91,034 research outputs found

    Silencing of cypx gene in the hypothalamus and its impact on whole-body metabolism

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    Cholesterol metabolism is tightly controlled in the brain, through an equilibrium of cholesterol de novo synthesis and cholesterol efflux. Most of the cholesterol in the brain is in myelin sheaths to insulate axons and to maintain their morphology and synaptic transmission. The importance of cholesterol in the brain is highlighted by the fact that dysfunctions in its metabolism homeostasis are correlated with different neurodegenerative disorders. The hypothalamus, specially the ARC, is the principal brain region in the control of whole-body energy homeostasis, having a crucial role in metabolic organ regulation. This project goal was to silence the expression of the Cyp46a1 mouse gene in the hypothalamus of C57BL/6J wild-type mice fed with Chow and HFD, and to investigate its impact in the whole-body metabolism homeostasis. As the ARC is implicated in the control of whole-body energy metabolism, and oxysterols levels are altered in obesity, we hypothesize that the silencing of Cyp46a1 gene could lead to an obesity and TIIDM phenotypes. In this study, C57BL/6J wild-type mice (n=45) were divided into two groups corresponding to two different diets. One group (n=24) had access to a Chow containing 10% of fat and the other group (n=21) had access to an HFD containing 60% of fat. This study was conducted during 12 weeks. In the 4th week, the two groups of mice (Chow and HFD) were divided into four subgroups, from which two were submitted to the stereotaxic injection delivering the AAV of the serotype 5, with the shRNA targeting the mouse Cyp46a1 gene in each side of the ARC and two remained non-injected, as control. The silencing of Cyp46a1 mouse gene in the ARC of mice fed with Chow and HFD leads to an increase of BW, changes in food and water intake, to a reduction of glucose tolerance, of insulin sensitivity and leads also to modifications in several metabolic organs, comparatively to the non-injected animals. In fact, in the Chow AAV5-shCyp46a1 animals, the impact of the silencing Cyp46a1 gene appears to mimic an HFD effect, whereas in HFD AAV5-shCyp46a1 mice this silencing exacerbates the phenotype of obesity. These results could suggest an important role of the cholesterol metabolism in the brain, specially of the Cyp46a1 enzyme in the control of whole-body homeostasis. Finally, additional studies are needed to continue this project and it would be interesting to perform the overexpression of Cyp46a1 in the ARC to investigate if this gene could be a potential target to further genetic therapies.No c√©rebro, o metabolismo do colesterol √© extremamente controlado atrav√©s do equil√≠brio entre a s√≠ntese de novo, convers√£o e efluxo do colesterol. Quando a s√≠ntese do colesterol excede a sua necessidade no c√©rebro, ocorre o processo hidroxila√ß√£o do colesterol em 24-hidroxicolesterol (24-OHC) atrav√©s da a√ß√£o da enzima CYP46A1, uma enzima codificada pelo gene CYP46A1. O oxisterol 24-OHC, ao contr√°rio do colesterol, possui a capacidade de atravessar a barreira hematoencef√°lica contribuindo desta forma para a homeostasia do colesterol no c√©rebro. Este est√° maioritariamente localizado nas bainhas de mielina, de modo a isolar os ax√≥nios e manter a sua morfologia e transmiss√£o sin√°ptica. Diversas doen√ßas neurodegenerativas, como a doen√ßa de Alzheimer e a doen√ßa de Huntington, foram correlacionados com disfun√ß√Ķes na homeostasia do colesterol no c√©rebro, destacando desta forma a sua import√Ęncia no organismo. O hipot√°lamo, especialmente o n√ļcleo arqueado, √© a regi√£o do c√©rebro respons√°vel pela regula√ß√£o da homeostasia energ√©tica corporal, apresentando um papel crucial no equil√≠brio entre o consumo e o gasto energ√©tico. O objetivo deste projeto foi silenciar a express√£o do gene Cyp46a1 no hipot√°lamo de ratinhos C57BL/6J wild-type alimentados com uma dieta de controlo com baixo teor em gordura (Chow ‚Äď low fat control diet) e com uma dieta com alto teor em gordura (HFD - high fat diet) e investigar o seu impacto na homeostasia energ√©tica corporal. Uma vez que o n√ļcleo arqueado est√° implicado regula√ß√£o da homeostasia energ√©tica corporal e que os n√≠veis de oxister√≥is encontram-se alterados na obesidade, criou-se a hip√≥tese de que o silenciamento do gene Cyp46a1 poderia resultar em um fen√≥tipo de obesidade e de diabetes mellitus do tipo 2. Desta forma, os ratinhos C57BL/6J (n=45) foram divididos em dois grupos correspondentes a dietas distintas. Um dos grupos (n=24) teve acesso a uma dieta Chow, contendo 10% de gordura, e o outro grupo (n=21) teve acesso a uma dieta HFD, contendo 60% de gordura. Este estudo foi conduzido durante o per√≠odo de 12 semanas, no qual, na quarta semana os dois grupos (Chow e HFD) foram divididos em quatro subgrupos. Dois grupos foram submetidos √† inje√ß√£o estereot√°xica bilateral no ARC, constituindo os grupos tratados, (Chow AAV5-shCyp46a1 e HFD AAV5-shCyp46a1) e os restantes grupos n√£o foram submetidos √† inje√ß√£o estereot√°xica, constituindo os grupos de controlo. O silenciamento do gene Cyp46a1 no n√ļcleo arqueado dos ratinhos C57BL/6J alimentados com uma dieta Chow resultou em um aumento no peso corporal, na ingest√£o de alimentos e de √°gua, na redu√ß√£o da toler√Ęncia √† glucose, na sensibilidade √† insulina e ainda, em modifica√ß√Ķes em v√°rios √≥rg√£os metab√≥licos. Da mesma forma, o silenciamento do gene Cyp46a1 dos ratinhos C57BL/6J alimentados com uma dieta HFD resultou em um aumento do peso corporal, em uma diminui√ß√£o da ingest√£o de alimentos, da sensibilidade √† insulina e em modifica√ß√Ķes nos √≥rg√£os metab√≥licos. O silenciamento do gene Cyp46a1 no n√ļcleo arqueado dos ratinhos C57BL/6J parece modificar a morfologia do tecido adiposo branco, do tecido adiposo castanho e do f√≠gado, nos animais submetidos √† inje√ß√£o estereot√°xica. Os grupos tratados apresentaram importantes modifica√ß√Ķes na acumula√ß√£o lip√≠dica nestes √≥rg√£os, bem como modifica√ß√Ķes nos n√≠veis de prote√≠na. Al√©m disso, o silenciamento do gene Cyp46a1 resulta em modifica√ß√Ķes no comportamento nos ratinhos C57BL/6J alimentados com dietas Chow e uma HFD. Nos animais Chow AAV5-shCyp46a1, o impacto do silenciamento parece mimetizar o efeito de uma HFD, enquanto nos ratinhos HFD AAV5-shCyp46a1 esse silenciamento parece exacerbar o fen√≥tipo da obesidade, uma vez que estes animais j√° se encontravam metabolicamente desregulados. Os resultados podem desta forma sugerir um papel importante do metabolismo do colesterol no c√©rebro, especialmente da enzima CYP46A1 no controlo da homeostasia energ√©tica corporal. Finalmente, estudos adicionais ainda s√£o necess√°rios e seria interessante realizar a sobre express√£o do gene Cyp46a1, no n√ļcleo arqueado, para investigar se este gene poder√° ser um potencial alvo terap√™utico

