RESUMEN
Obstructive sleep apnea (OSA) is an emerging risk factor for cancer occurrence and progression, mainly mediated by intermittent hypoxia (IH). Systemic IH, a main landmark of OSA, and local sustained hypoxia (SH), a classical feature at the core of tumors, may act separately or synergistically on tumor cells. Our aim was to compare the respective consequences of intermittent and sustained hypoxia on HIF-1, endothelin-1 and VEGF expression and on cell proliferation and migration in HepG2 liver tumor cells. Wound healing, spheroid expansion, proliferation and migration were evaluated in HepG2 cells following IH or SH exposure. The HIF-1α, endothelin-1 and VEGF protein levels and/or mRNA expression were assessed, as were the effects of HIF-1 (acriflavine), endothelin-1 (macitentan) and VEGF (pazopanib) inhibition. Both SH and IH stimulated wound healing, spheroid expansion and proliferation of HepG2 cells. HIF-1 and VEGF, but not endothelin-1, expression increased with IH exposure but not with SH exposure. Acriflavine prevented the effects of both IH and SH, and pazopanib blocked those of IH but not those of SH. Macitentan had no impact. Thus, IH and SH stimulate hepatic cancer cell proliferation via distinct signaling pathways that may act synergistically in OSA patients with cancer, leading to enhanced tumor progression.
Asunto(s)
Apnea Obstructiva del Sueño , Factor A de Crecimiento Endotelial Vascular , Humanos , Factor A de Crecimiento Endotelial Vascular/genética , Factor A de Crecimiento Endotelial Vascular/metabolismo , Células Hep G2 , Acriflavina , Hipoxia/metabolismo , Apnea Obstructiva del Sueño/metabolismo , Factor 1 Inducible por Hipoxia , Proliferación Celular , Subunidad alfa del Factor 1 Inducible por Hipoxia/genéticaRESUMEN
Zinc is involved in the expression and function of various transcription factors, including the hypoxia-inducible factor-1 (HIF-1). HIF-1 and its target gene endothelin-1 (ET-1) are activated by intermittent hypoxia (IH), one of the main consequences of obstructive sleep apnea (OSA), and both play a key role in the cardiovascular consequences of IH. Because OSA and IH are associated with zinc deficiency, we investigated the effect of zinc deficiency caused by chelation on the HIF-1/ET-1 pathway and its functional consequences in endothelial cells. Primary human microvascular endothelial cells (HMVEC) were incubated with submicromolar doses of the zinc-specific membrane-permeable chelator N,N,N',N'-tetrakis(2-pyridylmethyl)-ethylene diamine (TPEN, 0.5 µM) or ET-1 (0.01 µM) with or without bosentan, a dual ET-1-receptor antagonist. HIF-1α expression was silenced by transfection with specific siRNA. Nuclear HIF-1 content was assessed by immunofluorescence microscopy and Western blot. Migratory capacity of HMVEC was evaluated with a wound-healing scratch assay. Zinc chelation by TPEN exposure induced the translocation of the cytosolic HIF-1α subunit of HIF-1 to the nucleus as well as an HIF-1-mediated ET-1 secretion by HMVEC. Incubation with either TPEN or ET-1 increased endothelial wound-healing capacity. Both HIF-1α silencing or bosentan abolished this effect. Altogether, these results suggest that zinc deficiency upregulates ET-1 signaling through HIF-1 activation and stimulates endothelial cell migration, suggesting an important role of zinc in the vascular consequences of IH and OSA mediated by HIF-1-ET- signaling.
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Movimiento Celular/efectos de los fármacos , Quelantes/farmacología , Células Endoteliales/efectos de los fármacos , Endotelina-1/metabolismo , Etilenodiaminas/farmacología , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Zinc/deficiencia , Transporte Activo de Núcleo Celular , Bosentán/farmacología , Células Cultivadas , Células Endoteliales/metabolismo , Antagonistas de los Receptores de Endotelina/farmacología , Endotelina-1/genética , Humanos , Subunidad alfa del Factor 1 Inducible por Hipoxia/genética , Vías Secretoras , Transducción de SeñalRESUMEN
Performing hypoxia-reoxygenation cycles in cell culture with a cycle duration accurately reflecting what occurs in obstructive sleep apnea (OSA) patients is a difficult but crucial technical challenge. Our goal was to develop a novel device to expose multiple cell culture dishes to intermittent hypoxia (IH) cycles relevant to OSA with limited gas consumption. With gas flows as low as 200 ml/min, our combination of plate holders with gas-permeable cultureware generates rapid normoxia-hypoxia cycles. Cycles alternating 1 min at 20% O2 followed by 1 min at 2% O2 resulted in Po2 values ranging from 124 to 44 mmHg. Extending hypoxic and normoxic phases to 10 min allowed Po2 variations from 120 to 25 mmHg. The volume of culture medium or the presence of cells only modestly affected the Po2 variations. In contrast, the nadir of the hypoxia phase increased when measured at different heights above the membrane. We validated the physiological relevance of this model by showing that hypoxia inducible factor-1α expression was significantly increased by IH exposure in human aortic endothelial cells, murine breast carcinoma (4T1) cells as well as in a blood-brain barrier model (2.5-, 1.5-, and 6-fold increases, respectively). In conclusion, we have established a new device to perform rapid intermittent hypoxia cycles in cell cultures, with minimal gas consumption and the possibility to expose several culture dishes simultaneously. This device will allow functional studies of the consequences of IH and deciphering of the molecular biology of IH at the cellular level using oxygen cycles that are clinically relevant to OSA.
