Mechanism Of Uncoupling Research Articles

Efficacy of creatine nitrate supplementation on redox status and mitochondrial function in pectoralis major muscle of preslaughter transported broilers

This study was purposed to investigate the efficacy of dietary creatine nitrate (CrN) supplementation on redox status and mitochondrial function in pectoralis major (PM) muscle of broilers that experienced preslaughter transport. A total of 288 Arbor Acres broilers (28-day-old) were randomly assigned into five dietary treatments, including a basal diet or the basal diet supplemented with 600 mg/kg guanidinoacetic acid (GAA), 300, 600, or 900 mg/kg CrN for 14 days, respectively. On the transportation day, the basal diet group was divided into two groups on average, resulting in six groups. The control group was transported for 0.5 h and the other groups for 3 h (identified as Control, T3h, GAA600, CrN300, CrN600, and CrN900 group, respectively), and all crates were randomly placed on the truck travelling at an average speed of 80 km/h. Our results showed that GAA600 and CrN treatments decreased the muscle ROS level and MDA content (P < 0.05) and increased the mitochondrial membrane potential (P < 0.001), as well as a higher mRNA expression of avUCP (P < 0.001) and lower mRNA expressions of Nrf2 (P < 0.001), Nrf2 and PGC-1α (P < 0.05) compared with T3h group. Meanwhile, the mRNA and protein expressions of Nrf1, TFAM, and PGC-1α in CrN600 and CrN900 groups were lower than those in the T3h group (P < 0.05). Conclusively, dietary supplementation with GAA and CrN decreased muscle oxidative products and enhanced mitochondrial uncoupling mechanism and mtDNA copy number, which relieved muscle oxidative damage and maintained mitochondrial function.

Vascular Redox Signaling, Endothelial Nitric Oxide Synthase Uncoupling, and Endothelial Dysfunction in the Setting of Transportation Noise Exposure or Chronic Treatment with Organic Nitrates.

Significance: Cardiovascular disease and drug-induced health side effects are frequently associated with-or even caused by-an imbalance between the concentrations of reactive oxygen and nitrogen species (RONS) and antioxidants, respectively, determining the metabolism of these harmful oxidants. Recent Advances: According to the "kindling radical" hypothesis, the initial formation of RONS may further trigger the additional activation of RONS formation under certain pathological conditions. The present review specifically focuses on a dysfunctional, uncoupled endothelial nitric oxide synthase (eNOS) caused by RONS in the setting of transportation noise exposure or chronic treatment with organic nitrates, especially nitroglycerin (GTN). We further describe the various "redox switches" that are proposed to be involved in the uncoupling process of eNOS. Critical Issues: In particular, the oxidative depletion of tetrahydrobiopterin and S-glutathionylation of the eNOS reductase domain are highlighted as major pathways for eNOS uncoupling upon noise exposure or GTN treatment. In addition, oxidative disruption of the eNOS dimer, inhibitory phosphorylation of eNOS at the threonine or tyrosine residues, redox-triggered accumulation of asymmetric dimethylarginine, and l-arginine deficiency are discussed as alternative mechanisms of eNOS uncoupling. Future Directions: The clinical consequences of eNOS dysfunction due to uncoupling on cardiovascular disease are summarized also, providing a template for future clinical studies on endothelial dysfunction caused by pharmacological or environmental risk factors.

Abstract 2821: Mitigation of radiation-induced colon injuries in mice by oral sepiapterin (PTC923)

