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Na + /Ca 2+ exchangers (NCX) transport Ca 2+ in or out of cells in exchange for Na + . They are ubiquitously expressed and play an essential role in maintaining cytosolic Ca 2+ homeostasis. Although extensively studied, little is known about the global structural arrangement of eukaryotic NCXs and the structural mechanisms underlying their regulation by various cellular cues including cytosolic Na + and Ca 2+ . Here we present the cryo-EM structures of human cardiac NCX1 in both inactivated and activated states, elucidating key structural elements important for NCX ion exchange function and its modulation by cytosolic Ca 2+ and Na + . We demonstrate that the interactions between the ion-transporting transmembrane (TM) domain and the cytosolic regulatory domain define the activity of NCX. In the inward-facing state with low cytosolic [Ca 2+ ], a TM-associated four-stranded β-hub mediates a tight packing between the TM and cytosolic domains, resulting in the formation of a stable inactivation assembly that blocks the TM movement required for ion exchange function. Ca 2+ binding to the cytosolic second Ca 2+ -binding domain (CBD2) disrupts this inactivation assembly which releases its constraint on the TM domain, yielding an active exchanger. Thus, the current NCX1 structures provide an essential framework for the mechanistic understanding of the ion transport and cellular regulation of NCX family proteins. Here authors present the cryo-EM structures of human cardiac NCX1, elucidating key structural elements for ion exchange and modulation by cytosolic Ca2 + and Na + .

We previously reviewed our study of the pharmacological properties of cardiac Na+/Ca2+ exchange (NCX1) inhibitors among cardioprotective drugs, such as amiodarone, bepridil, dronedarone, cibenzoline, azimilide, aprindine, and benzyl-oxyphenyl derivatives (Watanabe et al. in J Pharmacol Sci 102:7–16, 2006). Since then we have continued our studies further and found that some cardioprotective drugs are NCX1 stimulators. Cardiac Na+/Ca2+ exchange current (INCX1) was stimulated by nicorandil (a hybrid ATP-sensitive K+ channel opener), pinacidil (a non-selective ATP-sensitive K+ channel opener), flecainide (an antiarrhythmic drug), and sodium nitroprusside (SNP) (an NO donor). Sildenafil (a phosphodiesterase-5 inhibitor) further increased the pinacidil-induced augmentation of INCX1. In paper, here I review the NCX stimulants that enhance NCX function among the cardioprotective agents we examined such as nicorandil, pinacidil, SNP, sildenafil and flecainide, in addition to atrial natriuretic (ANP) and dofetilide, which were reported by other investigators.

Nutraceuticals exert a series of health benefits, including protection against cardiovascular diseases. In this study, naringin, naringenin, and quercetin were tested for their safety and efficacy in ameliorating angiotensin (Ang) II-induced cardiac hypertrophy through carbonic anhydrase II (CA-II) inhibition. In silico molecular docking and MD simulations exhibited that naringin strongly binds CA-II with a docking score of -9.55 kcal/mol and hydrogen bonding energy of -6.07 kcal/mol. Naringin formed stable hydrogen bond interactions with Asn62, Trp5, and N-acetyl His4 via catalytic water molecule, and a continuous interaction via major water bridge with N-acetyl His4, His4, and Trp5. Moreover, naringin effectively inhibited CA-II activity with an IC 50 value of 82.99 ± 4.92 nM, followed by naringenin and quercetin. Of note, all the tested nutraceuticals were found to be safe as evident from the cell viability assays. Further, naringin effectively attenuated cardiac hypertrophy, as indicated by the reductions in the Ang II-induced increases in cell surface area of H9c2 cardio myoblasts (165.6 ± 1.26% Ang II vs. 109.8 ± 1.88% Ang II + naringin), followed by naringenin and quercetin. Furthermore, naringin significantly inhibited CA-II activity (191.77 ± 7.69% Ang II vs. 120.16 ± 5.52% Ang II + naringin) and suppressed Ang II-induced CA-II and Na + /H + exchanger 1 (NHE1) protein expression. Besides, naringin suppressed Ang II-induced CA-II, NHE1, Na + /Ca 2+ exchanger 1 (NCX1), and angiotensin-converting enzyme (ACE1) mRNA expression. Collectively, naringin when compared to naringenin and quercetin effectively attenuated Ang II-induced cardio myoblast hypertrophy, CA-II activity, CA-II, and NHE1 expression. The naringin-mediated attenuation of cardiac hypertrophy might be through the inhibition of CA-II enzyme activity, and the suppression of NHE1, and NCX1.