    Desarrollo de materiales bioactivos con potencial aplicación odontológica mediante impregnación asistida por CO2 supercrítico

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    Tesis (DCI)--FCEFN-UNC, 2021En esta tesis se estudi√≥ el proceso de incorporaci√≥n de eugenol en fibras de poliamida 6 (PA6) mediante la impregnaci√≥n asistida por CO2 supercr√≠tico para desarrollar un material con propiedades antimicrobianas con una potencial aplicaci√≥n odontol√≥gica. Para este prop√≥sito, se construy√≥ un equipo de alta presi√≥n en el que se llevaron a cabo m√ļltiples ensayos de impregnaci√≥n de eugenol y de sorci√≥n de CO2 en un hilo dental comercial de PA6 en distintas condiciones de presi√≥n y temperatura (40 ‚Äď 60 ¬įC y 8 ‚Äď 12 MPa). Con el fin de encontrar las mejores condiciones del proceso de impregnaci√≥n, se evalu√≥ la influencia de diferentes variables operativas (presi√≥n, temperatura, tiempo de contacto y velocidad de despresurizaci√≥n) sobre la cantidad de eugenol impregnada en el material. Adem√°s, se estudiaron los principales fen√≥menos difusivos que ocurren en el proceso de impregnaci√≥n del eugenol en condiciones supercr√≠ticas. Para ello, se hicieron ensayos de cin√©tica de sorci√≥n del CO2 y del eugenol en PA6 a diferentes condiciones de presi√≥n y temperatura y se determin√≥ el coeficiente de difusi√≥n aparente para ambas especies en este pol√≠mero. Por otra parte, se evaluaron las propiedades finales del material impregnado, analizando las propiedades mec√°nicas, t√©rmicas y morfol√≥gicas del material original, presurizado con CO2 e impregnado con eugenol. Adicionalmente se evalu√≥ la actividad antimicrobiana del material impregnado frente a dos bacterias comunes (Escherichia coli y Staphylococcus aureus). Asimismo, se estudi√≥ la migraci√≥n del compuesto activo impregnado en aire y en saliva artificial, obteniendo datos importantes para el potencial desarrollo de un producto comercial, como la estimaci√≥n de la vida √ļtil, el tipo de envase, y tipo de aplicaci√≥n del producto. Finalmente, se hizo un dise√Īo y dimensionamiento de un proceso industrial para la impregnaci√≥n de eugenol en bobinas de fibras de PA6 en CO2 supercr√≠tico, a partir de los datos de eficiencia de impregnaci√≥n y par√°metros difusivos del hilo impregnado con eugenol previamente obtenidos, realizando el dise√Īo de la bobina, el equipo impregnador y los c√°lculos de sus principales requerimientos de masa y energ√≠a.Fil: Mosquera Ruiz, Jos√© Euliser. Universidad Nacional de C√≥rdoba. Facultad de Ciencias Exactas, F√≠sicas y Naturales; Argentina.Fil: Mosquera Ruiz, Jos√© Euliser. Consejo Nacional de Investigaciones Cient√≠ficas y T√©cnicas. Instituto de Investigaci√≥n y Desarrollo en Ingenier√≠a de Procesos y Qu√≠mica Aplicada; Argentina

