Evaluación de la actividad de compuestos bioactivos microencapsulados en quitosano en modelos de inflamación intestinal

Tesis (Doctora en Ciencias Químicas) - - Universidad Nacional de Córdoba. Facultad de Ciencias Químicas, 2016Las enfermedades inflamatorias intestinales son de curso crónico y representan un factor de riesgo para el desarrollo de carcinoma colon-rectal. Los mecanismos por los que se inicia y perpetúa el daño inflamatorio en intestino se estudian empleando modelos experimentales como el de colitis inducida por la administración de dextrán sulfato de sodio (DSS). Numerosa evidencia clínica y experimental demuestra que la inflamación crónica se asocia con una exacerbada producción de especies reactivas de oxígeno (EROS), tales como radical anión superóxido (O2 •-), radical hidroxilo (HO•), oxígeno molecular singulete (O2 (1g)) y peróxido de hidrógeno (H2O2), que inducen un grado de estrés oxidativo crítico en la patofisiología de la inflamación intestinal. Una forma de reducir los efectos del estrés oxidativo es mediante el consumo o administración de compuestos bioactivos tales como flavonoides, que poseen capacidad antioxidante y efectos benéficos directos sobre la salud por su actividad anticancerígena, antitumoral y anti-inflamatoria. Desafortunadamente, estos compuestos son inestables y poco solubles en medios biológicos; una forma de estabilizarlos y vehiculizarlos es mediante la microencapsulación. Uno de los biopolímeros más empleados en la microencapsulación de compuestos bioactivos es el quitosano (Ch), un polisacárido mucoadhesivo, parcialmente desacetilado, derivado de la quitina, con numerosas aplicaciones en medicina y alimentación debido a que carece de toxicidad y alergenicidad, es biocompatible y biodegradable. Sin embargo, su empleo en medios biológicos se ve limitado por la reducida solubilidad a pH fisiológico, por lo que la funcionalización de la molécula con sustituyentes hidrofílicos se convierte en una alternativa viable para resolver tal inconveniente. En este contexto, se planteó como objetivo del presente trabajo, el estudio de la actividad de flavonoides microencapsulados en matrices de Ch en un modelo de colitis inducida por DSS. Para tal fin la metodología propuesta se estructuró en tres etapas: 1) Funcionalización de Ch con glucosamina mediante reacción de Maillard. Caracterización de Ch funcionalizado (ChF) mediante la determinación de grado de desacetilación, peso molecular, solubilidad a pH 7,4 y evaluación mediante espectroscopía UV-Vis y en el infrarrojo. Finalmente, se determinó la actividad antioxidante in vitro de Ch y ChF, mediante la evaluación de las eficiencias de desactivación de las EROS: O2 •-, HO•, y O2 (1g). 2) Microencapsulación de flavonoides en matrices de Ch nativo y ChF. Se obtuvieron microcápsulas (MC) de quercetina y genisteína, mediante la técnica de secado por aspersión (Spray-Drying). Las mismas fueron caracterizadas mediante la determinación de eficiencias y rendimientos de microencapsulación, morfología por microscopía electrónica de barrido, evaluación de liberación controlada de flavonoide bajo condiciones gastrointestinales simuladas, y capacidad antioxidante in vitro (comparativa entre de MC con y sin flavonoide y flavonoide en estado puro) mediante la desactivación de las EROS antes mencionadas. 3) Aplicación de MC de quercetina y genisteína en ChF en un modelo murino de colitis generada con DSS. A los animales bajo tratamiento se los evaluó clínicamente durante 10 días, posterior a su sacrificio se llevó a cabo el análisis histológico de colon, la evaluación de la actividad antiinflamatoria a través del ensayo de citoquinas y quimioquinas, la determinación de la actividad de mieloperoxidasa y N-acetilglucosaminidasa como indicadores de la infiltración de neutrófilos y macrófagos, respectivamente. Finalmente, se llevó a cabo la evaluación del estrés oxidativo mediante la determinación de la actividad de superóxido dismutasa y catalasa y la presencia de malondialdehído como producto de peroxidación lipídica de membrana. Se logró producir un derivado de Ch con solubilidad a pH fisiológico mejorada y capacidad antioxidante conservada, apto para la microencapsulación por Spray-Drying de flavonoides y su aplicación específica en un modelo murino de colitis. Se obtuvieron partículas gastro-resistentes con propiedades de liberación controlada que mejoraron la biodisponibilidad de los flavonoides en intestino, específicamente en colon, y actuaron aminorando los síntomas clínicos inducidos por el consumo de DSS, atenuando la inflamación y cooperando en el restablecimiento del balance de las especies oxidantes

