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Large conductance voltage-and calcium-activated K+ (BK) channel in health and disease.
Echeverría, F, Gonzalez-Sanabria, N, Alvarado-Sanchez, R, Fernández, M, Castillo, K, Latorre, R
Frontiers in pharmacology. 2024;:1373507
Abstract
Large Conductance Voltage- and Calcium-activated K+ (BK) channels are transmembrane pore-forming proteins that regulate cell excitability and are also expressed in non-excitable cells. They play a role in regulating vascular tone, neuronal excitability, neurotransmitter release, and muscle contraction. Dysfunction of the BK channel can lead to arterial hypertension, hearing disorders, epilepsy, and ataxia. Here, we provide an overview of BK channel functioning and the implications of its abnormal functioning in various diseases. Understanding the function of BK channels is crucial for comprehending the mechanisms involved in regulating vital physiological processes, both in normal and pathological conditions, controlled by BK. This understanding may lead to the development of therapeutic interventions to address BK channelopathies.
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The role of Eudragit® as a component of hydrogel formulations for medical devices.
Esporrín-Ubieto, D, Sonzogni, AS, Fernández, M, Acera, A, Matxinandiarena, E, Cadavid-Vargas, JF, Calafel, I, Schmarsow, RN, Müller, AJ, Larrañaga, A, et al
Journal of materials chemistry. B. 2023;(38):9276-9289
Abstract
Over the last decade, significant progress has been made in developing hydrogels as medical devices. By physically cross-linking pharmaceutically approved polymers into three-dimensional matrices, we can ensure their biocompatibility and facilitate their seamless transition from the laboratory to clinical applications. Moreover, the reversible nature of their physical cross-links allows hydrogels to dissolve in the presence of external stimuli. Particularly, their high degree of hydration, high molecular weight, and superior flexibility of the polymer chains facilitate their interaction with complex biological barriers (e.g., mucus layer), making them ideal candidates for mucosal drug delivery. However, fine-tuning the composition of the hydrogel formulations is of great importance to optimize the performance of the medical device and its therapeutic cargo. Herein, we investigated the influence of different Eudragits® on the properties of hydrogels based on polyvinylpyrrolidone (PVP), polyvinyl alcohol (PVA), and polyethylene glycol (PEG), which were originally proposed as ocular inserts in previous reports. Our research aims to determine the effects that including different Eudragits® have on the structure and protein ocular delivery ability of various hydrogel formulations. Properties such as matrix stability, protein encapsulation, release kinetics, mucoadhesion, and biocompatibility have been analyzed in detail. Our study represents a guideline of the features that Eudragits® have to exhibit to endow hydrogels with good adhesion to the eye's conjunctiva, biocompatibility, and structural strength to cope with the ocular biointerface and allow sustained protein release. This work has important implications for the design of new hydrogel materials containing Eudragits® in their composition, particularly in mucosal drug delivery.
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Differential Health Effects on Inflammatory, Immunological and Stress Parameters in Professional Soccer Players and Sedentary Individuals after Consuming a Synbiotic. A Triple-Blinded, Randomized, Placebo-Controlled Pilot Study.
Quero, CD, Manonelles, P, Fernández, M, Abellán-Aynés, O, López-Plaza, D, Andreu-Caravaca, L, Hinchado, MD, Gálvez, I, Ortega, E
Nutrients. 2021;13(4)
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Synbiotic, a mixture of prebiotics and probiotics, is known to improve neurotransmitter interactions, immune, inflammatory, and stress responses by modulating the gut microbial composition. It is also believed that physical activity plays an important role in the modulation of immune function and stress response. The purpose of this triple-blinded, randomized, placebo-controlled pilot study was to evaluate the health benefits of symbiotic intervention in fourteen sedentary students and thirteen soccer players, especially in terms of improving immunophysiological and metabolic parameters. The 300mg of symbiotic intervention contained Bifidobacterium lactis CBP-001010, Lactobacillus rhamnosus CNCM I-4036, Bifidobacterium longum ES1(109 colony-forming unit), and fructooligosaccharides (200 mg) plus 1.5 mg of zinc, 8.25 µg of selenium, 0.75 µg of vitamin, and maltodextrin. Following a one-month intervention with synbiotic formulation, soccer players showed improvements in anxiety, sleep quality and stress, a slight reduction in proinflammatory cytokine IL-1β, an exercise-induced significant increase in dopamine and a slight elevation of corticotropin-releasing hormone. For confirmation of results of this pilot study and to assess more significant effects of symbiotic intervention in athletes as well as in the general population, longer-term robust studies are required. The findings of this study can help healthcare professionals understand the extensive health benefits of synbiotic intervention and its relationship to physical activity.