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Barrera Hematoencefálica/metabolismo , Neoplasias de la Mama/metabolismo , Técnicas de Cultivo de Célula , Células Endoteliales/metabolismo , Hipoxia/metabolismo , Oxígeno/metabolismo , Apnea Obstructiva del Sueño/metabolismo , Animales , Técnicas de Cultivo de Célula/instrumentación , Hipoxia de la Célula , Línea Celular Tumoral , Medios de Cultivo/metabolismo , Diseño de Equipo , Femenino , Gases , Humanos , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Factores de Tiempo , Hipoxia TumoralRESUMEN
Obstructive sleep apnoea syndrome is characterized by repetitive episodes of upper airway collapse during sleep resulting in chronic intermittent hypoxia (IH). Obstructive sleep apnoea syndrome, through IH, promotes cardiovascular and metabolic disorders. Endothelin-1 (ET-1) secretion is upregulated by IH, and is able to modulate adipocyte metabolism. Therefore, the present study aimed to characterize the role of ET-1 in the metabolic consequences of IH on adipose tissue in vivo and in vitro. Wistar rats were submitted to 14 days of IH-cycles (30 s of 21% FiO2 and 30 s of 5% FiO2 ; 8 h day(-1) ) or normoxia (air-air cycles) and were treated or not with bosentan, a dual type A and B endothelin receptor (ETA-R and ETB-R) antagonist. Bosentan treatment decreased plasma free fatty acid and triglyceride levels, and inhibited IH-induced lipolysis in adipose tissue. Moreover, IH induced a 2-fold increase in ET-1 transcription and ETA-R expression in adipose tissue that was reversed by bosentan. In 3T3-L1 adipocytes, ET-1 upregulated its own and its ETA-R transcription and this effect was abolished by bosentan. Moreover, ET-1 induced glycerol release and inhibited insulin-induced glucose uptake. Bosentan and BQ123 inhibited these effects. Bosentan also reversed the ET-1-induced phosphorylation of hormone-sensitive lipase (HSL) on Ser(660) . Finally, ET-1-induced lipolysis and HSL phosphorylation were also observed under hypoxia. Altogether, these data suggest that ET-1 is involved in IH-induced lipolysis in Wistar rats, and that upregulation of ET-1 production and ETA-R expression by ET-1 itself under IH could amplify its effects. Moreover, ET-1-induced lipolysis could be mediated through ETA-R and activation of HSL by Ser(660) phosphorylation.
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Tejido Adiposo/metabolismo , Endotelina-1/metabolismo , Hipoxia/metabolismo , Lipólisis , Procesamiento Proteico-Postraduccional , Esterol Esterasa/metabolismo , Células 3T3 , Tejido Adiposo/efectos de los fármacos , Tejido Adiposo/patología , Animales , Bosentán , Antagonistas de los Receptores de Endotelina/farmacología , Endotelina-1/genética , Ácidos Grasos/sangre , Hipoxia/patología , Masculino , Ratones , Fosforilación , Ratas , Ratas Wistar , Receptores de Endotelina/genética , Receptores de Endotelina/metabolismo , Sulfonamidas/farmacología , Triglicéridos/sangreRESUMEN
NEW FINDINGS: What is the central question of this study? This study addresses the relative impact of obesity and intermittent hypoxia in the pathophysiological process of obstructive sleep apnoea by investigating the metabolic, inflammatory and cardiovascular consequences of intermittent hypoxia in lean and obese Zucker rats. What is the main finding and its importance? We found that obesity and intermittent hypoxia have mainly distinct consequences on the investigated inflammatory and cardiometabolic parameters in Zucker rats. This suggests that, for a given severity of sleep apnea, the association of obesity and obstructive sleep apnoea may not necessarily be deleterious. Obstructive sleep apnoea is associated with obesity with a high prevalence, and both co-morbidities are independent cardiovascular risk factors. Intermittent hypoxia (IH) is thought to be the main factor responsible for the obstructive sleep apnoea-related cardiometabolic alterations. The aim of this study was to assess the respective impact of obesity and IH on the inflammatory and cardiometabolic state in rats. Lean and obese Zucker rats were exposed to normoxia or chronic IH, and we assessed metabolic and inflammatory parameters, such as plasma lipids and glucose, serum leptin and adiponectin, liver cytokines, nuclear factor-κB activity and cardiac endothelin-1 levels. Myocardial infarct size was also evaluated following in vitro ischaemia-reperfusion. Circulating lipids, insulin, homeostasis model assessment of insulin resistance (HOMA-IR), leptin and adiponectin levels were higher in obese versus lean rats. Chronic IH did not have a significant impact on metabolic parameters in lean rats. In obese rats, IH increased glycaemia and HOMA-IR. Liver interleukin-6 and tumour necrosis factor-α levels were elevated in lean rats exposed to IH; obesity prevented the increase in interleukin-6 but not in tumour necrosis factor-α. Finally, IH exposure enhanced myocardial sensitivity to infarction in both lean and obese rats and increased cardiac endothelin-1 in lean but not obese rats. In conclusion, this study shows that the dyslipidaemia and insulin resistance induced by obesity of genetic origin does not enhance the deleterious cardiovascular response to IH and may even partly protect against IH-induced inflammation.
