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An increased flux of potassium ions into the mitochondrial matrix through the ATP-sensitive potassium channel (mitoKATP) has been shown to provide protection against ischemia-reperfusion injury. Recently, it was proposed that the mitochondrial-targeted isoform of the renal outer medullary potassium channel (ROMK) protein creates a pore-forming subunit of mitoKATP in heart mitochondria. Our research focuses on the properties of mitoKATP from heart-derived H9c2 cells. For the first time, we detected single-channel activity and describe the pharmacology of mitoKATP in the H9c2 heart-derived cells. The patch-clamping of mitoplasts from wild type (WT) and cells overexpressing ROMK2 revealed the existence of a potassium channel that exhibits the same basic properties previously attributed to mitoKATP. ROMK2 overexpression resulted in a significant increase of mitoKATP activity. The conductance of both channels in symmetric 150/150 mM KCl was around 97 ± 2 pS in WT cells and 94 ± 3 pS in cells overexpressing ROMK2. The channels were inhibited by 5-hydroxydecanoic acid (a mitoKATP inhibitor) and by Tertiapin Q (an inhibitor of both the ROMK-type channels and mitoKATP). Additionally, mitoKATP from cells overexpressing ROMK2 were inhibited by ATP/Mg2+ and activated by diazoxide. We used an assay based on proteinase K to examine the topology of the channel in the inner mitochondrial membrane and found that both termini of the protein localized to the mitochondrial matrix. We conclude that the observed activity of the channel formed by the ROMK protein corresponds to the electrophysiological and pharmacological properties of mitoKATP.

Coronavirus disease 2019 (COVID-19) causes pulmonary edema, which disrupts the lung alveoli–capillary barrier and leads to pulmonary cell apoptosis, the main cause of death. However, the molecular mechanism behind SARS-CoV-2’s apoptotic activity remains unknown. Here, we revealed that SARS-CoV-2-ORF-3a mediates the pulmonary pathology associated with SARS-CoV-2, which is demonstrated by the fact that it causes lung tissue damage. The in vitro results showed that SARS-CoV-2-ORF-3a triggers cell death via the disruption of mitochondrial homeostasis, which is modulated through the regulation of Mitochondrial ATP-sensitive Potassium Channel (MitoKATP). The addition of exogenous Potassium (K+) in the form of potassium chloride (KCl) attenuated mitochondrial apoptosis along with the inflammatory interferon response (IFN-β) triggered by SARS-ORF-3a. The addition of exogenous K+ strongly suggests that dysregulation of K+ ion channel function is the central mechanism underlying the mitochondrial dysfunction and stress response induced by SARS-CoV-2-ORF-3a. Our results designate that targeting the potassium channel or its interactions with ORF-3a may represent a promising therapeutic strategy to mitigate the damaging effects of infection with SARS-CoV-2.

Bronchial asthma is the common chronic inflammatory disease and is characterized by chronic airway inflammation, airway remodeling, and airway hyperreactivity (AHR). Aim of this study was to investigate the effects of mitochondrial ATP-sensitive potassium channels (MitoKATP) on the proliferation and secretion of human airway smooth muscle cells (HASMCs). HASMCs were treated with the serum from asthmatic patients to establish HASMCs asthma model of passive sensitization. Rhodamine 123 (R-123) and 2,7-dichloro-dihydrofluorescein diacetate (DCFH-DA) fluorescence staining were used to detect mitochondrial membrane potential (Δψm) and the content of reactive oxygen species (ROS) in the cells, respectively. The cell counting was used to detect cell proliferation, and RT-PCR was used to detect the expression of TGF-β1 mRNA. In the normal + Diazoxide group, the fluorescence intensity of R-123, ROS content, cell proliferation and TGF-β1 expression were enhanced, compared with the normal control group (p<0.05). There were no significant differences between the normal + 5-hydroxydecanoate (5-HD) group and the normal control group. In the asthma model control group, the fluorescence intensity of R-123, ROS content, cell proliferation and TGF-β1 expression were enhanced, compared with normal control group, (p<0.05). The aforementioned indices were enhanced in the asthma model + Diazoxide group, when compared with the asthma model control group, whereas these indices were attenuated in the asthma model + 5-HD group, when compared with the asthma model control group (p<0.05). In conclusion, asthma could activate MitoKATP channels in HASMCs, promote HASMC proliferation and TGF-β1 expression.