Abstract The rectum is the primary dose limiting structure for radiation therapy (RT) in treating prostate cancer. Two clinical forms of rectal injury have been described: early and late. Early injury typically develops during RT and occurs to some degree in most patients. The condition is usually self-limiting, but can predispose patients to the development of late injury. Late rectal injury occurs in up to 25% of patients months to years after pelvic RT and is more concerning, since it can be irreversible and can have a major impact on a patient’s lifestyle. Development of new pharmacologic approaches to eliminate the risk of RT-dependent rectal injury remains an important goal. Although detailed mechanisms underlying normal tissue toxicities vary from tissue to tissue, the central underlying mechanism is abnormal wound healing involving endothelial dysfunction (ED) and subsequently fibrosis. Regardless of its cause, ED is characterized by an “uncoupled” endothelial NOS (eNOS) activity that generates superoxide (O2 -) and peroxynitrite (ONOO-) rather than NO. One mechanism for uncoupling in inflammatory conditions is oxidation of the reduced cofactor tetrahydrobiopterin (BH4). The product of uncoupling, ONOO-, oxidizes BH4, initiating a feed forward mechanism to sustain uncoupling. Oral treatment with sepiapterin (PTC923), a tetrahydrobiopterin precursor, decreased infiltrating inflammatory cells and cytokine levels in mice with colitis. We therefore tested whether a synthetic PTC923 might mitigate radiation-induced colon injuries. C57L/J wild-type 6-8-week-old males mice received 13.5 Gy total-body irradiation (TBI). Starting from 24 h post-irradiation, mice were treated once daily with 1 mg/kg PTC923 for six days by oral gavage. Colon injuries were accessed on 6th day after IR by colonoscopy and scored by using distal-proximal endoscopic colitis scoring system (D-PECS). The system has a core inflammatory component of four non-co-linear parameters and decimal units to notable lesions or complications. Relative expression of cytokines IL-1β, IL-6, IL-17α, and TGF-β1 in colon endothelial cells were measured by RT-PCR 8 days after IR. According with D-PECS estimation, post-irradiation colon injury was significantly reduced by PTC923 treatment: on the 6th day after TBI mice demonstrated D-PECS score 5.8±1.48 vs 1.6±0.55 D-PECS score for animals received PTC923 treatment after TBI (p-value=0.00). Increase of cytokines IL-6 and TGF-β1 in the colon endothelium after TBI was effectively blocked by PTC923 treatment (IL-6: IR+Vehicle = 4.31±1.31 fold increase vs IR+PTC923 = 1.29±0.37 fold increase (p-value=0.0016); TGF-β1: IR+Vehicle = 5.18±0.66 fold increase vs IR+PTC923 = 2.01±0.75 fold increase (p-value=0.00029)). In conclusion, these findings support the proposal that oral treatment with sepiapterin is a potential mitigator of colon injury caused by RT. Citation Format: Vasily A. Yakovlev, Christopher S. Rabender, Ross B. Mikkelsen. Mitigation of radiation-induced colon injuries in mice by oral sepiapterin (PTC923) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2821.

Preliminary study to classify mechanisms of mitochondrial toxicity by in vitro metabolomics and bioinformatics

AimMitochondrial toxicity is one of the causes for drug-induced liver injury, and the classification of phenotypes or mitochondrial toxicity are highly required though there are no molecular-profiling approaches for classifying mitochondrial toxicity. Therefore, the aim of this study was to classify the mechanisms of mitochondrial toxicity by metabolic profiling in vitro and bioinformatics. Main methodsWe applied an established gas chromatography tandem mass spectrometry-based metabolomics to human hepatoma grade 2 (HepG2) cells that were exposed to mitochondrial toxicants, whose mechanisms are different, such as rotenone (0.1 μM), carbonyl cyanide-3-chlorophenylhydrazone (CCCP, 0.5 μM), nefazodone (20 μM), perhexiline (6.25 μM), or digitonin (positive cytotoxic substance, 4 μM). These concentrations were determined by the Mitochondrial ToxGlo Assay. Galactose medium was used for suppressing the Warburg effect in HepG2 cells, and the metabolome analysis successfully identified 125 metabolites in HepG2 cells. Multivariate, metabolic pathway and network analyses were performed by the R software. Key findingsMetabolic profiling enabled the classifying the mitochondrial toxicity mechanisms of RCC inhibition and uncoupling. The metabolic profiles of respiratory chain complex (RCC) inhibitors (rotenone and nefazodone) and an uncoupler (CCCP) were fully differentiated from those of other compounds. The metabolic pathway analysis revealed that the RCC inhibitors and the uncoupler mainly disrupted TCA-cycle and related metabolic pathways. In addition, the correlation-based network analysis revealed that succinic acid, β-alanine, and glutamic acid were potential metabolic indicators for RCC inhibition and uncoupling. SignificanceOur results provided new insights into classifying mechanisms of mitochondrial toxicity by in vitro metabolomics.

Repurposing nitazoxanide as a novel anti-atherosclerotic drug based on mitochondrial uncoupling mechanisms.

