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Changes in the Stoichiometry of Uniplex Decrease Mitochondrial Calcium Overload and Contribute to Tolerance of Cardiac Ischemia/Reperfusion Injury in Hypothyroidism.

ABSTRACT: Background: Thyroid hormone status in hypothyroidism (HT) downregulates key elements in Ca2+ handling within the heart, reducing contractility, impairing the basal energetic balance, and increasing the risk of cardiovascular disease. Mitochondrial Ca2+ transport is reduced in HT, and tolerance to reperfusion damage has been documented, but the precise mechanism is not well understood. Therefore, we aimed to determine the stoichiometry and activity of the mitochondrial Ca2+ uniporter or uniplex in an HT model and the relevance to the opening of the mitochondrial permeability transition pores (mPTP) during ischemia/reperfusion (I/R) injury. Methods: An HT model was established in Wistar rats by treatment with 6-propylthiouracil for 28 days. Uniplex composition and activity were determined in cardiac mitochondria. Hearts were perfused ex vivo to induce I/R injury, and functional parameters related to contractility and tissue viability were evaluated. Results: The cardiac stoichiometry between two subunits of the uniplex (MICU1/MCU) increased by 25% in animals with HT. The intramitochondrial Ca2+ content was reduced by 40% and was less prone to the mPTP opening. After I/R injury, ischemic contracture and the onset of ventricular fibrillation were delayed in animals with HT, concomitant with a reduction in oxidative damage and mitochondrial dysfunction. Conclusions: Our results suggest that HT is associated with an increase in the cardiac MICU1/MCU ratio, thereby changing the stoichiometry between these subunits to increase the threshold to cytosolic Ca2+ and reduce mitochondrial Ca2+ overload. Our results also demonstrate that this HT model can be used to explore the role of mitochondrial Ca2+ transport in cardiac diseases due to its induced tolerance to cardiac damage.

SUBMITTER: Chapoy-Villanueva H

PROVIDER: S-EPMC6918869 | biostudies-literature | 2019 Oct

REPOSITORIES: biostudies-literature

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Changes in the Stoichiometry of Uniplex Decrease Mitochondrial Calcium Overload and Contribute to Tolerance of Cardiac Ischemia/Reperfusion Injury in Hypothyroidism.

Chapoy-Villanueva Héctor H Silva-Platas Christian C Gutiérrez-Rodríguez Ana K AK García Noemí N Acuña-Morin Edgar E Elizondo-Montemayor Leticia L Oropeza-Almazán Yuriana Y Aguilar-Saenz Alejandro A García-Rivas Gerardo G

Thyroid : official journal of the American Thyroid Association 20191009 12

Background: Thyroid hormone status in hypothyroidism (HT) downregulates key elements in Ca2+ handling within the heart, reducing contractility, impairing the basal energetic balance, and increasing the risk of cardiovascular disease. Mitochondrial Ca2+ transport is reduced in HT, and tolerance to reperfusion damage has been documented, but the precise mechanism is not well understood. Therefore, we aimed to determine the stoichiometry and activity of the mitoc ...[more]

PMID: 31456501

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Project description:IntroductionIntracellular calcium overload is an important contributor to myocardial ischemia/reperfusion (MI/R) injury. Total saponins of the traditional Chinese medicinal plant Aralia elata (Miq.) Seem. (AS) are beneficial for treating MI/R injury, and Calenduloside E (CE) is the main active ingredient of AS.ObjectivesThis study aimed to investigate the effects of CE on MI/R injury and determine its specific regulatory mechanisms.MethodsTo verify whether CE mediated cardiac protection in vivo and in vitro, we performed MI/R surgery in SD rats and subjected neonatal rat ventricular myocytes (NRVMs) to hypoxia-reoxygenation (HR). CE's cardioprotective against MI/R injury was detected by Evans blue/TTC staining, echocardiography, HE staining, myocardial enzyme levels. Impedance and field potential recording, and patch-clamp techniques of human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) were used to detect the function of L-type calcium channels (LTCC). The mechanisms underlying between CE and LTCC was studied through western blot, immunofluorescence, and immunohistochemistry. Drug affinity responsive target stability (DARTS) and co-immunoprecipitation (co-IP) used to further clarify the effect of CE on LTCC and BAG3.ResultsWe found that CE protected against MI/R injury by inhibiting calcium overload. Furthermore, CE improved contraction and field potential signals of hiPSC-CMs and restored sarcomere contraction and calcium transient of adult rat ventricular myocytes (ARVMs). Moreover, patch-clamp data showed that CE suppressed increased L-type calcium current (ICa,L) caused by LTCC agonist, proving that CE could regulate calcium homeostasis through LTCC. Importantly, we found that CE promoted the interaction between LTCC and Bcl2-associated athanogene 3 (BAG3) by co-IP and DARTS.ConclusionOur results demonstrate that CE enhanced LTCC-BAG3 interaction to reduce MI/R induced-calcium overload, exerting a cardioprotective effect.

