Examinando por Autor "Blanco-Ruiz, Santos"

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Age-related changes of the nitric oxide system in the rat brain
(Elsevier BV, 2002) Siles, Eva; Martínez-Lara, Esther; Cañuelo, Ana; Sánchez, Marta; Hernández-Cobo, Raquel; López-Ramos, Juan Carlos; del-Moral, María Luisa; Esteban-Ruiz, Francisco José; Blanco-Ruiz, Santos; Pedrosa-Raya, Juan ángel; Rodrigo, José; Peinado-Herreros, María Ángeles
This work examines the age-related changes of the NO pathway in the central nervous system (CNS), analyzing nitric oxide synthase (NOS) isoform expression, the level of nitrotyrosine-modified proteins, and the NOS activity in the cerebral cortex, decorticated brain (basal ganglia, thalamus, hypothalamus, tegtum and tegmentum) and cerebellum of young, adult and aged rats. Our data demonstrate that the different NOS isoforms are not uniformly expressed across the CNS. In this sense, the nNOS and eNOS isoenzymes are expressed mainly in the cerebellum and decorticated brain, respectively, while the iNOS isoenzyme shows the highest level in cerebellum. Concerning age, in the cerebral cortex nNOS significantly increased its expression only in adult animals; meanwhile, in the cerebellum the eNOS expression decreased whereas iNOS increased in adult and aged rats. No age-related changes in any isoform were found in decorticated brain. NOS activity, determined by nitrate plus nitrite quantification, registered the highest levels in the cerebellum, where the significant increase detected with aging was probably related to iNOS activity. The number of nitrotyrosine-modified immunoreactive bands differed among regions; thus, the highest number was detected in the decorticated brain while the cerebellum showed the least number of bands. Finally, bulk protein nitration increased in cerebral cortex only in adult animal. No changes were found in the decorticated brain, and the decrease detected in the cerebellum of aged animals was not significant. According to these results, the NO pathway is differently modified with age in the three CNS regions analyzed.
APPLIED QUANTITATIVE PROTEOMICS ANALYSIS
(Jaén : Universidad de Jaén, 2020-04-03) Ovelleiro, David; Peinado-Herreros, María Ángeles; Blanco-Ruiz, Santos; Universidad de Jaén. Departamento de Biología Experimental
En esta tesis se ha aplicado el estado del arte en análisis cuantitativo en proteómica. Los datos analizados en este trabajo, provenientes de tres proyectos distintos, fueron obtenidos usando tres de las técnicas más utilizadas en proteómica: cuantificación label-free, marcaje isobárico y SWATH. Los resultados obtenidos en los diferentes proyectos son también interpretados mediante múltiples herramientas bioinformáticas. La cuantificación label-free es utilizada aquí para obtener la combinación óptima de software y parámetros usando un conjunto de datos públicos. El marcaje isobárico, usando TMT, se emplea en el estudio de los diferentes perfiles de expresión proteica, obtenidos con dos modelos de hipoxia de diferente severidad en cerebros de rata. La técnica SWATH se busca en la búsqueda de biomarcadores de síndorme de ovario poliquístico en plasma. Por último, los elementos necesarios para la implantación de una plataforma de análisis proteómica , en términos de software y hardware, se describen en forma detallada. In this thesis, the state of the art in quantitative proteomics analysis has been applied. The data analyzed in this work, coming from three different projects, were acquired using three of the most used techniques in proteomics: label-free, isobaric labeling and SWATH. The results obtained in the different projects are also interpreted using multiple bioinformatics tools. The label-free quantization is used here to asses the optimal combination of software and parameters using a public data set. Isobaric labeling, using TMT, is employed to study the different profiles in protein expression when two hypoxic models, with different severity, are applied in rat brains. The SWATH technique is used in the search of biomarkers for polycystic ovary syndrome in plasma. Finally, the elements required for setting up a platform for proteomics analysis, both in terms of hardware and software, are comprehensively described.
Are patients with fibromyalgia in a prothrombotic state?