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Enfermedades Cardiovasculares/metabolismo , Hipoxia/metabolismo , Inflamación/metabolismo , Obesidad/metabolismo , Adiponectina/metabolismo , Animales , Glucemia/metabolismo , Enfermedades Cardiovasculares/fisiopatología , Citocinas/metabolismo , Modelos Animales de Enfermedad , Endotelina-1/metabolismo , Insulina/metabolismo , Interleucina-6/metabolismo , Leptina/sangre , Lípidos/sangre , Hígado/metabolismo , Masculino , Miocardio/metabolismo , FN-kappa B/metabolismo , Ratas , Ratas Zucker , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
BACKGROUND AND OBJECTIVE: Randomized controlled trials (RCT) have shown that continuous positive airway pressure (CPAP) has only limited impact on blood pressure (BP). Alternative strategies for obstructive sleep apnoea (OSA)-associated hypertension are therefore needed. Endothelin-1 has been demonstrated a key player in the deleterious cardiovascular consequences of OSA. In OSA, CPAP treatment has never been compared with endothelin receptor antagonist medications. Thus, we assessed the respective efficacy of CPAP and bosentan in reducing 24-h diastolic BP (DBP) in patients with OSA never treated by either therapy. METHODS: In a crossover pilot study, 16 mildly hypertensive patients (office systolic BP (SBP)/DBP: 142 ± 7/85 ± 8 mm Hg) with severe OSA (55 ± 8 years; body mass index, 29.6 ± 4.2 kg/m(2) ; apnoea-hypopnoea index, 40.8 ± 20.2/h) were randomized to either CPAP (n = 7) or bosentan (125 mg/day, n = 9) first for 4 weeks. After 2-weeks of washout, the second 4-week period consisted of the alternative treatment (in crossover). The primary outcome was the 24-h mean DBP change after treatment. RESULTS: In intention-to-treat analysis, the mean difference in 24-h DBP measurements between treatments was -3.1 (-6.9/0.7) mm Hg (median, 25th/75th percentiles) (P = 0.101) with bosentan having a greater effect. CONCLUSION: In this RCT, in mildly hypertensive patients with OSA, bosentan did not modify 24-h DBP but only reduced office BP suggesting that Endothelin-1 blockade does not play a major role in treatment of OSA-related hypertension.
