Nonalcoholic steatohepatitis (NASH) is associated with caspase activation. However, a role for pro-inflammatory caspases or inflammasomes has not been explored in diet-induced liver injury. Our aims ...were to examine the role of caspase-1 in high fat-induced NASH. C57BL/6 wild-type and caspase 1-knockout (Casp1(-/-)) mice were placed on a 12-week high fat diet. Wild-type mice on the high fat diet increased hepatic expression of pro-caspase-1 and IL-1β. Both wild-type and Casp1(-/-) mice on the high fat diet gained more weight than mice on a control diet. Hepatic steatosis and TG levels were increased in wild-type mice on high fat diet, but were attenuated in the absence of caspase-1. Plasma cholesterol and free fatty acids were elevated in wild-type, but not Casp1(-/-) mice, on high fat diet. ALT levels were elevated in both wild-type and Casp1(-/-) mice on high fat diet compared to control. Hepatic mRNA expression for genes associated with lipogenesis was lower in Casp1(-/-) mice on high fat diet compared to wild-type mice on high fat diet, while genes associated with fatty acid oxidation were not affected by diet or genotype. Hepatic Tnfα and Mcp-1 mRNA expression was increased in wild-type mice on high fat diet, but not in Casp1(-/-) mice on high fat diet. αSMA positive cells, Sirius red staining, and Col1α1 mRNA were increased in wild-type mice on high fat diet compared to control. Deficiency of caspase-1 prevented those increases. In summary, the absence of caspase-1 ameliorates the injurious effects of high fat diet-induced obesity on the liver. Specifically, mice deficient in caspase-1 are protected from high fat-induced hepatic steatosis, inflammation and early fibrogenesis. These data point to the inflammasome as an important therapeutic target for NASH.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Aquaporin-4 (AQP4) plays an important role in regulating water exchange across the blood-brain barrier (BBB) and brain-cerebrospinal fluid interface. Studies on AQP-4 knockout mice (AQP4-KO) have ...reported considerable protection from brain edema induced by acute water intoxication and ischemic stroke, identifying AQP4 as a potential target for therapeutic interventions. However, the long-term effects of chronic AQP4 suppression are yet to be elucidated. In the current study, we evaluated the physiological and structural changes in adult AQP4-KO mice using magnetic resonance imaging (MRI) and immunohistochemical analysis. Water exchange across BBB was assessed by tracking an intravenous bolus injection of oxygen-17 (17O) water (H217O) using 17O-MRI. Cerebral blood flow (CBF) was quantified using arterial spin-labeling (ASL) MRI. Capillary density was determined by immunohistochemical staining for glucose transporter-1 (GLUT1). Compared to wildtype control mice, AQP4-KO mice showed a significant reduction in peak and steady-state H217O uptake despite unaltered CBF. Interestingly, a 22% increase in cortical capillary density was observed in AQP4-KO mice. These results suggest that increased cerebral vascularization may be an adaptive response to chronic reduction in water exchange across BBB in AQP4-KO mice.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Key points
Hyperammonaemia occurs in hepatic, cardiac and pulmonary diseases with increased muscle concentration of ammonia.
We found that ammonia results in reduced skeletal muscle mitochondrial ...respiration, electron transport chain complex I dysfunction, as well as lower NAD+/NADH ratio and ATP content.
During hyperammonaemia, leak of electrons from complex III results in oxidative modification of proteins and lipids.
Tricarboxylic acid cycle intermediates are decreased during hyperammonaemia, and providing a cell‐permeable ester of αKG reversed the lower TCA cycle intermediate concentrations and increased ATP content.
Our observations have high clinical relevance given the potential for novel approaches to reverse skeletal muscle ammonia toxicity by targeting the TCA cycle intermediates and mitochondrial ROS.
Ammonia is a cytotoxic metabolite that is removed primarily by hepatic ureagenesis in humans. Hyperammonaemia occurs in advanced hepatic, cardiac and pulmonary disease, and in urea cycle enzyme deficiencies. Increased skeletal muscle ammonia uptake and metabolism are the major mechanism of non‐hepatic ammonia disposal. Non‐hepatic ammonia disposal occurs in the mitochondria via glutamate synthesis from α‐ketoglutarate resulting in cataplerosis. We show skeletal muscle mitochondrial dysfunction during hyperammonaemia in a comprehensive array of human, rodent and cellular models. ATP synthesis, oxygen consumption, generation of reactive oxygen species with oxidative stress, and tricarboxylic acid (TCA) cycle intermediates were quantified. ATP content was lower in the skeletal muscle from cirrhotic patients, hyperammonaemic portacaval anastomosis rat, and C2C12 myotubes compared to appropriate controls. Hyperammonaemia in C2C12 myotubes resulted in impaired intact cell respiration, reduced complex I/NADH oxidase activity and electron leak occurring at complex III of the electron transport chain. Consistently, lower NAD+/NADH ratio was observed during hyperammonaemia with reduced TCA cycle intermediates compared to controls. Generation of reactive oxygen species resulted in increased content of skeletal muscle carbonylated proteins and thiobarbituric acid reactive substances during hyperammonaemia. A cell‐permeable ester of α‐ketoglutarate reversed the low TCA cycle intermediates and ATP content in myotubes during hyperammonaemia. However, the mitochondrial antioxidant MitoTEMPO did not reverse the lower ATP content during hyperammonaemia. We provide for the first time evidence that skeletal muscle hyperammonaemia results in mitochondrial dysfunction and oxidative stress. Use of anaplerotic substrates to reverse ammonia‐induced mitochondrial dysfunction is a novel therapeutic approach.
