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Inflammatory bowel disease (IBD) incidence has increased in the last decades due to changes in dietary habits. IBDs are characterized by intestinal epithelial barrier disruption, increased inflammatory mediator production and excessive tissue injury. Since the current treatments are not sufficient to achieve and maintain remission, complementary and alternative medicine (CAM) becomes a primary practice as a co-adjuvant for the therapy. Thus, the intake of functional food enriched in vegetal extracts represents a promising nutritional strategy. This study evaluates the anti-inflammatory effects of artichoke, caihua and fenugreek vegetal extract original blend (ACFB) in an in vitro model of gut barrier mimicking the early acute phases of the disease. Caco2 cells cultured on transwell supports were treated with digested ACFB before exposure to pro-inflammatory cytokines. The pre-treatment counteracts the increase in barrier permeability induced by the inflammatory stimulus, as demonstrated by the evaluation of TEER and CLDN-2 parameters. In parallel, ACFB reduces p65NF-κB pro-inflammatory pathway activation that results in the decrement of COX-2 expression as PGE2 and IL-8 secretion. ACFB properties might be due to the synergistic effects of different flavonoids, indicating it as a valid candidate for new formulation in the prevention/mitigation of non-communicable diseases.
Assuntos
Flavonoides , NF-kappa B , Extratos Vegetais , Humanos , Células CACO-2 , Extratos Vegetais/farmacologia , Flavonoides/farmacologia , NF-kappa B/metabolismo , Doenças Inflamatórias Intestinais/metabolismo , Doenças Inflamatórias Intestinais/tratamento farmacológico , Interleucina-8/metabolismo , Transdução de Sinais/efeitos dos fármacos , Anti-Inflamatórios/farmacologia , Ciclo-Oxigenase 2/metabolismo , Mucosa Intestinal/metabolismo , Mucosa Intestinal/efeitos dos fármacos , Trigonella/química , Dinoprostona/metabolismoRESUMO
The transition to higher education at University is a critical moment for young adults to acquire unhealthy habits regarding physical activity (PA) and adherence to a healthy diet. Negative behaviors might be maintained in the years to come with a major risk of suffering from a Non-Communicable Disease. This study aims to determine the relationship between diet and PA in the student community of University of Milano-Bicocca. Students between 18 and 30 years old completed an online survey (6949 students). Two analyses of covariance (ANCOVA), chi-square tests of independence, and a binomial logistic regression were performed to examine the relationship between adequacy of food consumption and PA, in association also with sociodemographic characteristics. Data show a strong correlation between behaviors analyzed, with a proportional positive association between PA and healthy diet. Nevertheless, a third of the sample students incur in incorrect habits for both diet and PA. Further, students performing intensive PA have the healthiest food consumption in general but the worst red meat and pork intake. Accordingly, men practice more PA but have a less adequate diet, exactly contrary to women. In conclusion, policies promoting consciousness of well-being would transform Universities into healthy hubs for virtuous habits.
Assuntos
Exercício Físico , Comportamento Alimentar , Estudantes , Humanos , Estudantes/estatística & dados numéricos , Estudantes/psicologia , Masculino , Feminino , Universidades , Adulto Jovem , Adulto , Adolescente , Estilo de Vida , Itália , Dieta , Inquéritos e Questionários , Comportamentos Relacionados com a SaúdeRESUMO
Epigenetic changes, host-gut microbiota interactions, and environmental factors contribute to inflammatory bowel disease (IBD) onset and progression. A healthy lifestyle may help to slow down the chronic or remitting/relapsing intestinal tract inflammation characteristic of IBD. In this scenario, the employment of a nutritional strategy to prevent the onset or supplement disease therapies included functional food consumption. Its formulation consists of the addition of a phytoextract enriched in bioactive molecules. A good candidate as an ingredient is the Cinnamon verum aqueous extract. Indeed, this extract, subjected to a process of gastrointestinal digestion simulation (INFOGEST), exhibits beneficial antioxidant and anti-inflammatory properties in an in vitro model of the inflamed intestinal barrier. Here, we deepen the study of the mechanisms related to the effect of digested cinnamon extract pre-treatment, showing a correlation between transepithelial electrical resistance (TEER) decrement and alterations in claudin-2 expression under Tumor necrosis factor-α/Interleukin-1ß (TNF-α/IL-1) ß cytokine administration. Our results show that pre-treatment with cinnamon extract prevents TEER loss by claudin-2 protein level regulation, influencing both gene transcription and autophagy-mediated degradation. Hence, cinnamon polyphenols and their metabolites probably work as mediators in gene regulation and receptor/pathway activation, leading to an adaptive response against renewed insults.
