Abstract
Mitochondrial Ca
2+
uptake through the recently discovered Mitochondrial Calcium Uniporter (MCU) is controlled by its gatekeeper Mitochondrial Calcium Uptake 1 (MICU1). However, the ...physiological and pathological role of MICU1 remains unclear. Here we show that MICU1 is vital for adaptation to postnatal life and for tissue repair after injury. MICU1 knockout is perinatally lethal in mice without causing gross anatomical defects. We used liver regeneration after partial hepatectomy as a physiological stress response model. Upon MICU1 loss, early priming is unaffected, but the pro-inflammatory phase does not resolve and liver regeneration fails, with impaired cell cycle entry and extensive necrosis. Ca
2+
overload-induced mitochondrial permeability transition pore (PTP) opening is accelerated in MICU1-deficient hepatocytes. PTP inhibition prevents necrosis and rescues regeneration. Thus, our study identifies an unanticipated dependence of liver regeneration on MICU1 and highlights the importance of regulating MCU under stress conditions when the risk of Ca
2+
overload is elevated.
Mitochondrial Ca2+ uptake has long been considered crucial for meeting the fluctuating energy demands of cells in the heart and other tissues. Increases in mitochondrial matrix Ca2+ drive ...mitochondrial ATP production via stimulation of Ca2+-sensitive dehydrogenases. Mitochondria-targeted sensors have revealed mitochondrial matrix Ca2+ rises that closely follow the cytoplasmic Ca2+ signals in many paradigms. Mitochondrial Ca2+ uptake is mediated by the Ca2+ uniporter (mtCU). Pharmacological manipulation of the mtCU is potentially key to understanding its physiological significance, but no specific, cell-permeable inhibitors were identified. In the past decade, as the molecular identity of the mtCU was brought to light, efforts have focused on genetic targeting. However, in the cells/animals that are able to survive impaired mtCU function, robust compensatory changes were found in the mtCU as well as other mechanisms. Thus, the discovery, through chemical library screens on normal and mtCU-deficient cells, of new small-molecule inhibitors with improved cell permeability and specificity might offer a better chance to test the relevance of mitochondrial Ca2+ uptake. Success with the development of small molecule mtCU inhibitors is also expected to have clinical impact, considering the growing evidence for the role of mitochondrial Ca2+ uptake in a variety of diseases, including heart attack, stroke and various neurodegenerative disorders. Here, we review the progress in pharmacological targeting of mtCU and illustrate the challenges in this field using data obtained with MCU-i11, a new small molecule inhibitor.
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•Pharmacological approach is needed to test mtCU's physiological and medical relevance.•Ruthenium compounds fail to inhibit the mtCU in intact cells.•MCU-i11 inhibits the mtCU in intact cells dependent on MICU1 expression.•Inhibition is inversely proportional to Ca2+ and fails to decrease overload injury.•Plan for testing the efficacy, specificity and reversibility of mtCU drug candidates.
Mitochondrial Ca2+ uptake through the Ca2+ uniporter supports cell functions, including oxidative metabolism, while meeting tissue-specific calcium signaling patterns and energy needs. The molecular ...mechanisms underlying tissue-specific control of the uniporter are unknown. Here, we investigated a possible role for tissue-specific stoichiometry between the Ca2+-sensing regulators (MICUs) and pore unit (MCU) of the uniporter. Low MICU1:MCU protein ratio lowered the Ca2+ threshold for Ca2+ uptake and activation of oxidative metabolism but decreased the cooperativity of uniporter activation in heart and skeletal muscle compared to liver. In MICU1-overexpressing cells, MICU1 was pulled down by MCU proportionally to MICU1 overexpression, suggesting that MICU1:MCU protein ratio directly reflected their association. Overexpressing MICU1 in the heart increased MICU1:MCU ratio, leading to liver-like mitochondrial Ca2+ uptake phenotype and cardiac contractile dysfunction. Thus, the proportion of MICU1-free and MICU1-associated MCU controls these tissue-specific uniporter phenotypes and downstream Ca2+ tuning of oxidative metabolism.