(Sage Publications Inc, 2019) Molina-Ortega, Francisco Javier; del-Moral, María Luisa; la-Rubia, Mercedes; Blanco-Ruiz, Santos; Carmona-Martos, Ramón; Rus-Martínez, Alma
Objectives: The aim of this study was to investigate thrombosis-related parameters (blood coagulation parameters, platelet indices, red blood cell [RBC] count, and inflammatory markers) in patients with fibromyalgia (FM). Method: We carried out a case–control study with 35 women with FM and 12 age-matched healthy volunteers to analyze fibrinogen levels, prothrombin time, cephaline time, platelet count, platelet distribution width (PDW), mean platelet volume (MPV), RBC count, neutrophil-tolymphocyte ratio, and platelet-to-lymphocyte ratio (PLR). Results: The results showed significantly increased fibrinogen levels (p < .05), platelet count (p < .05), PDW (p ¼ .059), RBC count (p < .05), and PLR (p < .05) in women with FM versus the healthy volunteers. Prothrombin time (p < .05) and MPV (p < .05) were significantly lower in patients with FM than in the controls. Conclusions: Elevated platelet and RBC counts, PDW values, and fibrinogen levels as well as decreased prothrombin time are all indicative of a prothrombotic state in FM patients, which may be enhanced by an increased inflammatory tone. This prothrombotic state may increase the risk of thrombosis-related cardiovascular disease in patients with FM.
Biological Implications of a Stroke Therapy Based in Neuroglobin Hyaluronate Nanoparticles. Neuroprotective Role and Molecular Bases
(MDPI, 2022) Peinado-Herreros, María Ángeles; Ovelleiro, David; del-Moral, María Luisa; Hernández-Cobo, Raquel; Martínez-Lara, Esther; Siles, Eva; Pedrajas, José Rafael; García-Martín, María Luisa; Caro, Carlos; Peralta, Sebastián; Morales-Hernández, María Encarnación; Ruiz-Martínez, María Adolfina; Blanco-Ruiz, Santos
Exogenous neuroprotective protein neuroglobin (Ngb) cannot cross the blood–brain barrier. To overcome this difficulty, we synthesized hyaluronate nanoparticles (NPs), able to deliver Ngb into the brain in an animal model of stroke (MCAO). These NPs effectively reached neurons, and were microscopically identified after 24 h of reperfusion. Compared to MCAO non-treated animals, those treated with Ngb-NPs showed survival rates up to 50% higher, and better neurological scores. Tissue damage improved with the treatment, but no changes in the infarct volume or in the oxidative/nitrosative values were detected. A proteomics approach (p-value < 0.02; fold change = 0.05) in the infarcted areas showed a total of 219 proteins that significantly changed their expression after stroke and treatment with Ngb-NPs. Of special interest, are proteins such as FBXO7 and NTRK2, which were downexpressed in stroke, but overexpressed after treatment with Ngb-NPs; and ATX2L, which was overexpressed only under the effect of Ngb. Interestingly, the proteins affected by the treatment with Ngb were involved in mitochondrial function and cell death, endocytosis, protein metabolism, cytoskeletal remodeling, or synaptic function, and in regenerative processes, such as dendritogenesis, neuritogenesis, or sinaptogenesis. Consequently, our pharmaceutical preparation may open new therapeutic scopes for stroke and possibly for other neurodegenerative pathologies.
Comparative proteomic study of early hypoxic response in the cerebral cortex of rats submitted to two different hypoxic models
(Wiley - V C H Verlag GmbH, 2017) Ovelleiro, David; Blanco-Ruiz, Santos; Hernández-Cobo, Raquel; Peinado-Herreros, María Ángeles
Purpose: The present study analyses and compares the cortical brain proteomic profiles of two different models of cerebral hypoxic insult in rats (HH: hypobaric hypoxia and HHI: ischemia followed by hypobaric hypoxia) in an attempt to describe the alterations of the early molecular hypoxic adaptive response underlying each one. Experimental Design: A quantitative proteomic profile of left-brain cortices of rats under HH, HHI, and control conditions was determined using isobaric labeling (Tandem Mass TagTM) on the protein extracts from pools of five individuals. Data are available via ProteomeXchange with identifier PXD004091. Results: Altogether, 339 proteins were confidently quantified, 99 of them showing significant variations in the hypoxic conditions with respect to the control. The HHI model presents a global effect of protein downregulation while HH produces an overall increase of the protein levels. While HH mainly affecting oxidative and energetic metabolism, HHI also interferes with synaptic transmission, neurotransmitter secretion, substantia nigra development, and triggers apoptosis through mitochondrial pathway. Conclusions and Clinical Relevance: The findings obtained show an overview of protein alterations under two hypoxic models of different aetiology and provide a basis for more detailed studies in order to unravel new specific mechanisms and therapies for hypoxic pathologies.