The anthelmintic drug nitazoxanide has a mitochondrial uncoupling effect. Mitochondrial uncouplers have been proven to inhibit smooth muscle cell proliferation and migration, inhibit NLRP3 inflammasome activation of macrophages and improve dyslipidaemia. Therefore, we aimed to demonstrate that nitazoxanide would protect against atherosclerosis. The mitochondrial oxygen consumption of cells was measured by using the high-resolution respirometry system, Oxygraph-2K. The proliferation and migration of A10 cells were measured by using Edu immunofluorescence staining, wound-induced migration and the Boyden chamber assay. Protein levels were measured by using the western blot technique. ApoE (-/-) mice were fed with a Western diet to establish an atherosclerotic model in vivo. The in vitro experiments showed that nitazoxanide and tizoxanide had a mitochondrial uncoupling effect and activated cellular AMPK. Nitazoxanide and tizoxanide inhibited serum- and PDGF-induced proliferation and migration of A10 cells. Nitazoxanide and tizoxanide inhibited NLRP3 inflammasome activation in RAW264.7 macrophages, the mechanism by which involved the AMPK/IκBα/NF-κB pathway. Nitazoxanide and tizoxanide also induced autophagy in A10 cells and RAW264.7 macrophages. The in vivo experiments demonstrated that oral administration of nitazoxanide reduced the increase in serum IL-1β and IL-6 levels and suppressed atherosclerosis in Western diet-fed ApoE (-/-) mice. Nitazoxanide inhibits the formation of atherosclerotic plaques in ApoE (-/-) mice fed on a Western diet. In view of nitazoxanide being an antiprotozoal drug already approved by the FDA, we propose it as a novel anti-atherosclerotic drug with clinical translational potential.

Ketones Improve the Mitochondrial Coupling Efficiency of Brown Adipocytes

Brown adipose tissue (BAT) thermogenesis is an energy‐demanding response, and this capacity can be leveraged to dissipate excess energy in obesity and its associated metabolic complications. Therefore, understanding the regulatory mechanisms of BAT mitochondrial uncoupling and thermogenesis is crucial. Ketone bodies are endogenous metabolites mainly found during starvation and carbohydrate restriction; however, their regulatory role on BAT thermogenesis is elusive. In this study, we aim to investigate the potential metabolic reprogramming induced by ketones on BAT. We hypothesize that ketones are alternative fuels for BAT during energy/carbohydrate shortage and decrease mitochondrial uncoupling to counterbalance the systemic energy deficit. In mice, ketogenic diet reduces core body temperature by 0.4˚C. Using Seahorse respirometry in differentiated brown adipocytes, we found that ketones improve mitochondrial coupling efficiency (ATP‐linked respiration/ basal respiration) by 30%, implying an attempt to decrease thermogenesis. The gene Oxct1, encoding succinyl‐CoA:3‐ketoacid CoA transferase that is required for ketolysis, is abundantly expressed by brown adipocytes. BAT‐specific Oxct1 knockout renders mice more resistant to cold‐induced decrease in core body temperature during fasting or ketogenic diet, via mechanisms independent of UCP1. Together, our data suggest that ketones potentially communicate hunger to brown adipocytes, thus promoting mitochondrial coupling efficiency for energy conservation.

Alkyl esters of umbelliferone-4-acetic acid as protonophores in bilayer lipid membranes and ALDH2-dependent soft uncouplers in rat liver mitochondria

A great variety of coumarin-related compounds, both natural and synthetic, being often brightly fluorescent, have shown themselves beneficial in medicine for both therapeutic and imaging purposes. Here, in search for effective uncouplers of oxidative phosphorylation, we synthesized a series of 7-hydroxycoumarin (umbelliferone, UB) derivatives combining rather high membrane affinity with the presence of a hydroxyl group deprotonable at physiological pH - alkyl esters of umbelliferone-4-acetic acid (UB-4 esters) differing in alkyl chain length. Addition of UB-4 esters to isolated rat liver mitochondria (RLM) resulted in their rapid depolarization, unexpectedly followed by membrane potential recovery on a minute time scale. According to TLC and HPLC data, incubation of RLM with UB-4 esters caused their hydrolysis, which led to disappearance of the uncoupling activity (recoupling). Both mitochondrial recoupling and hydrolysis of UB-4 esters were suppressed by inhibitors of mitochondrial aldehyde dehydrogenase (ALDH2), disulfiram and daidzin, thus pointing to the involvement of this enzyme in the recoupling of RLM incubated with UB-4 esters. The protonophoric mechanism of mitochondrial uncoupling by UB-4 esters was proved in experiments with artificial bilayer lipid membranes (BLM): these compounds induced proton-selective electrical current across planar BLM and caused dissipation of pH gradient on liposomes. UB-4 esters showed antibacterial activity against Bacillus subtilis, Staphylococcus aureus and Mycobacterium smegmatis.