Urinary creatinine excretion is an indicator of physical performance and function

Muscle mass is essential for performing physical activity, and low muscle mass (sarcopenia) has been found to have a strong association with all-cause mortality in patients with type 2 diabetes (T2D).1 However, muscle mass is not routinely assessed in clinical practice and low muscle mass can easily go unnoticed in obese patients (sarcopenic obesity), which was emphasized in previous DIALECT findings.2 The current definition of sarcopenia requires presence of either low muscle mass, low muscle strength or poor physical performance rather than low muscle mass alone.3 Two methods for estimating muscle mass independent of kidney function in clinical practice are the 24 h urinary creatinine excretion rate (CER) and bioelectric impedance analysis (BIA), but their association with physical performance and function is unclear.4 In this study we investigate whether CER or BIA-derived predicted muscle mass also indicate physical performance and function in patients with T2D, in order to indirectly screen patients on sarcopenia

Некоторые проблемы добычи полезных ископаемых на глубоких горизонтах недр

Cardiovascular screening may benefit middle-aged sportsmen, as coronary artery disease (CAD) is the main cause of exercise-related sudden cardiac death. Arterial stiffness, as measured by pulse wave velocity (PWV), may help identify sportsmen with subclinical CAD. We examined the additional value of PWV measurements to traditional CAD risk factors for identifying CAD.From the Measuring Athlete's Risk of Cardiovascular events (MARC) cohort of asymptomatic, middle-aged sportsmen who underwent low-dose Cardiac CT (CCT) after routine sports medical examination (SME), 193 consecutive sportsmen (aged 55 ± 6.6 years) were included with additional PWV measurements before CCT. Sensitivity, specificity and predictive values of PWV values (>8.3 and >7.5 m/s) assessed by Arteriograph were used to identify CAD (coronary artery calcium scoring ≥ 100 Agatston Units or coronary CT angiography luminal stenosis ≥ 50%) and to assess the additional diagnostic value of PWV to established cardiovascular risk factors.Forty-seven sportsmen (24%) had CAD on CCT. They were older (58.9 vs. 53.8 years, p<0.001), had more hypertension (17 vs. 4%, p=0.003), higher cholesterol levels (5.7 vs. 5.4 mmol/l) p=0.048), and more often were (ever) smokers (55 vs. 34%, p=0.008). Mean PWV was higher in those with CAD (8.9 vs. 8.0 m/s, p=0.017). For PWV >8.3m/s respectively >7.5 m/s sensitivity to detect CAD on CT was 43% and 74%, specificity 69% and 45%, positive predictive value 31% and 30%, and negative predictive value 79% and 84%. Adding PWV to traditional risk factor models did not change the area under the curve (from 0.78 (95% CI = 0.709-0.848)) to AUC 0.78 (95% CI 0.710-0.848, p = 0.99)) for prediction of CAD on CCT.Limited additional value was found for PWV on top of established risk factors to identify CAD. PWV might still have a role to identify CAD in middle-aged sportsmen if risk factors such as cholesterol are unknown

Effect of Q-switched laser surface texturing of titanium on osteoblast cell response

Titanium and its alloys are important biomedical materials. It is known that the surface texture of implanted medical devices affects cell response. Control of cell response has the potential to enhance fixation of implants into bone and, in other applications, to prevent undesired cell adhesion. The potential use of a 100W Q-switched YAG laser miller (DMG Lasertec 60 HSC) for texturing titanium is investigated. A series of regular features with dimensions of the order of tens of micrometers are generated in the surface of titanium samples and the cell response to these features is determined. Characterisation of the laser milled features reveals features with a lengthscale of a few microns superposed on the larger scale structures, this is attributed to resolidification of molten droplets generated and propelled over the surface by individual laser pulses. The laser textured samples are exposed to osteoblast cells and it is seen that cells do respond to the features in the laser textured surfaces

Low Physical Activity in Patients with Complicated Type 2 Diabetes Mellitus Is Associated with Low Muscle Mass and Low Protein Intake