Assuntos
Cinnamomum zeylanicum , Doenças Inflamatórias Intestinais , Humanos , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/metabolismo , Claudina-2 , Interleucina-1beta/genética , Casca de Planta/metabolismo , Doenças Inflamatórias Intestinais/tratamento farmacológico , Doenças Inflamatórias Intestinais/metabolismo , Expressão GênicaRESUMO
Particulate matter (PM) is a harmful component of urban air pollution and PM2.5, in particular, can settle in the deep airways. The RAS system plays a crucial role in the pathogenesis of pollution-induced inflammatory diseases: the ACE/AngII/AT1 axis activates a pro-inflammatory pathway counteracted by the ACE2/Ang(1-7)/MAS axis, which in turn triggers an anti-inflammatory and protective pathway. However, ACE2 acts also as a receptor through which SARS-CoV-2 penetrates host cells to replicate. COX-2, HO-1, and iNOS are other crucial proteins involved in ultrafine particles (UFP)-induced inflammation and oxidative stress, but closely related to the course of the COVID-19 disease. BALB/c male mice were subjected to PM2.5 sub-acute exposure to study its effects on ACE2 and ACE, COX-2, HO-1 and iNOS proteins levels, in the main organs concerned with the pathogenesis of COVID-19. The results obtained show that sub-acute exposure to PM2.5 induces organ-specific modifications which might predispose to greater susceptibility to severe symptomatology in the case of SARS-CoV-2 infection. The novelty of this work consists in using a molecular study, carried out in the lung but also in the main organs involved in the disease, to analyze the close relationship between exposure to pollution and the pathogenesis of COVID-19.
Assuntos
COVID-19 , Animais , Humanos , Masculino , Camundongos , Enzima de Conversão de Angiotensina 2 , Ciclo-Oxigenase 2 , Pandemias , Material Particulado , Peptidil Dipeptidase A/metabolismo , SARS-CoV-2RESUMO
Chronic noncommunicable diseases (NCDs) have become the major cause of morbidity and mortality in the Maldives, triggered by the nutrition transition to a "Western diet" that dramatically increases the prevalence of excess weight and hypertension. Our study aimed to evaluate dietary habits, blood pressure (BP) and body mass index in Maghoodoo Public School's students. A sample of 145 students (72 males and 73 females, age 9.37 ± 2.97 years) was enrolled. Factors causing excess weight were investigated through descriptive statistics. The relationship between blood pressure percentiles and possible influencing factors was investigated by a linear regression model.. Excess weight was present in 15.07% and 12.5% females and males, respectively. 15.18% of the subjects had elevated BP, with a significant difference according to gender detected only in the PAS z-score. Eating habits were investigated through a parent-filled questionnaire; 70.15% of the students consumed less than two portions of fruit per day, with a significant difference between gender (84.06% and 55.38% for boys and girls, respectively, p < 0.0001) and 71.64% ate less than two servings of vegetables per day. An alarming finding emerged for sweet snacks (30.6% of the students consumed 2-3 servings per day) and sugary drinks (2-3 servings per day for 32.84% of students) consumption. Our findings suggest that excess weight and hypertension in this population could be due to energy-rich, packaged-foods consumption. A nutrition education approach might thus help to reduce cardiovascular risk.