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Presión Sanguínea/fisiología , Presión de las Vías Aéreas Positiva Contínua/métodos , Hipertensión/terapia , Apnea Obstructiva del Sueño/terapia , Sulfonamidas/administración & dosificación , Índice de Masa Corporal , Bosentán , Estudios Cruzados , Relación Dosis-Respuesta a Droga , Antagonistas de los Receptores de Endotelina/administración & dosificación , Femenino , Humanos , Hipertensión/complicaciones , Hipertensión/fisiopatología , Masculino , Persona de Mediana Edad , Proyectos Piloto , Apnea Obstructiva del Sueño/complicaciones , Apnea Obstructiva del Sueño/fisiopatologíaAsunto(s)
Melanoma , Neoplasias Cutáneas , Apnea Obstructiva del Sueño , Antígeno B7-H1 , Biomarcadores , Humanos , HipoxiaRESUMEN
Remote ischemic preconditioning (RIPC) has emerged as a feasible and attractive therapeutic procedure for heart protection against ischemia/reperfusion (I/R) injury. However, its molecular mechanisms remain poorly understood. Hypoxia inducible factor-1α (HIF-1α) is a transcription factor that plays a key role in the cellular adaptation to hypoxia and ischemia. This study's aim was to test whether RIPC-induced cardioprotection requires HIF-1α upregulation to be effective. In the first study, wild-type mice and mice heterozygous for HIF1a (gene encoding the HIF-1α protein) were subjected to RIPC immediately before myocardial infarction (MI). RIPC resulted in a robust HIF-1α activation in the limb and acute cardioprotection in wild-type mice. RIPC-induced cardioprotection was preserved in heterozygous mice, despite the low HIF-1α expression in their limbs. In the second study, the role of HIF-1α in RIPC was evaluated using cadmium (Cd), a pharmacological HIF-1α inhibitor. Rats were subjected to MI (MI group) or to RIPC immediately prior to MI (R-MI group). Cd was injected 18 0min before RIPC (Cd-R-MI group). RIPC induced robust HIF-1α activation in rat limbs and significantly reduced infarct size (IS). Despite Cd's inhibition of HIF-1α activation, RIPC-induced cardioprotection was preserved in the Cd-R-MI group. RIPC applied immediately prior to MI increased HIF-1α expression and attenuated IS in rats and wild-type mice. However, RIPC-induced cardioprotection was preserved in partially HIF1a-deficient mice and in rats pretreated with Cd. When considered together, these results suggest that HIF-1α upregulation is unnecessary in acute RIPC.
Asunto(s)
Extremidades/irrigación sanguínea , Extremidades/patología , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Precondicionamiento Isquémico , Animales , Cadmio/farmacología , Cardiotónicos/metabolismo , Activación Enzimática/efectos de los fármacos , Glucógeno Sintasa Quinasa 3/metabolismo , Glucógeno Sintasa Quinasa 3 beta , Subunidad alfa del Factor 1 Inducible por Hipoxia/antagonistas & inhibidores , Subunidad alfa del Factor 1 Inducible por Hipoxia/deficiencia , Masculino , Ratones , Infarto del Miocardio/patología , Miocardio/enzimología , Miocardio/patología , Fosforilación/efectos de los fármacos , Ratas , Ratas Sprague-DawleyRESUMEN
The treatment of systemic sclerosis-related digital ulcers is challenging. Although the only effective drugs are prostacyclin analogs, their use is limited by vasodilation-related adverse reactions. In this study, we assessed the local iontophoresis administration of three soluble guanylate cyclase (A-350619 [3-[2-[(4-chlorophenyl)thiophenyl]-N-[4-(dimethylamino)butyl]-2-propenamide hydrochloride], SIN-1 [amino-3-morpholinyl-1,2,3-oxadiazolium chloride], and CFM 1571 [3-[3-(dimethylamino)propoxy]-N-(4-methoxyphenyl)-1-(phenylmethyl)-1H-pyrazole-5-carboxamide hydrochloride]) and two nonprostanoid prostaglandin I2 (prostacyclin) receptor agonists (MRE-269 [[4-[(5,6-diphenylpyrazinyl)(1-methylethyl)amino]butoxy]-acetic acid] and BMY 45778 [[3-(4,5-diphenyl[2,4'-bioxazol]-5'-yl)phenoxy]acetic acid]) to induce vasodilation onto the hindquarters of anesthetized rats. Skin blood flow was quantified using laser Doppler imaging during the whole experience, and safety was assessed by continuous recording of blood pressure and histopathological examination. Anodal iontophoresis of A-350619 (7.54 mM) induced a sustained increase in cutaneous blood flow (P = 0.008 vs. control). All other drugs exhibited poor or no effect on skin blood flow. Vasodilation with A-350619 iontophoresis was concentration-dependent (7.5, 0.75, and 0.075 mM; P < 0.001, Jonckheere-Terpstra trend test), and repeated administrations do not suggest any risk of tolerance. This study also compared continuous versus intermittent iontophoresis protocols. Continuous anodal iontophoresis of A-350619 at 7.5 mM increases cutaneous blood flow with good local tolerance. Iontophoresis of soluble guanylate cyclase stimulators should be investigated as potential local therapy for digital ulceration in patients with scleroderma.
