Mitochondrial ATP-sensitive Potassium Channel Research Articles

Mitochondrial Dysfunction and Ion Imbalance in a Rat Model of Hemodialysis-Induced Myocardial Stunning.

Background/Objectives: Hemodialysis-induced myocardial stunning (HIMS) is a frequent complication in patients undergoing maintenance hemodialysis, characterized by transient left ventricular dysfunction due to ischemic episodes. Mitochondrial dysfunction and fluctuations in key ions such as potassium (K+) and calcium (Ca2+) are implicated in the pathogenesis of HIMS. This study aims to investigate the role of mitochondrial dysfunction and the protective potential of mitochondrial ATP-sensitive potassium channels (mitoKATP) in mitigating HIMS. Methods: A 5/6 nephrectomy rat model was established to mimic chronic kidney disease and the subsequent HIMS. The effects of mitoKATP channel modulators were evaluated by administering diazoxide (DZX), a mitoKATP opener, and 5-hydroxydecanoate (5-HD), a mitoKATP blocker, before hemodialysis. Mitochondrial function was assessed by measuring membrane potential, ATP synthase activity, and intramitochondrial Ca2+ levels. Myocardial function was evaluated using speckle tracking echocardiography. Results: Rats undergoing hemodialysis exhibited significant reductions in left ventricular strain and synchrony. DZX administration significantly improved mitochondrial function and reduced myocardial strain compared to controls. Conversely, 5-HD worsened mitochondrial swelling and disrupted myocardial function. Higher K+ and Ca2+ concentrations in the dialysate were associated with improved mitochondrial energy metabolism and myocardial strain. Conclusions: Mitochondrial dysfunction and ion imbalances during hemodialysis are key contributors to HIMS. The activation of mitoKATP channels provides mitochondrial protection and may serve as a potential therapeutic strategy to mitigate HIMS.

Anti-Inflammatory Responses Produced with Nippostrongylus brasiliensis-Derived Uridine via the Mitochondrial ATP-Sensitive Potassium Channel and Its Anti-Atherosclerosis Effect in an Apolipoprotein E Gene Knockout Mouse Model.

Atherosclerosis (AS) has become the leading cause of cardiovascular disease worldwide. Our previous study had observed that Nippostrongylus brasiliensis (Nb) infection or its derived products could inhibit AS development by inducing an anti-inflammatory response. We performed a metabolic analysis to screen Nb-derived metabolites with anti-inflammation activity and evaluated the AS-prevention effect. We observed that the metabolite uridine had higher expression levels in mice infected with the Nb and ES (excretory-secretory) products and could be selected as a key metabolite. ES and uridine interventions could reduce the pro-inflammatory responses and increase the anti-inflammatory responses in vitro and in vivo. The apolipoprotein E gene knockout (ApoE-/-) mice were fed with a high-fat diet for the AS modeling. Following the in vivo intervention, ES products or uridine significantly reduced serum and liver lipid levels, alleviated the formation of atherosclerosis, and reduced the pro-inflammatory responses in serum or plaques, while the anti-inflammatory responses showed opposite trends. After blocking with 5-HD (5-hydroxydecanoate sodium) in vitro, the mRNA levels of M2 markers were significantly reduced. When blocked with 5-HD in vivo, the degree of atherosclerosis was worsened, the pro-inflammatory responses were increased compared to the uridine group, while the anti-inflammatory responses decreased accordingly. Uridine, a key metabolite from Nippostrongylus brasiliensis, showed anti-inflammatory and anti-atherosclerotic effects in vitro and in vivo, which depend on the activation of the mitochondrial ATP-sensitive potassium channel.

#604 Oral administration of UNI-494 ameliorates acute kidney injury in a rat model of delayed graft function

Abstract Background and Aims There are no FDA approved drugs for the treatment of acute kidney injury (AKI), which affects 10-15% of hospitalized patients and often results in renal transplantation or lifelong dialysis. UNI-494 is a novel nicotinamide ester derivative and selective mitochondrial ATP-sensitive potassium channel activator that may be beneficial for several disease states, including AKI. We present results from a study evaluating the in vivo efficacy of oral (PO) UNI-494 in the unilateral renal ischemia-reperfusion (I/R) rat model of AKI, which is a well-established model of delayed graft function (DGF) (Cavaille-Coll, 2013). Method Rats were anesthetized, the right kidney was removed, and I/R was induced in three groups of rats by clamping the renal vessels in the left kidney for 30 minutes; a fourth group had no surgery (sham group) without clamping the renal vessels. 50 mg/kg or 100 mg/kg UNI-494 were administered PO 30 minutes prior to I/R. One I/R group and the sham group were not treated. After 24 hours of reperfusion in metabolic cages, blood samples were collected for serum creatinine (sCr), and urinary samples were collected for albumin-creatinine ratio (ACR) and the tubular injury marker neutrophil gelatinase-associated lipocalin (NGAL). The clamped left kidney was collected and processed for histology for tubular injury scores. Results In this study, I/R induced significant increases of sCr, ACR, NGAL, and proximal convoluted tubular injury scores in the vehicle treated I/R group vs No I/R sham group. A single PO dose of UNI-494 at 50 mg/kg/PO or 100 mg/kg/PO reduced the kidney functional markers sCr and ACR, the tubular injury marker (NGAL), and proximal tubular injury scores (all p < 0.001, Fig. 1). Conclusion The results indicate that UNI-494 dose dependently prevented AKI markers and proximal tubular injury in a rat model of AKI and therefore is a potential candidate for the prevention of DGF and other AKI clinical conditions.