Assuntos
Dieta , Hipertensão , Masculino , Feminino , Humanos , Criança , Estudos Transversais , Maldivas , Comportamento Alimentar , Verduras , Estado Nutricional , Frutas , Inquéritos e Questionários , Hipertensão/epidemiologiaRESUMO
Age-related injuries are often connected to alterations in redox homeostasis. The imbalance between free radical oxygen species and endogenous antioxidants defenses could be associated with a growing risk of transient ischemic attack and stroke. In this context, a daily supply of dietary antioxidants could counteract oxidative stress occurring during ischemia/reperfusion injury (I/R), preventing brain damage. Here we investigated the potential antioxidant properties of coffee-derived circulating metabolites and a coffee pulp phytoextract, testing their efficacy as ROS scavengers in an in vitro model of ischemia. Indeed, the coffee fruit is an important source of phenolic compounds, such as chlorogenic acids, present both in the brewed seed and in the discarded pulp. Therefore, rat brain endothelial cells, subjected to oxygen and glucose deprivation (OGD) and recovery (ogR) to mimic reperfusion, were pretreated or not with coffee by-products. The results indicate that, under OGD/ogR, the ROS accumulation was reduced by coffee by-product. Additionally, the coffee extract activated the Nrf2 antioxidant pathway via Erk and Akt kinases phosphorylation, as shown by increased Nrf2 and HO-1 protein levels. The data indicate that the daily intake of coffee by-products as a dietary food supplement represents a potential nutritional strategy to counteract aging.
Assuntos
Antioxidantes/farmacologia , Coffea/química , Fator 2 Relacionado a NF-E2/agonistas , Fenóis/farmacologia , Extratos Vegetais/farmacologia , Traumatismo por Reperfusão/terapia , Animais , Antioxidantes/química , Isquemia Encefálica/metabolismo , Isquemia Encefálica/terapia , Linhagem Celular , Fator 2 Relacionado a NF-E2/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Fenóis/química , Extratos Vegetais/química , Ratos , Traumatismo por Reperfusão/metabolismoRESUMO
The contributing role of environmental factors to the development of neurodegenerative diseases has become increasingly evident. Here, we report that exposure of C6 glioma cells to diesel exhaust particles (DEPs), a major constituent of urban air pollution, causes intracellular reactive oxygen species (ROS) production. In this scenario, we suggest employing the possible protective role that coffee phenolic metabolites may have. Coffee is a commonly consumed hot beverage and a major contributor to the dietary intake of (poly) phenols. Taking into account physiological concentrations, we analysed the effects of two different coffee phenolic metabolites mixes consisting of compounds derived from bacterial metabolization reactions or phase II conjugations, as well as caffeic acid. The results showed that these mixes were able to counteract DEP-induced oxidative stress. The cellular components mediating the downregulation of ROS included extracellular signal-regulated kinase 1/2 (ERK1/2), nuclear factor erythroid 2-related factor 2 (Nrf2), heme oxygenase-1 (HO-1), and uncoupling protein 2 (UCP2). Contrary to coffee phenolic metabolites, the treatment with N-acetylcysteine (NAC), a known antioxidant, was found to be ineffective in preventing the DEP exposure oxidant effect. These results revealed that coffee phenolic metabolites could be promising candidates to protect against some adverse health effects of daily exposure to air pollution.
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A balanced diet is a fundamental component of athletes' health, training, and performance. The majority of athletes choose adequate quantities of macronutrients but, at the same time, do not respect World Health Organization dietary guidelines, eating a lot of discretionary food and not drinking enough water. Athletes need more nutritional education to improve the quality of their food choice. By modifying their eating habits, they could also enhance their performance. Our study aimed to evaluate the impact of nutritional intervention on eating habits in a group of Northern Italian athletes. A sample of 87 athletes (41 males and 46 females) aged 16.5 ± 2.9 was enrolled. We organized meetings and detected eating habits (before and after the meetings) using a food frequencies questionnaire. We found that nutritional intervention positively affected participants consumption of vegetables (p < 0.05), nuts (p < 0.001), legumes (p < 0.001), and fish (p < 0.05). Other aspects of the athletes' eating habits were not significantly improved. Some gender differences were found; males increased their consumption of vegetables (p < 0.05) and nuts (p < 0.001), while females increased their intake of legumes (p < 0.001). Our finding suggested that nutritional intervention could promote healthy eating habits among athletes. If sports nutrition experts, coaches, personal trainers, sports medicine experts, and athletes cooperated, they could guarantee athletes' health status.
