Extracellular superoxide dismutase is necessary to maintain renal blood flow during sepsis development

Background: Extracellular superoxide dismutase (ECSOD) protects nitric oxide (NO) bioavailability by decreasing superoxide levels and preventing peroxynitrite generation, which is important in maintaining renal blood flow and in preventing acute kidney injury. However, the profile of ECSOD expression after sepsis is not fully understood. Therefore, we intended to evaluate the content and gene expression of superoxide dismutase (SOD) isoforms in the renal artery and their relation to renal blood flow. Methods: Sepsis was induced in Wistar rats by caecal ligation and perforation. Several times after sepsis induction, renal blood flow (12, 24 and 48 h); the renal arterial content of SOD isoforms, nitrotyrosine, endothelial and inducible nitric oxide synthase (e-NOS and i-NOS), and phosphorylated vasodilator-stimulated phosphoprotein (pVASP); and SOD activity (3, 6 and 12 h) were measured. The influence of a SOD inhibitor was also evaluated. Results: An increase in ECSOD content was associated with decreased 3-nitrotyrosine levels. These events were associated with an increase in pVASP content and maintenance of renal blood flow. Moreover, previous treatment with a SOD inhibitor increased nitrotyrosine content and reduced renal blood flow. Conclusions: ECSOD appears to have a major role in decreasing peroxynitrite formation in the renal artery during the early stages of sepsis development, and its application can be important in renal blood flow control and maintenance during septic insult

Avalia??o da toxicidade causada pela exposi??o a IONPS utilizando zebrafish como organismo modelo