Constitutive nitric oxide synthases are responsible for the nitric oxide production in the ischemic aged cerebral cortex
(Elsevier BV, 2005) Martínez-Lara, Esther; Cañuelo, Ana; Siles, Eva; Hernández-Cobo, Raquel; del-Moral, María Luisa; Blanco-Ruiz, Santos; Pedrosa-Raya, Juan ángel; Rodrigo, José; Peinado-Herreros, María Ángeles
Aged brain shows reduced biological plasticity to meet emergency conditions such as ischemia, a process in which nitric oxide (NO) and apoptosis have been shown to play important roles. Using a model of transient global ischemia, we have analyzed the NO system and the p53, bax and bcl-2 response in the cerebral cortex of aged rats. Although immediately after ischemia the NO level is maintained, the reperfusion period increases NO concentrations together with the following: (i) greater bulk-protein nitration mainly due to a 50-kDa immunoreactive band; (ii) an increase in p53 protein; and (iii) an up-regulation of Bax together with a down-regulation of Bcl-2. These results match up with induced endothelial nitric oxide synthase expression immediately after ischemia and in neuronal nitric oxide synthase with the reperfusion. However, inducible nitric oxide synthase was not altered with ischemia/reperfusion. Altogether, these data suggest that NO production in cerebral cortex of aged ischemic animals is due to the constitutive NO synthase isoforms. This response is accompanied by the increased expression of pro-apoptotic proteins.
Crosstalk between hydroxytyrosol, a major olive oil phenol, and HIF-1 in MCF-7 breast cancer cells
(Nature Research, 2020) Calahorra, Jesús; Martínez-Lara, Esther; Granadino-Roldán, José Manuel; Martí, Juan Manuel; Cañuelo, Ana; Blanco-Ruiz, Santos; Oliver, Francisco Javier; Siles, Eva
Olive oil intake has been linked with a lower incidence of breast cancer. Hypoxic microenvironment in solid tumors, such as breast cancer, is known to play a crucial role in cancer progression and in the failure of anticancer treatments. HIF-1 is the foremost effector in hypoxic response, and given that hydroxytyrosol (HT) is one of the main bioactive compounds in olive oil, in this study we deepen into its modulatory role on HIF-1. Our results in MCF-7 breast cancer cells demonstrate that HT decreases HIF-1α protein, probably by downregulating oxidative stress and by inhibiting the PI3K/Akt/mTOR pathway. Strikingly, the expression of HIF-1 target genes does not show a parallel decrease. Particularly, adrenomedullin and vascular endothelial growth factor are up-regulated by high concentrations of HT even in HIF-1α silenced cells, pointing to HIF-1-independent mechanisms of regulation. In fact, we show, by in silico modelling and transcriptional analysis, that high doses of HT may act as an agonist of the aryl hydrocarbon receptor favoring the induction of these angiogenic genes. In conclusion, we suggest that the effect of HT in a hypoxic environment is largely affected by its concentration and involves both HIF-1 dependent and independent mechanisms.
Effects of a human milk oligosaccharide, 2′-fucosyllactose, on hippocampal long-term potentiation and learning capabilities in rodents
(Elsevier Science Inc, 2015) Vázquez, Enrique; Barranco, Alejandro; Ramírez, María; Gruart, Agnes; Delgado-García, José María; Martínez-Lara, Esther; Blanco-Ruiz, Santos; Martín-Martín, María Jesús; Castanys, Esther; Buck, Rachael; Prieto, Pedro; Rueda, Ricardo
Human milk oligosaccharides (HMOs) are unique with regard to their diversity, quantity and complexity, particularly in comparison to bovine milk oligosaccharides. HMOs are associated with functional development during early life, mainly related to immunity and intestinal health. Whether HMOs elicit a positive effect on cognitive capabilities of lactating infants remains an open question. This study evaluated the role of the most abundant HMO, 2′ fucosyllactose (2′-FL), in synaptic plasticity and learning capabilities in rodents. Mice and rats were prepared for the chronic recording of field excitatory postsynaptic potentials evoked at the hippocampal CA3–CA1 synapse. Following chronic oral administration of 2′-FL, both species showed improvements in input/output curves and in long-term potentiation (LTP) evoked experimentally in alert behaving animals. This effect on LTP was related to better performance of animals in various types of learning behavioral tests. Mice were tested for spatial learning, working memory and operant conditioning using the IntelliCage system, while rats were submitted to a fixed-ratio schedule in the Skinner box. In both cases, 2′-FL-treated animals performed significantly better than controls. In addition, chronic administration of 2′-FL increased the expression of different molecules involved in the storage of newly acquired memories, such as the postsynaptic density protein 95, phosphorylated calcium/calmodulin-dependent kinase II and brain-derived neurotrophic factor in cortical and subcortical structures. Taken together, the data show that dietary 2′-FL affects cognitive domains and improves learning and memory in rodents.
