RESUMEN
BACKGROUND: Intracerebral hemorrhage (ICH) is a severe type of stroke with high mortality. Persistent hyperglycemia following ICH is linked to deteriorated neurological functions and death. However, the exacerbating effect of hyperglycemia on ICH injury at the molecular level is still unclear. Therefore, this study explores the impact of diabetes on ICH injury using a non-obese diabetic (NOD) mouse model of type I diabetes mellitus. METHODS: NOD and non-diabetic (non-obese resistant) mice subjected to ICH by intrastriatal injection of collagenase were sacrificed three days following the ICH. Brains were collected for hematoma volume measurement and immunohistochemistry. Neurobehavioral assays were conducted 24 h before ICH and then repeated at 24, 48 and 72 h following ICH. RESULTS: NOD mice showed increased hematoma volume and impairment in neurological function, as revealed by rotarod and grip strength analyses. Immunohistochemical staining showed reduced glial cell activation, as indicated by decreased GFAP and Iba1 staining. Furthermore, the expression of oxidative/nitrosative stress markers represented by 3-nitrotyrosine and inducible nitric oxide synthase was reduced in the diabetic group. CONCLUSIONS: Overall, our findings support the notion that hyperglycemia exacerbates ICH injury and worsens neurological function and that the mechanism of injury varies depending on the type of diabetes model used.
RESUMEN
Objectives: Palliative treatment of cyanotic congenital heart disease (CCHD) uses systemic-to-pulmonary conduits, often a modified Blalock-Taussig-Thomas shunt (mBTTs). Expanded polytetrafluoroethylene (ePTFE) mBTTs have associated risks for thrombosis and infection. The Human Acellular Vessel (HAV) (Humacyte, Inc) is a decellularized tissue-engineered blood vessel currently in clinical trials in adults for vascular trauma, peripheral artery disease, and end-stage renal disease requiring hemodialysis. In addition to restoring blood flow, the engineered HAV demonstrates the capacity for host cellular remodeling into native-like vasculature. Here we report preclinical evaluation of a small-diameter (3.5 mm) HAV as a mBTTs in a non-human primate model. Methods: We implanted 3.5 mm HAVs as right subclavian artery to pulmonary artery mBTTs in non-immunosuppressed juvenile rhesus macaques (n = 5). HAV patency, structure, and blood flow were assessed by postoperative imaging from 1 week to 6 months. Histology of HAVs and surrounding tissues was performed. Results: Surgical procedures were well tolerated, with satisfactory anastomoses, showing feasibility of using the 3.5 mm HAV as a mBTTs. All macaques had some immunological reactivity to the human extracellular matrix, as expected in this xenogeneic model. HAV mBTTs remained patent for up to 6 months in animals, exhibiting mild immunoreactivity. Two macaques displaying more severe immunoreactivity to the human HAV material developed midgraft dilatation without bleeding or rupture. HAV repopulation by host cells expressing smooth muscle and endothelial markers was observed in all animals. Conclusions: These findings may support use of 3.5 mm HAVs as mBTTs in CCHD and potentially other pediatric vascular indications.
RESUMEN
Diabetes Mellitus (DM) is a major comorbid condition that increases susceptibility to stroke. Intracerebral hemorrhage (ICH), a devastating type of stroke, accounts for only 13% of the total stroke cases but is associated with higher mortality. Multimorbid models of DM and ischemic stroke have been widely studied; however, fewer pieces of evidence are available on the impact of DM on the outcomes of ICH injury. In this study, we investigated the effect of DM on ICH-induced injury and cognitive impairments. Streptozotocin (STZ) induced type-I DM (T1DM) animal model was used, and experimental ICH was induced by intrastriatal injection of collagenase. Our results demonstrated that DM is associated with a significant increase in hematoma volume and deficits in post-stroke locomotor, sensorimotor, and cognitive behavior in mice. The levels of neuroinflammation, oxidative/nitrosative stress, and glial cell activation were also increased in the diabetic mice following ICH injury. This study provides a better understanding of the influence of DM comorbidity on hemorrhagic stroke outcomes and uncovers the important pathological mechanisms underlying DM-induced exacerbation of ICH injury.