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Acrilamidas/farmacología , Guanilato Ciclasa/efectos de los fármacos , Nitrosaminas/farmacología , Pirazoles/farmacología , Flujo Sanguíneo Regional/efectos de los fármacos , Piel/irrigación sanguínea , Piel/efectos de los fármacos , Acetatos/efectos adversos , Acetatos/farmacología , Acrilamidas/administración & dosificación , Acrilamidas/efectos adversos , Animales , Interpretación Estadística de Datos , Estimulación Eléctrica , Electrodos , Técnicas In Vitro , Iontoforesis , Masculino , Nitrosaminas/administración & dosificación , Nitrosaminas/efectos adversos , Oxazoles/efectos adversos , Oxazoles/farmacología , Inhibidores de Agregación Plaquetaria/efectos adversos , Inhibidores de Agregación Plaquetaria/farmacología , Pirazinas/efectos adversos , Pirazinas/farmacología , Pirazoles/administración & dosificación , Pirazoles/efectos adversos , Ratas , Ratas Wistar , Receptores de Epoprostenol/agonistasRESUMEN
The association between obstructive sleep apnea (OSA) and cancer is still debated and data are scarce regarding the link between OSA and breast cancer progression. Since conclusive epidemiological studies require large sample sizes and sufficient duration of exposure before incident cancer occurrence, basic science studies represent the most promising approach to appropriately address the topic. Here we assessed the impact of intermittent hypoxia (IH), the major hallmark of OSA, on the development of breast cancer and explored the specific involvement of the endothelin signaling pathway. Original in vitro and in vivo models were used where 3D-spheroids or cultures of murine 4T1 breast cancer cells were submitted to IH cycles, and nude NMRI mice, orthotopically implanted with 4T1 cells, were submitted to chronic IH exposure before and after implantation. The role of the endothelin-1 in promoting cancer cell development was investigated using the dual endothelin receptor antagonist, macitentan. In vitro exposure to IH significantly increased 4T1 cell proliferation and migration. Meta-analysis of 4 independent in vivo experiments showed that chronic IH exposure promoted tumor growth, assessed by caliper measurement (overall standardized mean difference: 1.00 [0.45-1.55], p < 0.001), bioluminescence imaging (1.65 [0.59-2.71]; p < 0.01) and tumor weight (0.86 [0.31-1.41], p < 0.01), and enhanced metastatic pulmonary expansion (0.77 [0.12-1.42]; p = 0.01). Both in vitro and in vivo tumor-promoting effects of IH were reversed by macitentan. Overall, these findings demonstrate that chronic intermittent hypoxia exposure promotes breast cancer growth and malignancy and that dual endothelin receptor blockade prevents intermittent hypoxia-induced tumor development.
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Neoplasias , Apnea Obstructiva del Sueño , Animales , Endotelina-1/metabolismo , Hipoxia/metabolismo , Ratones , Receptor de Endotelina ARESUMEN
RATIONALE: Intermittent hypoxia (IH) is one of the main features of sleep-disordered breathing (SDB). Recent findings indicate that hypoxia inducible factor-1 (HIF-1) promotes cardiomyocytes apoptosis during chronic IH, but the mechanisms involved remain to be elucidated. Here, we hypothesize that IH-induced ER stress is associated with mitochondria-associated ER membrane (MAM) alteration and mitochondrial dysfunction, through HIF-1 activation. METHODS: Right atrial appendage biopsies from patients with and without SDB were used to determine HIF-1α, Grp78 and CHOP expressions. Wild-type and HIF-1α+/- mice were exposed to normoxia (N) or IH (21-5% O2, 60 cycles/h, 8 h/day) for 21 days. Expressions of HIF-1α, Grp78 and CHOP, and apoptosis, were measured by Western blot and immunochemistry. In isolated cardiomyocytes, we examined structural integrity of MAM by proximity ligation assay and their function by measuring ER-to-mitochondria Ca2+ transfer by confocal microscopy. Finally, we measured mitochondrial respiration using oxygraphy and calcium retention capacity (CRC) by spectrofluorometry. MAM structure was also investigated in H9C2 cells incubated with 1 mM CoCl2, a potent HIF-1α inducer. RESULTS: In human atrial biopsies and mice, IH induced HIF-1 activation, ER stress and apoptosis. IH disrupted MAM, altered Ca2+ homeostasis, mitochondrial respiration and CRC. Importantly, IH had no effect in HIF-1α+/- mice. Similar to what observed under IH, HIF-1α overexpression was associated with MAM alteration in H9C2. CONCLUSION: IH-induced ER stress, MAM alterations and mitochondrial dysfunction were mediated by HIF-1; all these intermediate mechanisms ultimately inducing cardiomyocyte apoptosis. This suggests that HIF-1 modulation might limit the deleterious cardiac effects of SDB.