Abstract
The main objective of this research was to carry out an experimental study, triple-blind, on the possible immunophysiological effects of a nutritional supplement (synbiotic, Gasteel Plus®, Heel España S.A.U.), containing a mixture of probiotic strains, such as Bifidobacterium lactis CBP-001010, Lactobacillus rhamnosus CNCM I-4036, and Bifidobacterium longum ES1, as well as the prebiotic fructooligosaccharides, on both professional athletes and sedentary people. The effects on some inflammatory/immune (IL-1β, IL-10, and immunoglobulin A) and stress (epinephrine, norepinephrine, dopamine, serotonin, corticotropin-releasing hormone (CRH), Adrenocorticotropic hormone (ACTH), and cortisol) biomarkers were evaluated, determined by flow cytometer and ELISA. The effects on metabolic profile and physical activity, as well as on various parameters that could affect physical and mental health, were also evaluated via the use of accelerometry and validated questionnaires. The participants were professional soccer players in the Second Division B of the Spanish League and sedentary students of the same sex and age range. Both study groups were randomly divided into two groups: a control group-administered with placebo, and an experimental group-administered with the synbiotic. Each participant was evaluated at baseline, as well as after the intervention, which lasted one month. Only in the athlete group did the synbiotic intervention clearly improve objective physical activity and sleep quality, as well as perceived general health, stress, and anxiety levels. Furthermore, the synbiotic induced an immunophysiological bioregulatory effect, depending on the basal situation of each experimental group, particularly in the systemic levels of IL-1β (increased significantly only in the sedentary group), CRH (decreased significantly only in the sedentary group), and dopamine (increased significantly only in the athlete group). There were no significant differences between groups in the levels of immunoglobulin A or in the metabolic profile as a result of the intervention. It is concluded that synbiotic nutritional supplements can improve anxiety, stress, and sleep quality, particularly in sportspeople, which appears to be linked to an improved immuno-neuroendocrine response in which IL-1β, CRH, and dopamine are clearly involved.
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The SARS-CoV-2 Ivermectin Navarra-ISGlobal Trial (SAINT) to Evaluate the Potential of Ivermectin to Reduce COVID-19 Transmission in low risk, non-severe COVID-19 patients in the first 48 hours after symptoms onset: A structured summary of a study protocol for a randomized control pilot trial.
