Calcium signaling from sarcoplasmic reticulum and mitochondria contact sites in acute myocardial infarction.

Acute myocardial infarction (AMI) is a serious condition that occurs when part of the heart is subjected to ischemia episodes, following partial or complete occlusion of the epicardial coronary arteries. The resulting damage to heart muscle cells have a significant impact on patient's health and quality of life. About that, recent research focused on the role of the sarcoplasmic reticulum (SR) and mitochondria in the physiopathology of AMI. Moreover, SR and mitochondria get in touch each other through multiple membrane contact sites giving rise to the subcellular region called mitochondria-associated membranes (MAMs). MAMs are essential for, but not limited to, bioenergetics and cell fate. Disruption of the architecture of these regions occurs during AMI although it is still unclear the cause-consequence connection and a complete overview of the pathological changes; for sure this concurs to further damage to heart muscle. The calcium ion (Ca2+) plays a pivotal role in the pathophysiology of AMI and its dynamic signaling between the SR and mitochondria holds significant importance. In this review, we tried to summarize and update the knowledge about the roles of these organelles in AMI from a Ca2+ signaling point of view. Accordingly, we also reported some possible cardioprotective targets which are directly or indirectly related at limiting the dysfunctions caused by the deregulation of the Ca2+ signaling.

Comprehensive Analysis of Mitochondrial Dynamics Alterations in Heart Diseases.

The most common alterations affecting mitochondria, and associated with cardiac pathological conditions, implicate a long list of defects. They include impairments of the mitochondrial electron transport chain activity, which is a crucial element for energy formation, and that determines the depletion of ATP generation and supply to metabolic switches, enhanced ROS generation, inflammation, as well as the dysregulation of the intracellular calcium homeostasis. All these signatures significantly concur in the impairment of cardiac electrical characteristics, loss of myocyte contractility and cardiomyocyte damage found in cardiac diseases. Mitochondrial dynamics, one of the quality control mechanisms at the basis of mitochondrial fitness, also result in being dysregulated, but the use of this knowledge for translational and therapeutic purposes is still in its infancy. In this review we tried to understand why this is, by summarizing methods, current opinions and molecular details underlying mitochondrial dynamics in cardiac diseases.

The Complex Relationship between Hypoxia Signaling, Mitochondrial Dysfunction and Inflammation in Calcific Aortic Valve Disease: Insights from the Molecular Mechanisms to Therapeutic Approaches.

Calcific aortic valve stenosis (CAVS) is among the most common causes of cardiovascular mortality in an aging population worldwide. The pathomechanisms of CAVS are such a complex and multifactorial process that researchers are still making progress to understand its physiopathology as well as the complex players involved in CAVS pathogenesis. Currently, there is no successful and effective treatment to prevent or slow down the disease. Surgical and transcatheter valve replacement represents the only option available for treating CAVS. Insufficient oxygen availability (hypoxia) has a critical role in the pathogenesis of almost all CVDs. This process is orchestrated by the hallmark transcription factor, hypoxia-inducible factor 1 alpha subunit (HIF-1α), which plays a pivotal role in regulating various target hypoxic genes and metabolic adaptations. Recent studies have shown a great deal of interest in understanding the contribution of HIF-1α in the pathogenesis of CAVS. However, it is deeply intertwined with other major contributors, including sustained inflammation and mitochondrial impairments, which are attributed primarily to CAVS. The present review aims to cover the latest understanding of the complex interplay effect of hypoxia signaling pathways, mitochondrial dysfunction, and inflammation in CAVS. We propose further hypotheses and interconnections on the complexity of these impacts in a perspective of better understanding the pathophysiology. These interplays will be examined considering recent studies that shall help us better dissect the molecular mechanism to enable the design and development of potential future therapeutic approaches that can prevent or slow down CAVS processes.

1,3,8-Triazaspiro[4.5]decane Derivatives Inhibit Permeability Transition Pores through a FO-ATP Synthase c Subunit Glu119-Independent Mechanism That Prevents Oligomycin A-Related Side Effects.