#157 UNI-494 Phase I tolerability and pharmacokinetics: trial in progress

Abstract Background and Aims Acute kidney injury (AKI) is a clinical syndrome defined by the sudden loss of kidney function, resulting in an inability to maintain electrolyte, acid-base and water balance. Currently, there are no effective treatments for AKI approved by the US Food and Drug Administration or the European Medicines Agency (EMA); management of the condition is primarily supportive. The most common cause of AKI was thought to be ischemia. However, more recently the focus has shifted from clinical causes to the underlying cellular processes inherent in these situations, in particular the dysfunction of mitochondria in AKI. UNI-494 is a novel nicotinamide ester derivative and selective mitochondrial ATP-sensitive potassium channel activator that improves mitochondrial function and may be beneficial for several disease states, including AKI. This study aims to evaluate the safety, tolerability, and pharmacokinetics of UNI-494. Method This is a single-center, double-blind, placebo-controlled, randomized single ascending dose (SAD) (Part 1) and multiple ascending dose (MAD) (Part 2) study in healthy male subjects and female subjects of non-childbearing potential. Part 1 will enroll up to approximately 40 subjects in 5 cohorts of 8 subjects each (randomized to a ratio of 6 active and 2 placebo per cohort). There will be an interim decision meeting after each cohort/period, to review the safety, tolerability, and PK data up to 48 h post-dose in order to decide the dose level for the subsequent cohort. Part 2 will enroll approximately 20 subjects in 2 cohorts of 10 subjects each, randomized to a ratio of 8 active treatment to 2 placebo who will be dosed for 5 days. The dose level for the Part 2 Cohort 1 will be selected based on the safety, tolerability and PK data from Part 1. Results We intend to present results elucidating safety, tolerability, and pharmacokinetics of UNI-494 in healthy subjects. Conclusion The safety, tolerability, and pharmacokinetics of UNI-494 in healthy subjects will be evaluated in this study.

Diazoxide attenuates DOX-induced cardiotoxicity in cultured rat myocytes.

Doxorubicin (DOX)-induced cardiotoxicity is a well known clinical problem, and many investigations have been made of its possible amelioration. We have investigated whether diazoxide (DIA), an agonist at mitochondrial ATP-sensitive potassium channels (mitoKATP), could reverse DOX-induced apoptotic myocardial cell loss, in cultured rat cardiomyocytes. The role of certain proteins in this pathway was also studied. The rat cardiomyocyte cell line (H9c2) was treated with DOX, and also co-treated with DOX and DIA, for 24 h. Distribution of actin filaments, mitochondrial membrane potential, superoxide dismutase (SOD) activity, total oxidant and antioxidant status (TOS and TAS, respectively), and some protein expressions, were assessed. DOX significantly decreased SOD activity, increased ERK1/2 protein levels, and depolarised the mitochondrial membrane, while DIA co-treatment inhibited such changes. DIA co-treatment ameliorated DOX-induced cytoskeletal changes via F-actin distribution and mitoKATP structure. Co-treatment also decreased ERK1/2 and cytochrome c protein levels. Cardiomyocyte loss due to oxidative stress-mediated apoptosis is a key event in DOX-induced cytotoxicity. DIA had protective effects on DOX-induced cardiotoxicity, via mitoKATP integrity, especially with elevated SUR2A levels; but also by a cascade including SOD/AMPK/ERK1/2. Therefore, DIA may be considered a candidate agent for protecting cardiomyocytes against DOX chemotherapy.

Exploring the Role of Surface and Mitochondrial ATP-Sensitive Potassium Channels in Cancer: From Cellular Functions to Therapeutic Potentials.