Chaccour, C, Ruiz-Castillo, P, Richardson, MA, Moncunill, G, Casellas, A, Carmona-Torre, F, Giráldez, M, Mota, JS, Yuste, JR, Azanza, JR, et al
Trials. 2020;(1):498
Abstract
OBJECTIVES The primary objective is to determine the efficacy of a single dose of ivermectin, administered to low risk, non-severe COVID-19 patients in the first 48 hours after symptom onset to reduce the proportion of patients with detectable SARS-CoV-2 RNA by Polymerase Chain Reaction (PCR) test from nasopharyngeal swab at day 7 post-treatment. The secondary objectives are: 1.To assess the efficacy of ivermectin to reduce the SARS-CoV-2 viral load in the nasopharyngeal swab at day 7 post treatment.2.To assess the efficacy of ivermectin to improve symptom progression in treated patients.3.To assess the proportion of seroconversions in treated patients at day 21.4.To assess the safety of ivermectin at the proposed dose.5.To determine the magnitude of immune response against SARS-CoV-2.6.To assess the early kinetics of immunity against SARS-CoV-2. TRIAL DESIGN SAINT is a single centre, double-blind, randomized, placebo-controlled, superiority trial with two parallel arms. Participants will be randomized to receive a single dose of 400 μg/kg ivermectin or placebo, and the number of patients in the treatment and placebo groups will be the same (1:1 ratio). PARTICIPANTS The population for the study will be patients with a positive nasopharyngeal swab PCR test for SARS-CoV-2, with non-severe COVID-19 disease, and no risk factors for progression to severity. Vulnerable populations such as pregnant women, minors (i.e.; under 18 years old), and seniors (i.e.; over 60 years old) will be excluded. Inclusion criteria 1. Patients diagnosed with COVID-19 in the emergency room of the Clínica Universidad de Navarra (CUN) with a positive SARS-CoV-2 PCR. 2. Residents of the Pamplona basin ("Cuenca de Pamplona"). 3. The patient must be between the ages of 18 and 60 years of age. 4. Negative pregnancy test for women of child bearing age*. 5. The patient or his/her representative, has given informed consent to participate in the study. 6. The patient should, in the PI's opinion, be able to comply with all the requirements of the clinical trial (including home follow up during isolation). Exclusion criteria 1. Known history of ivermectin allergy. 2. Hypersensitivity to any component of ivermectin. 3. COVID-19 pneumonia. Diagnosed by the attending physician.Identified in a chest X-ray. 4. Fever or cough present for more than 48 hours. 5. Positive IgG against SARS-CoV-2 by rapid diagnostic test. 6. Age under 18 or over 60 years. 7. The following co-morbidities (or any other disease that might interfere with the study in the eyes of the PI): Immunosuppression.Chronic Obstructive Pulmonary Disease.Diabetes.Hypertension.Obesity.Acute or chronic renal failure.History of coronary disease.History of cerebrovascular disease.Current neoplasm. 8. Recent travel history to countries that are endemic for Loa loa (Angola, Cameroon, Central African Republic, Chad, Democratic Republic of Congo, Ethiopia, Equatorial, Guinea, Gabon, Republic of Congo, Nigeria and Sudan). 9. Current use of CYP 3A4 or P-gp inhibitor drugs such as quinidine, amiodarone, diltiazem, spironolactone, verapamil, clarithromycin, erythromycin, itraconazole, ketoconazole, cyclosporine, tacrolimus, indinavir, ritonavir or cobicistat. Use of critical CYP3A4 substrate drugs such as warfarin. *Women of child bearing age may participate if they use a safe contraceptive method for the entire period of the study and at least one month afterwards. A woman is considered to not have childbearing capacity if she is post-menopausal (minimum of 2 years without menstruation) or has undergone surgical sterilization (at least one month before the study). The trial is currently planned at a single center, Clínica Universidad de Navarra, in Navarra (Spain), and the immunology samples will be analyzed at the Barcelona Institute for Global Health (ISGlobal), in Barcelona (Spain). Participants will be recruited by the investigators at the emergency room and/or COVID-19 area of the CUN. They will remain in the trial for a period of 28 days at their homes since they will be patients with mild disease. In the interest of public health and to contain transmission of infection, follow-up visits will be conducted in the participant's home by a clinical trial team comprising nursing and medical members. Home visits will assess clinical and laboratory parameters of the patients. INTERVENTION AND COMPARATOR Ivermectin will be administered to the treatment group at a 400μg/Kg dose (included in the