Glutathione S-transferase isoenzymatic response to aging in rat cerebral cortex and cerebellum
(Elsevier Inc., 2003) Martínez-Lara, Esther; Siles, Eva; Hernández-Cobo, Raquel; Cañuelo, Ana; del-Moral, María Luisa; Jiménez, Ana; Blanco-Ruiz, Santos; López-Ramos, Juan Carlos; Esteban-Ruiz, Francisco José; Pedrosa-Raya, Juan Ángel; Peinado-Herreros, María Ángeles
Aging is associated with increased oxidant generation. One mechanism involved in the defense of oxidative products is the family of glutathione transferases (GST). We have analyzed the activity, distribution and expression of GSTP1 and GSTA4 isoenzymes in the cerebral cortex and cerebellum of young, adult and aged rats. The total GST activity, measured with the universal substrate 1-chloro-2,4-dinitrobenzene (CDNB), increased only with the maturation process; however GSTA4 activity, using the specific substrate 4-hydroxynonenal (HNE), did show an age-dependent increase in both brain regions. Cellular location of GSTA4 in astrocytes was not changed except for young cerebral cortex and adult/aged cerebellum that also showed immunoreactivity in layer III pyramidal neurons and Bergman radial glia, respectively. Distribution of GSTP1 was similar among groups and only an increased number of positive oligodendrocytes was found in the Purkinje and granular layer of adult/aged cerebellum. The GSTA4 and GSTP1 expression increased from young to adult/aged brain and GSTA4 even augmented in the aged cerebral cortex. These results suggest a GST isoenzymatic response with aging, but above all with the maturation process.
Hyaluronate Nanoparticles as a Delivery System to Carry Neuroglobin to the Brain after Stroke
(MDPI, 2020) Blanco-Ruiz, Santos; Peralta, Sebastián; Morales-Hernández, María Encarnación; Martínez-Lara, Esther; Pedrajas, José Rafael; Castán, Herminia; Peinado-Herreros, María Ángeles; Ruiz-Martínez, María Adolfina
Therapies against stroke can restore the blood supply but cannot prevent the ischemic damage nor stimulate the recovery of the infarcted zone. The neuroglobin protein plays an important role in the neuro-regeneration process after stroke; however, the method for its effective systemic application has not been identified yet, as neuroglobin is unable to pass through the blood-brain barrier. Previously, we developed different types of sodium hyaluronate nanoparticles, which successfully cross the blood-brain barrier after stroke. In this work, these nanoparticles have been used to carry neuroglobin through the bloodstream to the nerve cells in rats submitted to stroke. We have biosynthesized rat-recombinant neuroglobin and determined the formulation of sodium hyaluronate nanoparticles loaded with neuroglobin, as well as its size and ζ-potential, encapsulation efficiently, in vitro release, and its kinetic of liberation. The results show that the formulation achieved is highly compatible with pharmaceutical use and may act as a delivery system to transport neuroglobin within the blood. We have found that this formulation injected intravenously immediately after stroke reached the damaged cerebral parenchyma at early stages (2 h). Neuroglobin colocalizes with its nanocarriers inside the nerve cells and remains after 24 h of reperfusion. In conclusion, the systemic administration of neuroglobin linked to nanoparticles is a potential neuroprotective drug-delivery strategy after stroke episodes.