Mild pentachlorophenol-mediated uncoupling of mitochondria depletes ATP but does not cause an oxidized redox state or dopaminergic neurodegeneration in Caenorhabditis elegans

AimsMitochondrial dysfunction is implicated in several diseases, including neurological disorders such as Parkinson’s disease. However, there is uncertainty about which of the many mechanisms by which mitochondrial function can be disrupted may lead to neurodegeneration. Pentachlorophenol (PCP) is an organic pollutant reported to cause mitochondrial dysfunction including oxidative stress and mitochondrial uncoupling. We investigated the effects of PCP exposure in Caenorhabditis elegans, including effects on mitochondria and dopaminergic neurons. We hypothesized that mild mitochondrial uncoupling by PCP would impair bioenergetics while decreasing oxidative stress, and therefore would not cause dopaminergic neurodegeneration. ResultsA 48-hour developmental exposure to PCP causing mild growth delay (∼10 % decrease in growth during 48 h, covering all larval stages) reduced whole-organism ATP content > 50 %, and spare respiratory capacity ∼ 30 %. Proton leak was also markedly increased. These findings suggest a main toxic mechanism of mitochondrial uncoupling rather than oxidative stress, which was further supported by a concomitant shift toward a more reduced cellular redox state measured at the whole organism level. However, exposure to PCP did not cause dopaminergic neurodegeneration, nor did it sensitize animals to a neurotoxic challenge with 6-hydroxydopamine. Whole-organism uptake and PCP metabolism measurements revealed low overall uptake of PCP in our experimental conditions (50 μM PCP in the liquid exposure medium resulted in organismal concentrations of < 0.25 μM), and no measurable production of the oxidative metabolites tetra-1,4-benzoquinone and tetrachloro-p-hydroquinone. InnovationThis study provides new insights into the mechanistic interplay between mitochondrial uncoupling, oxidative stress, and neurodegeneration in C. elegans. These findings support the premise of mild uncoupling-mediated neuroprotection, but are inconsistent with proposed broad “mitochondrial dysfunction”-mediated neurodegeneration models, and highlight the utility of the C. elegans model for studying mitochondrial and neurotoxicity. ConclusionsDevelopmental exposure to pentachlorophenol causes gross toxicological effects (growth delay and arrest) at high levels. At a lower level of exposure, still causing mild growth delay, we observed mitochondrial dysfunction including uncoupling and decreased ATP levels. However, this was associated with a more-reduced cellular redox tone and did not exacerbate dopaminergic neurotoxicity of 6-hydroxydopamine, instead trending toward protection. These findings may be informative of efforts to define nuanced mitochondrial dysfunction-related adverse outcome pathways that will differ depending on the form of initial mitochondrial toxicity.

Seismic performance of semi-rigid steel frame infilled with prefabricated damping wall panels

In this research, two innovative semi-rigid steel frames infilled with prefabricated damping wall panels in different types of connection are proposed. Working principles and main features of the developed infill system with both uncoupling mechanism and sliding joints are defined. A quasi-static test was conducted to explore the seismic response of the two systems, as compared with a bare steel frame. Test results confirm that the detrimental infill-frame interaction is prevented by the proposed wall panels. Both specimens infilled with prefabricated damping wall panels characterize a considerably large deformation capacity up to 3.33% drift, while significant crack or corner crushing in the wall panels is not observed, contributed to the configurations of the prefabricated damping wall panels. The infilled steel frames have stable seismic behavior similar to the bare steel frame without infill, indicating that the prefabricated damping wall panels can be designated as an additional safety stock with damping for the steel frame rather than the major component of strength, stiffness, and brittleness. Specifically, thanks to satisfactory energy dissipation capacity of the damping layers, dissipated energy of steel frame infilled with the prefabricated damping wall panels is about 23.3% larger than the bare steel frame after yielding. Moreover, it would be a more effective and controllable configuration that the lateral displacement of the upper wall panel is directly enforced by the top beam.

The Neurological Basis of Developmental Dyslexia and Related Disorders: A Reappraisal of the Temporal Hypothesis, Twenty Years on.