ATP-sensitive potassium (KATP) channels are found in plasma membranes and mitochondria. These channels are a type of ion channel that is regulated by the intracellular concentration of adenosine triphosphate (ATP) and other nucleotides. In cell membranes, they play a crucial role in linking metabolic activity to electrical activity, especially in tissues like the heart and pancreas. In mitochondria, KATP channels are involved in protecting cells against ischemic damage and regulating mitochondrial function. This review delves into the role of KATP channels in cancer biology, underscoring their critical function. Notably responsive to changes in cellular metabolism, KATP channels link metabolic states to electrical activity, a feature that becomes particularly significant in cancer cells. These cells, characterized by uncontrolled growth, necessitate unique metabolic and signaling pathways, differing fundamentally from normal cells. Our review explores the intricate roles of KATP channels in influencing the metabolic and ionic balance within cancerous cells, detailing their structural and operational mechanisms. We highlight the channels' impact on cancer cell survival, proliferation, and the potential of KATP channels as therapeutic targets in oncology. This includes the challenges in targeting these channels due to their widespread presence in various tissues and the need for personalized treatment strategies. By integrating molecular biology, physiology, and pharmacology perspectives, the review aims to enhance the understanding of cancer as a complex metabolic disease and to open new research and treatment avenues by focusing on KATP channels. This comprehensive approach provides valuable insights into the potential of KATP channels in developing innovative cancer treatments.

Effect of esaxerenone on ischemia and reperfusion injury in rat hearts.

In myocardial infarction, the addition of mineralocorticoid receptor blockers (MRBs) to standard therapies, such as angiotensin-converting enzyme inhibitors or beta-blockers, reportedly reduces mortality and cardiac events. We investigated whether the non-steroidal MRB esaxerenone has cardioprotective effects and its protective mechanisms. Isolated rat hearts were Langendorff-perfused (constant pressure, 80 mmHg) with oxygenated Krebs-Henseleit bicarbonate buffer and reperfused for 60 min; afterwards, recovery of function (left ventricular pressure, measured with an intraventricular balloon) and myocardial injury were measured. In a preliminary study, we determined the optimal concentration of esaxerenone required for myocardial protection. Next, esaxerenone was administered in the pre- and post-ischemic phases to determine the optimal timing of administration. In addition, we assessed coronary flow response to acetylcholine (ACh) with and without esaxerenone. We examined whether esaxerenone-induced cardioprotection was prevented by targeting putative components in the preconditioning manner (the mitochondrial ATP-sensitive potassium (KATP) channel). Myocardial protection by esaxerenone was observed when esaxerenone was administered before ischemia but not after ischemia. The coronary flow response to ACh was significantly better in the esaxerenone group than in the control group. The cardioprotective effect of esaxerenone was eliminated by the mitochondrial KATP channel blocker, 5-hydroxy decanoate. This study confirmed the myocardial protective effect of the pre-ischemic administration of esaxerenone. Esaxerenone may contribute to coronary endothelial protection and exert pharmacological preconditioning via the mitochondrial KATP channel.

Postconditioning with D-limonene exerts neuroprotection in rats via enhancing mitochondrial activity

Abstract Objectives The key component of neuroprotection after cerebral ischemia–reperfusion (I–R) injury is mitochondrial improvement. By focusing on the function of mitochondrial biogenesis and ATP-sensitive potassium (mK–ATP) channels and inflammatory responses, the current study assessed the neuroprotective potentials of lemon essential oil, D-limonene (LIM), in rats with cerebral I–R injury. Methods In order to simulate cerebral I–R injury, Sprague Dawley rats (n=72) were subjected to a two h local ischemia induced by middle cerebral artery blockage, followed by a 24 h reperfusion period. Five minutes before starting reperfusion, rats were intraperitoneally given LIM at doses of 10 or 100 mg/kg. Cerebral infarct volume was assessed by triphenyl-tetrazolium chloride staining, brain activity by behavioral tests and mitochondrial function/biogenesis, as well as proinflammatory cytokines by fluorometry, immunoblotting and other related techniques. Results When compared to the untreated control group, the administration of LIM substantially and dose-dependently decreased cerebral infarct volumes and neurological deficits (p<0.01). I–R injury-induced alterations in mitochondrial membrane depolarization, mitochondrial reactive oxygen species (mitoROS), and superoxide dismutase (mnSOD), as well as inflammatory cytokines TNF-α, IL-6 and IL-1β, were all significantly reversed after treatment with LIM 100 mg/kg (p<0.01). Additionally, this dose of LIM increased the expression of mitochondrial biogenesis proteins PGC-1α, TFAM, and NRF1. Interestingly, blockage of mK–ATP channels by 5-hydoxydecanoate diminished the effects of LIM on cerebral positive endpoints, cytokines production, and mitochondrial function/biogenesis. Conclusions Thus, the strong neuroprotective effects of LIM-postconditioning were mediated by an increase in mK–ATP channel activity, which improved mitochondrial biogenesis and suppressed inflammatory responses.