Submitted by PPG Medicina e Ci?ncias da Sa?de ([email protected]) on 2017-12-26T13:35:05Z No. of bitstreams: 1 GIOVANNA_MEDEIROS_TAVARES_DE_OLIVEIRA_TES.pdf: 3690052 bytes, checksum: e6e9b4fea2305637782a214e8f45af2d (MD5)Approved for entry into archive by Caroline Xavier ([email protected]) on 2017-12-29T10:45:02Z (GMT) No. of bitstreams: 1 GIOVANNA_MEDEIROS_TAVARES_DE_OLIVEIRA_TES.pdf: 3690052 bytes, checksum: e6e9b4fea2305637782a214e8f45af2d (MD5)Made available in DSpace on 2017-12-29T10:47:48Z (GMT). No. of bitstreams: 1 GIOVANNA_MEDEIROS_TAVARES_DE_OLIVEIRA_TES.pdf: 3690052 bytes, checksum: e6e9b4fea2305637782a214e8f45af2d (MD5) Previous issue date: 2017-08-28Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior - CAPESInitially used in magnetic resonance imaging in the late 1970s, iron oxide nanoparticles (IONPs) have wide application in the medical field today, in diagnostics, drug delivery, cellular therapies and theragnostic. The high biocompatibility, small size, functionalization and ability to respond to an applied magnetic field gives this nanoparticle great advantage over other nanomaterials. Studies have demonstrated the low toxicity and high applicability of this nanomaterial in the clinic, however some questions remain unanswered. When in contact with the in vivo metabolism, nanomaterials can behave in a way to degrade their coating and release the ions contained in their nucleus. In fact, side effects related to exposure to IONPs are mainly related to the release of its elemental nucleus; which, when endocyted, can be degraded inside the lysosomes and release [Fe] ions. Changes in iron homeostasis can be very damaging to the cell, causing inflammation, lipid peroxidation, and oxidative stress. Organs more sensitive to iron accumulation, such as the heart, may demonstrate apoptosis and tissue degeneration. Such toxic effects are easily detected in studies using zebrafish as an animal model. Protocols with zebrafish embryos for toxicological analysis have the advantage of allowing large-scale screening on development, survival, behavior, gene expression and cardiotoxicity. Under this scenario, this thesis aims to evaluate the toxicity of commertial and "in house" synthesis of Iron Oxide Nanoparticles in zebrafish. Behavioral analysis of locomotion and gene expression of zebrafish larvae exposed to uncoated and dextran-coated iron oxide nanoparticles indicated a toxicity at low concentrations of nanoparticle exposure, contrary to what is reported in the literature. In addition, changes in the apoptotic pathway suggest that this route is closely linked to the behavioral effects found. Subsequent analyzes, targeting cardiotoxicity, suggested that concentrations above 100 ?g/ml are damaging for the heart. Molecular analyzes in the groups exposed to the iron oxide nanoparticle and to iron solution helped to establish a parallel between the toxicity of these nanoparticles and the pathways of iron metabolism.Inicialmente utilizadas em exames de resson?ncia magn?tica no final dos anos 1970 as nanopart?culas de ?xido de ferro (IONPs) possuem hoje vasta aplica??o na ?rea m?dica, em exames de diagn?stico, sistema de envio de drogas, terapias celulares e como agente teragn?stico. Sua alta biocompatibilidade, pequeno tamanho, facilidade de manipula??o e capacidade de responder a aplica??o externa de campo magn?tico lhe oferece grande vantagem sobre outros nanomaterias. Estudos vem demostrando a baixa toxicidade e alta aplicabilidade deste nanomaterial na cl?nica, entretanto algumas quest?es ainda se encontram sem resposta. Quando em contato com o metabolismo in vivo, nanomateriais podem se comportar de forma a degradar sua estrtutura externa e liberar os ?oins contidos no seu n?cleo. De fato, efeitos adversos relacionado a exposi??o a IONPs est?o majoritariamente relacionadas a libera??o do seu n?cleo elementar; que, quando endocitado, pode ser degradado nos lisossomos e liberar ?ons [Fe]. Altera??es na homeostase de ferro podem ser muito prejudiciais ? c?lula, causando inflama??o, peroxida??o lip?dica e estresse oxidativo. ?rg?os mais sens?veis ao ac?mulo do ferro, como o cora??o, podem apresentar apoptose e degenera??o tecidual. Tais efeitos t?xicos s?o facilmente detectados em estudos utilizando zebrafish como animal modelo. Protocolos com embri?es de zebrafish para an?lise toxicol?gica possuem a vantagem de permitir estudos em grande escala de efeitos no desenvolvimento, sobreviv?ncia, comportamento, express?o g?nica e cardiotoxicidade. Sob esse cen?rio, esta tese tem como objetivo avaliar a toxicidade de Nanopart?culas de Oxido de Ferro (IONPs) de s?ntese pr?pria e comerciais no modelo experimental zebrafish. An?lise comportamental de locomo??o e express?o g?nica de larvas de zebrafish expostas a Nanopart?culas de ?xido de Ferro puras (sem envolt?rio) e revestidas com dextran indicou uma toxicidade em baixas concentra??es de exposi??o ? nanopart?culas, contrario ao que ? relatado na literatura. Al?m disso, altera??es na via apopt?tica sugere que esta rota esteja intimamente ligada aos efeitos comportamentais encontrados. An?lises posteriores, direcionadas ? cardiotoxicidade sugerem efeitos t?xicos acima de 100 ?g/mL. An?lises gen?mica de express?o nos grupos expostos ? nanopart?cula de ?xido de ferro e ? solu??o de ferro met?lico (usado como controle de positivo de excesso de ferro) permitiram a identifica??o de um paralelo entre toxicidade destas nanopart?culas e as vias de metaboliza??o do ferro

Modula??o da neurotransmiss?o colin?rgica como resposta aos efeitos causados pela exposi??o a nanopart?culas de ?xido de ferro revestidas com dextran-aminado utilizando zebrafish (Danio rerio) como modelo de estudo