Permeability transition pore (PTP) molecular composition and activity modulation have been a matter of research for several years, especially due to their importance in ischemia reperfusion injury (IRI). Notably, c subunit of ATP synthase (Csub) has been identified as one of the PTP-forming proteins and as a target for cardioprotection. Oligomycin A is a well-known Csub interactor that has been chemically modified in-depth for proposed new pharmacological approaches against cardiac reperfusion injury. Indeed, by taking advantage of its scaffold and through focused chemical improvements, innovative Csub-dependent PTP inhibitors (1,3,8-Triazaspiro[4.5]decane) have been synthetized in the past. Interestingly, four critical amino acids have been found to be involved in Oligomycin A-Csub binding in yeast. However, their position on the human sequence is unknown, as is their function in PTP inhibition. The aims of this study are to (i) identify for the first time the topologically equivalent residues in the human Csub sequence; (ii) provide their in vitro validation in Oligomycin A-mediated PTP inhibition and (iii) understand their relevance in the binding of 1,3,8-Triazaspiro[4.5]decane small molecules, as Oligomycin A derivatives, in order to provide insights into Csub interactions. Notably, in this study we demonstrated that 1,3,8-Triazaspiro[4.5]decane derivatives inhibit permeability transition pores through a FO-ATP synthase c subunit Glu119-independent mechanism that prevents Oligomycin A-related side effects.

Interorganellar calcium signaling in the regulation of cell metabolism: A cancer perspective.

Organelles were originally considered to be individual cellular compartments with a defined organization and function. However, recent studies revealed that organelles deeply communicate within each other via Ca2+ exchange. This communication, mediated by specialized membrane regions in close apposition between two organelles, regulate cellular functions, including metabolism and cell fate decisions. Advances in microscopy techniques, molecular biology and biochemistry have increased our understanding of these interorganelle platforms. Research findings suggest that interorganellar Ca2+ signaling, which is altered in cancer, influences tumorigenesis and tumor progression by controlling cell death programs and metabolism. Here, we summarize the available data on the existence and composition of interorganelle platforms connecting the endoplasmic reticulum with mitochondria, the plasma membrane, or endolysosomes. Finally, we provide a timely overview of the potential function of interorganellar Ca2+ signaling in maintaining cellular homeostasis.

Identification of small-molecule urea derivatives as PTPC modulators targeting the c subunit of F1/Fo-ATP synthase.

Maintaining a high percentage of living and functional cells in those pathologies in which excessive cell death occurs, such as neurodegenerative disorders and cardiovascular diseases, is one of the most intriguing challenges in the field of biochemical research for drug discovery. Here, mitochondrial permeability transition-driven regulated cell death is the main mechanism of mitochondrial impairment and cell fate; this pathway is still lacking of satisfying pharmacological treatments to counteract its becoming; for this reason, it needs continuous and intense research to find new compounds as modulator of the permeability transition pore complex (PTPC) activity. In this study, we report the identification of small-molecule urea derivatives able to inhibit PTPC opening following calcium overload and selected for future use in cytoprotection.

Aortic Valve Stenosis and Mitochondrial Dysfunctions: Clinical and Molecular Perspectives.

Calcific aortic stenosis is a disorder that impacts the physiology of heart valves. Fibrocalcific events progress in conjunction with thickening of the valve leaflets. Over the years, these events promote stenosis and obstruction of blood flow. Known and common risk factors are congenital defects, aging and metabolic syndromes linked to high plasma levels of lipoproteins. Inflammation and oxidative stress are the main molecular mediators of the evolution of aortic stenosis in patients and these mediators regulate both the degradation and remodeling processes. Mitochondrial dysfunction and dysregulation of autophagy also contribute to the disease. A better understanding of these cellular impairments might help to develop new ways to treat patients since, at the moment, there is no effective medical treatment to diminish neither the advancement of valve stenosis nor the left ventricular function impairments, and the current approaches are surgical treatment or transcatheter aortic valve replacement with prosthesis.

Mitochondrial permeability transition involves dissociation of F1FO ATP synthase dimers and C-ring conformation.