In a now-classic article published a couple of decades ago (Brain, 2000; 123: 2373–2399), I proposed an “extended temporal processing deficit hypothesis of dyslexia”, suggesting that a deficit in temporal processing could explain not only language-related peculiarities usually noticed in dyslexic children, but also a wider range of symptoms related to impaired processing of time in general. In the present review paper, I will revisit this “historical” hypothesis both in the light of a new clinical perspective, including the central yet poorly explained notion of comorbidity, and also taking a new look at the most recent experimental work, mainly focusing on brain imaging data. First, consistent with daily clinical practice, I propose to distinguish three groups of children who fail to learn to read, of fairly equal occurrence, who share the same initial presentation (difficulty in mastering the rules of grapheme–phoneme correspondence) but with differing associated signs and/or comorbid conditions (language disorders in the first group, attentional deficits in the second one, and motor coordination problems in the last one), thus suggesting, at least in part, potentially different triggering mechanisms. It is then suggested, in the light of brain imaging information available to date, that the three main clinical presentations/associations of cognitive impairments that compromise reading skills acquisition correspond to three distinct patterns of miswiring or “disconnectivity” in specific brain networks which have in common their involvement in the process of learning and their heavy reliance on temporal features of information processing. With reference to the classic temporal processing deficit of dyslexia and to recent evidence of an inability of the dyslexic brain to achieve adequate coupling of oscillatory brain activity to the temporal features of external events, a general model is proposed according to which a common mechanism of temporal uncoupling between various disconnected—and/or mis-wired—processors may account for distinct forms of specific learning disorders, with reading impairment being a more or less constant feature. Finally, the potential therapeutic implications of such a view are considered, with special emphasis on methods seeking to enhance cross-modal connectivity between separate brain systems, including those using rhythmic and musical training in dyslexic patients.

Adaptive mitochondrial response of the whiteleg shrimp Litopenaeus vannamei to environmental challenges and pathogens.

In most eukaryotic organisms, mitochondrial uncoupling mechanisms control ATP synthesis and reactive oxygen species production. One such mechanism is the permeability transition of the mitochondrial inner membrane. In mammals, ischemia-reperfusion events or viral diseases may induce ionic disturbances, such as calcium overload; this cation enters the mitochondria, thereby triggering the permeability transition. This phenomenon increases inner membrane permeability, affects transmembrane potential, promotes mitochondrial swelling, and induces apoptosis. Previous studies have found that the mitochondria of some crustaceans do not exhibit a calcium-regulated permeability transition. However, in the whiteleg shrimpLitopenaeus vannamei, contradictory evidence has prevented this phenomenon from being confirmed or rejected. Both the ability ofL. vannameimitochondria to take up large quantities of calcium through a putative mitochondrial calcium uniporter with conserved characteristics and permeability transition were investigated in this study by determining mitochondrial responses to cations overload. By measuring mitochondrial swelling and transmembrane potential, we investigated whether shrimp exposure to hypoxia-reoxygenation events or viral diseases may induce mitochondrial permeability transition. The results of this study demonstrate that shrimp mitochondria take up large quantities of calcium through a canonical mitochondrial calcium uniporter. Neither calcium nor other ions were observed to promote permeability transition. This phenomenon does not depend on the life cycle stage of shrimp, and it is not induced during hypoxia/reoxygenation events or in the presence of viral diseases. The absence of the permeability transition phenomenon and its adaptive meaning are discussed as a loss with biological advantages, possibly enabling organisms to survive under harsh environmental conditions.

Cognitive Defusion Is a Core Cognitive Mechanism for the Sensory-Affective Uncoupling of Pain During Mindfulness Meditation.

Mindfulness meditation can downregulate the experience of pain. However, its specific underlying regulatory mechanisms are still largely unknown. Here, we aimed to investigate the role of cognitive defusion-a form of psychological distancing from internal experiences-in mindfulness-based pain regulation. We implemented a thermal heat paradigm that was designed to amplify the cognitive-affective aspects of pain in 43 novice meditators (2-day formal training; 51.2% women; 53.2 ± 7.0 years old) and 27 expert meditators (>10,000-hour practice; 44.4% women; 51.9 ± 8.4 years old). We collected pain intensity and unpleasantness reports and trait measures of pain catastrophizing assessed by the Pain Catastrophizing Scale (PCS), cognitive defusion assessed by the Drexel Defusion Scale (DDS), and cognitive fusion assessed by the Cognitive Fusion Questionnaire, as well as of several other constructs commonly reported in the literature. Experts reported lower PCS (6.9 ± 5.2 versus 17.2 ± 8.5, p < .001) but higher DDS (39.4 ± 6.4 versus 28.9 ± 6.6, p < .001) than novices. Across participants, the PCS and DDS were negatively correlated and shared unique variance that survived adjusting for other mindfulness-related and cognitive-emotional constructs (β = -0.64, p < .001). Conversely, the relationships between PCS and other commonly reported constructs did not seem specific, as none of the relationships survived adjusting for DDS (adjusted β < 0.25, p > .05). Further supporting the relevance of DDS to pain, both the DDS and PCS specifically predicted pain unpleasantness as opposed to pain intensity. However, DDS seemed to be a more specific predictor of unpleasantness than PCS, as the relationship between DDS and unpleasantness survived adjusting for PCS (adjusted β = -0.33, p = .016), but not vice versa (adjusted β = 0.20, p = .162). We also found that the Cognitive Fusion Questionnaire showed a similar pattern of associations with PCS and pain self-reports to what was found for the DDS, although these associations were less consistent. Collectively, these findings highlight the central role of cognitive defusion in mindfulness-based pain regulation.