Hypobaric hypoxia and reoxygenation induce proteomic profile changes in the rat brain cortex
(Humana Press Inc, 2013) Hernández-Cobo, Raquel; Blanco-Ruiz, Santos; Peragón, Juan; Pedrosa-Raya, Juan Ángel; Peinado-Herreros, María Ángeles
Brain, due to its high metabolism, is severely affected by hypoxia/reoxygenation. In this study, cerebral cortexes from rats subjected to hypobaric hypoxia followed by several reoxygenation periods (0 h, 24 h, and 5 days) were compared with normobaric normoxic controls to identify protein-expression differences using proteomic approaches. Only 2-fold differences in spot abundance between controls and experimental groups from each reoxygenation period were considered. The proteins identified were grouped into categories, according to their similarity in function or to their involvement in the same metabolic pathway. We distinguished five groups: (1) glycolysis, including c-enolase (NSE), and glyceraldehyde 3-phosphate dehydrogenase (GAPDH); (2) tricarboxylic acid cycle, such as aconitate hydratase (ACO2); (3) oxidative phosphorylation, like F1-ATPase chains a and b; (4) cytoskeletal, including Spna2, a-tubulin, b-tubulin, b-actin, and microtubule-associated protein RP/EB family member 3 (EB3); and (5) chaperones, like heat-shock protein 72 kDa (HSP72). NSE was upregulated while GAPDH was downregulated, both peaking at 5 days post-hypoxia. ACO2 and F1-ATPase decreased in all the reoxygenation periods. Spna2 and EB3 were expressed neither in control nor at 0 h, but 5 days post-hypoxia new expression took place. The a- and b-tubulin levels significantly fell at 0 h, but after 24 h strongly increased. Also, b-actin and HSP72 were downregulated, and the last one reached the lowest level at 24 h of reoxygenation. We conclude that the molecular mechanisms underlying hypoxia/reoxygenation in the rat cortex might consist of a close relationship between energy metabolism, cytoskeleton, and chaperones. These findings may shed light on therapeutic targets against hypoxia-related damage.
Immunohistochemistry of neuronal nitric oxide synthase and protein nitration in the striatum of the aged rat
(John Wiley & Sons, Inc., 2004) del-Moral, María Luisa; Esteban-Ruiz, Francisco José; Hernández-Cobo, Raquel; Blanco-Ruiz, Santos; Molina-Ortega, Francisco Javier; Martínez-Lara, Esther; Siles, Eva; Viedma, Gloria; Ruiz, Alharilla; Pedrosa-Raya, Juan Ángel; Peinado-Herreros, María Ángeles
To ascertain the possible implications of the nitric oxide (NO● ) producing system in striatal senescence, and by using immunohistochemistry and image-processing approaches, we describe the presence of the enzyme nitric oxide synthase (NOS), the NADPH-diaphorase (NADPH-d) histochemical marker, and nitrotyrosine-derived complexes (N-Tyr) in the striatum of adult and aged rats. The results showed neuronal NOS immunoreactive (nNOS-IR) aspiny medium-sized neurons and nervous fibres in both age groups, with no variation in the percentage of immunoreactive area but a significant decrease in the intensity and in the number of somata with age, which were not related to the observed increase with age of the striatal bundles of the white matter. In addition, NADPH-d activity was detected in neurons with morphology similar to that of the nNOS-IR cells; a decrease in the percentage of area per field and in the number of cells, but an increase in the intensity of staining for the NADPH-d histochemical marker, were detected with age. The number of neuronal NADPH-d somata was higher than for the nNOS-IR ones in both age groups. Moreover, N-Tyr-IR complexes were observed in cells (neurons and glia) and fibres, with a significant increase in the percentage of the area of immunoreaction, related to the increase of white matter, but a decrease in intensity for the aged group. On the other hand, we did not detect the inducible isoform (iNOS) either in adult or in aged rats. Taken together, these results support the contention that NADPH-d staining is not such an unambiguous marker for nNOS, and that increased protein nitration may participate in striatal aging.
Is endothelial-nitric-oxide-synthase-derived nitric oxide involved in cardiac hypoxia/reoxygenation-related damage?