EU approved label of Stromectol and Scabioral). The control group will receive placebo. There is no current data on the efficacy of ivermectin against the virus in vivo, therefore the use of placebo in the control group is ethically justified. MAIN OUTCOMES Primary Proportion of patients with a positive SARS-CoV-2 PCR from a nasopharyngeal swab at day 7 post-treatment. Secondary 1.Mean viral load as determined by PCR cycle threshold (Ct) at baseline and on days 4, 7, 14, and 21.2.Proportion of patients with fever and cough at days 4, 7, 14, and 21 as well as proportion of patients progressing to severe disease or death during the trial.3.Proportion of patients with seroconversion at day 21.4.Proportion of drug-related adverse events during the trial.5.Median levels of IgG, IgM, IgA measured by Luminex, frequencies of innate and SARS-CoV-2-specific T cells assessed by flow cytometry, median levels of inflammatory and activation markers measured by Luminex and transcriptomics.6.Median kinetics of IgG, IgM, IgA levels during the trial, until day 28. RANDOMISATION Eligible patients will be allocated in a 1:1 ratio using a randomization list generated by the trial statistician using blocks of four to ensure balance between the groups. A study identification code with the format "SAINT-##" (##: from 01 to 24) will be generated using a sequence of random numbers so that the randomization number does not match the subject identifier. The sequence and code used will be kept in an encrypted file accessible only to the trial statistician. A physical copy will be kept in a locked cabinet at the CUN, accessible only to the person administering the drug who will not enrol or attend to patient care. A separate set of 24 envelopes for emergency unblinding will be kept in the study file. BLINDING (MASKING): The clinical trial team and the patients will be blinded. The placebo will not be visibly identical, but it will be administered by staff not involved in the clinical care or participant follow up. NUMBERS TO BE RANDOMISED (SAMPLE SIZE): The sample size is 24 patients: 12 participants will be randomised to the treatment group and 12 participants to the control group. TRIAL STATUS Current protocol version: 1.0 dated 16 of April 2020. Recruitment is envisioned to begin by May 14th and end by June 14th. TRIAL REGISTRATION EudraCT number: 2020-001474-29, registered April 1st. Clinicaltrials.gov: submitted, pending number FULL PROTOCOL The full protocol is attached as an additional file, accessible from the Trials website (Additional file 1). In the interest in expediting dissemination of this material, the familiar formatting has been eliminated; this Letter serves as a summary of the key elements of the full protocol.
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Determination of Ligand Profiles for Pseudomonas aeruginosa Solute Binding Proteins.
Fernández, M, Rico-Jiménez, M, Ortega, Á, Daddaoua, A, García García, AI, Martín-Mora, D, Torres, NM, Tajuelo, A, Matilla, MA, Krell, T
International journal of molecular sciences. 2019;(20)
Abstract
Solute binding proteins (SBPs) form a heterogeneous protein family that is found in all kingdoms of life. In bacteria, the ligand-loaded forms bind to transmembrane transporters providing the substrate. We present here the SBP repertoire of Pseudomonas aeruginosa PAO1 that is composed of 98 proteins. Bioinformatic predictions indicate that many of these proteins have a redundant ligand profile such as 27 SBPs for proteinogenic amino acids, 13 proteins for spermidine/putrescine, or 9 proteins for quaternary amines. To assess the precision of these bioinformatic predictions, we have purified 17 SBPs that were subsequently submitted to high-throughput ligand screening approaches followed by isothermal titration calorimetry studies, resulting in the identification of ligands for 15 of them. Experimentation revealed that PA0222 was specific for γ-aminobutyrate (GABA), DppA2 for tripeptides, DppA3 for dipeptides, CysP for thiosulphate, OpuCC for betaine, and AotJ for arginine. Furthermore, RbsB bound D-ribose and D-allose, ModA bound molybdate, tungstate, and chromate, whereas AatJ recognized aspartate and glutamate. The majority of experimentally identified ligands were found to be chemoattractants. Data show that the ligand class recognized by SPBs can be predicted with confidence using bioinformatic methods, but experimental work is necessary to identify the precise ligand profile.
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Myocardin-Dependent Kv1.5 Channel Expression Prevents Phenotypic Modulation of Human Vessels in Organ Culture.