The impact of the mitochondrial permeability transition (MPT) on cellular physiology is well characterized. In contrast, the composition and mode of action of the permeability transition pore complex (PTPC), the supramolecular entity that initiates MPT, remain to be elucidated. Specifically, the precise contribution of the mitochondrial F1FO ATP synthase (or subunits thereof) to MPT is a matter of debate. We demonstrate that F1FO ATP synthase dimers dissociate as the PTPC opens upon MPT induction. Stabilizing F1FO ATP synthase dimers by genetic approaches inhibits PTPC opening and MPT Specific mutations in the F1FO ATP synthase c subunit that alter C-ring conformation sensitize cells to MPT induction, which can be reverted by stabilizing F1FO ATP synthase dimers. Destabilizing F1FO ATP synthase dimers fails to trigger PTPC opening in the presence of mutants of the c subunit that inhibit MPT The current study does not provide direct evidence that the C-ring is the long-sought pore-forming subunit of the PTPC, but reveals that PTPC opening requires the dissociation of F1FO ATP synthase dimers and involves the C-ring.

Intersection of mitochondrial fission and fusion machinery with apoptotic pathways: Role of Mcl-1.

Mitochondria actively contribute to apoptotic cell death through mechanisms including the loss of integrity of the outer mitochondrial membrane, the release of intermembrane space proteins, such as cytochrome c, in the cytosol and the caspase cascade activation. This process is the result of careful cooperation not only among members of the Bcl-2 family but also dynamin-related proteins. These events are often accompanied by fission of the organelle, thus linking mitochondrial dynamics to apoptosis. Emerging evidences are suggesting a fine regulation of mitochondrial morphology by Bcl-2 family members and active participation of fission-fusion proteins in apoptosis. The debate whether in mitochondrial morphogenesis the role of Bcl-2 family members is functionally distinct from their role in apoptosis is still open and, above all, which morphological changes are associated with cell death sensitisation. This review will cover the findings on how the mitochondrial fission and fusion machinery may intersect apoptotic pathways focusing on recent advances on the key role played by Mcl-1.

Mitochondrial dysfunction and skeletal muscle atrophy: Causes, mechanisms, and treatment strategies.

Skeletal muscle, which accounts for approximately 40% of total body weight, is one of the most dynamic and plastic tissues in the human body and plays a vital role in movement, posture and force production. More than just a component of the locomotor system, skeletal muscle functions as an endocrine organ capable of producing and secreting hundreds of bioactive molecules. Therefore, maintaining healthy skeletal muscles is crucial for supporting overall body health. Various pathological conditions, such as prolonged immobilization, cachexia, aging, drug-induced toxicity, and cardiovascular diseases (CVDs), can disrupt the balance between muscle protein synthesis and degradation, leading to skeletal muscle atrophy. Mitochondrial dysfunction is a major contributing mechanism to skeletal muscle atrophy, as it plays crucial roles in various biological processes, including energy production, metabolic flexibility, maintenance of redox homeostasis, and regulation of apoptosis. In this review, we critically examine recent knowledge regarding the causes of muscle atrophy (disuse, cachexia, aging, etc.) and its contribution to CVDs. Additionally, we highlight the mitochondrial signaling pathways involvement to skeletal muscle atrophy, such as the ubiquitin-proteasome system, autophagy and mitophagy, mitochondrial fission-fusion, and mitochondrial biogenesis. Furthermore, we discuss current strategies, including exercise, mitochondria-targeted antioxidants, in vivo transfection of PGC-1α, and the potential use of mitochondrial transplantation as a possible therapeutic approach.

Perspectives on mitochondrial relevance in cardiac ischemia/reperfusion injury.

Cardiovascular disease is the most common cause of death worldwide and in particular, ischemic heart disease holds the most considerable position. Even if it has been deeply studied, myocardial ischemia-reperfusion injury (IRI) is still a side-effect of the clinical treatment for several heart diseases: ischemia process itself leads to temporary damage to heart tissue and obviously the recovery of blood flow is promptly required even if it worsens the ischemic injury. There is no doubt that mitochondria play a key role in pathogenesis of IRI: dysfunctions of these important organelles alter cell homeostasis and survival. It has been demonstrated that during IRI the system of mitochondrial quality control undergoes alterations with the disruption of the complex balance between the processes of mitochondrial fusion, fission, biogenesis and mitophagy. The fundamental role of mitochondria is carried out thanks to the finely regulated connection to other organelles such as plasma membrane, endoplasmic reticulum and nucleus, therefore impairments of these inter-organelle communications exacerbate IRI. This review pointed to enhance the importance of the mitochondrial network in the pathogenesis of IRI with the aim to focus on potential mitochondria-targeting therapies as new approach to control heart tissue damage after ischemia and reperfusion process.

