Defective dimerization of FoF1-ATP synthase secondary to glycation favors mitochondrial energy deficiency in cardiomyocytes during aging.

Aged cardiomyocytes develop a mismatch between energy demand and supply, the severity of which determines the onset of heart failure, and become prone to undergo cell death. The FoF1-ATP synthase is the molecular machine that provides >90% of the ATP consumed by healthy cardiomyocytes and is proposed to form the mitochondrial permeability transition pore (mPTP), an energy-dissipating channel involved in cell death. We investigated whether aging alters FoF1-ATP synthase self-assembly, a fundamental biological process involved in mitochondrial cristae morphology and energy efficiency, and the functional consequences this may have. Purified heart mitochondria and cardiomyocytes from aging mice displayed an impaired dimerization of FoF1-ATP synthase (blue native and proximity ligation assay), associated with abnormal mitochondrial cristae tip curvature (TEM). Defective dimerization did not modify the in vitro hydrolase activity of FoF1-ATP synthase but reduced the efficiency of oxidative phosphorylation in intact mitochondria (in which membrane architecture plays a fundamental role) and increased cardiomyocytes' susceptibility to undergo energy collapse by mPTP. High throughput proteomics and fluorescence immunolabeling identified glycation of 5 subunits of FoF1-ATP synthase as the causative mechanism of the altered dimerization. In vitro induction of FoF1-ATP synthase glycation in H9c2 myoblasts recapitulated the age-related defective FoF1-ATP synthase assembly, reduced the relative contribution of oxidative phosphorylation to cell energy metabolism, and increased mPTP susceptibility. These results identify altered dimerization of FoF1-ATP synthase secondary to enzyme glycation as a novel pathophysiological mechanism involved in mitochondrial cristae remodeling, energy deficiency, and increased vulnerability of cardiomyocytes to undergo mitochondrial failure during aging.

Mitochondrial ROS and mitochondria-targeted antioxidants in the aged heart.

Excessive mitochondrial ROS production has been causally linked to the pathophysiology of aging in the heart and other organs, and plays a deleterious role in several age-related cardiac pathologies, including myocardial ischemia-reperfusion injury and heart failure, the two worldwide leading causes of death and disability in the elderly. However, ROS generation is also a fundamental mitochondrial function that orchestrates several signaling pathways, some of them exerting cardioprotective effects. In cardiac myocytes, mitochondria are particularly abundant and are specialized in subcellular populations, in part determined by their relationships with other organelles and their cyclic calcium handling activity necessary for adequate myocardial contraction/relaxation and redox balance. Depending on their subcellular location, mitochondria can themselves be differentially targeted by ROS and display distinct age-dependent functional decline. Thus, precise mitochondria-targeted therapies aimed at counteracting unregulated ROS production are expected to have therapeutic benefits in certain aging-related heart conditions. However, for an adequate design of such therapies, it is necessary to unravel the complex and dynamic interactions between mitochondria and other cellular processes.

Cardiomyocyte ageing and cardioprotection: consensus document from the ESC working groups cell biology of the heart and myocardial function.

Advanced age is a major predisposing risk factor for the incidence of coronary syndromes and comorbid conditions which impact the heart response to cardioprotective interventions. Advanced age also significantly increases the risk of developing post-ischaemic adverse remodelling and heart failure after ischaemia/reperfusion (IR) injury. Some of the signalling pathways become defective or attenuated during ageing, whereas others with well-known detrimental consequences, such as glycoxidation or proinflammatory pathways, are exacerbated. The causative mechanisms responsible for all these changes are yet to be elucidated and are a matter of active research. Here, we review the current knowledge about the pathophysiology of cardiac ageing that eventually impacts on the increased susceptibility of cells to IR injury and can affect the efficiency of cardioprotective strategies.

Enhancing Glycolysis Protects against Ischemia-Reperfusion Injury by Reducing ROS Production.

After myocardial ischemia-reperfusion, fatty acid oxidation shows fast recovery while glucose oxidation rates remain depressed. A metabolic shift aimed at increasing glucose oxidation has shown to be beneficial in models of myocardial ischemia-reperfusion. However, strategies aimed at increasing glucose consumption in the clinic have provided mixed results and have not yet reached routine clinical practice. A better understanding of the mechanisms underlying the protection afforded by increased glucose oxidation may facilitate the transfer to the clinic. The purpose of this study was to evaluate if the modulation of reactive oxygen species (ROS) was involved in the protection afforded by increased glucose oxidation. Firstly, we characterized an H9C2 cellular model in which the use of glucose or galactose as substrates can modulate glycolysis and oxidative phosphorylation pathways. In this model, there were no differences in morphology, cell number, or ATP and PCr levels. However, galactose-grown cells consumed more oxygen and had an increased Krebs cycle turnover, while cells grown in glucose had increased aerobic glycolysis rate as demonstrated by higher lactate and alanine production. Increased aerobic glycolysis was associated with reduced ROS levels and protected the cells against simulated ischemia-reperfusion injury. Furthermore, ROS scavenger N-acetyl cysteine (NAC) was able to reduce the amount of ROS and to prevent cell death. Lastly, cells grown in galactose showed higher activation of mTOR/Akt signaling pathways. In conclusion, our results provide evidence indicating that metabolic shift towards increased glycolysis reduces mitochondrial ROS production and prevents cell death during ischemia-reperfusion injury.