Key points
Hyperammonaemia occurs in hepatic, cardiac and pulmonary diseases with increased muscle concentration of ammonia.
We found that ammonia results in reduced skeletal muscle mitochondrial respiration, electron transport chain complex I dysfunction, as well as lower NAD+/NADH ratio and ATP content.
During hyperammonaemia, leak of electrons from complex III results in oxidative modification of proteins and lipids.
Tricarboxylic acid cycle intermediates are decreased during hyperammonaemia, and providing a cell‐permeable ester of αKG reversed the lower TCA cycle intermediate concentrations and increased ATP content.
Our observations have high clinical relevance given the potential for novel approaches to reverse skeletal muscle ammonia toxicity by targeting the TCA cycle intermediates and mitochondrial ROS.
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FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
Abstract There is significant interest in recent years in developing magnetic nanoparticles (MNPs) having multifunctional characteristics with complimentary roles. In this study, we investigated the ...drug delivery and magnetic resonance imaging (MRI) properties of our novel oleic acid-coated iron-oxide and pluronic-stabilized MNPs. The drug incorporation efficiency of doxorubicin and paclitaxel (alone or in combination) in MNPs was 74–95%; the drug release was sustained and the incorporated drugs had marginal effects on physical (size and zeta potential) and magnetization properties of the MNPs. The drugs in combination incorporated in MNPs demonstrated highly synergistic antiproliferative activity in MCF-7 breast cancer cells. The T2 relaxivity ( r2 ) was higher for our MNPs than Feridex IV™, whereas the T1 relaxivity ( r1 ) was better for Feridex IV than for our MNPs, suggesting greater sensitivity of our MNPs than Feridex IV in T2 weighted imaging. The circulation half-life ( t1/2 ), determined from the changes in the MRI signal intensity in carotid arteries in mice, was longer for our MNPs than Feridex IV ( t1/2 = 31.2 vs. 6.4 min). MNPs with combined characteristics of MRI and drug delivery could be of high clinical significance in the treatment of various disease conditions.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UL, UM, UPCLJ, UPUK
A suite of imaging tools for detecting specific chemicals in the central nervous system could accelerate the understanding of neural signaling events critical to brain function and disease. Here, we ...introduce a class of nanoparticle sensors for the highly specific detection of acetylcholine in the living brain using magnetic resonance imaging. The nanosensor is composed of acetylcholine-catalyzing enzymes and pH-sensitive gadolinium contrast agents co-localized onto the surface of polymer nanoparticles, which leads to changes in T 1 relaxation rate (1/T 1). The mechanism of the sensor involves the enzymatic hydrolysis of acetylcholine leading to a localized decrease in pH which is detected by the pH-sensitive gadolinium chelate. The concomitant change in 1/T 1 in vitro measured a 20% increase from 0 to 10 μM acetylcholine concentration. The applicability of the nanosensors in vivo was demonstrated in the rat medial prefrontal cortex showing distinct changes in 1/T 1 induced by pharmacological stimuli. The highly specific acetylcholine nanosensor we present here offers a promising strategy for detection of cholinergic neurotransmission and will facilitate our understanding of brain function through chemical imaging.
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IJS, KILJ, NUK, PNG, UL, UM
Current paradigms suggest that two macrophage subsets, termed M1 and M2, are involved in inflammation and host defense. While the distinct functions of M1 and M2 macrophages have been intensively ...studied - the former are considered proinflammatory and the latter antiinflammatory - the determinants of their speciation are incompletely understood. Here we report our studies that identify Krüppel-like factor 4 (KLF4) as a critical regulator of macrophage polarization. Macrophage KLF4 expression was robustly induced in M2 macrophages and strongly reduced in M1 macrophages, observations that were recapitulated in human inflammatory paradigms in vivo. Mechanistically, KLF4 was found to cooperate with Stat6 to induce an M2 genetic program and inhibit M1 targets via sequestration of coactivators required for NF-κB activation. KLF4-deficient macrophages demonstrated increased proinflammatory gene expression, enhanced bactericidal activity, and altered metabolism. Furthermore, mice bearing myeloid-specific deletion of KLF4 exhibited delayed wound healing and were predisposed to developing diet-induced obesity, glucose intolerance, and insulin resistance. Collectively, these data identify KLF4 as what we believe to be a novel regulator of macrophage polarization.