Some Insights into the Regulation of Cardiac Physiology and Pathology by the Hippo Pathway.

The heart is one of the most fascinating organs in living beings. It beats up to 100,000 times a day throughout the lifespan, without resting. The heart undergoes profound anatomical, biochemical, and functional changes during life, from hypoxemic fetal stages to a completely differentiated four-chambered cardiac muscle. In the middle, many biological events occur after and intersect with each other to regulate development, organ size, and, in some cases, regeneration. Several studies have defined the essential roles of the Hippo pathway in heart physiology through the regulation of apoptosis, autophagy, cell proliferation, and differentiation. This molecular route is composed of multiple components, some of which were recently discovered, and is highly interconnected with multiple known prosurvival pathways. The Hippo cascade is evolutionarily conserved among species, and in addition to its regulatory roles, it is involved in disease by drastically changing the heart phenotype and its function when its components are mutated, absent, or constitutively activated. In this review, we report some insights into the regulation of cardiac physiology and pathology by the Hippo pathway.

Impairment of mitophagy and autophagy accompanies calcific aortic valve stenosis favouring cell death and the severity of disease.

AIMS: In the last 15 years, some observations tried to shed light on the dysregulation of the cellular self-digestion process in calcific aortic valve stenosis (CAVS), but the results obtained remain still controversial. This work is aimed to definitively establish the trend of autophagy in patients affected by CAVS, to analyse the putative involvement of other determinants, which impact on the mitochondrial quality control mechanisms and to explore possible avenues for pharmacological interventions in the treatment of CAVS. METHODS AND RESULTS: This observational study, performed exclusively in ex vivo human samples (cells and serum), by using biochemical approaches and correlations with clinical data, describes new biological features of the calcified valve in terms of mitochondrial dysfunctions. In detail, we unveiled a significant deficiency in mitochondrial respiration and in ATP production coupled to increase production of lactates. In addition, mitochondrial population in the pathologic group is aged with significant alterations in biogenesis and mitophagy pathways. We are also reporting an updated view about autophagy accompanying the calcification process and advanced stages of the disease. We provided evidence for a rapamycin-based therapeutic strategy to revert the calcified phenotype to the wild type one. CONCLUSION: Our data suggest that the CAVS phenotype is featured by defects in mitochondrial quality control mechanisms and that autophagy is not activated enough to counteract cell death and sustain cell functions. Thus, boosting autophagy and mitophagy from short- to long-term reverts quite all pathological phenotypes.

A naturally occurring mutation in ATP synthase subunit c is associated with increased damage following hypoxia/reoxygenation in STEMI patients.

Preclinical models of ischemia/reperfusion injury (RI) demonstrate the deleterious effects of permeability transition pore complex (PTPC) opening in the first minutes upon revascularization of the occluded vessel. The ATP synthase c subunit (Csub) influences PTPC activity in cells, thus impacting tissue injury. A conserved glycine-rich domain in Csub is classified as critical because, when mutated, it modifies ATP synthase properties, protein interaction with the mitochondrial calcium (Ca2+) uniporter complex, and the conductance of the PTPC. Here, we document the role of a naturally occurring mutation in the Csub-encoding ATP5G1 gene at the G87 position found in two ST-segment elevation myocardial infarction (STEMI) patients and how PTPC opening is related to RI in patients affected by the same disease. We report a link between the expression of ATP5G1G87E and the response to hypoxia/reoxygenation of human cardiomyocytes, which worsen when compared to those expressing the wild-type protein, and a positive correlation between PTPC and RI.

Methods to Monitor Mitophagy and Mitochondrial Quality: Implications in Cancer, Neurodegeneration, and Cardiovascular Diseases.