The Na + -Ca 2+ exchanger (NCX1) is the dominant Ca 2+ extrusion mechanism in cardiac myocytes. NCX1 activity is inhibited by intracellular Na + via a process known as Na + -dependent inactivation. A central question is whether this inactivation plays a physiological role in heart function. Using CRISPR/Cas9, we inserted the K229Q mutation in the gene ( Slc8a1 ) encoding for NCX1. This mutation removes the Na + -dependent inactivation while preserving transport properties and other allosteric regulations. NCX1 mRNA levels, protein expression, and protein localization are unchanged in K229Q male mice. However, they exhibit reduced left ventricular ejection fraction and fractional shortening, while displaying a prolonged QT interval. K229Q ventricular myocytes show enhanced NCX1 activity, resulting in action potential prolongation, higher incidence of aberrant action potentials, a faster decline of Ca 2+ transients, and depressed cell shortening. The results demonstrate that NCX1 Na + -dependent inactivation plays an essential role in heart function by affecting both cardiac excitability and contractility. The sodium-calcium exchanger (NCX1) is the primary calcium extrusion mechanism of cardiac myocytes. Here, the authors show that removal of a long questioned allosteric regulation of NCX1 by intracellular sodium alters cardiac excitation-contraction coupling.

Aims Cancer‐induced bone pain (CIBP) is a chronic and refractory pain condition characterized by neuronal hyperexcitability, calcium dysregulation, and neuroinflammation. Voltage‐gated sodium channels (VGSCs) and sodium/calcium exchangers (NCXs) are crucial in regulating sensory neuron sodium–calcium homeostasis, influencing nociceptive signaling and neuroinflammatory responses. This study focused on exploring how Nav1.7 from the VGSC family and NCX1 from the NCX family influence nociceptive signaling and neuroinflammation in CIBP. Methods CIBP was induced in mice. Nav1.7 and NCX1 expression and colocalization in DRG neurons were analyzed by qPCR, western blotting, and immunofluorescence. Calcium overload and neuronal excitability were assessed using calcium imaging and electrophysiological recordings. Neuroinflammation markers were detected by qPCR and western blotting. Results Among the VGSC and NCX subtypes, Nav1.7 and NCX1 were notably upregulated and colocalized in the DRG neurons of CIBP mice. Combined inhibition of these channels demonstrated a synergistic analgesic effect and markedly reduced neuronal calcium overload and hyperexcitability. Furthermore, the combined inhibition substantially alleviated neuroinflammation by inhibiting the p38 MAPK/NF‐κB pathway and lowering proinflammatory cytokine levels. Conclusions The combined inhibition of Nav1.7 and NCX1 enhances analgesic effects and reduces neuroinflammation, presenting a potential therapeutic approach for CIBP and other cancer‐associated pain disorders. Nav1.7‐mediated sodium influx promotes NCX1 reverse transport, increasing sensory neuron excitability and causing calcium overload, which activates the p38 MAPK/NF‐κB pathway and induces neuroinflammation. Dual inhibition reduces hyperexcitability, restores calcium homeostasis, suppresses neuroinflammation, and ultimately alleviates cancer‐induced bone pain (CIBP).