Baicalin alleviates bleomycin-induced early pulmonary fibrosis in mice via the mitoKATP signaling pathway

Bleomycin (BLM), a frequently employed chemotherapeutic agent, exhibits restricted clinical utility owing to its pulmonary toxicity. Meanwhile, baicalin (BA)—an active ingredient extracted from the roots of Scutellaria baicalensis Georgi —has been shown to alleviate BLM-induced pulmonary fibrosis (PF). Hence, the objective of this study was to examine the protective effects of BA in the context of BLM-induced early PF in mice and elucidate the underlying mechanism(s). We established an in vivo BLM (3.5 mg/kg)-induced PF murine model and in vitro BLM (35 μM)-damaged MLE-12 cell model. On Day 14 of treatment, the levels of fibrosis and apoptosis were evaluated in mouse lungs via hydroxyproline analysis, western blotting (COL1A1, TGF-β, Bax, Bcl-2, cleaved caspase-3), and Masson, immunohistochemical (α-SMA, AIF, Cyto C), and TUNEL staining. Additionally, in vitro, apoptosis was assessed in MLE-12 cells exposed to BLM for 24 h using the Annexin V/PI assay and western blotting (Bax, Bcl-2, cleaved caspase-3, AIF, Cyto C). To elucidate the role of the mitochondrial ATP-sensitive potassium channel (mitoKATP) in the protective effect of BA, we utilised diazoxide (DZX)—a mitoKATP agonist—and 5-hydroxydecanoate sodium (5-HD)—a mitoKATP inhibitor. Results revealed the involvement of mitoKATP in the protective effect of BA in BLM-induced PF. More specifically, mitoKATP activation can attenuate BLM-induced PF progression and mitigate alveolar epithelial type II cell death by reducing mitochondrial ROS, maintaining the mitochondrial membrane potential, and impeding the mitochondrial apoptotic pathway. Collectively, the findings offer pharmacological support to use BA for the treatment or prevention of BLM-induced PF and suggest that mitoKATP might serve as an effective therapeutic target for this condition.

#6485 UNI-494 LOWERS URINARY B2-MICROGLOBULIN LEVELS IN RATS

Abstract Background and Aims Mitochondrial dysfunction in renal cells play a critical role in the pathophysiology of acute kidney injury (AKI) and chronic kidney disease (CKD). The proximal tubule is the primary sensor and effector in CKD progression and AKI. The measurement of urine Β2-microglobulin (Β2M) is a sensitive assay for proximal tubule injury. Nicorandil, a selective mitochondrial ATP-sensitive potassium channel activator, may be a promising AKI treatment, but its clinical use is limited by serious gastrointestinal side effects and rapid absorption and elimination. UNI-494, a novel nicorandil prodrug designed to improve its pharmacologic properties, may increase the short half-life and improve the safety profile of nicorandil. This study presents the efficacy data on the effects of UNI-494 in a rat ischemia-reperfusion (I/R) model, evaluating biomarkers of renal injury. Method 49 male Sprague-Dawley rats were randomly assigned to 4 groups to evaluate the in vivo efficacy of UNI-494 in a bilateral renal (I/R) model. Group 1, the Sham group, consisted of 10 subjects while groups 2–4 each had 13. Group 1 received no treatment, Group 2 a vehicle, Group 3 10mg/kg of UNI-494, and Group 4 20mg/kg of UNI-494. Treatments were administered as a single dose on Day 0, 1 hour prior to modeling. Body weights were measured on Days -1, 0, and 1. Urine Β2M levels and urinary albumin-to-creatinine ratio (UACR) samples were collected within 24 hours after the surgery using the metabolic cages. Serum blood urea nitrogen (BUN) and creatinine samples were collected on Day -1 (pre-modeling) and 24 hours after modeling. T-tests were used to evaluate statistical differences between groups. Results Β2M levels were significantly lower for the 20 mg/kg UNI-494 dose group compared to the vehicle group, and Β2M content in both UNI-494 dose groups (10 and 20 mg/kg) was significantly lower compared to the vehicle group (Fig. 1). Β2M levels and content were lower for the 20 mg/kg UNI-494 dose group than for the 10 mg/kg dose group (Fig. 1). There was no difference in body weight changes between the UNI-494 dose groups and the vehicle group. There were no statistically significant differences among all groups for serum creatinine and BUN pre-modeling and in UACR. Conclusion Lower levels of urine Β2M levels in the higher UNI-494 dose group compared to the other groups indicate that UNI-494 may have a renoprotective effect. Additionally, the lower levels of Β2M in the higher dose group compared to the lower dose group indicate a dose-response trend. Our study results are consistent with previously published data with nicorandil in a similar rat model of I/R. This potential renoprotective effect should be further investigated.