    What is the importance of sperm subpopulations?

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    .The study of sperm subpopulations spans three decades. The origin, meaning, and practical significance, however, are less clear. Current technology for assessing sperm morphology (CASA-Morph) and motility (CASA-Mot) has enabled the accurate evaluation of these features, and there are many options for data classification. Subpopulations could occur as a result of the stage of development of each spermatozoon in the subpopulation. Spermatogenesis might contribute to the production of these subpopulations. Insights from evolutionary biology and recent molecular research are indicative of the diversity among male gametes that could occur from unequal sharing of transcripts and other elements through cytoplasmic bridges between spermatids. Sperm cohorts exiting the gonads would contain different RNA and protein contents, affecting the spermatozoon physiology and associations with the surrounding environmental milieu. Subsequently, these differences could affect how spermatozoa interact with the environmental milieu (maturation, mixing with seminal plasma, and interacting with the environmental milieu, or female genital tract and female gamete). The emergence of sperm subpopulations as an outcome of evolution, related to the reproductive strategies of the species, genital tract structures, and copulatory and fertilization processes. This kind of approach in determining the importance of sperm subpopulations in fertilization capacity should have a practical impact for conducting reproductive technologies, inspiring and enabling new ways for the more efficient use of spermatozoa in the medical, animal breeding, and conservation fields. This manuscript is a contribution to the Special Issue in memory of Dr. Duane GarnerS

    RELATIONSHIP BETWEEN BODY-SEAT INTERFACE PRESSURE AND DISCOMFORT DURING ROWING

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    Discomfort and pressure-related tissue injury to the buttocks are common complaints among rowers. The soft tissues of the buttocks are non-uniformly loaded during rowing. The current state of literature on seating discomfort is inconclusive as to a desirable body-seat interface pressure pattern. The purpose of this study was to determine whether localising pressure under bony protuberances or diffusing pressure over soft tissues would result in the least amount of discomfort. Force sensing arrays were used to measure body-seat interface pressures in 11 elite female rowers during rowing. Peak pressure measures were identified and pressure gradients were calculated. Discomfort was quantified using a questionnaire, and pressure data were then correlated with discomfort scores.Discomfort was weakly correlated with each of maximal pressure gradient (r=0.45) and peak pressure (r=0.43). The findings indicate pressure should be redistributed in order to avoid concentrating pressure under the bony protuberances o f the buttocks

    Exploring the effects of spinal cord stimulation for freezing of gait in parkinsonian patients