RESUMEN
Sleep Apnea Syndrome (SAS) is one of the most common chronic diseases, affecting nearly one billion people worldwide. The repetitive occurrence of abnormal respiratory events generates cyclical desaturation-reoxygenation sequences known as intermittent hypoxia (IH). Among SAS metabolic sequelae, it has been established by experimental and clinical studies that SAS is an independent risk factor for the development and progression of non-alcoholic fatty liver disease (NAFLD). The principal goal of this study was to decrypt the molecular mechanisms at the onset of IH-mediated liver injury. To address this question, we used a unique mouse model of SAS exposed to IH, employed unbiased high-throughput transcriptomics and computed network analysis. This led us to examine hepatic mitochondrial ultrastructure and function using electron microscopy, high-resolution respirometry and flux analysis in isolated mitochondria. Transcriptomics and network analysis revealed that IH reprograms Nuclear Respiratory Factor- (NRF-) dependent gene expression and showed that mitochondria play a central role. We thus demonstrated that IH boosts the oxidative capacity from fatty acids of liver mitochondria. Lastly, the unbalance between oxidative stress and antioxidant defense is tied to an increase in hepatic ROS production and DNA damage during IH. We provide a comprehensive analysis of liver metabolism during IH and reveal the key role of the mitochondria at the origin of development of liver disease. These findings contribute to the understanding of the mechanisms underlying NAFLD development and progression during SAS and provide a rationale for novel therapeutic targets and biomarker discovery.
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BACKGROUND: Chronic intermittent hypoxia (IH), the hallmark feature of obstructive sleep apnoea syndrome, contributes to infarct size enhancement after myocardial ischemia-reperfusion (I/R). Curcumin (Curc), the natural pigment of Curcuma longa, has been demonstrated to be beneficial in the context of myocardial injury. In this study, we assessed the effects of Curc on the maladaptive cardiac response to IH, and particularly on IH-induced hypoxia inducible factor-1 (HIF-1) expression, oxidative stress, inflammation, endoplasmic reticulum (ER) stress and apoptosis. METHODS: Swiss/SV129 mice were exposed to normoxia or IH (21-5% FiO2, 60 s cycles, 8 h per day, for 21 days) and treated orally with Curc (100 mg kg-1 day-1, oral gavage) or its vehicle. Mice were then either euthanised for heart sampling in order to perform biochemical and histological analysis, or subjected to an in vivo ischemia-reperfusion protocol in order to measure infarct size. RESULTS: IH increased nuclear HIF-1α expression and superoxide anion (O2 .-) production as well as nuclear factor kappa B (NF-kB) p65, glucose-regulated protein (Grp78) and C/EBP homologous protein (CHOP) expression. IH also induced apoptosis and increased infarct size after I/R . The IH-induced HIF-1 activation, oxidative stress, inflammation, ER stress and apoptosis were abolished by chronic Curc treatment. Curc also significantly decreased infarct size only in mice exposed to IH. CONCLUSION: Curc prevents IH-induced myocardial cell death signalling. Curc might be used as a combined therapy with continuous positive airway pressure in sleep apnoea patients with high cardiovascular risk.
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Artemisinin and its derivatives are currently recommended by World Health Organization for the treatment of malaria. Severe malaria requires a parenteral administration of artemisinin-based formulations. However, the effective use of artemisinin is limited by the pharmacokinetic characteristics of the drug (low water solubility, poor bioavailability and short half-life). To overcome some of these drawbacks, artemisinin-loaded surface-decorated nanoparticles were prepared by co-nanoprecipitation of γ-cyclodextrin bioesterified with C10 alkyl chains and polyethylene glycol (PEG) derivatives (polysorbate 80 and DMPE-mPEG2000). Using a single dose (1.5â¯mgâ¯kg-1 or 2â¯mgâ¯kg-1)â¯by intravenous administration, we investigated the in vivo pharmacokinetic properties in healthy rats of two types of artemisinin-loaded nanoparticle formulations, namely, nanosphere and nanoreservoir systems versus an ethanolic-aqueous solution of artemisinin as reference. Significantly enhanced pharmacokinetic parameters were obtained with artemisinin-loaded nanoparticles. In comparison to reference formulation, the geometric mean exposures in plasma (AUC0-t) exhibited 2.35 and 3.26-fold increases when artemisinin was loaded in nanoreservoir and nanosphere systems, respectively. Its plasma half-life increased 4.00 and 6.25-fold and its clearance decreased up to 2.5 and 4.72-fold. Artemisinin was successfully administered intravenously by means of surface-decorated amphiphilic γ-cyclodextrin nanostructures and showed a longer elimination half-life with respect to an artemisinin solution in ethanol. Therefore, these systems are likely to provide significant advantages for the intravenous treatment of severe malaria.