#6489 UNI-494 DOES NOT SIGNIFICANTLY AFFECT RAT RESPIRATORY FUNCTION

Abstract Background and Aims Nicorandil, a selective mitochondrial ATP-sensitive potassium channel activator, may be beneficial for several disease states, including acute kidney injury. However, its clinical use is limited by serious gastrointestinal side effects and rapid absorption and elimination. UNI-494, a novel nicorandil prodrug designed to improve its pharmacologic properties, may increase the short half-life and improve the safety profile of nicorandil. We present safety results from a safety pharmacology study evaluating the effects of UNI-494 on respiratory function in a rat model. Method The effects of UNI-494 on respiratory function (peak inspiratory and expiratory flow, inspiratory and expiratory time, minute volume, and respiratory rate) were evaluated using whole body plethysmography test in 4 groups of male Wistar Han rats (n = 8 per group). In addition to a control group, 3 test groups were administered doses of 2, 20, or 200 mg UNI-494/kg. Airway function parameter data generated were acquired using specialized EMKA Technologies software (IOX version 2.9.5; DATANLYST version 2.6.1) connected to Emka plethysmographic equipment. Recordings were taken 3 hours before and 4 hours after administration of the control and test items. Effects were reported at 0, 10, 15, 20, 30, 60, 90, 120, 180, and 240 minutes after administration. Results UNI-494 had no relevant effects on rat respiratory function over the 240-minute test period at any dose (2, 20, or 200 mg UNI-494/kg). There were small but insignificant differences in peak inspiratory and expiratory flow (Figure 1, 2), minute volume, respiratory rate, and inspiratory and expiratory time between the control group and the 200 mg UNI-494/kg group. Conclusion Rat respiratory function was not significantly affected by UNI-494. Future studies should continue to evaluate the safety and efficacy of this novel drug.

Competitive interaction between ATP and GTP regulates mitochondrial ATP-sensitive potassium channels

Mitochondrial ATP-sensitive K+ channels (mitoKATP) have been recently characterized structurally, and possess a protein through which K+ enters mitochondria (MitoKIR), and a regulatory subunit (mitoSUR). The mitoSUR regulatory subunit is an ATP-binding cassette (ABC) protein isoform 8 (ABCB8). Opening these channels is known to be cardioprotective, but the molecular and physiological mechanisms that activate them are not fully known. Here, to better understand the molecular and physiological mechanisms of activators (GTP) and inhibitors (ATP) on the activity of mitoKATP, we exposed isolated mitochondria to both nucleotides. We also used molecular docking directed to the nucleotide-binding domain of human ABCB8/mitoSUR to test a comparative model of ATP and GTP effects. As expected, we find that ATP dose-dependently inhibits mitoKATP activity (IC50 = 21.24 ± 1.4 μM). However, simultaneous exposure of mitochondria to GTP dose-dependently (EC50 = 13.19 ± 1.33 μM) reversed ATP inhibition. Pharmacological and computational studies suggest that GTP reverses ATP activity competitively. Docking directed to the site of crystallized ADP reveals that both nucleotides bind to mitoSUR with high affinity, with their phosphates directed to the Mg2+ ion and the walker A motif of the protein (SGGGKTT). These effects, when combined, result in GTP binding, ATP displacement, mitochondrial ATP-sensitive K+ transport, and lower formation of reactive oxygen species. Overall, our findings demonstrate the basis for ATP and GTP binding in mitoSUR using a combination of biochemical, pharmacological, and computational experiments. Future studies may reveal the extent to which the balance between ATP and GTP actions contributes toward cardioprotection against ischemic events.

Modulators of mitochondrial ATP-sensitive potassium channel affect cytotoxicity of heavy metals: Action on isolated rat liver mitochondria and AS-30D ascites hepatoma cells