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    Dopaminergic replacement therapies (e.g. levodopa) provide limited to no response for axial motor symptoms including gait dysfunction and freezing of gait (FOG) in Parkinson’s disease (PD) and Richardson’s syndrome progressive supranuclear palsy (PSP-RS) patients. Dopaminergic-resistant FOG may be a sensorimotor processing issue that does not involve basal ganglia (nigrostriatal) impairment. Recent studies suggest that spinal cord stimulation (SCS) has positive yet variable effects for dopaminergic-resistant gait and FOG in parkinsonian patients. Further studies investigating the mechanism of SCS, optimal stimulation parameters, and longevity of effects for alleviating FOG are warranted. The hypothesis of the research described in this thesis is that mid-thoracic, dorsal SCS effectively reduces FOG by modulating the sensory processing system in gait and may have a dopaminergic effect in individuals with FOG. The primary objective was to understand the relationship between FOG reduction, improvements in upper limb visual-motor performance, modulation of cortical activity and striatal dopaminergic innervation in 7 PD participants. FOG reduction was associated with changes in upper limb reaction time, speed and accuracy measured using robotic target reaching choice tasks. Modulation of resting-state, sensorimotor cortical activity, recorded using electroencephalography, was significantly associated with FOG reduction while participants were OFF-levodopa. Thus, SCS may alleviate FOG by modulating cortical activity associated with motor planning and sensory perception. Changes to striatal dopaminergic innervation, measured using a dopamine transporter marker, were associated with visual-motor performance improvements. Axial and appendicular motor features may be mediated by non-dopaminergic and dopaminergic pathways, respectively. The secondary objective was to demonstrate the short- and long-term effects of SCS for alleviating dopaminergic-resistant FOG and gait dysfunction in 5 PD and 3 PSP-RS participants without back/leg pain. SCS programming was individualized based on which setting best improved gait and/or FOG responses per participant using objective gait analysis. Significant improvements in stride velocity, step length and reduced FOG frequency were observed in all PD participants with up to 3-years of SCS. Similar gait and FOG improvements were observed in all PSP-RS participants up to 6-months. SCS is a promising therapeutic option for parkinsonian patients with FOG by possibly influencing cortical and subcortical structures involved in locomotion physiology

    Examination of a Brief, Self-Paced Online Self-Compassion Intervention Targeting Intuitive Eating and Body Image Outcomes among Men and Women