Arévalo-Martínez, M, Cidad, P, García-Mateo, N, Moreno-Estar, S, Serna, J, Fernández, M, Swärd, K, Simarro, M, de la Fuente, MA, López-López, JR, et al
Arteriosclerosis, thrombosis, and vascular biology. 2019;(12):e273-e286
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Abstract
OBJECTIVE We have previously described that changes in the expression of Kv channels associate to phenotypic modulation (PM), so that Kv1.3/Kv1.5 ratio is a landmark of vascular smooth muscle cells phenotype. Moreover, we demonstrated that the Kv1.3 functional expression is relevant for PM in several types of vascular lesions. Here, we explore the efficacy of Kv1.3 inhibition for the prevention of remodeling in human vessels, and the mechanisms linking the switch in Kv1.3 /Kv1.5 ratio to PM. Approach and Results: Vascular remodeling was explored using organ culture and primary cultures of vascular smooth muscle cells obtained from human vessels. We studied the effects of Kv1.3 inhibition on serum-induced remodeling, as well as the impact of viral vector-mediated overexpression of Kv channels or myocardin knock-down. Kv1.3 blockade prevented remodeling by inhibiting proliferation, migration, and extracellular matrix secretion. PM activated Kv1.3 via downregulation of Kv1.5. Hence, both Kv1.3 blockers and Kv1.5 overexpression inhibited remodeling in a nonadditive fashion. Finally, myocardin knock-down induced vessel remodeling and Kv1.5 downregulation and myocardin overexpression increased Kv1.5, while Kv1.5 overexpression inhibited PM without changing myocardin expression. CONCLUSIONS We demonstrate that Kv1.5 channel gene is a myocardin-regulated, vascular smooth muscle cells contractile marker. Kv1.5 downregulation upon PM leaves Kv1.3 as the dominant Kv1 channel expressed in dedifferentiated cells. We demonstrated that the inhibition of Kv1.3 channel function with selective blockers or by preventing Kv1.5 downregulation can represent an effective, novel strategy for the prevention of intimal hyperplasia and restenosis of the human vessels used for coronary angioplasty procedures.
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Prevalence of eosinophilic esophagitis: A multicenter study on a pediatric population evaluated at thirty-six Latin American gastroenterology centers.
Pierre, R, Vieira, M, Vázquez, R, Ninomiya, I, Messere, G, Daza, W, Dadan, S, Higuera, M, Sifontes, L, Harris, P, et al
Revista de gastroenterologia de Mexico (English). 2019;(4):427-433
Abstract
INTRODUCTION AND OBJECTIVE Eosinophilic esophagitis is a chronic, immune-mediated disease described in case series and publications worldwide. Over the past twenty years, the authors of different studies have attempted to evaluate its incidence and prevalence. The objetive of the present study was to estimate the prevalence of eosinophilic esophagitis in a group of children seen at 36 pediatric gastroenterology centers in ten Latin American countries. MATERIALS AND METHODS A multicenter, observational, and cross-sectional study was conducted that estimated the period prevalence of eosinophilic esophagitis in children seen at outpatient consultation and that underwent diagnostic upper gastrointestinal endoscopy for any indication at 36 centers in 10 Latin American countries, within a 3-month time frame. RESULTS Between April and June 2016, 108 cases of eosinophilic esophagitis were evaluated. Likewise, an average of 29,253 outpatient consultations and 4,152 diagnostic upper gastrointestinal endoscopies were carried out at the 36 participating centers. The period prevalence of eosinophilic esophagitis in the population studied (n=29,253) was 3.69 cases×1,000 (95% CI: 3.04 to 4.44), and among the children that underwent routine upper gastrointestinal endoscopy (n=4,152), it was 26x1,000 (95% CI: 22.6 to 29.4). CONCLUSIONS The general period prevalence of eosinophilic esophagitis in a group of children evaluated at 36 Latin American pediatric gastroenterology centers was 3.69×1,000, and in the children that underwent endoscopy, it was 26×1,000. There was important prevalence variability between the participating countries and centers. The present analysis is the first study conducted on the prevalence of pediatric eosinophilic esophagitis in Latin America.
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Enterococcus faecalis Bacteriophage 156 Is an Effective Biotechnological Tool for Reducing the Presence of Tyramine and Putrescine in an Experimental Cheese Model.