Heavy metals are ubiquitous environmental pollutants that are extremely dangerous for public health, but the molecular mechanisms of their cytotoxic action are still not fully understood. In the present work, the possible contribution of the mitochondrial ATP-sensitive potassium channel (mK(ATP)), which is usually considered protective for the cell, to hepatotoxicity caused by heavy metals was investigated using polarography and swelling techniques as well as flow cytometry. Using isolated liver mitochondria from adult male Wistar rats and various potassium media containing or not containing penetrating anions (KNO3, KSCN, KAcet, KCl), we studied the effect of mK(ATP) modulators, namely its blockers (5-hydroxydecanoate, glibenclamide, ATP, ADP) and activators (diazoxide, malonate), on respiration and/or membrane permeability in the presence of hepatotoxins such as Cd2+, Hg2+, and Cu2+. It has been shown for the first time that, contrary to Hg2+ and depending on media used, the mK(ATP) modulators affect Cd2+- and/or Cu2+-induced alterations in mitochondrial swelling and respiration rates, although differently, nevertheless, in the ways compatible with mK(ATP) participation in both these cases. On rat AS-30D ascites hepatoma cells, it was found that, unlike Cd2+, an increase in the production of reactive oxygen species was observed with the simultaneous use of Cu2+ and diazoxide; in addition, there was no protective effect of diazoxide against cell death, which also occurred in the presence of Cu2+. In conclusion, the relationships (functional, structural and/or regulatory) between mK(ATP), components of the mitochondrial electron transport chain (CI, CII-CIII and/or ATP synthase, CV) and mitochondrial permeability transition pores were discussed, as well as the role of these molecular structures in the mechanisms of the cytotoxic action of heavy metals.

Electrochemical studies of the mitochondrial ROMK2 potassium channel activity reconstituted into the free-standing and tethered bilayer lipid membranes

The renal-outer-medullary‑potassium (ROMK2) channel modulates potassium transport in the kidney. It has been postulated that the ROMK2 is the pore-forming subunit of the mitochondrial ATP-sensitive potassium channel as a mediator of cardioprotection. In this study, cell-free synthesis of the ROMK2 was performed in presence of membrane scaffold protein (MSP1D1) nanodiscs. Activity measurements were achieved after channel reconstitution into the planar lipid bilayer and tethered bilayer lipid membranes. Both methods allowed for monitoring of channel function, verified with channel blocking and activation/re-activation experiments. The primary function of the mitochondrial potassium channels is to regulate the potential of the mitochondrial membrane, which allows them to play an important role in cytoprotection. This work focuses on obtaining the ROMK2 using a cell-free expression system, followed by the incorporation of the channel protein into the lipid bilayer and studying the influence of voltage changes and molecular modulators on channel activity. Channel activity was measured after its reconstitution into two models of lipid bilayers - BLM (Bilayer Lipid Membrane) and tBLM (Tethered Bilayer Lipid Membrane) deposited on a solid gold electrode. These two model membranes and electrochemical measurements made it possible to measure the flux of K+ ions in the presence of channel modulators.

Effects of Mitochondrial ATP-Sensitive Potassium Channel in Rats with Acute Myocardial Infarction and Its Association with the AKT/mTOR Pathway.

Myocardial infarction is associated with the autophagy and apoptosis of cardiomyocytes, and the protein kinase B/mammalian target of rapamycin (AKT/mTOR) pathway plays a crucial role in this mechanism. Acute myocardial infarction rat models were assessed 0.5, 2, 4, and 6 hours after the induction of the myocardial infarction using hematoxylin and eosin staining, triphenyl tetrazolium chloride staining, myocardial enzyme measurements, and levels of autophagic activity. Additionally, diazoxide, 5-hydroxydecanoate, and LY294002 were intraperitoneally administered to rat models at peak myocardial injury to assess their effects on cardiac injury. The expression levels of autophagy-related and apoptosis-related proteins, as well as p-AKT and p-mTOR, were measured. Electron microscopy was used to assess the ultrastructure and the number of autophagosomes in the cardiac tissue. We demonstrated that the degree of myocardial injury and the level of autophagy were significantly elevated in the experimental cohort compared with the control cohort. In addition, the myocardial infarct size was significantly smaller in diazoxide-treated acute myocardial infarction rats compared with untreated rats. Diazoxide also decreased the levels of myocardial injury markers, autophagy, and apoptosis, while it also induced the levels of AKT and mTOR phosphorylation, decreased the number of autophagosomes, and improved the myocardial ultrastructure of the acute myocardial infarction rats. 5-Hydroxydecanoate treatment resulted in an opposite effect to those observed upon diazoxide treatment. LY294002 was also able to reverse diazoxide treatment effects. Peak levels of myocardial tissue injury and autophagy were observed 2 hours post-acute myocardial infarction induction in rats. Diazoxide treatment inhibited myocardial autophagy and apoptosis while protecting cardiac tissue from ischemic injury, which is likely to have proceeded through activation of the AKT/mTOR pathway.

Urocortin I Protects against Myocardial Ischemia/Reperfusion Injury by Sustaining Respiratory Function and Cardiolipin Content via Mitochondrial ATP-Sensitive Potassium Channel Opening.