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Antimaláricos/farmacocinética , Artemisininas/química , Nanopartículas/química , gamma-Ciclodextrinas/química , Administración Intravenosa , Animales , Antimaláricos/sangre , Antimaláricos/química , Artemisininas/sangre , Artemisininas/farmacocinética , Portadores de Fármacos/química , Semivida , Masculino , Tamaño de la Partícula , Polietilenglicoles/química , Ratas , Ratas Wistar , Propiedades de SuperficieRESUMEN
Chronic intermittent hypoxia (CIH) occurring during sleep apnea amplifies infarct size owing to ischemia-reperfusion. CIH activates hypoxia-inducible factor 1 (HIF-1) and activating transcription factor 4 (ATF4). However, whether HIF-1 and ATF4 interact to promote cardiomyocyte death remains unexplored. For the first time, we observed that in myocardium from apneic patients, CCAAT enhancer-binding protein homologous protein (CHOP) expression is increased and HIF-1α expression is correlated with sleep apnea severity. In mice, single-allele deletion of HIF-1α prevents CIH increase in CHOP expression and infarct size. We uncovered a physical interaction between HIF-1α and ATF4 in CIH that may represent a novel cardiomyocyte death complex.
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Factor de Transcripción Activador 4/metabolismo , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Hipoxia , Infarto del Miocardio , Síndromes de la Apnea del Sueño , Animales , Proteínas Potenciadoras de Unión a CCAAT/metabolismo , Muerte Celular , Humanos , Hipoxia/complicaciones , Hipoxia/etiología , Hipoxia/metabolismo , Ratones , Infarto del Miocardio/etiología , Infarto del Miocardio/metabolismo , Infarto del Miocardio/patología , Miocitos Cardíacos/metabolismo , Miocitos Cardíacos/patología , Síndromes de la Apnea del Sueño/complicaciones , Síndromes de la Apnea del Sueño/metabolismoRESUMEN
BACKGROUND Sleep-disordered breathing is associated with a poor prognosis (mortality) in patients with ischemic cardiomyopathy. The understanding of mechanisms linking intermittent hypoxia (IH), the key feature of sleep-disordered breathing, to ischemic cardiomyopathy progression is crucial for identifying specific actionable therapeutic targets. The aims of the present study were (1) to evaluate the impact of IH on the time course evolution of cardiac remodeling and contractile dysfunction in a rat model of ischemic cardiomyopathy; and (2) to determine the impact of IH on sympathetic activity, hypoxia inducible factor-1 activation, and endoplasmic reticulum stress in the time course of ischemic cardiomyopathy progression. METHODS AND RESULTS Ischemic cardiomyopathy was induced by a permanent ligature of the left coronary artery in male Wistar rats (rats with myocardial infarction). Rats with myocardial infarction were then exposed to either IH or normoxia for up to 12 weeks. Cardiac remodeling and function were analyzed by Sirius red and wheat germ agglutinin staining, ultrasonography, and cardiac catheterization. Sympathetic activity was evaluated by spectral analysis of blood pressure variability. Hypoxia-inducible factor-1α activation and burden of endoplasmic reticulum stress were characterized by Western blots. Long-term IH exposure precipitated cardiac remodeling (hypertrophy and interstitial fibrosis) and contractile dysfunction during the time course evolution of ischemic cardiomyopathy in rodents. Among associated mechanisms, we identified the early occurrence and persistence of sympathetic activation, associated with sustained hypoxia-inducible factor-1α expression and a delayed pro-apoptotic endoplasmic reticulum stress. CONCLUSIONS Our data provide the demonstration of the deleterious impact of IH on post-myocardial infarction remodeling and contractile dysfunction. Further studies are needed to evaluate whether targeting sympathetic nervous system or HIF-1 overactivities could limit these effects and improve management of coexisting ischemic cardiomyopathy and sleep-disordered breathing.
Asunto(s)
Cardiomiopatías/fisiopatología , Hipoxia/fisiopatología , Contracción Miocárdica/fisiología , Síndromes de la Apnea del Sueño/fisiopatología , Remodelación Ventricular/fisiología , Animales , Presión Sanguínea/fisiología , Cardiomiopatías/complicaciones , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Estrés del Retículo Endoplásmico/fisiología , Corazón , Hipoxia/complicaciones , Factor 1 Inducible por Hipoxia/metabolismo , Pulmón/patología , Masculino , Infarto del Miocardio/patología , Isquemia Miocárdica/etiología , Miocardio/patología , Ratas , Ratas Wistar , Síndromes de la Apnea del Sueño/complicaciones , Factores de TiempoRESUMEN
Myocardial infarction (MI) followed by left ventricular (LV) remodeling is the most frequent cause of heart failure. Lebetin 2 (L2), a snake venom-derived natriuretic peptide, exerts cardioprotection during acute myocardial ischemia-reperfusion (IR) ex vivo. However, its effects on delayed consequences of IR injury, including post-MI inflammation and fibrosis have not been defined. Here, we determined whether a single L2 injection exerts cardioprotection in IR murine models in vivo, and whether inflammatory response to ischemic injury plays a role in L2-induced effects. We quantified infarct size (IS), fibrosis, inflammation, and both endothelial cell and cardiomyocyte densities in injured myocardium and compared these values with those induced by B-type natriuretic peptide (BNP). Both L2 and BNP reduced IS, fibrosis, and inflammatory response after IR, as evidenced by decreased leukocyte and proinflammatory M1 macrophage infiltrations in the infarcted area compared to untreated animals. However, only L2 increased anti-inflammatory M2-like macrophages. L2 also induced a higher density of endothelial cells and cardiomyocytes. Our data show that L2 has strong, acute, prolonged cardioprotective effects in post-MI that are mediated, at least in part, by the modulation of the post-ischemic inflammatory response and especially, by the enhancement of M2-like macrophages, thus reducing IR-induced necrotic and fibrotic effects.