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•Abundance of MICU1 relative to MCU directly reflects their association•Proportion of MICU1-bound MCU is limited by tissue-specific MICU1 availability•MICU1:MCU ratio affects mitochondrial Ca2+ uptake in liver and muscle•Liver-like MICU1:MCU ratio in heart leads to contractile dysfunction
Paillard et al. report that the relative abundance of the pore-forming protein of the mitochondrial Ca2+ uniporter (MCU) and its Ca2+-sensing regulator (MICU1) define the proportion of MCU complexes with or without MICU1. This ratio is central to programming tissue-specific mitochondrial Ca2+ uptake phenotypes in the heart and liver.
BACKGROUND—Under physiological conditions, Ca transfer from the endoplasmic reticulum (ER) to mitochondria might occur at least in part at contact points between the 2 organelles and involves the ...VDAC1/Grp75/IP3R1 complex. Accumulation of Ca into the mitochondrial matrix may activate the mitochondrial chaperone cyclophilin D (CypD) and trigger permeability transition pore opening, whose role in ischemia/reperfusion injury is well recognized. We questioned here whether the transfer of Ca from ER to mitochondria might play a role in cardiomyocyte death after hypoxia-reoxygenation.
METHODS AND RESULTS—We report that CypD interacts with the VDAC1/Grp75/IP3R1 complex in cardiomyocytes. Genetic or pharmacological inhibition of CypD in both H9c2 cardiomyoblasts and adult cardiomyocytes decreased the Ca transfer from ER to mitochondria through IP3R under normoxic conditions. During hypoxia-reoxygenation, the interaction between CypD and the IP3R1 Ca channeling complex increased concomitantly with mitochondrial Ca content. Inhibition of either CypD, IP3R1, or Grp75 decreased protein interaction within the complex, attenuated mitochondrial Ca overload, and protected cells from hypoxia-reoxygenation. Genetic or pharmacological inhibition of CypD provided a similar effect in adult mice cardiomyocytes. Disruption of ER-mitochondria interaction via the downregulation of Mfn2 similarly reduced the interaction between CypD and the IP3R1 complex and protected against hypoxia-reoxygenation injury.
CONCLUSIONS—Our data (1) point to a new role of CypD at the ER-mitochondria interface and (2) suggest that decreasing ER-mitochondria interaction at reperfusion can protect cardiomyocytes against lethal reperfusion injury through the reduction of mitochondrial Ca overload via the CypD/VDAC1/Grp75/IP3R1 complex.
Type 2 diabetic cardiomyopathy features Ca
2+
signaling abnormalities, notably an altered mitochondrial Ca
2+
handling. We here aimed to study if it might be due to a dysregulation of either the ...whole Ca
2+
homeostasis, the reticulum–mitochondrial Ca
2+
coupling, and/or the mitochondrial Ca
2+
entry through the uniporter. Following a 16-week high-fat high-sucrose diet (HFHSD), mice developed cardiac insulin resistance, fibrosis, hypertrophy, lipid accumulation, and diastolic dysfunction when compared to standard diet. Ultrastructural and proteomic analyses of cardiac reticulum–mitochondria interface revealed tighter interactions not compatible with Ca
2+
transport in HFHSD cardiomyocytes. Intramyocardial adenoviral injections of Ca
2+
sensors were performed to measure Ca
2+
fluxes in freshly isolated adult cardiomyocytes and to analyze the direct effects of in vivo type 2 diabetes on cardiomyocyte function. HFHSD resulted in a decreased IP3R–VDAC interaction and a reduced IP3-stimulated Ca
2+
transfer to mitochondria, with no changes in reticular Ca
2+
level, cytosolic Ca
2+
transients, and mitochondrial Ca
2+
uniporter function. Disruption of organelle Ca
2+
exchange was associated with decreased mitochondrial bioenergetics and reduced cell contraction, which was rescued by an adenovirus-mediated expression of a reticulum-mitochondria linker. An 8-week diet reversal was able to restore cardiac insulin signaling, Ca
2+
transfer, and cardiac function in HFHSD mice. Therefore, our study demonstrates that the reticulum–mitochondria Ca
2+
miscoupling may play an early and reversible role in the development of diabetic cardiomyopathy by disrupting primarily the mitochondrial bioenergetics. A diet reversal, by counteracting the MAM-induced mitochondrial Ca
2+
dysfunction, might contribute to restore normal cardiac function and prevent the exacerbation of diabetic cardiomyopathy.