Objective Our experiments were aimed at probing whether urocortin I postconditioning was beneficial for maintaining the mitochondrial respiratory function and inhibiting the surging of reactive oxygen species. In addition, our experiments also intended to reveal the relationships between urocortin I postconditioning and mitochondrial ATP-sensitive potassium channel. Methods Langendorff and MPA perfusion systems were used to establish myocardial ischemia-reperfusion injury model and cardiomyocytes hypoxia-reoxygenation injury model in rats, respectively. Isolated hearts and cardiomyocytes were randomly divided into normal group, ischemia-reperfusion/hypoxia-reoxygenation group, urocortin I postconditioning group, and 5-hydroxysolanoic acid (5-HD)+urocortin I group. At the end of balance (T1) and reperfusion (T2), cardiac functions, mitochondrial state3 respiratory, respiratory control ratio, mitochondrial respiratory enzyme activity, and mitochondrial cardiolipin content were measured. Our experiments also observed the ultrastructure of myocardium. The changes of cardiomyocyte mitochondrial permeability transition pore, mitochondrial membrane potential, reactive oxygen species, expression of apoptosis protein, and cardiomyocytes activity were detected at the end of reoxygenation. Results The cardiac functions, mitochondrial respiratory function, and enzyme activity of the normal group were better than other three groups at T2, and urocortin I postconditioning group was better than the IR group and 5-HD+urocortin I group. LVEDP, +dp/dtmax, mitochondrial respiratory function, and enzyme activity of IR group were worse than 5-HD+urocortin I group. Cardiolipin content of the normal group was higher than the other three groups at T2, urocortin I postconditioning group was higher than the IR group and 5-HD+urocortin I group, and 5-HD+urocortin I group was still higher than the IR group. The ultrastructure of the normal group maintained the most integrated than the other groups, IR group suffered the most serious damage, and ultrastructure of the urocortin I postconditioning group was better than the IR group and 5-HD+urocortin I group. At the end of reoxygenation, activity of mitochondrial permeability transition pore and generation of reactive oxygen species of normal group were lower than the other groups, HR group and 5-HD+urocortin I group were higher than the urocortin I postconditioning group, and 5-HD+urocortin I group was still higher than the urocortin I postconditioning group. Normal group had the highest level of mitochondrial membrane potential at the end of reoxygenation, and the urocortin I postconditioning group was higher than the HR group and 5-HD+urocortin I group. The normal group had the lowest expression level of Bax and the highest expression level of Bcl-2 at the end of reoxygenation. Urocortin I postconditioning group had lower Bax expression but higher Bcl-2 expression than the HR and 5-HD+urocortin I group. Accordingly, the normal group had the highest activity of cardiomyocytes, and the urocortin I postconditioning group was higher than the HR group and 5-HD+urocortin I group. Conclusions Urocortin I postconditioning can protect the activity of cardiomyocytes after hypoxia-reoxygenation injury, improve the mitochondrial respiratory function, and enhance the contractility of isolated heart after myocardial ischemia-reperfusion injury. The alleviation of myocardial injury relates to the opening of mitochondrial ATP-sensitive potassium channel.

Association Between Specificity of Sulfonylureas to Cardiac Mitochondrial KATP Channels and the Risk of Major Adverse Cardiovascular Events in Type 2 Diabetes.

Previous studies have revealed an intraclass difference in major adverse cardiovascular events (MACE) among sulfonylureas. In vitro and ex vivo studies reported several sulfonylureas to exhibit high-affinity blockage of cardiac mitochondrial ATP-sensitive potassium (mitoKATP) channels and could interfere with ischemic preconditioning, the most important mechanism of self-cardiac protection. However, no studies have examined whether these varying binding affinities of sulfonylureas could account for their intraclass difference in MACE. We compared mitoKATP channel high-affinity versus low-affinity sulfonylureas regarding the MACE risk in real-world settings. Using the Taiwan nationwide health care claims database, patients with type 2 diabetes initiating sulfonylurea monotherapy between 2007 and 2016 were included in the cohort study. A total of 33,727 new mitoKATP channel high-affinity (glyburide and glipizide) and low-affinity (gliclazide and glimepiride) sulfonylurea users, respectively, were identified after 1:1 propensity score matching. Cox proportional hazard models were used to estimate adjusted hazard ratios (aHRs) and 95% CI. MitoKATP channel high-affinity sulfonylureas were associated with a significantly increased risk of three-point MACE (aHR 1.21 [95% CI 1.03-1.44]), ischemic stroke (aHR 1.23 [95% CI 1.02-1.50]), and cardiovascular death (aHR 2.61 [95% CI 1.31-5.20]), but not with that of myocardial infarction (aHR 1.04 [95% CI 0.75-1.46]). The duration-response analyses revealed the highest MACE risk to be within 90 days of therapy (aHR 4.67 [95% CI 3.61-6.06]). Cardiac mitoKATP channel high-affinity sulfonylureas were associated with an increased MACE risk compared with low-affinity sulfonylureas in a nationwide population with diabetes.