Asunto(s)
Hemorragia Cerebral/metabolismo , Disfunción Cognitiva/metabolismo , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Tipo 1/metabolismo , Estrés Oxidativo/fisiología , Accidente Cerebrovascular/metabolismo , Animales , Hemorragia Cerebral/inducido químicamente , Disfunción Cognitiva/inducido químicamente , Diabetes Mellitus Experimental/inducido químicamente , Diabetes Mellitus Tipo 1/inducido químicamente , Fuerza de la Mano/fisiología , Mediadores de Inflamación/metabolismo , Locomoción/efectos de los fármacos , Locomoción/fisiología , Masculino , Aprendizaje por Laberinto/fisiología , Ratones , Ratones Endogámicos C57BL , Estreptozocina/toxicidad , Accidente Cerebrovascular/inducido químicamenteRESUMEN
Intracerebral hemorrhage (ICH) resulting from the rupture of the blood vessels in the brain is associated with significantly higher mortality and morbidity. Clinical studies focused on alleviating the primary injury, hematoma formation and expansion, were largely ineffective, suggesting that secondary injury-induced inflammation and the formation of reactive species also contribute to the overall injury process. In this study, we explored the effects of cofilin knockdown in a mouse model of ICH. Animals given stereotaxic injections of cofilin siRNA, 72-h prior to induction of ICH by collagenase injection within the area of siRNA administration showed significantly decreased cofilin expression levels and lower hemorrhage volume and edema, and the animals performed significantly better in neurobehavioral tasks i.e., rotarod, grip strength and neurologic deficit scores. Cofilin siRNA knocked-down mice had reduced ICH-induced DNA fragmentation, blood-brain barrier disruption and microglial activation, with a concomitant increase in astrocyte activation. Increased expression of pro-survival proteins and decreased markers of oxidative stress were also observed in cofilin siRNA-treated mice possibly due to the reduced levels of cofilin. Our results suggest that cofilin plays a major role in ICH-induced secondary injury, and could become a potential therapeutic target.
Asunto(s)
Factores Despolimerizantes de la Actina/metabolismo , Hemorragias Intracraneales/fisiopatología , Microglía/patología , Estrés Oxidativo/fisiología , Animales , Técnicas de Silenciamiento del Gen , Hemorragias Intracraneales/metabolismo , Hemorragias Intracraneales/patología , Masculino , Ratones , Ratones Endogámicos C57BLRESUMEN
Ischemic stroke is caused by a blockage of cerebral blood flow resulting in neuronal and glial hypoxia leading to inflammatory and reactive oxygen species (ROS)-mediated cell death. Nitric oxide (NO) formed by NO synthase (NOS) is known to be protective in ischemic stroke, however NOS has been shown to 'uncouple' under oxidative conditions to instead produce ROS. Nitrones are antioxidant molecules that are shown to trap ROS to then decompose and release NO. In this study, the nitrone 5 was designed such that its decomposition product is a NOS inhibitor, 6, effectively leading to NOS inhibition specifically at the site of ROS production. The ability of 5 to spin-trap radicals and decompose to 6 was observed using EPR and LC-MS/MS. The pro-drug concept was tested in vitro by measuring cell viability and 6 formation in SH-SY5Y cells subjected to oxygen glucose deprivation (OGD). 5 was found to be more efficacious and more potent than PBN, and was able to increase phospho-Akt while reducing nitrotyrosine and cleaved caspase-3 levels. 6 treatment, but not 5, was found to decrease NO production in LPS-stimulated microglia. Doppler flowmetry on anesthetized mice showed increased cerebral blood flow upon intravenous administration of 1mg/kg of 5, but a return to baseline upon administration of 10mg/kg, likely due to its dual nature of antioxidant/NO-donor and NOS-inhibition. Mice treated with 5 after permanent ischemia exhibited a >30% reduction in infarct volume, and higher formation of 6 in ischemic tissue resulting in region specific effects limited to the infarct area.