Diabetic cardiomyopathy (DCM) is a leading complication in type 2 diabetes patients. Recently, we have shown that the reticulum-mitochondria Ca2+ uncoupling is an early and reversible trigger of the ...cardiac dysfunction in a diet-induced mouse model of DCM. Metformin is a first-line antidiabetic drug with recognized cardioprotective effect in myocardial infarction. Whether metformin could prevent the progression of DCM remains not well understood. We therefore investigated the effect of a chronic 6-week metformin treatment on the reticulum-mitochondria Ca2+ coupling and the cardiac function in our high-fat high-sucrose diet (HFHSD) mouse model of DCM. Although metformin rescued the glycemic regulation in the HFHSD mice, it did not preserve the reticulum-mitochondria Ca2+ coupling either structurally or functionally. Metformin also did not prevent the progression towards cardiac dysfunction, i.e., cardiac hypertrophy and strain dysfunction. In summary, despite its cardioprotective role, metformin is not sufficient to delay the progression to early DCM.
Background
The immune system, composed of organs, tissues, cells, and proteins, is the key to protecting the body from external biological attacks and inflammation. The latter occurs in several ...pathologies, such as cancers, type 1 diabetes, and human immunodeficiency virus infection. Immunophenotyping by flow cytometry is the method of choice for diagnosing these pathologies. Under inflammatory conditions, the peripheral blood mononuclear cells (PBMCs) are partially activated and generate intracellular pathways involving Ca
2+
-dependent signaling cascades leading to transcription factor expression. Ca
2+
signaling is typically studied by microscopy in cell lines but can present some limitations to explore human PBMCs, where flow cytometry can be a good alternative.
Objective
In this review, we dived into the research field of inflammation and Ca
2+
signaling in PBMCs. We aimed to investigate the structure and evolution of this field in a physio-pathological context, and then we focused our review on flow cytometry analysis of Ca
2+
fluxes in PBMCs.
Methods
From 1984 to 2022, 3865 articles on inflammation and Ca
2+
signaling in PBMCs were published, according to The Clarivate Web of Science (WOS) database used in this review. A bibliometric study was designed for this collection and consisted of a co-citation and bibliographic coupling analysis.
Results
The co-citation analysis was performed on 133 articles: 4 clusters highlighted the global context of Ca
2+
homeostasis, including chemical probe development, identification of the leading players in Ca
2+
signaling, and the link with chemokine production in immune cell function. Next, the bibliographic coupling analysis combined 998 articles in 8 clusters. This analysis outlined the mechanisms of PBMC activation, from signal integration to cellular response. Further explorations of the bibliographic coupling network, focusing on flow cytometry, revealed 21 articles measuring cytosolic Ca
2+
in PBMCs, with only 5 since 2016. This final query showed that Ca
2+
signaling analysis in human PBMCs using flow cytometry is still underdeveloped and investigates mainly the cytosolic Ca
2+
compartment.
Conclusion
Our review uncovers remaining knowledge gaps of intracellular players involved in Ca
2+
signaling in PBMCs, such as reticulum and mitochondria, and presents flow cytometry as a solid option to supplement gold-standard microscopy studies.