Asunto(s)
Antiinflamatorios/uso terapéutico , Daño por Reperfusión Miocárdica/tratamiento farmacológico , Péptido Natriurético Encefálico/uso terapéutico , Venenos de Víboras/uso terapéutico , Animales , Presión Sanguínea/efectos de los fármacos , Fibrosis , Frecuencia Cardíaca/efectos de los fármacos , Masculino , Ratones Endogámicos C57BL , Daño por Reperfusión Miocárdica/patología , Daño por Reperfusión Miocárdica/fisiopatología , Ratas WistarRESUMEN
Muscle wasting reduces functional capacity and increases cardiometabolic risk in chronic disease. Neuromuscular electrical stimulation (NMES) of the lower limb has been shown to reverse muscle wasting in these patients but its effect on cardiometabolic health is unclear. We investigated a mouse model of in-vivo non-invasive chronic NMES on muscle mass, insulin sensitivity and arterial blood pressure (BP). Twenty-three C57BL6 mice underwent unilateral NMES or sham training over 2.5 weeks while anesthetized by isoflurane. Lower limb muscle mass and the stimulated limb to non-stimulated limb muscle mass ratio were compared between groups (NMES vs. sham). Insulin sensitivity was assessed 48 h after training using an intraperitoneal insulin tolerance test (ITT) and BP was assessed before and after training using the tail-cuff technique. After training, muscle mass increased in NMES vs. sham (416 ± 6 vs. 397 ± 6 mg, p = 0.04) along with the ratio of muscle mass (+3 ± 1% vs. -1 ± 1% p = 0.04). Moreover, insulin sensitivity improved in NMES vs. sham (average blood glucose during ITT: 139.6 ± 8.5 vs. 161.9 ± 9.0 mg/dl blood, p = 0.01). BP was decreased in both groups, although it is likely that the effect of NMES on BP was dampened by repetitive anesthesia. The metabolic benefit of NMES training could be of great utility in patients with chronic disease. Moreover, the clinical-like mouse model of NMES is an effective tool to investigate the systemic effects of local muscle strengthening.
Asunto(s)
Resistencia a la Insulina/fisiología , Músculo Esquelético/fisiopatología , Animales , Estimulación Eléctrica/métodos , Terapia por Estimulación Eléctrica/métodos , Extremidad Inferior/fisiología , Ratones , Ratones Endogámicos C57BL , Fuerza Muscular/fisiología , Atrofia Muscular/fisiopatologíaRESUMEN
We investigated the effects of intermittent hypoxia (IH), such as that encountered in severe obstructive sleep apnea (OSA) patients, on the development and severity of myocardial ischemia-related ventricular arrhythmias. Rats were exposed to 14 days of IH (30 s at 5%O2 and 30 s at 21%O2, 8 h·day-1) or normoxia (N, similar air-air cycles) and submitted to a 30-min coronary ligature. Arterial blood pressure (BP) and ECG were recorded for power spectral analysis, ECG interval measurement and arrhythmia quantification. Left ventricular monophasic action potential duration (APD) and expression of L-type calcium (LTCC) and transient receptor potential (TRPC) channels were assessed in adjacent epicardial and endocardial sites. Chronic IH enhanced the incidence of ischemic arrhythmias, in particular ventricular fibrillation (66.7% vs. 33.3% in N rats, p < 0.05). IH also increased BP and plasma norepinephine levels along with increased low-frequency (LF), decreased high-frequency (HF) and increased LF/HF ratio of heart rate and BP variability. IH prolonged QTc and Tpeak-to-Tend intervals, increased the ventricular APD gradient and upregulated endocardial but not epicardial LTCC, TRPC1 and TRPC6 (p < 0.05). Chronic IH, is a major risk factor for sudden cardiac death upon myocardial ischemia through sympathoactivation and alterations in ventricular repolarization, transmural APD gradient and endocardial calcium channel expression.