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    Ideals for appearance and body image are pervasive in Western culture in which men and women are portrayed with unrealistic and often unattainable standards (Ferguson, 2013; Martin, 2010). Exposure and reinforcement have created a culture of social acceptance and internalization of these ideals, contributing to pervasive body image disturbance (i.e., body dissatisfaction; Fallon et al., 2014; Stice, 2001; Thompson & Stice, 2001; Thompson et al., 1999). Research has suggested that body dissatisfaction is expressed differently across sexes (Grossbard et al., 2008), with attention to thin ideals among women and muscular ideals among men. Body dissatisfaction has been linked to numerous poor outcomes, including dieting, unhealthy weight control behaviors, disordered eating, and increased psychopathology. Although dieting is one of the primary mechanisms employed to reduce body dissatisfaction (Thompson & Stice, 2001), research has shown that such efforts are contraindicated as dieting predicts weight gain over time (Pietil√ɬ§inen et al., 2012) as well as preoccupation with food, disordered eating, eating disorders, emotional distress, and higher body dissatisfaction (Grabe et al., 2007; Johnson & Wardle, 2005; Neumark- Sztianer et al., 2006; Paxton et al., 2006; Tiggemann, 2005). Restrictive dietary behaviors suppress physiological cues to eat (e.g., hunger) that presents a vulnerability to eating in response to alternative cues, both internal (e.g., emotions) and external (e.g., availability of food). Intuitive eating is a non-restrictive approach to eating that encourages adherence to internal physiological cues to indicate when, what, and how much to eat (Tylka, 2006) and has demonstrated an inverse relationship with disordered eating, restrained eating, food preoccupation, dieting, body dissatisfaction, and negative affect (Bruce & Ricciardelli, 2016). Self-compassion, relating to oneself in a caring and supportive manner (Neff, 2003a), has been proposed as a pathway to increase intuitive eating and reduce body dissatisfaction (Neff & Knox, 2017; Schoenefeld & Webb, 2013; Webb & Hardin, 2016). Research has highlighted the efficacy of self-compassion interventions in addressing weight-related concerns (Rahimi-Ardabili et al., 2018) as well as brief experiential exercises for reducing body dissatisfaction (Moffitt et al., 2018). Additionally, there is a growing body of evidence supporting the efficacy of internet-based self-compassion interventions (Mak et al., 2018; Kelman et al., 2018; Nadeau et al., 2020). The purpose of the current study was to examine the effectiveness of a brief, self-paced online self-compassion intervention targeting body image and adaptive eating behaviors and potential mechanisms of change (e.g., self-compassion and psychological flexibility) among undergraduate men and women. This study also examined outcomes among men and women in the area of self-compassion, body dissatisfaction, and intuitive eating as research has highlighted the need to determine who benefits more from self-compassion interventions (Rahimi-Ardabili et al., 2018). The study compared a one-hour, self-guided online self-compassion intervention to an active control condition. The intervention was comprised of psychoeducation, experiential exercises, and mindfulness practice designed to increase self-compassion surrounding body image and eating behaviors. In contrast, the active control condition consisted of self-care recommendations and self-assessments for nutrition, exercise, and sleep. The study was administered over three parts (e.g., baseline, intervention, and follow-up) in which variables of interest were assessed at each time point. Outcome variables included self-compassion, intuitive eating, disordered eating, body appreciation, muscle dysmorphia, internalized weight bias, fear of self-compassion, and psychological inflexibility. Participants were randomized on a 2:1 intervention to control ratio at the second time point in order to make comparisons between groups while simultaneously having sufficient power for examining mediation and moderation within the treatment condition. Overall, 1023 individuals (64% women, Mage = 18.9, 67.4% white) signed informed consent and participated in at least one part of the study whereas 101 participants (71% women, Mage = 19.3, 71% white) completed all three study portions. As predicted, self-compassion was correlated with all variables of interest, and all study variables were correlated with each other (p < .01). In contrast to hypothesized outcomes, the self-compassion condition failed to demonstrate improvements across time or between conditions on all study outcomes. These results persisted when participants were screened for levels of intuitive eating as well. Contrary to prediction, internalized weight bias, muscle dysmorphia, and fear of self-compassion demonstrated increased levels within the intervention condition and decreases in the control condition. There were significant gender differences on multiple outcome variables, with men demonstrating higher levels of self-compassion and body appreciation whereas women endorsed higher levels of disordered eating, internalized weight bias, muscle dysmorphia, and psychological inflexibility. Additionally, there were significant gender interactions for internalized weight bias, body appreciation, and muscle dysmorphia. The interactions existed such that men demonstrated increased internalized weight bias and muscle dysmorphia across time whereas women displayed decreased weight bias and muscle dysmorphia. The opposite pattern was found within body appreciation; women demonstrated increased body appreciation across time while men reported decreased levels of body appreciation. Despite this study√Ę‚ā¨‚ĄĘs intent to examine underlying mechanisms of change, the condition in which participants were randomly selected did not have any relationship, positive or negative, with the outcome variables of interest. As such, mediation within the current study was not conducted as it would violate statistical assumptions required to examine this hypothesis. Finally, upon examining the moderating relationship of fear of self-compassion between self-compassion and outcome variables, there were main effects for self-compassion on intuitive eating, emotional eating, internalized weight bias, body appreciation, and psychological inflexibility as well as main effects of fear of self-compassion on psychological inflexibility. There were significant interactions for intuitive eating and emotional eating, such that as fear of self-compassion increased, the effect of self-compassion on intuitive eating decreased, and the effect of self-compassion on reducing emotional eating behaviors decreased. Overall, the brief, self-paced online intervention delivered in the current study did not prove to be an effective means for improving self-compassion, intuitive eating, body appreciation, disordered eating, muscle dysmorphia, and psychological inflexibility. Nevertheless, the relationships between self-compassion and outcome variables of interest throughout the study mirror that of the existing literature. Findings from this study, in general, were also consistent with differences between men and women despite a gap in the research for intervention outcomes. Although fear of self-compassion demonstrated a moderating effect on the relationship between self-compassion and intuitive eating as well as emotional eating, this does not account for the lack of significant findings. The context surrounding this study, such as the COVID-19 pandemic, provided a considerable challenge to examining the efficacy of the current intervention. However, the findings of this study suggest future research will likely need to identify ways to enhance the delivery of experiential exercises that encourage engagement, provide a safe and warm environment for participants, and create flexibility and willingness surrounding painful and difficult experiences in order to undermine internalized and socially accepted beliefs about body image and eating behaviors

    Hepatic glycogen participates in the regulation of hypothalamic pAkt/Akt ratio in high-sugar/high-fat diet-induced obesity

