Lipid (per) oxidation in mitochondria:an emerging target in the ageing process?

Lipids are essential for physiological processes such as maintaining membrane integrity, providing a source of energy and acting as signalling molecules to control processes including cell proliferation, metabolism, inflammation and apoptosis. Disruption of lipid homeostasis can promote pathological changes that contribute towards biological ageing and age-related diseases. Several age-related diseases have been associated with altered lipid metabolism and an elevation in highly damaging lipid peroxidation products; the latter has been ascribed, at least in part, to mitochondrial dysfunction and elevated ROS formation. In addition, senescent cells, which are known to contribute significantly to age-related pathologies, are also associated with impaired mitochondrial function and changes in lipid metabolism. Therapeutic targeting of dysfunctional mitochondrial and pathological lipid metabolism is an emerging strategy for alleviating their negative impact during ageing and the progression to age-related diseases. Such therapies could include the use of drugs that prevent mitochondrial uncoupling, inhibit inflammatory lipid synthesis, modulate lipid transport or storage, reduce mitochondrial oxidative stress and eliminate senescent cells from tissues. In this review, we provide an overview of lipid structure and function, with emphasis on mitochondrial lipids and their potential for therapeutic targeting during ageing and age-related disease

Age Related Bioenergetics Profiles in Isolated Rat Cardiomyocytes Using Extracellular Flux Analyses.

Mitochondrial dysfunction is increasingly recognized and studied as a mediator of heart disease. Extracellular flux analysis (XF) has emerged as a powerful tool to investigate cellular bioenergetics in the context of cardiac health and disease, however its use and interpretation requires improved understanding of the normal metabolic differences in cardiomyocytes (CM) at various stages of maturation. This study standardized XF analyses methods (mitochondrial stress test, glycolytic stress test and palmitate oxidation test) and established age related differences in bioenergetics profiles of healthy CMs at newborn (NB1), weaning (3WK), adult (10WK) and aged (12-18MO) time points. Findings show that immature CMs demonstrate a more robust and sustained glycolytic capacity and a relative inability to oxidize fatty acids when compared to older CMs. The study also highlights the need to recognize the contribution of CO2 from the Krebs cycle as well as lactate from anaerobic glycolysis to the proton production rate before interpreting glycolytic capacity in CMs. Overall, this study demonstrates that caution should be taken to assure that translatable developmental time points are used to investigate mitochondrial dysfunction as a cause of cardiac disease. Specifically, XF analysis of newborn CMs should be reserved to study fetal/neonatal disease and older CMs (≥10 weeks) should be used to investigate adult disease pathogenesis. Knowledge gained will aid in improved investigation of developmentally programmed heart disease and stress the importance of discerning maturational differences in bioenergetics when developing mitochondrial targeted preventative and therapeutic strategies for cardiac disease

Mitochondrial stress test of cardiomyocytes at various maturational ages.

<p>XF trace (A) and bar graph (B) illustrates OCR changes (from baseline) in NB1, 3WK, 10WK, and 12–18MNTH CMs during mitochondrial stress testing. For group comparisons OCR was normalized to baseline prior to addition of Oligomycin, FCCP and Rotenone/Antimycin A where indicated. Respiratory control ratio (C) and coupling efficiency (D) as calculated from traces are compared between groups. Data are expressed as means ± SEM of 4–7 independent experiments. *p<0.05 by one-way ANOVA and Tukey’s post-test.</p

Palmitate oxidation test of cardiomyocytes at various maturational ages.

<p>OCR was measured in NB1, 3WK, 10WK, and 12–18MNTH CMs and normalized to baseline rates to interrogate differences in response among the groups. OCR response to exogenous Palmitate-BSA and Etomoxir (carnitine palmitoyl transport inhibitor) are illustrated in trace (A). Bar graphs illustrate group differences in OCR response from baseline conditions (B). Data are expressed as means ± SEM of 3–4 independent experiments. *p<0.05 by one-way ANOVA and Tukey’s post-test.</p

Proton production rate during palmitate oxidation test.

<p>The positive or notionally negative contribution of lactate from both anaerobic glycolysis (grey) and from CO₂/respiration (white) to the sum (black) proton production rate (PPR) are demonstrated for cardiomyocytes from each maturational time-point. The PPR is demonstrated in response to palmitate (A) and etomoxir (B). Data shown are means ± SEM of 3–5 independent experiments. *p<0.05 by one way ANOVA and Tukey’s post-test.</p

Glycolysis stress test of cardiomyocytes at various maturational ages.

<p>Extra-cellular acidification rate (ECAR) traces (A) and bar graphs (B) show the glycolytic response of NB1, 3WK, 10WK and 12–18MTH old cardiomyocytes in response to glucose and oligomycin injection where indicated. Data shown are means ± SEM of 3–5 independent experiments. *p<0.05 by one way ANOVA and Tukey’s post-test.</p

XF Analyses of Isolated Cardiomyocytes.

<p>Examples of real-time trace oxygen consumption rate (OCR) and extracellular acidification rate (ECAR) during extracellular flux analyses using (A) mitochondrial stress test, (B) glycolytic stress test and (C) palmitate oxidation test. Data points represent mean of three readings taken at each time-point. Drug injections are shown for each test and boxes visually depict computed data.</p

Proton production rate during glycolytic stress test.

<p>The positive or notionally negative contribution of lactate from both anaerobic glycolysis (grey) and from CO₂/respiration (white) to the sum (black) proton production rate (PPR) are demonstrated for cardiomyocytes from each maturational time-point. The PPR is demonstrated in response to glucose (A) and oligomycin (B). Data shown are means ± SEM of 3–5 independent experiments. *p<0.05 by one way ANOVA and Tukey’s post-test.</p