Asunto(s)
Antioxidantes/uso terapéutico , Isquemia/tratamiento farmacológico , Microglía/fisiología , Neuronas/fisiología , Óxido Nítrico Sintasa/antagonistas & inhibidores , Accidente Cerebrovascular/tratamiento farmacológico , Animales , Antioxidantes/síntesis química , Caspasa 3/metabolismo , Supervivencia Celular , Células Cultivadas , Modelos Animales de Enfermedad , Humanos , Ratones , Ratones Endogámicos C57BL , Microglía/efectos de los fármacos , Neuronas/efectos de los fármacos , Óxido Nítrico/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Tirosina/análogos & derivados , Tirosina/metabolismoRESUMEN
Ginkgo biloba extract is an alternative medicine available as a standardized formulation, EGb 761(®), which consists of ginkgolides, bilobalide, and flavonoids. The individual constituents have varying therapeutic mechanisms that contribute to the pharmacological activity of the extract as a whole. Recent studies show anxiolytic properties of ginkgolide A, migraine with aura treatment by ginkgolide B, a reduction in ischemia-induced glutamate excitotoxicity by bilobalide, and an alternative antihypertensive property of quercetin, among others. These findings have been observed in EGb 761 as well and have led to clinical investigation into its use as a therapeutic for conditions such as cognition, dementia, cardiovascular, and cerebrovascular diseases. This review explores the therapeutic mechanisms of the individual EGb 761 constituents to explain the pharmacology as a whole and its clinical application to cardiovascular and neurological disorders, in particular ischemic stroke.
RESUMEN
Reactive oxygen species (ROS) are important in regulating normal cell physiological functions, but when produced in excess lead to the augmented pathogenesis of various diseases. Among these, ischemia reperfusion injury, Alzheimer's disease and rheumatoid arthritis are particularly important. Since ROS can be counteracted by a variety of antioxidants, natural and synthetic antioxidants have been developed. However, due to the ubiquitous production of ROS in living systems, poor in vivo efficiency of these agents and lack of target specificity, the current clinical modalities to treat oxidative stress damage are limited. Advances in the developing field of nanomedicine have yielded nanoparticles that can prolong antioxidant activity, and target specificity of these agents. This article reviews recent advances in antioxidant nanoparticles and their applications to manage oxidative stress-mediated diseases. FROM THE CLINICAL EDITOR: Production of reactive oxygen species (ROS) is a purely physiological process in many disease conditions. However, excessive and uncontrolled production will lead to oxidative stress and further tissue damage. Advances in nanomedicine have provided many novel strategies to try to combat and counteract ROS. In this review article, the authors comprehensively highlighted the current status and future developments in using nanotechnology for providing novel therapeutic options in this field.
Asunto(s)
Antioxidantes/administración & dosificación , Antioxidantes/uso terapéutico , Sistemas de Liberación de Medicamentos/métodos , Estrés Oxidativo/efectos de los fármacos , Especies Reactivas de Oxígeno/metabolismo , Animales , Antioxidantes/química , Portadores de Fármacos/química , Humanos , Nanomedicina/métodos , Nanopartículas/química , Nanotecnología/métodosRESUMEN
In this work, a series of α-phenyl-N-tert-butyl nitrones bearing one, two, or three substituents on the tert-butyl group was synthesized. Cyclic voltammetry (CV) was used to investigate their electrochemical properties and showed a more pronounced substituent effect for oxidation than for reduction. Rate constants of superoxide radical (O2(â¢-)) reactions with nitrones were determined using a UV-vis stopped-flow method, and phenyl radical (Ph(â¢)) trapping rate constants were measured by EPR spectroscopy. The effect of N-tert-butyl substitution on the charge density and electron density localization of the nitronyl carbon as well as on the free energies of nitrone reactivity with O2(â¢-) and HO2(â¢) were computationally rationalized at the PCM/B3LYP/6-31+G**//B3LYP/6-31G* level of theory. Theoretical and experimental data showed that the rates of the reaction correlate with the nitronyl carbon charge density, suggesting a nucleophilic nature of O2(â¢-) and Ph(â¢) addition to the nitronyl carbon atom. Finally, the substituent effect was investigated in cell cultures exposed to hydrogen peroxide and a correlation between the cell viability and the oxidation potential of the nitrones was observed. Through a combination of computational methodologies and experimental methods, new insights into the reactivity of free radicals with nitrone derivatives have been proposed.