Assuntos
Ingestão de Energia , Comportamento Alimentar , Atletas , Dieta , Feminino , Preferências Alimentares , Humanos , Itália , MasculinoRESUMO
The strong spread of COVID-19 and the significant number of deaths associated with it could be related to improper lifestyles, which lead to a low-grade inflammation (LGI) that not only increases the risk of chronic diseases, but also the risk of facing complications relating to infections and a greater susceptibility to infections themselves. Recently, scientific research has widely demonstrated that the microbiota plays a fundamental role in modulating metabolic responses in the immune system. There is, in fact, a two-way interaction between lifestyle, infection, and immunity. The immune response is compromised if nutrition is unbalanced or insufficient, because diet affects the intestinal flora predisposing people to infections and, at the same time, the nutritional state can be aggravated by the immune response itself to the infection. We evaluate the link between balanced diet, the efficiency of the immune system, and microbiota with the aim of providing some practical advice for individuals, with special attention to the elderly. A correct lifestyle that follows the Mediterranean model, which is especially rich in plant-based foods along with the use of extra-virgin olive oil, are the basis of preventing LGI and other chronic pathologies, directly influencing the intestinal microbiota and consequently the immune response.
Assuntos
COVID-19 , Dieta Mediterrânea , Viroses , Idoso , Dieta , Humanos , Sistema Imunitário , Inflamação , Azeite de Oliva , SARS-CoV-2RESUMO
The aim of this study was to investigate the relationship between endothelin-1, nitric oxide, insulin resistance, and blood pressure in young subjects with a high prevalence of excess weight and/or elevated blood pressure. In a cohort of 238 children (mean age = 11.1 years), height, weight, waist circumference, and blood pressure were assessed. Body mass index, waist-to-height ratio, and blood pressure percentiles were calculated, and the children were classified as having excess weight and elevated blood pressure according to the International Obesity Task Force and the US blood pressure nomograms specific for gender, age and height, respectively. Endothelin-1 and nitric oxide production were assessed, and the homeostatic model assessment index was calculated. Forty-three percent of children were male, 71% had excess weight, and 37% had systolic and/or diastolic values above the ninetieth percentile. Plasma endothelin-1 and nitric oxide production were independently correlated (p < 0.05). In multivariate analyses, the HOMA index was associated with systolic and diastolic blood pressure (p = 0.01), and nitric oxide was independently related to diastolic blood pressure (p = 0.04), even after adjustment for measures of body composition. By using the waist-to-height ratio instead of BMI in the statistical model, the association between the homeostatic model assessment index and blood pressure was attenuated, while the results remained similar for nitric oxide. No correlation was found between endothelin-1 and blood pressure. In our study population, the correlation between nitric oxide and blood pressure and the lack of a relationship between endothelin-1 and blood pressure could be explained by an increase in the vasodilator effect of local and systemic nitric oxide, which counteracts the possible hypertensive effect of endothelin-1.
Assuntos
Pressão Sanguínea , Endotelina-1/sangue , Hipertensão/etiologia , Resistência à Insulina , Óxido Nítrico/sangue , Adolescente , Criança , Estudos Transversais , Feminino , Humanos , Hipertensão/sangue , Masculino , Sobrepeso/complicaçõesRESUMO
Over the past 60 years there has been a dramatic increase in the prevalence of overweight in children and adolescents, ranging from 4% in 1975 to 18% in 2016. Recent estimates indicate that overweight or obese children and adolescents are more than 340 million. Obesity is often associated with hypertension, which is an important cardiovascular risk factor. Recent studies show that the presence of hypertension is a frequent finding in the pediatric age. Hypertensive children easily become hypertensive adults. This phenomenon contributes to increasing cardiovascular risk in adulthood. Primary hypertension is a growing problem especially in children and adolescents of western countries, largely because of its association with the ongoing obesity epidemic. Recently, it has been hypothesized that a dietary link between obesity and elevated blood pressure (BP) values could be simple carbohydrate consumption, particularly fructose, both in adults and in children. Excessive intake of fructose leads to increased serum uric acid (SUA) and high SUA values are independently associated with the presence of hypertension and weaken the efficacy of lifestyle modifications in children. The present review intends to provide an update of existing data regarding the relationship between BP, simple carbohydrates (particularly fructose), and uric acid in pediatric age. In addition, we analyze the national policies that have been carried out over the last few years, in order to identify the best practices to limit the socio-economic impact of the effects of excessive sugar consumption in children.