BackgroundClassically activated M1 macrophages, characterized by aberrant glycolysis and secretion of inflammatory cytokines, play pivotal roles in inflammatory diseases, including inflammatory bowel disease (IBD). Recently, sodium-glucose co-transporter 2 (SGLT2) inhibitors were shown to suppress Na+/H+ exchanger 1 (NHE1) and Na+/Ca2+ exchanger 1 (NCX1) activity, regulating downstream intracellular Ca2+ concentrations in cardiomyocytes. However, whether SGLT2 inhibitors regulate M1 macrophage polarization by downregulating NHE1 and NCX1 remains unknown.MethodsWe analyzed cellular responses to SGLT2 inhibitors using mouse bone marrow-derived macrophages and peritoneal macrophages treated with lipopolysaccharide (LPS). To induce IBD, we used a dextran sulfate sodium salt-induced colitis mouse model.ResultsWe observed that NHE1 and NCX1 were overexpressed in LPS-treated macrophages, leading to M1 macrophage polarization. Mechanistically, NHE1 and NCX1-mediated Ca2+ accumulation in the macrophage resulted in enhanced glycolysis by promoting PI3K/AKT/mTORC1 signaling. SGLT2 inhibitors suppressed both the expression levels and activities of NHE1 and NCX1, and consequently downregulated PI3K/AKT/mTORC1 signaling and glycolysis in LPS-treated macrophages. We observed inhibition of LPS-stimulated M1 polarization and cytokine production by SGLT2 inhibitors in vitro, ex vivo, and in an IBD mouse model.ConclusionsNHE1 promotes M1 macrophage polarization and SGLT2 inhibitors are a novel strategy to treat M1 macrophage-mediated inflammatory diseases, including IBD.

Plasma membrane Na + /Ca 2+ exchanger 1 (NCX1) is a bidirectional ion transporter to operate in Ca 2+ entry or exit modes, and TRPC1 is Ca 2+ -permeable channel. Both NCX1 and TRPC1 play critical roles in maintaining cytosolic free Ca 2+ ([Ca 2+ ] cyt ) homeostasis in mammalian cells. Although either TRPC1 channel or Ca 2+ entry mode of NCX1 is implicated in some tumorigenesis, it has not been explored if a coordination of NCX1 and TRPC1 involves in the pathogenesis of H. pylori- associated human gastric cancer (GC). Here we found the protein expression of NCX1 was significantly enhanced in human GC specimens, which correlated with tumor progression and poor survival in GC patients. TRPC1 and NCX1 were parallelly enhanced, co-localized and bound in human GC cells. By a functional coupling, TRPC1 drives NCX1 to the Ca 2+ entry mode, raising [Ca 2+ ] cyt in GC cells. Moreover, CaCl 2 , H. pylori and their virulence factors all enhanced expressions and activities of NCX1 and TRPC1, and evoked aberrant Ca 2+ entry to promote proliferation, migration, and invasion of GC cells through AKT/β-catenin pathway. Tumor growth and metastasis also depended on the enhanced expression of NCX1 in subcutaneously xenografted GC mouse model. Overall, our findings indicate that TRPC1/NCX1 coupling may promote H. pylori- associated GC through the Ca 2+ /AKT/β-catenin pathway. Since the Ca 2+ exit mode and the Ca 2+ entry mode of NCX1 play different roles under mostly physiological and pathological conditions respectively, targeting TRPC1/NCX1 coupling could be a novel strategy for selectively blocking Ca 2+ entry mode to potentially treat digestive cancer with less side effect.