Asunto(s)
Óxidos de Nitrógeno/síntesis química , Conformación Molecular , Óxidos de Nitrógeno/química , Teoría CuánticaRESUMEN
Reactive nitrogen/oxygen species (ROS/RNS) at low concentrations play an important role in regulating cell function, signaling, and immune response but in unregulated concentrations are detrimental to cell viability. While living systems have evolved with endogenous and dietary antioxidant defense mechanisms to regulate ROS generation, ROS are produced continuously as natural by-products of normal metabolism of oxygen and can cause oxidative damage to biomolecules resulting in loss of protein function, DNA cleavage, or lipid peroxidation, and ultimately to oxidative stress leading to cell injury or death. Superoxide radical anion (O2â¢-) is the major precursor of some of the most highly oxidizing species known to exist in biological systems such as peroxynitrite and hydroxyl radical. The generation of O2â¢- signals the first sign of oxidative burst, and therefore, its detection and/or sequestration in biological systems is important. In this demonstration, O2â¢- was generated from polymorphonuclear neutrophils (PMNs). Through chemotactic stimulation with phorbol-12-myristate-13-acetate (PMA), PMN generates O2â¢- via activation of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase. Nitric oxide (NO) synthase which comes in three isoforms, as inducible-, neuronal- and endothelial-NOS, or iNOS, nNOS or eNOS, respectively, catalyzes the conversion of L- arginine to L-citrulline, using NADPH to produce NO. Here, we generated NO from endothelial cells. Under oxidative stress conditions, eNOS for example can switch from producing NO to O2â¢- in a process called uncoupling, which is believed to be caused by oxidation of heme or the co-factor, tetrahydrobiopterin (BH4). There are only few reliable methods for the detection of free radicals in biological systems but are limited by specificity and sensitivity. Spin trapping is commonly used for the identification of free radicals and involves the addition reaction of a radical to a spin trap forming a persistent spin adduct which can be detected by electron paramagnetic resonance (EPR) spectroscopy. The various radical adducts exhibit distinctive spectrum which can be used to identify the radicals being generated and can provide a wealth of information about the nature and kinetics of radical production. The cyclic nitrones, 5,5-dimethyl-pyrroline-N-oxide, DMPO, the phosphoryl-substituted DEPMPO, and the ester-substituted, EMPO and BMPO, have been widely employed as spin traps--the latter spin traps exhibiting longer half-lives for O2â¢- adduct. Iron (II)-N-methyl-D-glucamine dithiocarbamate, Fe(MGD)2 is commonly used to trap NO due to high rate of adduct formation and the high stability of the spin adduct.
Asunto(s)
Espectroscopía de Resonancia por Spin del Electrón/métodos , Óxido Nítrico/química , Detección de Spin/métodos , Superóxidos/química , Animales , Bovinos , Células Endoteliales/química , Óxido Nítrico/análisis , Óxidos de Nitrógeno/química , Superóxidos/análisisRESUMEN
Reactive nitrogen species (RNS) such as nitrogen dioxide ((â¢)NO(2)), peroxynitrite (ONOO(-)), and nitrosoperoxycarbonate (ONOOCO(2)(-)) are among the most damaging species present in biological systems due to their ability to cause modification of key biomolecular systems through oxidation, nitrosylation, and nitration. Nitrone spin traps are known to react with free radicals and nonradicals via electrophilic and nucleophilic addition reactions and have been employed as reagents to detect radicals using electron paramagnetic resonance (EPR) spectroscopy and as pharmacological agents against oxidative stress-mediated injury. This study examines the reactivity of cyclic nitrones such as 5,5-dimethylpyrroline N-oxide (DMPO) with (â¢)NO(2), ONOO(-), ONOOCO(2)(-), SNAP, and SIN-1 using EPR. The thermochemistries of nitrone reactivity with RNS and isotropic hfsc's of the addition products were also calculated at the PCM(water)/B3LYP/6-31+G**//B3LYP/6-31G* level of theory with and without explicit water molecules to rationalize the nature of the observed EPR spectra. Spin trapping of other RNS such as azide ((â¢)N(3)), nitrogen trioxide ((â¢)NO(3)), amino ((â¢)NH(2)) radicals and nitroxyl (HNO) were also theoretically and experimentally investigated by EPR spin trapping and mass spectrometry. This study also shows that other spin traps such as 5-carbamoyl-5-methyl-pyrroline N-oxide, 5-ethoxycarbonyl-5-methyl-pyrroline N-oxide, and 5-(diethoxyphosphoryl)-5-methyl-1-pyrroline N-oxide can react with radical and nonradical RNS, thus making spin traps suitable probes as well as antioxidants against RNS-mediated oxidative damage.