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PURPOSE: Mesoporous silica nanoparticles (MSNPs) are excellent candidates for biomedical applications and drug delivery to different human body areas, the brain included. Although toxicity at cellular level has been investigated, we are still far from using MSNPs in the clinic, because the mechanisms involved in the cellular responses activated by MSNPs have not yet been elucidated. MATERIALS AND METHODS: This study used an in vitro multiparametric approach to clarify relationships among size, dose, and time of exposure of MSNPs (0.05-1 mg/mL dose range), and cellular responses by analyzing the morphology, viability, and functionality of human vascular endothelial cells and neurons. RESULTS: The results showed that 24 hours of exposure of endothelial cells to 250 nm MSNPs exerted higher toxicity in terms of mitochondrial activity and membrane integrity than 30 nm MSN at the same dose. This was due to induced cell autophagy (in particular mitophagy), probably consequent to MSNP cellular uptake (>20%). Interestingly, after 24 hours of treatment with 30 nm MSNPs, very low MSNP uptake (<1%) and an increase in nitric oxide production (30%, P<0.01) were measured. This suggests that MSNPs were able to affect endothelial functionality from outside the cells. These differences could be attributed to the different protein-corona composition of the MSNPs used, as suggested by sodium dodecyl sulfate polyacrylamide-gel electrophoresis analysis of the plasma proteins covering the MSNP surface. Moreover, doses of MSNPs up to 0.25 mg/mL perturbed network activity by increasing excitability, as detected by multielectrode-array technology, without affecting neuronal cell viability. CONCLUSION: These results suggest that MSNPs may be low-risk if prepared with a diameter <30 nm and if they reach human tissues at doses <0.25 mg/mL. These important advances could help the rational design of NPs intended for biomedical uses, demonstrating that careful toxicity evaluation is necessary before using MSNPs in patients.
Assuntos
Mitofagia/efeitos dos fármacos , Nanopartículas/toxicidade , Neurônios/efeitos dos fármacos , Autofagia/efeitos dos fármacos , Linhagem Celular , Membrana Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Sistemas de Liberação de Medicamentos , Eletroforese em Gel de Poliacrilamida , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/patologia , Enzimas/metabolismo , Células Endoteliais da Veia Umbilical Humana/efeitos dos fármacos , Humanos , Nanopartículas/administração & dosagem , Nanopartículas/química , Neurônios/patologia , Óxido Nítrico/metabolismo , Dióxido de Silício/química , Dióxido de Silício/toxicidadeRESUMO
Surface functionalization with antitransferrin receptor (TfR) mAbs has been suggested as the strategy to enhance the transfer of nanoparticles (NPs) across the blood-brain barrier (BBB) and to carry nonpermeant drugs from the blood into the brain. However, the efficiency of BBB crossing is currently too poor to be used in vivo. In the present investigation, we compared 6 different murine mAbs specific for different epitopes of the human TfR to identify the best performing one for the functionalization of NPs. For this purpose, we compared the ability of mAbs to cross an in vitro BBB model made of human brain capillary endothelial cells (hCMEC/D3). Liposomes functionalized with the best performing mAb (MYBE/4C1) were uptaken, crossed the BBB in vitro, and facilitated the BBB in vitro passage of doxorubicin, an anticancer drug, 3.9 folds more than liposomes functionalized with a nonspecific IgG, as assessed by confocal microscopy, radiochemical techniques, and fluorescence, and did not modify the cell monolayer structural or functional properties. These results show that MYBE/4C1 antihuman TfR mAb is a powerful resource for the enhancement of BBB crossing of NPs and is therefore potentially useful in the treatment of neurologic diseases and disorders including brain carcinomas.