Cu-Mn oxygen carrier with improved mechanical resistance: Analyzing performance under CLC and CLOU environments

Chemical Looping Combustion process allows combustion of gaseous, liquid or solid fuels with CO2 capture. The oxygen necessary for combustion can be supplied using lattice oxygen (CLC) or oxygen uncoupling (CLOU) mechanisms. The present work studies the effects of kaolin addition on CuMn oxygen carrier behavior for CLC and CLOU processes. CuMn oxygen carrier was prepared by granulation with a composition: 27.2 wt% CuO, 52.8 wt% Mn3O4 and 20 wt% kaolin. Oxygen release rates and fluidization behavior were analyzed by TGA and batch fluidized bed reactor. The oxygen carrier was studied for CH4 and synthetic biogas combustion in a 500 Wth CLC continuous unit for 50 h of combustion at temperatures up to 930 °C. No agglomeration problems were observed. Results found during biogas combustion were similar to methane combustion. The addition of 20 wt% kaolin changed the mixed oxide chemical composition generating Cu1.2Mn1.8O4 and improved significantly the oxygen carrier mechanical resistance, increasing the extrapolated lifetime to 19,000 h, 3.6 times over the value found for any Cu based oxygen carrier in CLC. However, the oxygen carrier reactivity had an important decrease with respect to a similar oxygen carrier without kaolin addition, whose fraction in oxygen carrier must be optimized.

Mitochondrial uncoupling and the disruption of the metabolic network in hepatocellular carcinoma.

Background: Hepatocellular Carcinoma (HCC) is the third most common cause of cancer related death worldwide. Adequate treatment options for patients with advanced HCC are currently limited.Materials and Methods: We studied the anti-HCC effect of FH535 and a novel derivative Y3, on proliferation, mitochondrial function and cellular metabolism focusing on the three key substrates, glutamine, glucose, and fatty acids.Results: FH535 and Y3 disrupted mitochondrial redox control in HCC cells that resulted from uncoupling mechanisms that increased proton leakage and decreased ATP production leading to apoptosis. The uncoupling effects of the sulfonamides in HCC cells were supported by the loss of activity of the methylated analogs. The accumulation of ROS significantly contributed to cell damage after the impaired autophagic machinery. These sulfonamides, FH535 and Y3, targeted glutamine and fatty acid metabolism and caused HCC cell reprograming towards the preferential use of glucose and the glycolytic pathway.Conclusions: FH535, and Y3, demonstrated potent anti-HCC activity by targeting OXPHOS, increasing dangerous levels of ROS and reducing ATP production. These sulfonamides target glutamine and FA metabolic pathways significantly increasing the cellular dependency on glycolysis.

Functional characterization of the mitochondrial uncoupling proteins from the white shrimp Litopenaeus vannamei

Mitochondrial uncoupling proteins (UCPs) play an essential role in dissipating the proton gradient and controlling the mitochondrial inner membrane potential. When active, UCPs promote proton leak across the inner membrane, oxidative phosphorylation uncoupling, oxygen uptake increase and decrease the ATP synthesis. Invertebrates possess only isoforms UCP4 and UCP5, however, the role of these proteins is not clear in most species since it may depend on the physiological needs of each animal. This study presents the first functional characterization of crustacean uncoupling proteins from the white shrimp Litopenaeus vannamei LvUCP4 and LvUCP5. Free radicals production in various shrimp organs/tissues was first evaluated, and mitochondria were isolated from shrimp pleopods. The oxygen consumption rate, membrane potential and proton transport of the isolated non-phosphorylating mitochondria were used to determine LvUCPs activation/inhibition. Results indicate that UCPs activity is stimulated in the presence of 4-hydroxyl-2-nonenal (HNE) and myristic acid, and inhibited by the purine nucleotide GDP. A hypoxia/re-oxygenation assay was conducted to determine whether UCPs participate in shrimp mitochondria response to oxidative stress. Isolated mitochondria from shrimp at re-oxygenation produced large quantities of hydrogen peroxide and higher levels of both LvUCPs were immunodetected. Results suggest that, besides the active response of the shrimp antioxidant system, UCP-like activity is activated after hypoxia exposure and during re-oxygenation. LvUCPs may represent a mild uncoupling mechanism, which may be activated before the antioxidant system of cells, to early control reactive oxygen species production and oxidative damage in shrimp.