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    The hypothalamus is a major integrating centre that controls energy homeostasis and plays a major role in hepatic glycogen (HGlyc) turnover. Not only do hypothalamic and hepatic Akt levels influence glucose homeostasis and glycogen synthesis, but exposure to high-sugar/high-fat diets (HSHF) can also lead to hypothalamic inflammation and HGlyc accumulation. HSHF withdrawal overall restores energy and glucose homeostasis, but the actual relationship between hypothalamic inflammation and HGlyc after short-term HSHF withdrawal has not yet been fully elucidated. Here we investigated the short-term effects of HSHF withdrawal preceded by a 30-day HSHF intake on the liver-hypothalamus crosstalk and glucose homeostasis. Sixty-day old male Wistar rats were fed for 30 days a control chow (n‚ÄČ=‚ÄČ10) (Ct), or an HSHF diet (n‚ÄČ=‚ÄČ20). On the 30th day of dietary intervention, a random HSHF subset (n‚ÄČ=‚ÄČ10) had their diets switched to control chow for 48 h (Hw) whilst the remaining HSHF rats remained in the HSHF diet (n‚ÄČ=‚ÄČ10) (Hd). All rats were anaesthetized and euthanized at the end of the protocol. We quantified HGlyc, Akt phosphorylation, inflammation and glucose homeostasis biomarkers. We also assessed the effect of propensity to obesity on those biomarkers, as detailed previously. Hd rats showed impaired glucose homeostasis, higher HGlyc and hypothalamic inflammation, and lower pAkt/Akt. Increased HGlyc was significantly associated with HSHF intake on pAkt/Akt lowered levels. We also found that HGlyc breakdown may have prevented a further pAkt/Akt drop after HSHF withdrawal. Propensity to obesity showed no apparent effect on hypothalamic inflammation or glucose homeostasis. Our findings suggest a comprehensive role of HGlyc as a structural and functional modulator of energy metabolism, and such roles may come into play relatively rapidly

    Bioinformatic characterization of a triacylglycerol lipase produced by Aspergillus flavus isolated from the decaying seed of Cucumeropsis mannii

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    Lipases are enzymes of industrial importance responsible for the hydrolysis of ester bonds of triglycerides. A lipolytic fungus was isolated and subsequently identified based on the ITS sequence analysis as putative Aspergillus flavus with accession number LC424503. The gene coding for extracellular triacylglycerol lipase was isolated from Aspergillus flavus species, sequenced, and characterised using bioinformatics tools. An open reading frame of 420 amino acid sequence was obtained and designated as Aspergillus flavus lipase (AFL) sequence. Alignment of the amino acid sequence with other lipases revealed the presence GHSLG sequence which is the lipase consensus sequence Gly-X1-Ser-X2-Gly indicating that it a classical lipase. A catalytic active site lid domain composed of TYITDTIIDLS amino acids sequence was also revealed. This lid protects the active site, control the catalytic activity and substrate selectivity in lipases. The 3-Dimensional structural model shared 34.08% sequence identity with a lipase from Yarrowia lipolytica covering 272 amino acid residues of the template model. A search of the lipase engineering database using AFL sequence revealed that it belongs to the class GX-lipase, superfamily abH23 and homologous family abH23.02, molecular weight and isoelectric point values of 46.95‚ÄČKDa and 5.7, respectively. N-glycosylation sites were predicted at residues 164, 236 and 333, with potentials of 0.7250, 0.7037 and 0.7048, respectively. O-glycosylation sites were predicted at residues 355, 358, 360 and 366. A signal sequence of 37 amino acids was revealed at the N-terminal of the polypeptide. This is a short peptide sequence that marks a protein for transport across the cell membrane and indicates that AFL is an extracellular lipase. The findings on the structural and molecular properties of Aspergillus flavus lipase in this work will be crucial in future studies aiming at engineering the enzyme for biotechnology applications

    Medicina personalizada en lactantes con cáncer: Estudio farmacogenético de polimorfismos relacionados con toxicidad y respuesta a la quimioterapia