Assuntos
Anticorpos Bloqueadores/farmacologia , Barreira Hematoencefálica/efeitos dos fármacos , Receptores da Transferrina/antagonistas & inibidores , Animais , Antibióticos Antineoplásicos/administração & dosagem , Antibióticos Antineoplásicos/farmacocinética , Capilares/efeitos dos fármacos , Capilares/metabolismo , Doxorrubicina/administração & dosagem , Doxorrubicina/farmacocinética , Sistemas de Liberação de Medicamentos , Células Endoteliais , Epitopos , Humanos , Imunoglobulina G/química , Lipossomos , Camundongos , Tamanho da PartículaRESUMO
BACKGROUND: Amyloid ß (Aß) peptide aggregation is the main molecular mechanism underlying the development of Alzheimer's disease, the most widespread form of senile dementia worldwide. Increasing evidence suggests that the key factor leading to impaired neuronal function is accumulation of water-soluble Aß oligomers rather than formation of the senile plaques created by the deposition of large fibrillary aggregates of Aß. However, several questions remain about the preliminary steps and the progression of Aß oligomerization. METHODS: We show that the initial stages of the aggregation of fluorescently labeled Aß can be determined with a high degree of precision and at physiological (i.e., nanomolar) concentrations by using either steady-state fluorimetry or time-correlated single-photon counting. RESULTS: We study the dependence of the oligomerization extent and rate on the Aß concentration. We determine the chemical binding affinity of fluorescently labeled Aß for liposomes that have been recently shown to be pharmacologically active in vivo, reducing the Aß burden within the brain. We also probe their capacity to hinder the Aß oligomerization process in vitro. CONCLUSIONS: We introduced a fluorescence assay allowing investigation of the earliest steps of Aß oligomerization, the peptide involved in Alzheimer's disease. The assay proved to be sensitive even at Aß concentrations as low as those physiologically observed in the cerebrospinal fluid. GENERAL SIGNIFICANCE: This work represents an extensive and quantitative study on the initial events of Aß oligomerization at physiological concentration. It may enhance our comprehension of the molecular mechanisms leading to Alzheimer's disease, thus paving the way to novel therapeutic strategies.
Assuntos
Peptídeos beta-Amiloides/química , Lipossomos/química , Fragmentos de Peptídeos/química , Agregação Patológica de Proteínas , Doença de Alzheimer/metabolismo , Peptídeos beta-Amiloides/metabolismo , Humanos , Fragmentos de Peptídeos/metabolismo , Espectrometria de FluorescênciaRESUMO
In the search of new drug delivery carriers for the brain, self-assembled nanoparticles (NP) were prepared from poly(N,N-dimethylacrylamide)-block-polystyrene polymer. NP displayed biocompatibility on cultured endothelial cells, macrophages and differentiated SH-SY5Y neuronal-like cells. The surface-functionalization of NP with a modified fragment of human Apolipoprotein E (mApoE) enhanced the uptake of NP by cultured human brain capillary endothelial cells, as assessed by confocal microscopy, and their permeability through a Transwell Blood Brain Barrier model made with the same cells, as assessed by fluorescence. Finally, mApoE-NP embedding doxorubicin displayed an enhanced release of drug at low pH, suggesting the potential use of these NP for the treatment of brain tumors.
Assuntos
Acrilamidas/química , Apolipoproteínas E , Barreira Hematoencefálica/metabolismo , Doxorrubicina , Portadores de Fármacos , Células Endoteliais da Veia Umbilical Humana/metabolismo , Nanopartículas/química , Poliestirenos/química , Apolipoproteínas E/química , Apolipoproteínas E/farmacocinética , Apolipoproteínas E/farmacologia , Linhagem Celular , Doxorrubicina/química , Doxorrubicina/farmacocinética , Doxorrubicina/farmacologia , Portadores de Fármacos/química , Portadores de Fármacos/farmacocinética , Portadores de Fármacos/farmacologia , HumanosRESUMO
Targeting amyloid-ß peptide (Aß) within the brain is a strategy actively sought for therapy of Alzheimer's disease (AD). We investigated the ability of liposomes bi-functionalized with phosphatidic acid and with a modified ApoE-derived peptide (mApoE-PA-LIP) to affect Aß aggregation/disaggregation features and to cross in vitro and in vivo the blood-brain barrier (BBB). Surface plasmon resonance showed that bi-functionalized liposomes strongly bind Aß (kD=0.6 µM), while Thioflavin-T and SDS-PAGE/WB assays show that liposomes inhibit peptide aggregation (70% inhibition after 72 h) and trigger the disaggregation of preformed aggregates (60% decrease after 120 h incubation). Moreover, experiments with dually radiolabelled LIP suggest that bi-functionalization enhances the passage of radioactivity across the BBB either in vitro (permeability=2.5×10(-5) cm/min, 5-fold higher with respect to mono-functionalized liposomes) or in vivo in healthy mice. Taken together, our results suggest that mApoE-PA-LIP are valuable nanodevices with a potential applicability in vivo for the treatment of AD. From the clinical editor: Bi-functionalized liposomes with phosphatidic acid and a modified ApoE-derived peptide were demonstrated to influence Aß aggregation/disaggregation as a potential treatment in an Alzheimer's model. The liposomes were able to cross the blood-brain barrier in vitro and in vivo. Similar liposomes may become clinically valuable nanodevices with a potential applicability for the treatment of Alzheimer's disease.