Decreased Hand Motor Resting-State Functional Connectivity in Patients with Glioma: Analysis of Factors including Neurovascular Uncoupling.

Background Resting-state functional MRI holds substantial potential for clinical application, but limitations exist in current understanding of how tumors exert local effects on resting-state functional MRI readings. Purpose To investigate the association between tumors, tumor characteristics, and changes in resting-state connectivity, to explore neurovascular uncoupling as a mechanism underlying these changes, and to evaluate seeding methodologies as a clinical tool. Materials and Methods Institutional review board approval was obtained for this HIPAA-compliant observational retrospective study of patients with glioma who underwent MRI and resting-state functional MRI between January 2016 and July 2017. Interhemispheric symmetry of connectivity was assessed in the hand motor region, incorporating tumor position, perfusion, grade, and connectivity generated from seed-based correlation. Statistical analysis was performed by using one-tailed t tests, Wilcoxon rank sum tests, one-way analysis of variance, Pearson correlation, and Spearman rank correlation, with significance at P < .05. Results Data in a total of 45 patients with glioma (mean age, 51.3 years ± 14.3 [standard deviation]) were compared with those in 10 healthy control subjects (mean age, 50.3 years ± 17.2). Patients showed loss of symmetry in measures of hand motor resting-state connectivity compared with control subjects (P < .05). Tumor distance from the ipsilateral hand motor (IHM) region correlated with the degree (R = 0.38, P = .01) and strength (R = 0.33, P = .03) of resting-state connectivity. In patients with World Health Organization grade IV glioblastomas 40 mm or less from the IHM region, loss of symmetry in strength of resting-state connectivity was correlated with tumor perfusion (R = 0.74, P < .01). In patients with gliomas 40 mm or less from the IHM region, seeding the nontumor hemisphere yielded less asymmetric hand motor resting-state connectivity than seeding the tumor hemisphere (connectivity seeded:contralateral = 1.34 nontumor vs 1.38 tumor hemisphere seeded; P = .03, false discovery rate threshold = 0.01). Conclusion Hand motor resting-state connectivity was less symmetrical in a tumor distance-dependent manner in patients with glioma. Differences in resting-state connectivity may be false-negative results driven by a neurovascular uncoupling mechanism. Seeding from the nontumor hemisphere may attenuate asymmetry in patients with tumors near ipsilateral hand motor cortices. © RSNA, 2020 Online supplemental material is available for this article.

Calmodulin-Mediated Regulation of Gap Junction Channels.

Evidence that neighboring cells uncouple from each other as one dies surfaced in the late 19th century, but it took almost a century for scientists to start understanding the uncoupling mechanism (chemical gating). The role of cytosolic free calcium (Ca2+i) in cell–cell channel gating was first reported in the mid-sixties. In these studies, only micromolar [Ca2+]i were believed to affect gating—concentrations reachable only in cell death, which would discard Ca2+i as a fine modulator of cell coupling. More recently, however, numerous researchers, including us, have reported the effectiveness of nanomolar [Ca2+]i. Since connexins do not have high-affinity calcium sites, the effectiveness of nanomolar [Ca2+]i suggests the role of Ca-modulated proteins, with calmodulin (CaM) being most obvious. Indeed, in 1981 we first reported that a CaM-inhibitor prevents chemical gating. Since then, the CaM role in gating has been confirmed by studies that tested it with a variety of approaches such as treatments with CaM-inhibitors, inhibition of CaM expression, expression of CaM mutants, immunofluorescent co-localization of CaM and gap junctions, and binding of CaM to peptides mimicking connexin domains identified as CaM targets. Our gating model envisions Ca2+-CaM to directly gate the channels by acting as a plug (“Cork” gating model), and probably also by affecting connexin conformation.

Seasonal patterns of fungal colonisation in Australian native plants of different ages

Plant fungal relationships should vary with abiotic and biotic factors to minimise plant stress and are likely to vary seasonally and with age. We investigated how fungal colonisation, specifically arbuscular mycorrhizal fungi and dark septate fungal endophytes, would vary with species identity and season, and how these interactions change with ontogeny. Plant roots of adults and seedlings of 9 species were collected from heathland and coastal dune habitats along the Australian east coast in New South Wales. Roots were stained and investigated for arbuscular mycorrhizal fungi and dark septate endophyte structures to determine colonisation strength. Species identity was the most important factor driving colonisation strength, while low rainfall and heatwaves were associated with declining arbuscular mycorrhizal fungi colonisation in the warmest sampling period. AMF colonisation may be supressed by plants under heat and water stress as a way of avoiding loss of limited photosynthates. Dark septate endophyte colonisation was more common in this time period and may assist with the stress of the warmer, drier conditions. Colonisation by arbuscular mycorrhizal fungi differed with age but in unpredictable ways and, along with dark septate endophytes, was evident even in plants that are considered non-mycorrhizal, although more extensive in known mycorrhizal species. The lack of arbuscular mycorrhizal fungi colonisation and the increase in dark septate endophyte colonisation during the most stressful period suggest an uncoupling mechanism in the symbiotic relationship which needs further investigation.