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    Introducci√≥n La farmacogen√©tica es una herramienta de la ‚ÄúMedicina personalizada‚ÄĚ que contribuye a optimizar los tratamientos antineopl√°sicos, adapt√°ndolos a las caracter√≠sticas gen√©ticas particulares de cada individuo, maximizando su eficacia y minimizando su toxicidad. El lactante con c√°ncer es un paciente de particular vulnerabilidad y sus comorbilidades tiene una especial repercusi√≥n vital. El estudio farmacogen√©tico en esta poblaci√≥n resulta pionero y novedoso en nuestro medio. La identificaci√≥n de marcadores predictivos espec√≠ficos permitir√° individualizar m√°s la terap√©utica en esta poblaci√≥n tan fr√°gil y mejorar en consecuencia su calidad de vida y su pron√≥stico vital. Material y m√©todos Estudio de cohortes ambispectivo de pacientes oncol√≥gicos entre 1 y 18 meses de edad, receptores de quimioterapia en el Hospital La Fe de Valencia, en el periodo comprendido entre enero 2007 y agosto 2019. La parte retrospectiva comprende hasta diciembre 2015, el estudio prospectivo desde enero 2016 hasta agosto 2019. En primer lugar, se realiza un an√°lisis descriptivo de variables epidemiol√≥gicas, cl√≠nico/biol√≥gicas, terap√©uticas y pron√≥sticas de 72 pacientes con dichas caracter√≠sticas. Se describe la toxicidad derivada de sus 578 ciclos de quimioterapia (37 variables cl√≠nicas), el tiempo de seguimiento y su supervivencia (meses). En segundo lugar, se realiza un estudio anal√≠tico cuyo objetivo es correlacionar los polimorfismos gen√©ticos relacionados con la quimioterapia de 64 de los pacientes, la toxicidad grave secundaria al tratamiento (‚Č• grado 3 seg√ļn CTAE 4.0) y su supervivencia. El genotipado se realiza en el Centro Nacional de Genotipado (CEGEN) mediante la tecnolog√≠a MassArray (AgenaBioscience), previa configuraci√≥n de un panel farmacogen√©tico pedi√°trico en base a las evidencias recogidas en la base de datos PharmGKB, fichas t√©cnicas de los medicamentos y consorcios internacionales expertos. El an√°lisis estad√≠stico descriptivo se realiza con los programas Excel 2016 y R: las variables cualitativas con el recuento num√©rico (porcentaje) y las variables cuantitativas como mediana +/- rango intercuart√≠lico ante ausencia de normalidad en la distribuci√≥n de los datos (p <0,05, prueba de Kolmogorov-Smirnov). En el an√°lisis de supervivencia se utiliza el estimador Kaplan-Meier. El an√°lisis estad√≠stico anal√≠tico de correlaci√≥n entre polimorfismos y toxicidad se realiza mediante regresi√≥n log√≠stica penalizada por Elastic Net empleando R. El an√°lisis estad√≠stico anal√≠tico de correlaci√≥n entre polimorfismos y reca√≠da/muerte se realiza mediante regresi√≥n de Cox penalizada por Elastic Net. Resultados Las variables epidemiol√≥gicas, cl√≠nico/biol√≥gicas y terap√©uticas de los pacientes de la muestra son consecuentes con las descritas en la literatura del lactante con c√°ncer. Las neoplasias con mayor impacto negativo en la supervivencia son la leucemia mielobl√°stica aguda y los tumores del sistema nervioso central. La toxicidad m√°s prevalente es hematol√≥gica, digestiva e infecciosa. Existe correlaci√≥n entre la toxicidad grave secundaria a los quimioter√°picos en forma de anemia, neutropenia y/o trombopenia y 46 polimorfismos gen√©ticos diferentes. As√≠ mismo se encuentra asociaci√≥n estad√≠sticamente significativa entre la supervivencia global y la supervivencia libre de enfermedad y ciertos polimorfismos gen√©ticos (26 y 13 respectivamente). Los polimorfismos obtenidos pertenecen a genes encargados del transporte (6 genes) y metabolismo (17 genes) de los f√°rmacos, de la reparaci√≥n del material gen√©tico y la supresi√≥n tumoral (5 genes) y de otras funciones biol√≥gicas (4 genes). Conclusi√≥n Los resultados del presente estudio muestran correlaci√≥n estad√≠stica entre 46 polimorfismos de genes implicados en la cin√©tica farmacol√≥gica y la reparaci√≥n del material gen√©tico y la variabilidad en la toxicidad quimioter√°pica y supervivencia de pacientes lactantes con c√°ncer. En definitiva, aportaciones de la farmacogen√©tica que pueden contribuir a la optimizaci√≥n del tratamiento antineopl√°sico en esta poblaci√≥n particular y a la predicci√≥n de sus riesgos, de especial impacto en los supervivientes del c√°ncer infantil

    Uso de las histonas circulantes y sus modificaciones post-traduccionales como biomarcadores en sepsis y shock séptico