Assuntos
Doença de Alzheimer/terapia , Apolipoproteínas E/química , Barreira Hematoencefálica , Lipossomos , Peptídeos/química , Ácidos Fosfatídicos/química , Apolipoproteínas E/administração & dosagem , Western Blotting , Eletroforese em Gel de Poliacrilamida , Humanos , Ácidos Fosfatídicos/administração & dosagem , Ressonância de Plasmônio de SuperfícieRESUMO
BACKGROUND: As part of a project designing nanoparticles for the treatment of Alzheimer's disease, we have synthesized and characterized a small library of nanoparticles binding with high affinity to the ß-amyloid peptide and showing features of biocompatibility in vitro, which are important properties for administration in vivo. In this study, we focused on biocompatibility issues, evaluating production of nitric oxide by cultured human umbilical vein endothelial cells and macrophages, used as models of cells which would be exposed to nanoparticles after systemic administration. METHODS: The nanoparticles tested were liposomes and solid lipid nanoparticles carrying phosphatidic acid or cardiolipin, and PEGylated poly(alkyl cyanoacrylate) nanoparticles (PEG-PACA). We measured nitric oxide production using the Griess method as well as phosphorylation of endothelial nitric oxide synthase and intracellular free calcium, which are biochemically related to nitric oxide production. MTT viability tests and caspase-3 detection were also undertaken. RESULTS: Exposure to liposomes did not affect the viability of endothelial cells at any concentration tested. Increased production of nitric oxide was detected only with liposomes carrying phosphatidic acid or cardiolipin at the highest concentration (120 µg/mL), together with increased synthase phosphorylation and intracellular calcium levels. Macrophages exposed to liposomes showed a slightly dose-dependent decrease in viability, with no increase in production of nitric oxide. Exposure to solid lipid nanoparticles carrying phosphatidic acid decreased viability in both cell lines, starting at the lowest dose (10 µg/mL), with increased production of nitric oxide detected only at the highest dose (1500 µg/mL). Exposure to PEG-PACA affected cell viability and production of nitric oxide in both cell lines, but only at the highest concentration (640 µg/mL). CONCLUSION: Liposomal and PEG-PACA nanoparticles have a limited effect on vascular homeostasis and inflammatory response, rendering them potentially suitable for treatment of Alzheimer's disease. Moreover, they highlight the importance of testing such nanoparticles for production of nitric oxide in vitro in order to identify a therapeutic dose range suitable for use in vivo.
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Peptídeos beta-Amiloides/metabolismo , Células Endoteliais/efeitos dos fármacos , Macrófagos/efeitos dos fármacos , Nanopartículas , Óxido Nítrico/metabolismo , Peptídeos beta-Amiloides/química , Animais , Cálcio/análise , Caspase 3/química , Caspase 3/metabolismo , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Células Endoteliais/citologia , Células Endoteliais/metabolismo , Células Endoteliais da Veia Umbilical Humana , Humanos , Espaço Intracelular/química , Macrófagos/citologia , Macrófagos/metabolismo , Teste de Materiais , Camundongos , Óxido Nítrico/análise , Óxido Nítrico Sintase Tipo III/química , Óxido Nítrico Sintase Tipo III/metabolismo , Fosforilação/efeitos dos fármacosRESUMO
Mutations in the presenilin 1 (PS1) gene are associated with early onset familial Alzheimer's disease (FAD). In this study, we found that the expression of mutant-PS1 in stable transfectants of SH-SY5Y neuroblastoma cells results in a reduction of the biosynthesis and steady-state levels of glucosylceramide. As an in vivo corroboration of these data, there was a significant reduction of brain glucosylceramide and gangliosides in an animal model of FAD. In mutant-PS1-transfectants (I143T, G384A), immunocytochemistry disclosed a remarkable reduction of glucosylceramide synthase (GlcT-1)-like immunoreactivity in the cells when compared with those of mock- and wild-PS1 transfectants. Immunoprecipitation of GlcT-1 protein from mutant-PS1 transfectants demonstrated a marked reduction in GlcT-1 protein, but there was no reduction in the levels of GlcT-1 mRNA. Both coprecipitation and γ-secretase inhibition experiments suggest that mutant-PS1 seems to form a complex with GlcT-1 protein and to be involved in GlcT-1 degradation, which was never found in other cell types. Thus, mutations in the PS1 gene result in profound glycosphingolipids abnormalities by abnormal molecular interaction with GlcT-1.