Asunto(s)
Modelos Moleculares , Óxidos de Nitrógeno/química , Especies de Nitrógeno Reactivo/química , Carbonatos/química , Óxidos N-Cíclicos/química , Espectroscopía de Resonancia por Spin del Electrón , Espectrometría de Masas , Nitratos/química , Oxidación-Reducción , Ácido Peroxinitroso/química , Detección de Spin , TermodinámicaRESUMEN
Nitrone therapeutics has been employed in the treatment of oxidative stress-related diseases such as neurodegeneration, cardiovascular disease and cancer. The nitrone-based compound NXY-059, which is the first drug to reach clinical trials for the treatment of acute ischemic stroke, has provided promise for the development of more robust pharmacological agents. However, the specific mechanism of nitrone bioactivity remains unclear. In this review, we present a variety of nitrone chemistry and biological activity that could be implicated for the nitrone's pharmacological activity. The chemistries of spin trapping and spin adduct reveal insights on the possible roles of nitrones for altering cellular redox status through radical scavenging or nitric oxide donation, and their biological effects are presented. An interdisciplinary approach towards the development of novel synthetic antioxidants with improved pharmacological properties encompassing theoretical, synthetic, biochemical and in vitro/in vivo studies is covered.
Asunto(s)
Bencenosulfonatos/metabolismo , Depuradores de Radicales Libres/metabolismo , Marcadores de Spin , Superóxidos/metabolismo , Bencenosulfonatos/químicaRESUMEN
Nitrone spin traps have been employed as probes for the identification of transient radical species in chemical and biological systems using electron paramagnetic resonance (EPR) spectroscopy and have exhibited pharmacological activity against oxidative-stress-mediated diseases. Since superoxide radical anion (O2(â¢-)) is a major precursor to most reactive oxygen species and calix[4]pyrroles have been shown to exhibit high affinity to anions, a cyclic nitrone conjugate of calix[4]pyrrole (CalixMPO) was designed, synthesized, and characterized. Computational studies at the PCM/B3LYP/6-31+G(d,p)//B3LYP/6-31G(d) level suggest a pendant-type linkage between the calix[4]pyrrole and the nitrone to be the most efficient design for spin trapping of O2(â¢-), giving exoergic reaction enthalpies (ΔH(298K,aq)) and free energies (ΔG(298K,aq)) of -16.9 and -2.1 kcal/mol, respectively. (1)H NMR study revealed solvent-dependent conformational changes in CalixMPO leading to changes in the electronic properties of the nitronyl group upon H-bonding with the pyrrole groups as also confirmed by calculations. CalixMPO spin trapping of O2(â¢-) exhibited robust EPR spectra. Kinetic analysis of O2(â¢-) adduct formation and decay in polar aprotic solvents using UV-vis stopped-flow and EPR methods gave a larger trapping rate constant for CalixMPO and a longer half-life for its O2(â¢-) adduct compared to the commonly used nitrones. The unusually high reactivity of CalixMPO with O2(â¢-) was rationalized to be due to the synergy between the α-effect and electrostatic effect by the calix[4]pyrrole moiety on O2(â¢-) and the nitrone, respectively. This work demonstrates for the first time the application of an anion receptor for the detection of one of the most important radical intermediates in biological and chemical systems (i.e., O2(â¢-)).