Made available in DSpace on 2015-04-14T14:51:29Z (GMT). No. of bitstreams: 1 451666.pdf: 1289575 bytes, checksum: 022edd4e2e930676c57fc69c0dc3e849 (MD5) Previous issue date: 2013-07-24Superparamagnetic iron oxide nanoparticles (SPIONs) are of great interest in nanomedicine due to their capability to act simultaneously as a contrast agent in magnetic resonance imaging and as a targeted drug delivery system with good biocompatibility. At present, one of the biggest concerns about the use of SPIONs remains around its toxicity. For this reason, it is important to establish the safe upper limit for each use. In the present study, SPION coated with cross-linked and aminated dextran (CLIO-NH2) were synthesized by the co-precipitation method and characterized by transmission electron microscopy (TEM) and light scattering dynamics. The analysis of the elementary composition of the solution at the end of the synthesis was also performed. The magnetite core size was 5.5 ? 1.4 nm, on a concentration of 10 mg Fe-NPs/ml. We have evaluated the effects of different CLIO-NH2 doses (20, 50, 100, 140 and 200 mg/kg) as a function of time after exposure (one, 16, 24 and 48 hours) on AChE activity and ache expression in zebrafish brain. At tested concentrations, only the animals exposed to 200 mg/kg, and assessed 24 h after administration of the nanoparticles, have shown decreased AChE activity. These values returned to control levels after 48 h of exposition, indicating a transitory toxical effect. The RT-qPCR results suggested that inhibition of brain AChE is not directly related with the transcriptional control, and it was probably due to post-transcriptional events. Once ACh is recognized to play an important role in the regulation of locomotor control, we further evaluated parameters of zebrafish swimming activity. CLIO-NH2 at 200mg/kg, evaluated after 24 h, also impaired all the tested parameters of zebrafish swimming activity, i.e. decreased traveled distance, mean speed, number of line crossings, and turn angle. We further investigated the iron accumulation in zebrafish brain by ICP-MS, and a significant higher level of ferric iron was found in zebrafish brains exposed to CLIO-NH2. In summary, the results presented herein provide further experimental evidence that exposure to high doses of dextran-coated iron oxide nanoparticles can be transiently neurotoxic.Nanopart?culas superparamagn?ticas de ?xido de ferro (SPIONs) s?o de grande interesse na nanomedicina, devido ? sua capacidade para agir, simultaneamente, como um agente de contraste em imagem por resson?ncia magn?tica e como um sistema de entrega de f?rmacos espec?ficos, possuindo boa biocompatibilidade. Atualmente, uma das maiores preocupa??es com a utiliza??o de SPIONs permanece em torno da sua toxicidade e, por esta raz?o, ? importante estabelecer um limite de seguran?a para sua utiliza??o. Neste estudo, SPIONs revestidas com dextran-aminado (CLIO-NH2) foram sintetizados pelo m?todo de co-precipita??o e caracterizadas por microscopia eletr?nica de transmiss?o (MET) e din?mica de espalhamento de luz. A an?lise da composi??o elementar da solu??o no final da s?ntese foi tamb?m realizada. O tamanho do n?cleo de magnetite foi de 5,5 ? 1,4 nm, com uma concentra??o de 10 mg Fe-NPs/ml. Foram avaliados o efeito de diferentes doses de CLIO-NH2 (20, 50, 100, 140 e 200 mg/kg), ap?s a exposi??o de CLIO-NH2 (em uma, 16, 24 e 48 h), sobre a atividade de AChE e sobre a express?o de ache em c?rebro de peixe-zebra. Nas concentra??es testadas, apenas os animais expostos a 200 mg/kg, testados 24 h ap?s a administra??o das nanopart?culas, demonstraram diminui??o da atividade da AChE. Quarenta e oito horas ap?s exposi??o a CLIO-NH2 os valores voltaram aos n?veis controle, indicando um efeito t?xico transit?rio. Os resultados de RT-qPCR sugerem que a inibi??o da AChE no c?rebro n?o est? diretamente relacionada com o controle da transcri??o e que esta relacionada com a modula??o de eventos p?s-transcricionais. Uma vez que a ACh ? reconhecida por desempenhar um papel importante na regula??o do controle locomotor, foram avaliados par?metros da atividade da nata??o do peixe-zebra. CLIO-NH2, ? 200 mg/kg, ap?s 24 h de tratamento, tamb?m prejudicou todos os par?metros testados de comportamento de nado, ocorrendo diminui??o da dist?ncia percorrida, velocidade, n?mero de cruzamentos de linha e ?ngulo de virada. Ainda foi analisado o ac?mulo de ferro no c?rebro de peixe-zebra, por ICP-MS, onde um n?vel significativamente maior de ferro f?rrico foi encontrado nos c?rebros de peixe-zebra expostos a CLIO-NH2 nas mesmas condi??es experimentais (200 mg/kg e 24 h de exposi??o). Concluindo, os resultados apresentados fornecem evid?ncias de que doses elevadas de nanopart?culas de ?xido de ferro, revestidas com dextran-aminado, podem produzir neurotoxicidade transit?ria, coincidente com n?veis cerebrais elevados de ferro, bem como as altera??es comportamentais

Iron from haemoglobin and haemin modulates nucleotide hydrolysis in Trichomonas vaginalis