Objective: In order to promote physical activity (PA) in patients with complicated type 2 diabetes, a better understanding of daily movement is required. We (1) objectively assessed PA in patients with type 2 diabetes, and (2) studied the association between muscle mass, dietary protein intake, and PA. Methods: We performed cross-sectional analyses in all patients included in the Diabetes and Lifestyle Cohort Twente (DIALECT) between November 2016 and November 2018. Patients were divided into four groups: = 10,000 steps/day. We studied the association between muscle mass (24 h urinary creatinine excretion rate, CER) and protein intake (by Maroni formula), and the main outcome variable PA (steps/day, Fitbit Flex device) using multivariate linear regression analyses. Results: In the 217 included patients, the median steps/day were 6118 (4115-8638). Of these patients, 48 patients (22%) took 7000-9999 steps/day, 37 patients (17%) took >= 10,000 steps/day, and 78 patients (36%) took = 10,000 steps/day, a higher body mass index (BMI) (33 +/- 6 vs. 30 +/- 5 kg/m(2), p = 0.009), lower CER (11.7 +/- 4.8 vs. 14.8 +/- 3.8 mmol/24 h, p = 0.001), and lower protein intake (0.84 +/- 0.29 vs. 1.08 +/- 0.22 g/kg/day, p < 0.001). Both creatinine excretion (beta = 0.26, p < 0.001) and dietary protein intake (beta = 0.31, p < 0.001) were strongly associated with PA, which remained unchanged after adjustment for potential confounders. Conclusions: Prevalent insufficient protein intake and low muscle mass co-exist in obese patients with low physical activity. Dedicated intervention studies are needed to study the role of sufficient protein intake and physical activity in increasing or maintaining muscle mass in patients with type 2 diabetes

Accelerated in vivo proliferation of memory phenotype CD4+ T-cells in human HIV-1 infection irrespective of viral chemokine co-receptor tropism.

CD4(+) T-cell loss is the hallmark of HIV-1 infection. CD4 counts fall more rapidly in advanced disease when CCR5-tropic viral strains tend to be replaced by X4-tropic viruses. We hypothesized: (i) that the early dominance of CCR5-tropic viruses results from faster turnover rates of CCR5(+) cells, and (ii) that X4-tropic strains exert greater pathogenicity by preferentially increasing turnover rates within the CXCR4(+) compartment. To test these hypotheses we measured in vivo turnover rates of CD4(+) T-cell subpopulations sorted by chemokine receptor expression, using in vivo deuterium-glucose labeling. Deuterium enrichment was modeled to derive in vivo proliferation (p) and disappearance (d*) rates which were related to viral tropism data. 13 healthy controls and 13 treatment-naive HIV-1-infected subjects (CD4 143-569 cells/ul) participated. CCR5-expression defined a CD4(+) subpopulation of predominantly CD45R0(+) memory cells with accelerated in vivo proliferation (p = 2.50 vs 1.60%/d, CCR5(+) vs CCR5(-); healthy controls; P<0.01). Conversely, CXCR4 expression defined CD4(+) T-cells (predominantly CD45RA(+) naive cells) with low turnover rates. The dominant effect of HIV infection was accelerated turnover of CCR5(+)CD45R0(+)CD4(+) memory T-cells (p = 5.16 vs 2.50%/d, HIV vs controls; P<0.05), naïve cells being relatively unaffected. Similar patterns were observed whether the dominant circulating HIV-1 strain was R5-tropic (n = 9) or X4-tropic (n = 4). Although numbers were small, X4-tropic viruses did not appear to specifically drive turnover of CXCR4-expressing cells (p = 0.54 vs 0.72 vs 0.44%/d in control, R5-tropic, and X4-tropic groups respectively). Our data are most consistent with models in which CD4(+) T-cell loss is primarily driven by non-specific immune activation

Cigarette smoke induces β2-integrin-dependent neutrophil migration across human endothelium