Abstract
Despite advances in cardioprotection, new therapeutic strategies capable of preventing ischemia-reperfusion injury of patients are still needed. Here, we discover that ...sarcoplasmic/endoplasmic reticulum Ca
2+
ATPase (SERCA2) phosphorylation at serine 663 is a clinical and pathophysiological event of cardiac function. Indeed, the phosphorylation level of SERCA2 at serine 663 is increased in ischemic hearts of patients and mouse. Analyses on different human cell lines indicate that preventing serine 663 phosphorylation significantly increases SERCA2 activity and protects against cell death, by counteracting cytosolic and mitochondrial Ca
2+
overload. By identifying the phosphorylation level of SERCA2 at serine 663 as an essential regulator of SERCA2 activity, Ca
2+
homeostasis and infarct size, these data contribute to a more comprehensive understanding of the excitation/contraction coupling of cardiomyocytes and establish the pathophysiological role and the therapeutic potential of SERCA2 modulation in acute myocardial infarction, based on the hotspot phosphorylation level of SERCA2 at serine 663 residue.
Abstract Background Type 2 diabetes (T2D) is a frequent comorbidity encountered in patients with severe aortic stenosis (AS), leading to an adverse left ventricular (LV) remodeling and dysfunction. ...Metabolic alterations have been suggested as contributors of the deleterious effect of T2D on LV remodeling and function in patients with severe AS, but so far, the underlying mechanisms remain unclear. Mitochondria play a central role in the regulation of cardiac energy metabolism. Objectives We aimed to explore the mitochondrial alterations associated with the deleterious effect of T2D on LV remodeling and function in patients with AS, preserved ejection fraction, and no additional heart disease. Methods We combined an in-depth clinical, biological and echocardiography phenotype of patients with severe AS, with (n = 34) or without (n = 50) T2D, referred for a valve replacement, with transcriptomic and histological analyses of an intra-operative myocardial LV biopsy. Results T2D patients had similar AS severity but displayed worse cardiac remodeling, systolic and diastolic function than non-diabetics. RNAseq analysis identified 1029 significantly differentially expressed genes. Functional enrichment analysis revealed several T2D-specific upregulated pathways despite comorbidity adjustment, gathering regulation of inflammation, extracellular matrix organization, endothelial function/angiogenesis, and adaptation to cardiac hypertrophy. Downregulated gene sets independently associated with T2D were related to mitochondrial respiratory chain organization/function and mitochondrial organization. Generation of causal networks suggested a reduced Ca 2+ signaling up to the mitochondria, with the measured gene remodeling of the mitochondrial Ca 2+ uniporter in favor of enhanced uptake. Histological analyses supported a greater cardiomyocyte hypertrophy and a decreased proximity between the mitochondrial VDAC porin and the reticular IP3-receptor in T2D. Conclusions Our data support a crucial role for mitochondrial Ca 2+ signaling in T2D-induced cardiac dysfunction in severe AS patients, from a structural reticulum-mitochondria Ca 2+ uncoupling to a mitochondrial gene remodeling. Thus, our findings open a new therapeutic avenue to be tested in animal models and further human cardiac biopsies in order to propose new treatments for T2D patients suffering from AS. Trial registration URL: https://www.clinicaltrials.gov ; Unique Identifier: NCT01862237. Graphical abstract
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Endoplasmic reticulum (ER) and mitochondria are functionally distinct with regard to membrane protein biogenesis and oxidative energy production, respectively, but cooperate in several essential cell ...functions, including lipid biosynthesis, cell signaling and organelle dynamics. The interorganellar cooperation requires local communication that can occur at the strategically positioned and dynamic associations between ER and mitochondria. Calcium is locally transferred from ER to mitochondria at the associations and exerts regulatory effects on numerous proteins. A common Ca2+ sensing mechanism is the EF-hand Ca2+ binding domain, many of which can be found in proteins of the mitochondria, including Miro1&2, MICU1,2&3, LETM1 and mitochondrial solute carriers. Recently, these proteins have triggered much interest and were described in reports with diverging conclusions. The present essay focuses on their shared features and established specific functions.
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