Cyclophilins A and B oppositely regulate renal tubular epithelial cell phenotype.

Restoration of kidney tubular epithelium following sublethal injury sequentially involves partial epithelial-mesenchymal transition (pEMT), proliferation, and further redifferentiation into specialized tubule epithelial cells (TECs). Because the immunosuppressant cyclosporine-A produces pEMT in TECs and inhibits the peptidyl-prolyl isomerase (PPIase) activity of cyclophilin (Cyp) proteins, we hypothesized that cyclophilins could regulate TEC phenotype. Here we demonstrate that in cultured TECs, CypA silencing triggers loss of epithelial features and enhances transforming growth factor ß (TGFß)-induced EMT in association with upregulation of epithelial repressors Slug and Snail. This pro-epithelial action of CypA relies on its PPIase activity. By contrast, CypB emerges as an epithelial repressor, because CypB silencing promotes epithelial differentiation, prevents TGFß-induced EMT, and induces tubular structures in 3D cultures. In addition, in the kidneys of CypB knockout mice subjected to unilateral ureteral obstruction, inflammatory and pro-fibrotic events were attenuated. CypB silencing/knockout leads to Slug, but not Snail, downregulation. CypB support of Slug expression depends on its endoplasmic reticulum location, where it interacts with calreticulin, a calcium-buffering chaperone related to Slug expression. As CypB silencing reduces ionomycin-induced calcium release and Slug upregulation, we suggest that Slug expression may rely on CypB modulation of calreticulin-dependent calcium signaling. In conclusion, this work uncovers new roles for CypA and CypB in modulating TEC plasticity and identifies CypB as a druggable target potentially relevant in promoting kidney repair.

Ryanodine Receptor Glycation Favors Mitochondrial Damage in the Senescent Heart.

BACKGROUND: Senescent cardiomyocytes exhibit a mismatch between energy demand and supply that facilitates their transition toward failing cells. Altered calcium transfer from sarcoplasmic reticulum (SR) to mitochondria has been causally linked to the pathophysiology of aging and heart failure. METHODS: Because advanced glycation-end products accumulate throughout life, we investigated whether intracellular glycation occurs in aged cardiomyocytes and its impact on SR and mitochondria. RESULTS: Quantitative proteomics, Western blot and immunofluorescence demonstrated a significant increase in advanced glycation-end product-modified proteins in the myocardium of old mice (≥20months) compared with young ones (4-6months). Glyoxalase-1 activity (responsible for detoxification of dicarbonyl intermediates) and its cofactor glutathione were decreased in aged hearts. Immunolabeling and proximity ligation assay identified the ryanodine receptor (RyR2) in the SR as prominent target of glycation in aged mice, and the sites of glycation were characterized by quantitative mass spectrometry. RyR2 glycation was associated with more pronounced calcium leak, determined by confocal microscopy in cardiomyocytes and SR vesicles. Interfibrillar mitochondria-directly exposed to SR calcium release-from aged mice had increased calcium content compared with those from young ones. Higher levels of advanced glycation-end products and reduced glyoxalase-1 activity and glutathione were also present in atrial appendages from surgical patients ≥75 years as compared with the younger ones. Elderly patients also exhibited RyR2 hyperglycation and increased mitochondrial calcium content that was associated with reduced myocardial aerobic capacity (mitochondrial O2 consumption/g) attributable to less respiring mitochondria. In contracting HL-1 cardiomyocytes, pharmacological glyoxalase-1 inhibition recapitulated RyR2 glycation and defective SR-mitochondria calcium exchange of aging. CONCLUSIONS: Mitochondria from aging hearts develop calcium overload secondary to SR calcium leak. Glycative damage of RyR2, favored by deficient dicarbonyl detoxification capacity, contributes to calcium leak and mitochondrial damage in the senescent myocardium.

Evolution and bad prognostic value of advanced glycation end products after acute heart failure: relation with body composition.

AIM: The role of advanced glycation end products (AGEs) and their soluble receptor (sRAGE) on the progression and prognosis of acute heart failure (HF) was analysed in relation with metabolic parameters as body composition and nutritional status. METHODS: A hundred and fifty consecutive patients were included in a prospective clinical study during hospitalization by acute HF. Detailed medical history, physical examination, electrocardiogram, echocardiogram and vein peripheral blood were taken for all patients. During the follow-up period [297 days (88-422 days)] blood samples for biochemical measurements were obtained 1 and 6 months after the inclusion. Dual-energy X-ray absorptiometry analyses were performed 1 week after discharge. RESULTS: AGEs and sRAGE levels continuously increased, up to 6 months, after acute HF, but AGEs increase was mainly observed in those patients with incident HF. Both AGEs and sRAGE levels were related with bad renal function and clinical malnutrition (CONUT score) and they were negatively related with body mass index or percentage of body fat. AGEs levels (≥40 a.u.) 1 month after discharge and basal sRAGE levels (>1000 pg/mL) were related with worse prognosis in terms of patient death and HF readmission (Log-rank <0.05 in Kaplan-Meier survival test), independently of age, gender, body mass index and other risk factors. Regression models also corroborated this finding. CONCLUSIONS: AGEs and sRAGE are bad prognostic biomarkers for HF and useful markers of HF progression. Since their levels seem to be related with clinical malnutrition and body composition these parameters could serve to modulate them.