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.

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.

Neuronal pyroptosis serves an important role in the progress of neurologic dysfunction following subarachnoid hemorrhage (SAH), which is predominantly caused by a ruptured aneurysm. Hydrogen gas has been previously reported to be an effective anti-inflammatory agent against ischemia-associated diseases by regulating mitochondrial function. The objective of the present study was to investigate the potential neuroprotective effects of hydrogen gas post-conditioning against neuronal pyroptosis after SAH, with specific focus on the mitochondrial ATP-sensitive K+ (mitoKATP) channels. Following SAH induction by endovascular perforation, rats were treated with inhalation of 2.9% hydrogen gas for 2 h post-perforation. Neurologic deficits, brain water content, reactive oxygen species (ROS) levels, neuronal pyroptosis, phosphorylation of ERK1/2, p38 MAPK and pyroptosis-associated proteins IL-1β and IL-18 were evaluated 24 h after perforation by a modified Garcia method, ratio of wet/dry weight, 2',7'-dichlorofluorescin diacetate, immunofluorescence and western blot assays, respectively. An inhibitor of the mitoKATP channel, 5-hydroxydecanoate sodium (5-HD), was used to assess the potential role of the mitoKATP-ERK1/2-p38 MAPK signal pathway. Hydrogen gas post-conditioning significantly alleviated brain edema and improved neurologic function, reduced ROS production and neuronal pyroptosis, suppressed the expression of IL-1β and IL-18 whilst upregulating ERK1/2 phosphorylation, but downregulated p38 MAPK activation 24 h post-SAH. These aforementioned effects neuroprotective were partially reversed by 5-HD treatment. Therefore, these observations suggest that post-conditioning with hydrogen gas ameliorated SAH-induced neuronal pyroptosis at least in part through the mitoKATP/ERK1/2/p38 MAPK signaling pathway.

It has previously been demonstrated that hyperglycemia-induced oxidative stress and inflammation are closely associated with the development of diabetic complications, including diabetic neuropathy. Additionally, mitochondrial ATP-sensitive potassium (Mito-K-ATP) channels play a homeostatic role on blood glucose regulation in organisms. Molecular hydrogen (H2) exhibits anti-inflammatory, anti-antioxidative and anti-apoptotic properties and can be used to treat more than 71 diseases safely. In addition, the diabetes animal models which are set up using streptozotocin (STZ) injection, is a type of high long-term stability, low animal mortality rate and security method. The aim of the current study was to assess the value of hydrogen-rich saline (HS) in diabetic peripheral neuropathy (DPN) treatment and to determine its associated mechanisms in STZ-induced diabetic experimental rats. Additionally, the effects of the Mito-K-ATP channels, oxidative stress, inflammatory cytokines and apoptosis on DPN were also evaluated. From week 5 of STZ injections, HS (2.5, 5 and 10 ml/kg) was injected into the rat abdominal cavity every day for a period of 4 weeks. The results of the current study demonstrated that HS significantly reduced behavioral, biochemical and molecular effects caused by DPN. However, 5-hydroxydecanoate, a selective Mito-K-ATP channels general pathway inhibitor, partially eliminated the therapeutic effect of HS on DPN. These results indicated that the use of HS may be a novel strategy to treat DPN by activating the Mito-K-ATP pathway and reducing oxidative stress, inflammatory cytokines and apoptosis.

The aim of the study reported here was to evaluate whether the mitochondrial ATP-sensitive potassium (mitoK ATP ) channel could participate in the effect of dexmedetomidine on cerebral ischemia-reperfusion (I/R) rats. Forty rats were randomly assigned into 5 groups: sham operation (S) group; cerebral I/R group; dexmedetomidine (D) group; 5-hydroxydecanoate (5-HD) group; 5-HD + D group. The cerebral I/R were produced by 2 h right middle cerebral artery occlusion followed by 24 h reperfusion. Dexmedetomidine (50μg/kg) was injected intraperitoneally before ischemia and after the onset of reperfusion. 5-HD (30 mg/kg) was injected intraperitoneally at 1 h before ischemia. The neurological deficit score (NDS) and the levels of super oxide dismutase (SOD), malondialdehyde (MDA), myeloperoxidase (MPO), Interleukin 6 (IL-6) and tumor necrosis factor-α (TNF-α) were evaluated. Compared to group S, NDS and the levels of MDA, MPO, IL-6 and TNF-α were significantly higher, and SOD levels were significantly lower in the other groups ( P  < 0.05). Compared to group I/R,NDS and the levels of MDA, MPO, IL-6 and TNF-α were significantly lower, and SOD level was significantly higher in group D ( P  < 0.05). Compared to group D, NDS and the levels of MDA, MPO, IL-6 and TNF-α were significantly higher, and SOD level was significantly lower in group5-HD + D ( P  < 0.05). The activation of the mitoK ATP channel could contribute to the protective effect of dexmedetomidine on rats induced by focal cerebral ischemia-reperfusion injury.