<p>Abstract</p> <p>Background</p> <p>Cigarette smoking induces peripheral inflammatory responses in all smokers and is the major risk factor for neutrophilic lung disease such as chronic obstructive pulmonary disease. The aim of this study was to investigate the effect of cigarette smoke on neutrophil migration and on β₂-integrin activation and function in neutrophilic transmigration through endothelium.</p> <p>Methods and results</p> <p>Utilizing freshly isolated human PMNs, the effect of cigarette smoke on migration and β₂-integrin activation and function in neutrophilic transmigration was studied. In this report, we demonstrated that cigarette smoke extract (CSE) dose dependently induced migration of neutrophils <it>in vitro</it>. Moreover, CSE promoted neutrophil adherence to fibrinogen. Using functional blocking antibodies against CD11b and CD18, it was demonstrated that Mac-1 (CD11b/CD18) is responsible for the cigarette smoke-induced firm adhesion of neutrophils to fibrinogen. Furthermore, neutrophils transmigrated through endothelium by cigarette smoke due to the activation of β₂-integrins, since pre-incubation of neutrophils with functional blocking antibodies against CD11b and CD18 attenuated this transmigration.</p> <p>Conclusion</p> <p>This is the first study to describe that cigarette smoke extract induces a direct migratory effect on neutrophils and that CSE is an activator of β₂-integrins on the cell surface. Blocking this activation of β₂-integrins might be an important target in cigarette smoke induced neutrophilic diseases.</p

Commissioning of inline ECE system within waveguide based ECRH transmission systems on ASDEX upgrade

A CW capable inline electron cyclotron emission (ECE) separation system for feedback control, featuring oversized corrugated waveguides, is commissioned on ASDEX upgrade (AUG). The system is based on a combination of a polarization independent, non-resonant, Mach-Zehnder diplexer equipped with dielectric plate beam splitters [2, 3] employed as corrugated oversized waveguide filter, and a resonant Fast Directional Switch, FADIS [4, 5, 6, 7] as ECE/ECCD separation system. This paper presents an overview of the system, the low power characterisation tests and first high power commissioning on AUG

Dose response of the 16p11.2 distal copy number variant on intracranial volume and basal ganglia.

Carriers of large recurrent copy number variants (CNVs) have a higher risk of developing neurodevelopmental disorders. The 16p11.2 distal CNV predisposes carriers to e.g., autism spectrum disorder and schizophrenia. We compared subcortical brain volumes of 12 16p11.2 distal deletion and 12 duplication carriers to 6882 non-carriers from the large-scale brain Magnetic Resonance Imaging collaboration, ENIGMA-CNV. After stringent CNV calling procedures, and standardized FreeSurfer image analysis, we found negative dose-response associations with copy number on intracranial volume and on regional caudate, pallidum and putamen volumes (β = -0.71 to -1.37; P &lt; 0.0005). In an independent sample, consistent results were obtained, with significant effects in the pallidum (β = -0.95, P = 0.0042). The two data sets combined showed significant negative dose-response for the accumbens, caudate, pallidum, putamen and ICV (P = 0.0032, 8.9 × 10-6, 1.7 × 10-9, 3.5 × 10-12 and 1.0 × 10-4, respectively). Full scale IQ was lower in both deletion and duplication carriers compared to non-carriers. This is the first brain MRI study of the impact of the 16p11.2 distal CNV, and we demonstrate a specific effect on subcortical brain structures, suggesting a neuropathological pattern underlying the neurodevelopmental syndromes

Heritability of fractional anisotropy in human white matter: a comparison of Human Connectome Project and ENIGMA-DTI data

The degree to which genetic factors influence brain connectivity is beginning to be understood. Large-scale efforts are underway to map the profile of genetic effects in various brain regions. The NIH-funded Human Connectome Project (HCP) is providing data valuable for analyzing the degree of genetic influence underlying brain connectivity revealed by state-of-the-art neuroimaging methods. We calculated the heritability of the fractional anisotropy (FA) measure derived from diffusion tensor imaging (DTI) reconstruction in 481 HCP subjects (194/287 M/F) consisting of 57/60 pairs of mono- and dizygotic twins, and 246 siblings. FA measurements were derived using (Enhancing NeuroImaging Genetics through Meta-Analysis) ENIGMA DTI protocols and heritability estimates were calculated using the SOLAR-Eclipse imaging genetic analysis package. We compared heritability estimates derived from HCP data to those publicly available through the ENIGMA-DTI consortium, which were pooled together from five-family based studies across the US, Europe, and Australia. FA measurements from the HCP cohort for eleven major white matter tracts were highly heritable (h2 = 0.53–0.90, p < 10− 5), and were significantly correlated with the joint-analytical estimates from the ENIGMA cohort on the tract and voxel-wise levels. The similarity in regional heritability suggests that the additive genetic contribution to white matter microstructure is consistent across populations and imaging acquisition parameters. It also suggests that the overarching genetic influence provides an opportunity to define a common genetic search space for future gene-discovery studies. Uniquely, the measurements of additive genetic contribution performed in this study can be repeated using online genetic analysis tools provided by the HCP ConnectomeDB web application