(Indian Academy of Sciences, 2011) Rus-Martínez, Alma; Peinado-Herreros, María Ángeles; Blanco-Ruiz, Santos; del-Moral, María Luisa
Nitric oxide (NO) has been reported to act both as a destructive and a protective agent in the pathogenesis of the injuries that occur during hypoxia/reoxygenation (H/R). It has been suggested that this dual role of NO depends directly on the isoform of NO synthase (NOS) involved. In this work, we investigate the role that NO derived from endothelial NOS (eNOS) plays in cardiac H/R-induced injury. Wistar rats were submitted to H/R (hypoxia for 30 min; reoxygenation of 0 h, 12 h and 5 days), with or without prior treatment using the selective eNOS inhibitor L-NIO (20 mg/kg). Lipid peroxidation, apoptosis and protein nitration, as well as NO production (NOx), were analysed. The results showed that L-NIO administration lowered NOx levels in all the experimental groups. However, no change was found in the lipid peroxidation level, the percentage of apoptotic cells or nitrated protein expression, implying that eNOS-derived NO may not be involved in the injuries occurring during H/R in the heart. We conclude that LNIO would not be useful in alleviating the adverse effects of cardiac H/R.
Lipid nanoparticles for the transport of drugs like dopamine through the blood–brain barrier
(Springer, 2021) Ortega-Escandell, Elena; Blanco-Ruiz, Santos; Ruiz-Martínez, María Adolfina; Peinado-Herreros, María Ángeles; Peralta, Sebastián; Morales-Hernández, María Encarnación
Diseases and disorders of the nervous system, like Parkinson disease (PD) and other neurodegenerative pathologies, are widespread in our society. The arsenal of treatments against these pathologies continues to increase, but in many cases, its use is limited. This is due to the blood-brain barrier (BBB), which acts by limiting the penetration of drugs into the brain. To overcome this handicap, in the current research, solid lipid nanoparticles (SLNPs) able to encapsulate drugs and to cross the blood-brain barrier have been designed to transport and release these drugs into their targets. These SLNPs were synthesized by a sonication method and high agitation process searching the most adequate physicochemical profile to achieve the objectives set. Today, the most efficient treatment for PD consists of providing the dopamine (DP) that is lost by neurodegeneration; however, the nature of this neurotransmitter prevents its crossing of the BBB. Therefore, DP may be considered as a good candidate to be encapsulated in SLNPs while studying how the loading drug could affect such nanoparticles. Based on these antecedents, in this research, both empty and DP-charged SLNPs were characterized physicochemically. The results obtained indicated a great stability of the nanoparticles loaded with DP when drug was used at 0.2 to 0.05%; these concentrations barely affected its size, polydispersity, and ζ-potential, and the SLNPs elaborated in this research were high appropriate to be injected systemically. Finally, empty SLNPs labeled and administered systemically to adult male Wistar rats demonstrate their penetration ability into the brain parenchyma.
Melatonin influences NO/NOS pathway and reduces oxidative and nitrosative stress in a model of hypoxic-ischemic brain damage
(Academic Press Inc Elsevier Science, 2017) Blanco-Ruiz, Santos; Hernández-Cobo, Raquel; Franchelli, Gustavo; Ramos-Álvarez, Manuel Miguel; Peinado-Herreros, María Ángeles
In this work, using a rat model combining ischemia and hypobaric hypoxia (IH), we evaluate the relationships between the antioxidant melatonin and the cerebral nitric oxide/nitric oxide synthase (NO/ NOS) system seeking to ascertain whether melatonin exerts its antioxidant protective action by balancing this key pathway, which is highly involved in the cerebral oxidative and nitrosative damage underlying these pathologies. The application of the IH model increases the expression of the three nitric oxide synthase (NOS) isoforms, as well as nitrogen oxide (NOx) levels and nitrotyrosine (n-Tyr) impacts on the cerebral cortex. However, melatonin administration before IH makes nNOS expression response earlier and stronger, but diminishes iNOS and n-Tyr expression, while both eNOS and NOx remain unchanged. These results were corroborated by nicotine adenine dinucleotide phosphate diaphorase (NADPH-d) staining, as indicative of in situ NOS activity. In addition, the rats previously treated with melatonin exhibited a reduction in the oxidative impact evaluated by thiobarbituric acid reactive substances (TBARS). Finally, IH also intensified glial fibrillary acidic protein (GFAP) expression, reduced hypoxia-inducible factor-1alpha (HIF-1a), but did not change nuclear factor kappa B (NF-kB); meanwhile, melatonin did not significantly affect any of these patterns after the application of the IH model. The antioxidant melatonin acts on the NO/NOS system after IH injury balancing the release of NO, reducing peroxynitrite formation and protecting from nitrosative/oxidative damage. In addition, this paper raises questions concerning the classical role of some controversial molecules such as NO, which are of great consequence in the final fate of hypoxic neurons. We conclude that melatonin protects the brain from hypoxic/ischemic-derived damage in the first steps of the ischemic cascade, influencing the NO/NOS pathway and reducing oxidative and nitrosative stress.