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    La sepsis es una afecci√≥n potencialmente mortal causada por una respuesta anormal del hu√©sped a una infecci√≥n, produciendo respuestas fisiol√≥gicas alteradas que da√Īan los propios tejidos del paciente y pueden provocar disfunci√≥n org√°nica e incluso la muerte. Asimismo, algunos pacientes s√©pticos progresan a shock s√©ptico, caracterizado por alteraciones circulatorias, celulares y metab√≥licas sustanciales que aumentan el riesgo de mortalidad. A pesar de que la sepsis se caracteriza por un mal funcionamiento del sistema inmunol√≥gico, lo que a su vez conduce a una respuesta inmune alterada e inmunosupresi√≥n, la alta complejidad de la fisiopatolog√≠a de la sepsis requiere una mayor investigaci√≥n para comprender las respuestas inmunes que ocurren durante la sepsis. Asimismo, las histonas extracelulares circulantes han ganado relevancia como mediadores citot√≥xicos en la sepsis, ya que act√ļan como patrones moleculares asociados a da√Īo, que inducen estr√©s oxidativo y activan el inflamasoma NLRP3. Estos mecanismos median la activaci√≥n de la piroptosis, un mecanismo de muerte celular programada que produce inflamaci√≥n mediante la expresi√≥n de IL-18, IL-1ő≤ and IL-1őĪ. Sin embargo, a pesar de la evidencia de activaci√≥n del inflamasoma en las c√©lulas inmunes durante la sepsis, se desconoce si las histonas extracelulares son capaces de activar los inflamasomas endoteliales y sus consecuencias. En este trabajo destacamos el papel previamente desconocido de las histonas extracelulares, mediando la activaci√≥n del inflamasoma NLRP3 y la piroptosis en las c√©lulas endoteliales, contribuyendo a la disfunci√≥n endotelial y la desregulaci√≥n de la respuesta inmune mediada por el endotelio. Asimismo, tambi√©n demostramos c√≥mo la acetilaci√≥n de histonas disminuye la activaci√≥n de la piroptosis. Adem√°s, demostramos que la piroptosis se produce en pacientes con shock s√©ptico y los niveles de histonas circulantes se correlacionan con la expresi√≥n de citoquinas proinflamatorias y citoquinas piropt√≥ticas, la liberaci√≥n de factores de adhesi√≥n endotelial y la gravedad de la enfermedad. Proponemos la piroptosis mediada por histonas como un nuevo objetivo para desarrollar intervenciones cl√≠nicas. De manera similar, hemos analizado las respuestas inmunorelacionadas que ocurren durante las primeras etapas de la sepsis con el objetivo de proporcionar nuevos datos comparando las cantidades de citoquinas, inmunomoduladores y otros mediadores endoteliales en pacientes cr√≠ticamente enfermos no s√©pticos, s√©pticos y de shock s√©ptico. Nuestro enfoque ayudar√° a caracterizar r√°pidamente las respuestas inmunes alteradas en pacientes s√©pticos y de shock s√©ptico ingresados en la Unidad de Cuidados Intensivos. Finalmente analizamos el papel de la metilaci√≥n del ADN en el control del sistema inmune s√©ptico. Nuestros resultados demostraron el papel central de la metilaci√≥n del ADN modulando la respuesta molecular en los pacientes de shock s√©ptico y contribuyendo a la inmunosupresi√≥n, a trav√©s de la alteraci√≥n de los patrones de metilaci√≥n de los promotores de IL-10 y TREM-2.Sepsis is a life-threatening condition caused by an abnormal host response to an infection that produce altered physiological responses which damages own tissues of the patient and can result in organ dysfunction and in some cases death. Likewise, a subset of septic patients progresses to septic shock, characterized by substantial circulatory, cellular and metabolic abnormalities, which substantially increase the risk of mortality. Sepsis is characterized by a malfunction of the immune system and it can lead to an altered immune response and immunosuppression. Moreover, the high complexity of the pathophysiology of sepsis requires of further investigation to characterize the immune responses in sepsis and septic shock. Likewise, circulating extracellular histones have gained relevance as cytotoxic mediators in sepsis pathophysiology, since they act as damage-associated molecular patterns, which induce oxidative stress and activate NLRP3 inflammasome. Subsequently, inflammasome mediates pyroptosis activation, a programmed cell death mechanism that produces inflammation through the release of IL-18, IL-1ő≤ and IL-1őĪ. However, despite inflammasome activation may occur in immune cells during sepsis, it is unknown if this process also takes place in endothelial cells and particularly whether extracellular histones are capable of activating endothelial inflammasomes and their consequences. In this work we highlight a previously unknown role for extracellular histones, that mediates the activation of NLRP3 inflammasome and pyroptosis in endothelial cells by contributing to endothelial dysfunction and the dysregulation of the immune response mediated by endothelium. Likewise, we demonstrated how histone acetylation decreases pyroptosis activation. Furthermore, we show how pyroptosis occurs in septic shock patients and how circulating histone levels correlate with the expression of pro-inflammatory and pyroptotic cytokines, the release of endothelial adhesion factors and septic shock severity. We propose histone-mediated pyroptosis as a new target to develop clinical interventions. Similarly, we have analyzed the immune-related responses occurring during the early stages of sepsis with the aim of providing new data by comparing the amounts of cytokines, immune modulators and other endothelial mediators in critically-ill non-septic patients, septic and septic shock patients. Our approach will help to rapidly characterize the altered immune responses in septic and septic shock patients admitted in the Intensive Care Unit. Finally, we also analyzed the role of DNA methylation in the control of septic immune system. Our results demonstrated the central role of DNA methylation modulating the molecular response in septic shock patients and contributing to immunosuppression, through the alteration of DNA methylation patterns of IL-10 and TREM2 promoters
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