Del Rio, B, Sánchez-Llana, E, Redruello, B, Magadan, AH, Fernández, M, Martin, MC, Ladero, V, Alvarez, MA
Frontiers in microbiology. 2019;:566
Abstract
Biogenic amines (BA) - nitrogenous compounds of low molecular weight - are the result of metabolism of certain amino acids. They are biologically present in all living organisms and play essential physiological roles. However, their accumulation in foodstuffs due to the metabolic activity of certain microorganisms represents a toxicological risk. Containing such microorganisms, and with an abundance of precursor substrate amino acids, fermented foods in general, and cheeses in particular, provide an ideal matrix for the accumulation of these toxic compounds. Unfortunately, the main microorganisms responsible for BA accumulation are members of the lactic acid bacteria (LAB) group, which are also essential for the development of the organoleptic characteristics of the final product. The methods used to reduce the BA content of cheese, such as milk pasteurization, commonly fail to do so, and affect desirable non-BA-producing LAB as well. Bacteriophages have been proposed as biotechnological tools for diminishing the presence of undesirable microorganisms in dairy products. Given their specificity, they could be used to target the population of BA-producing bacteria. In this work, we aimed to explore the use of Enterococcus faecalis infecting phages as a tool to reduce the content of BA in dairy products. For this, we proceeded to the isolation and characterization of E. faecalis bacteriophage 156, a member of the family Myoviridae. Its genome was sequenced and compared with that of E. faecalis family Myoviridae phages available in public databases. Its capacity to decrease the accumulation of the BA tyramine and putrescine in an experimental laboratory-scale cheese model was proven.
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Functional Annotation of Bacterial Signal Transduction Systems: Progress and Challenges.
Martín-Mora, D, Fernández, M, Velando, F, Ortega, Á, Gavira, JA, Matilla, MA, Krell, T
International journal of molecular sciences. 2018;(12)
Abstract
Bacteria possess a large number of signal transduction systems that sense and respond to different environmental cues. Most frequently these are transcriptional regulators, two-component systems and chemosensory pathways. A major bottleneck in the field of signal transduction is the lack of information on signal molecules that modulate the activity of the large majority of these systems. We review here the progress made in the functional annotation of sensor proteins using high-throughput ligand screening approaches of purified sensor proteins or individual ligand binding domains. In these assays, the alteration in protein thermal stability following ligand binding is monitored using Differential Scanning Fluorimetry. We illustrate on several examples how the identification of the sensor protein ligand has facilitated the elucidation of the molecular mechanism of the regulatory process. We will also discuss the use of virtual ligand screening approaches to identify sensor protein ligands. Both approaches have been successfully applied to functionally annotate a significant number of bacterial sensor proteins but can also be used to study proteins from other kingdoms. The major challenge consists in the study of sensor proteins that do not recognize signal molecules directly, but that are activated by signal molecule-loaded binding proteins.
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High-Throughput Screening to Identify Chemoreceptor Ligands.
Fernández, M, Ortega, Á, Rico-Jiménez, M, Martín-Mora, D, Daddaoua, A, Matilla, MA, Krell, T
Methods in molecular biology (Clifton, N.J.). 2018;:291-301
Abstract
The majority of bacterial chemoreceptors remain functionally un-annotated. The knowledge of chemoreceptor function, however, is indispensable to understanding the evolution of the chemotaxis system in bacteria with different lifestyles. Significant progress in the annotation of chemoreceptor function has been made using experimental strategies that are based on the individual, genetically engineered ligand binding domain (LBD) of chemoreceptors. There is now evidence that all major classes of LBDs can be produced as individual domains that retain their ligand binding activity. Here, we provide a protocol for the combined use of high-throughput ligand screening using Differential Scanning Fluorimetry followed by Isothermal Titration Calorimetry to identify and characterize ligands that bind to recombinant chemoreceptor LBDs. This approach has been shown to be very efficient for determining the function of novel chemoreceptors.