Interaction of nitric oxide synthase and mitochondrial ATP-sensitive potassium channels in protective impacts of combination therapy with irisin-preconditioning and melatonin-postconditioning in myocardial ischemia/reperfusion in aging

ObjectivesThe main challenge for cardioprotection in patients with cardiac ischemia–reperfusion (I/R) damage is the existence of negatively-acting multiple risk factors such as aging. This work aimed to evaluate the efficacy of combined therapy with irisin and melatonin on cardiac I/R injury in aged rats by focusing on the involvement of mitochondrial ATP-sensitive potassium (mitoKATP) channels, and the eNOS/NO pathway. MethodsMale aged Wistar rats (78 rats, 400–450 g) were experienced LAD ligation for 30 min followed by 3 h or 2 weeks reperfusion. Irisin (0.5 mg/kg/day) was administered for one week before ischemic insult. Melatonin (20 mg/kg) was administered 10 min before the onset of reperfusion. The releases of CK and LDH, mitochondrial function, inflammatory and apoptosis responses, the expression of eNOS, the levels of NO, cardiac function, and infarct size (IS) were evaluated. ResultsCombined therapy with irisin and melatonin significantly improved heart function, reduced IS, and limited the releases of CK and LDH (P < 0.01 for all). This combined therapy improved mitochondrial function, decreased inflammatory cytokines and apoptosis response, and increased eNOS/NO activity (P < 0.05 for all). Monotherapies had no significant impacts. Inhibition of mitoKATP channels and eNOS enzyme by 5HD and L-NAME, respectively, reversed the cardioprotective effects of combination therapy (P < 0.05), however, the effect of 5HD was more potent as compared to L-NAME (P < 0.05). ConclusionsIt can be suggested that combined therapy with irisin and melatonin can be proposed as a potential candidate to mitigate the I/R severity of aged hearts via activation of mitoKATP channels partly through the eNOS/NO signaling pathway.

Role of caveolin-eNOS platform and mitochondrial ATP-sensitive potassium channel in abrogated cardioprotective effect of ischemic preconditioning in postmenopausal women

Role of caveolin-eNOS platform and mitochondrial ATP-sensitive potassium channel in abrogated cardioprotective effect of ischemic preconditioning in postmenopausal women

Ginsenoside-Mc1 reduces cerebral ischemia-reperfusion injury in hyperlipidemia through mitochondrial improvement and attenuation of oxidative/endoplasmic reticulum stress

The neuroprotective effect of ginsenoside-Mc1 (GMc1) in hyperlipidemic rats in the setting of cerebral ischemiareperfusion injury (I/RI), as well as the role of mitochondrial ATP-sensitive potassium (mitoKATP) channels and oxidative/ endoplasmic reticulum (ER) stress, was investigated. Hyperlipidemia (8 weeks) was induced by a high-fat diet in Sprague Dawley rats. GMc1 (10 mg/kg, i.p.) was given to hyperlipidemic rats daily for one month before I/RI. Rat brains were subjected to 2 h of local ischemia followed by 24 h reperfusion. The cerebral infarcted injury was measured by triphenyltetrazolium chloride staining and the levels of oxidative stress indicators were detected by ELISA and spectrophotometry. A fluorometric technique was employed to evaluate mitochondrial function. Western blotting was used to detect changes in the expression of ER stress proteins. GMc1 reduced cerebral infarct volume in hyperlipidemic rats in comparison to untreated ones (P&lt;0.01). GMc1 reduced cerebral infarct volume in hyperlipidemic rats as compared to untreated rats (P&lt;0.01). GMc1 significantly decreased mitochondrial membrane depolarization, mitochondrial reactive oxygen species (mitoROS) and malondialdehyde levels (P&lt;0.01), while increasing the activity of superoxide dismutase (SOD), catalase (CAT) and glutathione-peroxidase (GPx) (P&lt;0.001). GMc1 administration reduced the expression of ER stress markers, including phosphorylated (p)-endoplasmic reticulum kinase (PERK), p-eukaryotic translation initiation factor 2 subunit 1 (elF2?), and C/EBP homologous protein (CHOP). Inhibition of mitoKATP channels with hydroxydecanoate significantly eliminated the protective impacts of GMc1 in hyperlipidemic rats subjected to cerebral I/RI. The neuroprotective effect of GMc1 preconditioning was remarkably improved by increasing mitoKATP channel activity and decreasing oxidative and ER stress levels in hyperlipidemic rats, implying that this compound could be an appropriate candidate for reducing cerebral I/RI in comorbidities.