We previously demonstrated in rats that noninvasive delayed limb ischemic preconditioning (LIPC) induced by three cycles of 5-min occlusion and 5-min reperfusion of the left hind limb per day for three days confers the same cardioprotective effect as local ischemic preconditioning of the heart, but the mechanism has not been studied in depth. The aim of this project was to test the hypothesis that delayed LIPC enhances myocardial antioxidative ability during ischemia-reperfusion by a mitochondrial KATP channel (mito KATP)-dependent mechanism. Rats were randomized to five groups: ischemia-reperfusion (IR)-control group, myocardial ischemic preconditioning (MIPC) group, LIPC group, IR-5HD group and LIPC-5HD group. The MIPC group underwent local ischemic preconditioning induced by three cycles of 5-min occlusion and 5-min reperfusion of the left anterior descending coronary arteries. The LIPC and LIPC-5HD groups underwent LIPC induced by three cycles of 5-min occlusion and 5-min reperfusion of the left hind limb using a modified blood pressure aerocyst per day for three days. All rats were subjected to myocardial ischemia-reperfusion injury. The IR-5HD and LIPC-5HD groups received the mito KATP channel blocker 5-hydroxydecanoate Na (5-HD) before and during the myocardial ischemia-reperfusion injury. Compared with the IR-control group, both the LIPC and MIPC groups showed an amelioration of ventricular arrhythmia, reduced myocardial infarct size, increased activities of total superoxide dismutase, manganese-superoxide dismutase (Mn-SOD) and glutathione peroxidase, increased expression of Mn-SOD mRNA and decreased xanthine oxidase activity and malondialdehyde concentration. These beneficial effects of LIPC were prevented by 5-HD. In conclusion, delayed LIPC offers similar cardioprotection as local IPC. These results support the hypothesis that the activation of mito KATP channels enhances myocardial antioxidative ability during ischemia-reperfusion, thereby contributing, at least in part, to the anti-arrhythmic and anti-infarct effects of delayed LIPC.

Autophagy has emerged as a powerful process in the response to cellular injury. The present study was designed to investigate signal transduction pathways in angiotensin II (Ang II)-induced autophagy. Rat vascular smooth muscle cells (VSMCs) were stimulated with different doses of Ang II (10 −9 –10 −5  mol/L) for different time periods (6–72 h). Incubation with Ang II increased the production of reactive oxygen species (ROS), increased the LC3-II to LC3-I ratio, increased beclin-1 expression, and decreased SQSTM1/p62 expression in a dose- and time-dependent manner. In addition, Ang II increased autophagosome formation. Increased ROS production induced by Ang II was inhibited by Ang II type 1 receptor (AT1) blockers (Olmesartan and Candesartan, ARB), a NADPH Oxidase inhibitor (apocynin), and mitochondrial K ATP channels inhibitor (5-hydroxydecanoate, 5HD). Ang II (10 −7  mol/L, 48 h)-induced increase in the LC3-II to LC3-I ratio, the formation of autophagosomes, expression of beclin-1 and decrease in the expression of SQSTM1/p62 were also inhibited by pretreatment with 3-methyladenine or bafilomycin A1 (inhibitors of autophagy), olmesartan and candesartan (in dose-dependent manners), apocynin, 5HD, and siRNA Atg5. Our results indicate that Ang II increases autophagy levels via activation of AT1 receptor and NADPH oxidase. Mitochondrial K ATP channels also play an important role in Ang II-induced autophagy. Our results may provide a new strategy for treatment of cardiovascular diseases with Ang II.