Project description:IntroductionSurgery, a crucial therapeutic modality in the treatment of solid tumors, can induce sterile inflammatory processes which can result in metastatic progression. Liver ischemia and reperfusion (I/R) injury, an inevitable consequence of hepatic resection of metastases, has been shown to foster hepatic capture of circulating cancer cells and accelerate metastatic growth. Efforts to reduce these negative consequences have not been thoroughly investigated. Drag reducing polymers (DRPs) are blood-soluble macromolecules that can, in nanomolar concentrations, increase tissue perfusion, decrease vascular resistance and decrease near-wall microvascular concentration of neutrophils and platelets thereby possibly reducing the inflammatory microenvironment. We hypothesize that DRP can potentially be used to ameliorate metastatic capture of tumor cells and tumor growth within the I/R liver.MethodsExperiments were performed utilizing a segmental ischemia model of mice livers. Five days prior or immediately prior to ischemia, murine colon adenocarcinoma cells (MC38) were injected into the spleen. DRP (polyethylene oxide) or a control of low-molecular-weight polyethylene glycol without drag reducing properties were administered intraperitoneally at the onset of reperfusion.ResultsAfter three weeks from I/R, we observed that liver I/R resulted in an increased ability to capture and foster growth of circulating tumor cells; in addition, the growth of pre-existing micrometastases was accelerated three weeks later. These effects were significantly curtailed when mice were treated with DRPs at the time of I/R. Mechanistic investigations in vivo indicated that DRPs protected the livers from I/R injury as evidenced by significant decreases in hepatocellular damage, neutrophil recruitment into the liver, formation of neutrophil extracellular traps, deposition of platelets, formation of microthrombi within the liver sinusoids and release of inflammatory cytokines.ConclusionsDRPs significantly attenuated metastatic tumor development and growth. DRPs warrant further investigation as a potential treatment for liver I/R injury in the clinical setting to improve cancer-specific outcomes.

Project description:RationaleWhile thrombin is the key protease in thrombus formation, other coagulation proteases, such as fXa (factor Xa) or aPC (activated protein C), independently modulate intracellular signaling via partially distinct receptors.ObjectivesTo study the differential effects of fXa or fIIa (factor IIa) inhibition on gene expression and inflammation in myocardial ischemia-reperfusion injury.Methods and resultsMice were treated with a direct fIIa inhibitor (fIIai) or direct fXa inhibitor (fXai) at doses that induced comparable anticoagulant effects ex vivo and in vivo (tail-bleeding assay and FeCl3-induced thrombosis). Myocardial ischemia-reperfusion injury was induced via left anterior descending ligation. We determined infarct size and in vivo aPC generation, analyzed gene expression by RNA sequencing, and performed immunoblotting and ELISA. The signaling-only 3K3A-aPC variant and inhibitory antibodies that blocked all or only the anticoagulant function of aPC were used to determine the role of aPC. Doses of fIIai and fXai that induced comparable anticoagulant effects resulted in a comparable reduction in infarct size. However, unbiased gene expression analyses revealed marked differences, including pathways related to sterile inflammation and inflammasome regulation. fXai but not fIIai inhibited sterile inflammation by reducing the expression of proinflammatory cytokines (IL [interleukin]-1β, IL-6, and TNFα [tumor necrosis factor alpha]), as well as NF-κB (nuclear factor kappa B) and inflammasome activation. This anti-inflammatory effect was associated with reduced myocardial fibrosis 28 days post-myocardial ischemia-reperfusion injury. Mechanistically, in vivo aPC generation was higher with fXai than with fIIai. Inhibition of the anticoagulant and signaling properties of aPC abolished the anti-inflammatory effect associated with fXai, while inhibiting only the anticoagulant function of aPC had no effect. Combining 3K3A-aPC with fIIai reduced the inflammatory response, mimicking the fXai-associated effect.ConclusionsWe showed that specific inhibition of coagulation via direct oral anticoagulants had differential effects on gene expression and inflammation, despite comparable anticoagulant effects and infarct sizes. Targeting individual coagulation proteases induces specific cellular responses unrelated to their anticoagulant effect.

Dataset Information

Changes in the Stoichiometry of Uniplex Decrease Mitochondrial Calcium Overload and Contribute to Tolerance of Cardiac Ischemia/Reperfusion Injury in Hypothyroidism.

Publications

Changes in the Stoichiometry of Uniplex Decrease Mitochondrial Calcium Overload and Contribute to Tolerance of Cardiac Ischemia/Reperfusion Injury in Hypothyroidism.

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