Thermochemical assessment of chemical looping assisted by oxygen uncoupling with a MnFe-based oxygen carrier

A previously developed manganese-iron mixed oxide doped with TiO2 shows promising properties for the combustion of solid fuels with intrinsic CO2 capture by Chemical Lopping assisted by Oxygen Uncoupling (CLaOU) owing to its oxygen uncoupling capability, i.e. ability to generate gaseous oxygen at high temperature, and to oxidize gaseous fuels such as H2, CO or CH4 by reacting with lattice oxygen. In this work, a window of suitable operating conditions was determined to facilitate the use of this material in a CLaOU unit. For this purpose, mass and enthalpy balances were performed for the whole CLaOU unit considering theoretical values of thermochemical parameters, such as the equilibrium constant of relevant reactions in CLaOU and the formation enthalpies of reacting solids. An air reactor temperature of 1148 K and a high air excess ratio are suggested in order to promote the transfer of oxygen via the oxygen uncoupling mechanism, which would be preferable for the combustion of solid fuels. Auto-thermal operation of the CLaOU unit is guaranteed for air excess ratio values λ < 3. The fuel reactor temperature is highly influenced by the oxygen carrier-to-fuel ratio (ϕ) and the oxygen carrier regeneration in the air reactor. Assuming a 50–75% regeneration of the oxygen uncoupling capability, and ϕ = 4–6, the fuel reactor temperature would be between 35 and 50 K above the air reactor temperature. The higher temperature in the fuel reactor could promote the fuel conversion, while oxygen carrier regeneration would be guaranteed by the lower air reactor temperature.

Site and mechanism of uncoupling of nitric-oxide synthase: Uncoupling by monomerization and other misconceptions

Nitric oxide synthase (NOS) catalyzes the transformation of l-arginine, molecular oxygen (O2), and NADPH-derived electrons to nitric oxide (NO) and l-citrulline. Under some conditions, however, NOS catalyzes the reduction of O2 to superoxide (O2−) instead, a phenomenon that is generally referred to as uncoupling. In principle, both the heme in the oxygenase domain and the flavins in the reductase domain could catalyze O2− formation. In the former case the oxyferrous (Fe(II)O2) complex that is formed as an intermediate during catalysis would dissociate to heme and O2−; in the latter case the reduced flavins would reduce O2 to O2−. The NOS cofactor tetrahydrobiopterin (BH4) is indispensable for coupled catalysis. In the case of uncoupling at the heme this is explained by the essential role of BH4 as an electron donor to the oxyferrous complex; in the case of uncoupling at the flavins it is assumed that the absence of BH4 results in NOS monomerization, with the monomers incapable to sustain NO synthesis but still able to support uncoupled catalysis. In spite of little supporting evidence, uncoupling at the reductase after NOS monomerization appears to be the predominant hypothesis at present. To set the record straight we extended prior studies by determining under which conditions uncoupling of the neuronal and endothelial isoforms (nNOS and eNOS) occurred and if a correlation exists between uncoupling and the monomer/dimer equilibrium. We determined the rates of coupled/uncoupled catalysis by measuring NADPH oxidation spectrophotometrically at 340 nm and citrulline synthesis as the formation of [3H]-citrulline from [3H]-Arg. The monomer/dimer equilibrium was determined by FPLC and, for comparison, by low-temperature polyacrylamide gel electrophoresis. Uncoupling occurred in the absence of Arg and/or BH4, but not in the absence of Ca2+ or calmodulin (CaM). Since omission of Ca2+/CaM will completely block heme reduction while still allowing substantial FMN reduction, this argues against uncoupling by the reductase domain. In the presence of heme-directed NOS inhibitors uncoupling occurred to the extent that these compound allowed heme reduction, again arguing in favor of uncoupling at the heme. The monomer/dimer equilibrium showed no correlation with uncoupling. We conclude that uncoupling by BH4 deficiency takes place exclusively at the heme, with virtually no contribution from the flavins and no role for NOS monomerization.