Biomarcadores de respuesta al interferón beta en la esclerosis múltiple

La esclerosis múltiple (EM) es una enfermedad crónica del sistema nervioso central (SNC) que se caracteriza por presentar desmielinización, inflamación y daño axonal. El Interferón beta (IFN-beta) fue el primer fármaco inmunomodulador usado en el tratamiento de la esclerosis múltiple remitente-recidivante (EMRR). Es un fármaco seguro y eficaz, capaz de disminuir la actividad de la enfermedad. Sin embargo, la respuesta a este tratamiento no es uniforme en todos los pacientes de EM. Por ello, dado el gran abanico de opciones terapéuticas que existen en la actualidad, la búsqueda de biomarcadores que permitan identificar de manera precoz a los pacientes con alta probabilidad de mostrar una respuesta óptima al interferón beta es de gran relevancia para poder realizar una medicina personalizada en la EM. Hasta ahora son los marcadores clínicos y radiológicos los que más se utilizan para establecer la eficacia de las distintas opciones terapéuticas. Sin embargo, esto implica un cierto deterioro para los pacientes no respondedores que puede repetirse con distintas terapias. Objetivos El primer objetivo del estudio fue identificar antes del inicio del tratamiento, biomarcadores biológicos que predigan la respuesta a IFN-beta. En segundo lugar comparamos los niveles de todos los posibles marcadores incluidos en el estudio con los de los controles sanos. Nuestro último objetivo fue analizar los cambios que el tratamiento podría inducir en todos los factores estudiados..

Public Cord Blood Banks as a source of starting material for clinical grade HLA-homozygous induced pluripotent stem cells

Background The increasing number of clinical trials for induced pluripotent stem cell (iPSC)-derived cell therapy products makes the production on clinical grade iPSC more and more relevant and necessary. Cord blood banks are an ideal source of young, HLA-typed and virus screened starting material to produce HLA-homozygous iPSC lines for wide immune-compatibility allogenic cell therapy approaches. The production of such clinical grade iPSC lines (haplolines) involves particular attention to all steps since donor informed consent, cell procurement and a GMP-compliant cell isolation process. Methods Homozygous cord blood units were identified and quality verified before recontacting donors for informed consent. CD34+ cells were purified from the mononuclear fraction isolated in a cell processor, by magnetic microbeads labelling and separation columns. Results We obtained a median recovery of 20.0% of the collected pre-freezing CD34+, with a final product median viability of 99.1% and median purity of 83.5% of the post-thawed purified CD34+ population. Conclusions Here we describe our own experience, from unit selection and donor reconsenting, in generating a CD34+ cell product as a starting material to produce HLA-homozygous iPSC following a cost-effective and clinical grade-compliant procedure. These CD34+ cells are the basis for the Spanish bank of haplolines envisioned to serve as a source of cell products for clinical research and therapy

Evaluation of the Spanish population coverage of a prospective HLA haplobank of induced pluripotent stem cells

Background: iPSC (induced pluripotent stem cells) banks of iPSC lines with homozygous HLA (human leukocyte antigen) haplotypes (haplobanks) are proposed as an affordable and off-the-shelf approach to allogeneic transplantation of iPSC derived cell therapies. Cord blood banks offer an extensive source of HLA-typed cells suitable for reprogramming to iPSC. Several initiatives worldwide have been undertaken to create national and international iPSC haplobanks that match a significant part of a population. Methods: To create an iPSC haplobank that serves the Spanish population (IPS-PANIA), we have searched the Spanish Bone Marrow Donor Registry (REDMO) to identify the most frequently estimated haplotypes. From the top ten donors identified, we estimated the population coverage using the criteria of zero mismatches in HLA-A, HLA-B, and HLA-DRB1 with different stringencies: high resolution, low resolution, and beneficial mismatch. Results: We have calculated that ten cord blood units from homozygous donors stored at the Spanish cord blood banks can provide HLA-A, HLA-B, and HLA-DRB1 matching for 28.23% of the population. Conclusion: We confirm the feasibility of using banked cord blood units to create an iPSC haplobank that will cover a significant percentage of the Spanish and international population for future advanced therapy replacement strategies

Translocated LPS Might Cause Endotoxin Tolerance in Circulating Monocytes of Cystic Fibrosis Patients

Cystic Fibrosis (CF) is an inherited pleiotropic disease that results from abnormalities in the gene codes of a chloride channel. The lungs of CF patients are chronically infected by several pathogens but bacteraemia have rarely been reported in this pathology. Besides that, circulating monocytes in CF patients exhibit a patent Endotoxin Tolerance (ET) state since they show a significant reduction of the inflammatory response to bacterial stimulus. Despite a previous description of this phenomenon, the direct cause of ET in CF patients remains unknown. In this study we have researched the possible role of microbial/endotoxin translocation from a localized infection to the bloodstream as a potential cause of ET induction in CF patients. Plasma analysis of fourteen CF patients revealed high levels of LPS compared to healthy volunteers and patients who suffer from Chronic Obstructive Pulmonary Disease. Experiments in vitro showed that endotoxin concentrations found in plasma of CF patients were enough to induce an ET phenotype in monocytes from healthy controls. In agreement with clinical data, we failed to detect bacterial DNA in CF plasma. Our results suggest that soluble endotoxin present in bloodstream of CF patients causes endotoxin tolerance in their circulating monocytes

Mitigation of greenhouse gases in dairy cattle via genetic selection: 1. Genetic parameters of direct methane using noninvasive methods and proxies of methane