New Strategies for Stroke Therapy: Nanoencapsulated Neuroglobin
(MDPI, 2022) Blanco-Ruiz, Santos; Martínez-Lara, Esther; Siles, Eva; Peinado-Herreros, María Ángeles
Stroke is a global health and socio-economic problem. However, no efficient preventive and/or palliative treatments have yet been found. Neuroglobin (Ngb) is an endogen neuroprotective protein, but it only exerts its beneficial action against stroke after increasing its basal levels. Therefore, its systemic administration appears to be an efficient therapy applicable to stroke and other neurodegenerative pathologies. Unfortunately, Ngb cannot cross the blood-brain barrier (BBB), making its direct pharmacological use unfeasible. Thus, the association of Ngb with a drug delivery system (DDS), such as nanoparticles (NPs), appears to be a good strategy for overcoming this handicap. NPs are a type of DDS which efficiently transport Ngb and increase its bioavailability in the infarcted area. Hence, we previously built hyaluronate NPS linked to Ngb (Ngb-NPs) as a therapeutic tool against stroke. This nanoformulation induced an improvement of the cerebral infarct prognosis. However, this innovative therapy is still in development, and a more in-depth study focusing on its long-lasting neuroprotectant and neuroregenerative capabilities is needed. In short, this review aims to update the state-of-the-art of stroke therapies based on Ngb, paying special attention to the use of nanotechnological drug-delivering tools.
Steatosis recovery after treatment with a balanced sunflower or olive oil-based diet: involvement of perisinusoidal stellate cells
(Baishideng Publishing Group Co., Limited, 2005) Hernández-Cobo, Raquel; Martínez-Lara, Esthe; Cañuelo, Ana; del-Moral, María Luisa; Blanco-Ruiz, Santos; Siles, Eva; Jiménez, Ana; Pedrosa-Raya, Juan Ángel; Peinado-Herreros, María Ángeles
AIM: To analyze the relationship between perisinusoidal stellate cell (PSC) activation and the dietary fat quantity and composition in the treatment of hepatic steatosis. METHODS: Using an experimental rat model of steatosis based on the intake of a hyperlipidic diet (14% fat as olive oil or sunfl ower oil, HL-O and HL-S, respectively), we analyzed the liver’s capability of recovery after the treatment with a normal-lipidic diet (5% fat as olive oil or sunfl ower oil, NL-O and NL-S, respectively) by immunocytochemical and Western blot analysis of glial fi brillary acidic protein (GFAP) expression in PSCs, collagen quantification and serum aminotransferase determination. RESULTS: The fatty infi ltration in the steatotic livers decreased after the treatment with both NL diets, indicating liver recovery. This decrease was accompanied with a lower collagen deposition and aminotransferase level as well as changes in the PSC population that increased the GFAP expression. The above-mentioned effects were more pronounced in animals fed on NL-O based diet. CONCLUSION: Treatment with a balanced diet enriched in olive oil contributes to the liver recovery from a steatotic process. The PSC phenotype is a marker of this hepatic-recovery model.
Study of the nitric oxide system in the rat cerebellum during aging
(BioMed Central Ltd, 2010) Blanco-Ruiz, Santos; Molina-Ortega, Francisco Javier; Castro-Ortega, Lourdes; del-Moral, María Luisa; Hernández-Cobo, Raquel; Jiménez, Ana; Rus-Martínez, Alma; Martínez-Lara, Esther; Siles, Eva; Peinado-Herreros, María Ángeles
Background: The cerebellum is the neural structure with the highest levels of nitric oxide, a neurotransmitter that has been proposed to play a key role in the brain aging, although knowledge concerning its contribution to cerebellar senescence is still unclear, due mainly to absence of integrative studies that jointly evaluate the main factors involved in its cell production and function. Consequently, in the present study, we investigate the expression, location, and activity of nitric oxide synthase isoenzymes; the protein nitration; and the production of nitric oxide in the cerebellum of adult and old rats. Results: Our results show no variation in the expression of nitric oxide synthase isoforms with aging, although, we have detected some changes in the cellular distribution pattern of the inducible isoform particularly in the cerebellar nuclei. There is also an increase in nitric oxide synthase activity, as well as greater protein-nitration levels, and maintenance of nitrogen oxides (NOx) levels in the senescent cerebellum. Conclusions: The nitric oxide/nitric oxide syntahses system suffers from a number of changes, mainly in the inducible nitric oxide synthase distribution and in overall nitric oxide synthases activity in the senescent cerebellum, which result in an increase of the protein nitration. These changes might be related to the oxidative damage detected with aging in the cerebellum.