Adjuvant radiotherapy is frequently prescribed to treat cancer. To minimize radiation-related damage to healthy tissue, it requires high precision in tumor localization and radiation dose delivery. ...This can be achieved by MR guidance and targeted amplification of radiation dose selectively to tumors by using radiosensitizers. Here, we demonstrate prostate cancer-targeted gold nanoparticles (AuNPs) for MR-guided radiotherapy to improve the targeting precision and efficacy. By conjugating Gd(III) complexes and prostate-specific membrane antigen (PSMA) targeting ligands to AuNP surfaces, we found enhanced uptake of AuNPs by PSMA-expressing cancer cells with excellent MR contrast and radiation therapy outcome in vitro and in vivo. The AuNPs binding affinity and r 1 relaxivity were dramatically improved and the combination of Au and Gd(III)provided better tumor suppression after radiation. The precise tumor localization by MR and selective tumor targeting of the PSMA-1-targeted AuNPs could enable precise radiotherapy, reduction in irradiating dose, and minimization of healthy tissue damage.
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IJS, KILJ, NUK, PNG, UL, UM
Autosomal-dominant polycystic kidney disease (ADPKD) is a common genetic disorder that frequently leads to renal failure. Mutations in polycystin-1 (PC1) underlie most cases of ADPKD, but the ...function of PC1 has remained poorly understood. No preventive treatment for this disease is available. Here, we show that the cytoplasmic tail of PC1 interacts with tuberin, and the mTOR pathway is inappropriately activated in cyst-lining epithelial cells in human ADPKD patients and mouse models. Rapamycin, an inhibitor of mTOR, is highly effective in reducing renal cystogenesis in two independent mouse models of PKD. Treatment of human ADPKD transplant-recipient patients with rapamycin results in a significant reduction in native polycystic kidney size. These results indicate that PC1 has an important function in the regulation of the mTOR pathway and that this pathway provides a target for medical therapy of ADPKD.
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BFBNIB, NMLJ, NUK, PNG, SAZU, UL, UM, UPUK
Recent studies have suggested the glymphatic system as a key mechanism of waste removal in the brain. Dynamic contrast-enhanced MRI (DCE-MRI) using intracisternally administered contrast agents is a ...promising tool for assessing glymphatic function in the whole brain. In this study, we evaluated the transport kinetics and distribution of three MRI contrast agents with vastly different molecular sizes in mice. Our results demonstrate that oxygen-17 enriched water (H217O), which has direct access to parenchymal tissues via aquaporin-4 water channels, exhibited significantly faster and more extensive transport compared to the two gadolinium-based contrast agents (Gd-DTPA and GadoSpin). Time-lagged correlation and clustering analyses also revealed different transport pathways for Gd-DTPA and H217O. Furthermore, there were significant differences in transport kinetics of the three contrast agents to the lateral ventricles, reflecting the differences in forces that drive solute transport in the brain. These findings suggest the size-dependent transport pathways and kinetics of intracisternally administered contrast agents and the potential of DCE-MRI for assessing multiple aspects of solute transport in the glymphatic system.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
PURPOSEWe aimed to determine the immediacy of exercise intervention on liver-specific metabolic processes in nonalcoholic fatty liver disease.
METHODSWe undertook a short-term (7-d) exercise training ...study (60 min·d treadmill walking at 80%–85% of maximal heart rate) in obese adults (N = 13, 58 ± 3 yr, 34.3 ± 1.1 kg·m, >5% hepatic lipid by H-magnetic resonance spectroscopy). Insulin sensitivity index was estimated by oral glucose tolerance test using the Soonthorpun model. Hepatic insulin extraction (HIE) was calculated as the molar difference in area under the curve (AUC) for insulin and C-peptide (HIE = 1 − (AUCInsulin/AUCC-Pep)).
RESULTSThe increases in HIE, V˙O2max, and insulin sensitivity index after the intervention were 9.8%, 9.8%, and 34%, respectively (all, P < 0.05). Basal fat oxidation increased (pre47 ± 6 mg·min vs post65 ± 6 mg·min, P < 0.05) and carbohydrate oxidation decreased (pre160 ± 20 mg·min vs post112 ± 15 mg·min, P < 0.05) with exercise training. After the intervention, HIE correlated positively with adiponectin (r = 0.56, P < 0.05) and negatively with TNF-α (r = −0.78, P < 0.001).
CONCLUSIONSBy increasing HIE along with peripheral insulin sensitivity, aerobic exercise training rapidly reverses some of the underlying physiological mechanisms associated with nonalcoholic fatty liver disease, in a weight loss-independent manner. This reversal could potentially act through adipokine-related pathways.
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