Assuntos
Doença de Alzheimer/genética , Doença de Alzheimer/metabolismo , Glicoesfingolipídeos/biossíntese , Proteínas Mutantes/genética , Proteínas Mutantes/metabolismo , Presenilina-1/genética , Presenilina-1/metabolismo , Substituição de Aminoácidos , Secretases da Proteína Precursora do Amiloide/antagonistas & inibidores , Animais , Sequência de Bases , Encéfalo/metabolismo , Carbamatos/farmacologia , Linhagem Celular , Primers do DNA/genética , Dipeptídeos/farmacologia , Inibidores Enzimáticos/farmacologia , Glucosiltransferases/genética , Glucosiltransferases/metabolismo , Humanos , Camundongos , Camundongos Transgênicos , Mutação , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , TransfecçãoRESUMO
Although the diverse triggers of AD are still under debate, the hypothesis of the contribution of cerebrovascular deficiencies has emerged in recent years. Cerebrovascular dysfunction may precede cognitive decline and onset of neurodegeneration. Indeed, the toxic Aß(42) aggregates constituting senile plaques, one of AD hallmarks, is often detected as amorphous material or fine fibrils in the brain capillary of AD patients. Aß(42) causing cerebral microangiopathy might originate either from the circulating blood, the vessel wall itself or the brain parenchyma. In the present investigation we show, for the first time, that in rat brain capillary endothelial cells (RBE4), in vitro oxygen glucose deprivation treatment elicits 250% of Aß(42) peptide production increase through a mechanism that involves the hypoxia inducible factor-1-mediated ß-secretase (BACE1) up-regulation. Furthermore, we observed a time dependent increase of amyloid protein precursor (AßPP) gene and protein expression, confirming previous reports which established the existence of AßPP in the cerebrovascular domain. Our experimental evidences point out that ischemic events may directly contribute in brain capillary endothelial cells to the enhancement of the amyloidogenic metabolism, leading to intracellular deposition of Aß(42). This events may contribute to the impairment of Aß brain clearance and AD related blood brain barrier dysfunctions.
Assuntos
Peptídeos beta-Amiloides/biossíntese , Barreira Hematoencefálica/metabolismo , Hipóxia Celular/fisiologia , Células Endoteliais/metabolismo , Hipóxia-Isquemia Encefálica/metabolismo , Fragmentos de Peptídeos/biossíntese , Doença de Alzheimer/metabolismo , Secretases da Proteína Precursora do Amiloide/biossíntese , Animais , Ácido Aspártico Endopeptidases/biossíntese , Western Blotting , Encéfalo/irrigação sanguínea , Encéfalo/metabolismo , Capilares/metabolismo , Ensaio de Imunoadsorção Enzimática , Perfilação da Expressão Gênica , Glucose/deficiência , Subunidade alfa do Fator 1 Induzível por Hipóxia/biossíntese , Neurônios/metabolismo , RNA Mensageiro/análise , Ratos , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
We investigated whether the toxicity of oligomeric amyloid-beta peptide (Abeta1-42) upon differentiated human neuroblastoma SH-SY5Y cells, can be affected by changes of membrane lipid composition. An immunostaining technique, using lipids extracted from the cells and separated by thin layer chromatography, suggested that Abeta preferentially binds to phosphatidylethanolamine (PE), one of the major lipids in the cell extract. For this reason, we utilized treatments with putative inhibitors of phosphatidylethanolamine biosynthesis (choline, phosphocholine, R59949) to decrease its proportion in the cell membrane; choline treatment (2.5 mM, 24 h) showed the best performance, reducing phosphatidylethanolamine content from 5.7 to 3.3 µg phosphorous/mg protein. Either the extent of Abeta binding or its toxicity decreased onto choline-treated cells. These data may open the possibility to develop future strategies aiming to reduce Abeta toxicity in Alzheimer disease.