Synthesis and characterization of different sodium hyaluronate nanoparticles to transport large neurotherapheutic molecules through blood brain barrier after stroke
(Pergamon-Elsevier Science LTD, 2019) Peralta, Sebastián; Blanco-Ruiz, Santos; Hernández-Cobo, Raquel; Castán, Herminia; Siles, Eva; Martínez-Lara, Esther; Morales-Hernández, María Encarnación; Peinado-Herreros, María Ángeles; Ruiz-Martínez, María Adolfina
Some biological drugs with proven neuroprotective capacity are unable to cross the blood brain barrier (BBB), preventing its use in neuroregenerative diseases such as stroke. The use of nanoparticles as a delivery system to transport large therapeutic molecules to the cerebral parenchyma may be a good option to overcome this limitation. To achieve this goal, we have designed some polymer nanoparticles (NPs) by two ionic gelation methods of synthesis: external (M1) and internal (M2), both using sodium hyaluronate (SH) as polymer but with differences in the elaboration of their core. Additionally, both SH-NPs were coated with chitosan and glycerol tripalmitin in order to improve their penetration capabilities into cells. The nanoparticles were characterized by size, shape and charge. Then, an experimental approach was carried out in animals submitted to a stroke model, where NPs penetration into the brain was studied and analysed after its systemic administration. All types of NPs assayed were able to cross the BBB and were endocytosed by neurons; however, the SH-NPs obtained by M2 are lightly more efficient in the rate of penetration than those obtained by M1. There were not visible differences between coated and non-coated NPs obtained by both gelation methods. This may be due to the fact that not only the size, shape and charge of NPs, but also its chemical structure influences its cellular capture by endocytic mechanisms.
Upregulation of endothelial nitric oxide synthase maintains nitric oxide production in the cerebellum of thioacetamide cirrhotic rats
(Pergamon Press, 2004) Hernández-Cobo, Raquel; Martínez-Lara, Esther; del-Moral, María Luisa; Blanco-Ruiz, Santos; Cañuelo, Ana; Siles, Eva; Esteban-Ruiz, Francisco José; Pedrosa-Raya, Juan Ángel; Peinado-Herreros, María Ángeles
This study examines the expression and cellular distribution pattern of nitric oxide synthase (NOS) isoforms, nitrotyrosine-derived complexes, and the nitric oxide (NO) production in the cerebellum of rats with cirrhosis induced by thioacetamide (TAA). The results showed local changes in the tissue distribution pattern of the NOS isoforms and nitrated proteins in the cerebellum of these animals. Particularly, eNOS immunoreactivity in perivascular glial cells of the white matter was detected only in TAA-treated animals. In addition, although neither neuronal NOS (nNOS) nor inducible NOS (iNOS) cerebellar protein levels appeared to be affected, the endothelial NOS (eNOS) isoform significantly increased its expression, and NO production slightly augmented in TAA-treated rats. These NOS/NO changes may contribute differently to the evolution of the hepatic disease either by maintaining the guanosine monophosphate–NO signal transduction pathways and the physiological cerebellar functions or by inducing oxidative stress and cell damage. This model gives rise to the hypothesis that the upregulation of the eNOS maintains the physiological production of NO, while the iNOS is silenced and the nNOS remains unchanged. The differential NOS-distribution and expression pattern may be one of the mechanisms involved to balance cerebellar NO production in order to minimize TAA toxic injury. These data help elucidate the role of the NOS/NO system in the development and progress of hepatic encephalopathy associated with TAA cirrhosis.

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Examinando por Autor "Blanco-Ruiz, Santos"