Extracellular ATP may act as a danger signalling molecule, inducing inflammation and immune responses in infection sites. The ectonucleotidases NTPDase and ecto-5’-nucleotidase are enzymes that modulate extracellular nucleotide levels; these enzymes have been previously characterised in Trichomonas vaginalis. Iron plays an important role in the complex trichomonal pathogenesis. Herein, the effects of iron on growth, nucleotide hydrolysis and NTPDase gene expression in T. vaginalis isolates from female and male patients were evaluated. Iron from different sources sustained T. vaginalis growth. Importantly, iron from haemoglobin (HB) and haemin (HM) enhanced NTPDase activity in isolates from female patients and conversely reduced the enzyme activity in isolates from male patients. Iron treatments could not alter the NTPDase transcript levels in T. vaginalis. Furthermore, our results reveal a distinct ATP, ADP and AMP hydrolysis profile between isolates from female and male patients influenced by iron from HB and HM. Our data indicate the participation of NTPDase and ecto-5’-nucleotidase in the establishment of trichomonas infection through ATP degradation and adenosine production influenced by iron

Iron from haemoglobin and haemin modulates nucleotide hydrolysis in Trichomonas vaginalis

Extracellular ATP may act as a danger signalling molecule, inducing inflammation and immune responses in infection sites. The ectonucleotidases NTPDase and ecto-5’-nucleotidase are enzymes that modulate extracellular nucleotide levels; these enzymes have been previously characterised in Trichomonas vaginalis. Iron plays an important role in the complex trichomonal pathogenesis. Herein, the effects of iron on growth, nucleotide hydrolysis and NTPDase gene expression in T. vaginalis isolates from female and male patients were evaluated. Iron from different sources sustained T. vaginalis growth. Importantly, iron from haemoglobin (HB) and haemin (HM) enhanced NTPDase activity in isolates from female patients and conversely reduced the enzyme activity in isolates from male patients. Iron treatments could not alter the NTPDase transcript levels in T. vaginalis. Furthermore, our results reveal a distinct ATP, ADP and AMP hydrolysis profile between isolates from female and male patients influenced by iron from HB and HM. Our data indicate the participation of NTPDase and ecto-5’-nucleotidase in the establishment of trichomonas infection through ATP degradation and adenosine production influenced by iron

Effects of chlorogenic acid, caffeine and coffee on components of the purinergic system of streptozotocin-induced diabetic rats

We evaluated the effect of chlorogenic acid (CGA), caffeine (CA) and coffee (CF) on components of the purinergic system from the cerebral cortex and platelets of streptozotocin-induced diabetic rats. Animals were divided into eight groups: control animals treated with (I) water (WT), (II) CGA (5 mg/kg), (III) CA (15 mg/kg) and (IV) CF (0.5 g/kg), and diabetic animals treated with (V) WT, (VI) CGA (5 mg/kg), (VII) CA (15 mg/kg) and (VIII) CF (0.5 g/kg). Our results showed an increase (173%) in adenosine monophosphate (AMP) hydrolysis in the cerebral cortex of diabetic rats. In addition, CF treatment increased adenosine diphosphate (ADP) and AMP hydrolysis in group VIII synaptosomes. Platelets showed an increase in ectonucleotidase activity in group V, and all treatments reduced the increase in adenosine triphosphate and ADP hydrolysis. Furthermore, there was an increase in platelet aggregation of 72% in the diabetic rats, and CGA and CF treatment reduced platelet aggregation by nearly 60% when compared to diabetic rats. In this context, we can suggest that CGA and CF treatment should be considered a therapeutic and scientific target to be investigated in diseases associated with hyperglycemia

Regulation of lung oxidative damage by endogenous superoxide dismutase in sepsis

Background: The purpose of this research is to study the relationship between superoxide dismutase (SOD) and lung redox state in an animal model of sepsis. Methods: Sepsis was induced in rats by the cecal ligation and perforation model (CLP). After 3, 6, and 12 h, CLP protein content and expression of SOD1, SOD2, and SOD3 were evaluated, and SOD activity was assessed. Oxidative damage was determined by quantifying nitrotyrosine content. Lung localization of SOD3 was performed by immunohistochemistry. The protective effect of a SOD mimetic on oxidative damage, inflammation, and lung permeability was assessed 12 and 24 h after sepsis induction. Results: Lung levels of SOD1 decreased 3 and 12 h after sepsis, but SOD2 and SOD3 increased, as well as SOD activity. These alterations were not associated with changes in sod gene expression. Nitrotyrosine levels increased 3 and 12 h after sepsis. The administration of a SOD mimetic decreased nitrotyrosine and proinflammatory cytokine levels and improved lung permeability. Conclusions: SOD2 and SOD3 increased after sepsis induction, but this was insufficient to protect the lung. Treatments based on SOD mimetics could have a role in lung injury associated with sepsis