Mitochondria are dynamic organelles that participate in a broad array of molecular functions within the cell. They are responsible for maintaining the appropriate energetic levels and control the cellular homeostasis throughout the generation of intermediary metabolites. Preserving a healthy and functional mitochondrial population is of fundamental importance throughout the life of the cells under pathophysiological conditions. Hence, cells have evolved fine-tuned mechanisms of quality control that help to preserve the right amount of functional mitochondria to meet the demand of the cell. The specific recycling of mitochondria by autophagy, termed mitophagy, represents the primary contributor to mitochondrial quality control. During this process, damaged or unnecessary mitochondria are recognized and selectively degraded. In the past few years, the knowledge in mitophagy has seen rapid progress, and a growing body of evidence confirms that mitophagy holds a central role in controlling cellular functions and the progression of various human diseases.In this chapter, we will discuss the pathophysiological roles of mitophagy and provide a general overview of the current methods used to monitor and quantify mitophagy. We will also outline the main established approaches to investigate the mitochondrial function, metabolism, morphology, and protein damage.

Measurement of ATP concentrations in mitochondria of living cells using luminescence and fluorescence approaches.

Adenosine 5'-triphosphate (ATP) is the central metabolite in the energy metabolism of cells and is hydrolyzed to ADP and inorganic phosphate to provide free energy in various cellular processes. ATP also functions as an intracellular signaling molecule. Thus, it is important to know the ATP concentration within cells to understand cellular activities. Here, we describe two methods to detect ATP concentrations in the cytoplasm and mitochondrial matrix using genetically encoded luminescent or fluorescent biosensors. These methods enable quantitative investigation of ATP concentration dynamics in living cells, single cells and cell populations.

Mitophagy in Cardiovascular Diseases.

Cardiovascular diseases are one of the leading causes of death. Increasing evidence has shown that pharmacological or genetic targeting of mitochondria can ameliorate each stage of these pathologies, which are strongly associated with mitochondrial dysfunction. Removal of inefficient and dysfunctional mitochondria through the process of mitophagy has been reported to be essential for meeting the energetic requirements and maintaining the biochemical homeostasis of cells. This process is useful for counteracting the negative phenotypic changes that occur during cardiovascular diseases, and understanding the molecular players involved might be crucial for the development of potential therapies. Here, we summarize the current knowledge on mitophagy (and autophagy) mechanisms in the context of heart disease with an important focus on atherosclerosis, ischemic heart disease, cardiomyopathies, heart failure, hypertension, arrhythmia, congenital heart disease and peripheral vascular disease. We aim to provide a complete background on the mechanisms of action of this mitochondrial quality control process in cardiology and in cardiac surgery by also reviewing studies on the use of known compounds able to modulate mitophagy for cardioprotective purposes.

The role of mitochondria-associated membranes in cellular homeostasis and diseases.

Mitochondria and endoplasmic reticulum (ER) are fundamental in the control of cell physiology regulating several signal transduction pathways. They continuously communicate exchanging messages in their contact sites called MAMs (mitochondria-associated membranes). MAMs are specific microdomains acting as a platform for the sorting of vital and dangerous signals. In recent years increasing evidence reported that multiple scaffold proteins and regulatory factors localize to this subcellular fraction suggesting MAMs as hotspot signaling domains. In this review we describe the current knowledge about MAMs' dynamics and processes, which provided new correlations between MAMs' dysfunctions and human diseases. In fact, MAMs machinery is strictly connected with several pathologies, like neurodegeneration, diabetes and mainly cancer. These pathological events are characterized by alterations in the normal communication between ER and mitochondria, leading to deep metabolic defects that contribute to the progression of the diseases.

Physiopathology of the Permeability Transition Pore: Molecular Mechanisms in Human Pathology.

Mitochondrial permeability transition (MPT) is the sudden loss in the permeability of the inner mitochondrial membrane (IMM) to low-molecular-weight solutes. Due to osmotic forces, MPT is paralleled by a massive influx of water into the mitochondrial matrix, eventually leading to the structural collapse of the organelle. Thus, MPT can initiate outer-mitochondrial-membrane permeabilization (MOMP), promoting the activation of the apoptotic caspase cascade and caspase-independent cell-death mechanisms. The induction of MPT is mostly dependent on mitochondrial reactive oxygen species (ROS) and Ca2+, but is also dependent on the metabolic stage of the affected cell and signaling events. Therefore, since its discovery in the late 1970s, the role of MPT in human pathology has been heavily investigated. Here, we summarize the most significant findings corroborating a role for MPT in the etiology of a spectrum of human diseases, including diseases characterized by acute or chronic loss of adult cells and those characterized by neoplastic initiation.