Neutrophil-driven inflammation is central to the pathogenesis of chronic obstructive pulmonary disease (COPD). Emerging evidence suggests that Ca²⁺ signaling is critical in regulating neutrophil activation, recruitment and tissue residency. In this study, we investigated the function of Na⁺/Ca²⁺ exchanger 1 (NCX1), a Ca²⁺/cation membrane transporter, in neutrophils during COPD pathogenesis. Analysis of human specimens show that NCX1 is primarily upregulated in neutrophils from patients with mixed chronic bronchitis and emphysema. Cigarette smoke exposure induces NCX1 upregulation and promotes its reverse-mode transport activity, leading to elevated intracellular Ca²⁺ levels and enhanced NETs formation. Neutrophil-specific genetic deletion of Slc8a1 or pharmacological inhibition of NCX1 reverse transport effectively suppresses Ca²⁺ influx, NETs release, and neutrophil accumulation and retention, thereby ameliorating chronic bronchitis and emphysematous changes. Collectively, our findings identify NCX1 as a regulator of Ca²⁺-dependent NETs release in neutrophils. Targeting NCX1-mediated Ca²⁺ influx or NETs formation represents a potential therapeutic strategy for neutrophilic inflammation in COPD. Neutrophils are known to be important in COPD pathogenesis and calcium signaling is involved in this process. Here the authors examine the function of Na + Ca 2+ exchanger 1 (NCX1) in neutrophil function in mouse models of chronic cigarette smoke exposure and show that inhibition of NCX1 reduces neutrophil accumulation in the lungs and formation of neutrophil extracellular traps (NETs).

Inflammation and infection can trigger local tissue Na+ accumulation. This Na+-rich environment boosts proinflammatory activation of monocyte/macrophage-like cells (MΦs) and their antimicrobial activity. Enhanced Na+-driven MΦ function requires the osmoprotective transcription factor nuclear factor of activated T cells 5 (NFAT5), which augments nitric oxide (NO) production and contributes to increased autophagy. However, the mechanism of Na+ sensing in MΦs remained unclear. High extracellular Na+ levels (high salt [HS]) trigger a substantial Na+ influx and Ca2+ loss. Here, we show that the Na+/Ca2+ exchanger 1 (NCX1, also known as solute carrier family 8 member A1 [SLC8A1]) plays a critical role in HS-triggered Na+ influx, concomitant Ca2+ efflux, and subsequent augmented NFAT5 accumulation. Moreover, interfering with NCX1 activity impairs HS-boosted inflammatory signaling, infection-triggered autolysosome formation, and subsequent antibacterial activity. Taken together, this demonstrates that NCX1 is able to sense Na+ and is required for amplifying inflammatory and antimicrobial MΦ responses upon HS exposure. Manipulating NCX1 offers a new strategy to regulate MΦ function.

Pancreatic stellate cells (PSCs) that can co-metastasize with cancer cells shape the tumor microenvironment (TME) in pancreatic ductal adenocarcinoma (PDAC) by producing an excessive amount of extracellular matrix. This leads to a TME characterized by increased tissue pressure, hypoxia, and acidity. Moreover, cells within the tumor secrete growth factors. The stimuli of the TME trigger Ca2+ signaling and cellular Na+ loading. The Na+/Ca2+ exchanger (NCX) connects the cellular Ca2+ and Na+ homeostasis. The NCX is an electrogenic transporter, which shuffles 1 Ca2+ against 3 Na+ ions over the plasma membrane in a forward or reverse mode. Here, we studied how the impact of NCX activity on PSC migration is modulated by cues from the TME. NCX expression was revealed with qPCR and Western blot. [Ca2+]i, [Na+]i, and the cell membrane potential were determined with the fluorescent indicators Fura-2, Asante NaTRIUM Green-2, and DiBAC4(3), respectively. PSC migration was quantified with live-cell imaging. To mimic the TME, PSCs were exposed to hypoxia, pressure, acidic pH (pH 6.6), and PDGF. NCX-dependent signaling was determined with Western blot analyses. PSCs express NCX1.3 and NCX1.9. [Ca2+]i, [Na+]i, and the cell membrane potential are 94.4 nmol/l, 7.4 mmol/l, and − 39.8 mV, respectively. Thus, NCX1 usually operates in the forward (Ca2+ export) mode. NCX1 plays a differential role in translating cues from the TME into an altered migratory behavior. When NCX1 is operating in the forward mode, its inhibition accelerates PSC migration. Thus, NCX1-mediated extrusion of Ca2+ contributes to a slow mode of migration of PSCs.