg/kg or 5.7 mg/kg), when delivered upon R, significantly reduced infarct size by 34% and improved LV function following CAL (55 min I/2 or 6 hr R) in MO OVX. While age-associated elevations in cyclophilin D (CypD) and mitochondrial acetylation (p<0.001) were unaffected by nec-1, profound reductions in serum cytokines (p<0.05) and cardiac immune cell infiltration were observed in MO OVX but not adult, suggesting a distinct protective mechanism involving RIP1 signaling and post-I/R inflammatory responses. In the third study (Chapter 5), mitochondrial targets of ethanol (EtOH)-induced cardiac dysfunction were determined in adult and aged rats. Chronic EtOH consumption is known to predispose the aged female heart to the development of alcoholic cardiomyopathy, and as such served as an additional model system in which to evaluate age-related changes in cardiac mitochondrial adaptive responses to a superimposed stress. Mitochondrial proteomic changes were evaluated using isobaric tags for relative and absolute quantification (iTRAQ) 8plex labeling and mass spectrometry. Adult, adult OVX, aged, and aged OVX rats were fed a control or Lieber-DeCarli ‘all liquid’ EtOH diet (36% of total kcals) for 20 weeks. After EtOH, significant differences were observed in proteins involved in the electron transport chain (ETC), lipid metabolism, and cellular defense, suggesting a possible link to congestive heart failure. Additionally, EtOH significantly reduced SSM state 3 respiration in all groups, yet only reduced respiratory control index (RCI) in the aged OVX rat heart (p<0.05). In summary, key findings are as follows. 1) A sex-specific mitochondrial phenotype was identified whereby it is hypothesized that reductions in both SSM and IFM function may play an additive role in the enhanced susceptibility to I/R injury and myocardial infarction in aged females. Moreover, novel insight into altered mitochondrial quality control with age highlights a potentially important regulatory role of mitochondrial dynamics on sustaining respiratory function in the aged female heart. 2) PN was identified as the dominant cell death mechanism in the aged female heart. PN inhibition was found to be cardioprotective in adult and aged females through distinct cellular mechanisms. Reductions in RIP1-mediated pro-inflammatory cytokines were specific to MO OVX. 3) Chronic EtOH consumption exacerbates cardiac dysfunction in E2-deficient hearts. Furthermore, the combination of mitochondrial protein alterations and reduced mitochondrial function in the aged OVX suggest a possible link to congestive heart failure, whereas the adult heart was virtually unaffected by EtOH-induced changes in the mitochondrial proteome. Collectively, these data emphasize the vulnerability of the aged, E2-deficient female heart to cardiac injury, and characterize distinctive cell death mechanisms that ultimately rely on acute or chronic mitochondrial dysfunction in the adult and aged, respectively.