11 Pág.Records of methane emissions from 1,501 cows on 14 commercial farms in 4 regions of Spain were collected from May 2018 to June 2019. Methane concentrations (MeC) were measured using a nondispersive infrared methane detector installed within the feed bin of the automatic milking system during 14- to 21-d periods. Rumination time (RT; min/d) was collected using collars with a tag that registered time (minutes) spent eating and ruminating. The means of MeC and methane production (MeP) were 1,254.28 ppm and 182.49 g/d, respectively; mean RT was 473.38 min/d. Variance components for MeC, MeP, and RT were estimated with REML using pedigree and genomic information in a single-step model. Heritabilities for MeC and MeP were 0.11 and 0.12, respectively. Rumination time showed a slightly larger heritability estimate (0.17). The genetic correlation between MeP and MeC was high (>0.95), suggesting that selection on either trait would lead to a positive correlated response on the other. Negative correlations were estimated between RT and MeC (-0.24 ± 0.38) and MeP (-0.43 ± 0.35). Methane concentration and MeP had slightly positive correlations with milk yield (0.17 ± 0.39 and 0.21 ± 0.36), protein percentage (0.08 ± 0.32 and 0.30 ± 0.45), protein yield (0.22 ± 0.41 and 0.31 ± 0.35), fat percentage (0.02 ± 0.40 and 0.27 ± 0.36), and fat yield (0.27 ± 0.28 and 0.29 ± 0.28) from bivariate analyses. Rumination time had positive correlations with milk yield (0.41 ± 0.75) and protein yield (0.26 ± 0.57) and negative correlations with fat yield (-0.45 ± 0.32), protein percentage (-0.15 ± 0.38), and fat percentage (-0.40 ± 0.47). A positive approximated genetic correlation was estimated between fertility and MeC (0.10 ± 0.05) and MeP (0.18 ± 0.05), resulting in slightly higher CH4 production when selecting for better fertility [days open estimated breeding values (EBV) are expressed with mean 100 and SD 10, inversely related to days from calving to conception; that is, greater days open EBV implies better fertility]. Positive correlations were also estimated for stature with MeC and MeP (0.30 ± 0.04 and 0.43 ± 0.04, respectively). Other type traits (chest width, udder depth, angularity, and capacity) were positively correlated with methane traits, possibly because of higher milk yield and higher feed intake from these animals. Rumination time showed positive EBV correlations with production traits and type traits, and negative correlations with somatic cell count and body condition score. Based on the genetic correlations and heritabilities estimated in this study, methane is measurable and heritable, and estimates of genetic correlations suggest no strong opposition to current breeding objectives in Spanish Holsteins.This research was funded by RTA2015-00022-C03 (METALGEN) project from the Spanish National Plan of Research, development and innovation 2013-2020 and the Department of Economic Development and Competitiveness (Basque Government, Vitoria-Gasteiz, Spain). Farmers and breeders' associations (EFRIFE, AFNA, GIFE, AFCA) are acknowledge for data sharing and infrastructure. Data were analyzed using the CESGA Supercomputing infrastructure (Galicia, Spain). The editor and three reviewers are acknowledged for thorough and insightful comments. The authors have not stated any conflicts of interest.Peer reviewe

Mitigation of greenhouse gases in dairy cattle via genetic selection: 1. Genetic parameters of direct methane using noninvasive methods and proxies of methane

11 Pág.Records of methane emissions from 1,501 cows on 14 commercial farms in 4 regions of Spain were collected from May 2018 to June 2019. Methane concentrations (MeC) were measured using a nondispersive infrared methane detector installed within the feed bin of the automatic milking system during 14- to 21-d periods. Rumination time (RT; min/d) was collected using collars with a tag that registered time (minutes) spent eating and ruminating. The means of MeC and methane production (MeP) were 1,254.28 ppm and 182.49 g/d, respectively; mean RT was 473.38 min/d. Variance components for MeC, MeP, and RT were estimated with REML using pedigree and genomic information in a single-step model. Heritabilities for MeC and MeP were 0.11 and 0.12, respectively. Rumination time showed a slightly larger heritability estimate (0.17). The genetic correlation between MeP and MeC was high (>0.95), suggesting that selection on either trait would lead to a positive correlated response on the other. Negative correlations were estimated between RT and MeC (-0.24 ± 0.38) and MeP (-0.43 ± 0.35). Methane concentration and MeP had slightly positive correlations with milk yield (0.17 ± 0.39 and 0.21 ± 0.36), protein percentage (0.08 ± 0.32 and 0.30 ± 0.45), protein yield (0.22 ± 0.41 and 0.31 ± 0.35), fat percentage (0.02 ± 0.40 and 0.27 ± 0.36), and fat yield (0.27 ± 0.28 and 0.29 ± 0.28) from bivariate analyses. Rumination time had positive correlations with milk yield (0.41 ± 0.75) and protein yield (0.26 ± 0.57) and negative correlations with fat yield (-0.45 ± 0.32), protein percentage (-0.15 ± 0.38), and fat percentage (-0.40 ± 0.47). A positive approximated genetic correlation was estimated between fertility and MeC (0.10 ± 0.05) and MeP (0.18 ± 0.05), resulting in slightly higher CH4 production when selecting for better fertility [days open estimated breeding values (EBV) are expressed with mean 100 and SD 10, inversely related to days from calving to conception; that is, greater days open EBV implies better fertility]. Positive correlations were also estimated for stature with MeC and MeP (0.30 ± 0.04 and 0.43 ± 0.04, respectively). Other type traits (chest width, udder depth, angularity, and capacity) were positively correlated with methane traits, possibly because of higher milk yield and higher feed intake from these animals. Rumination time showed positive EBV correlations with production traits and type traits, and negative correlations with somatic cell count and body condition score. Based on the genetic correlations and heritabilities estimated in this study, methane is measurable and heritable, and estimates of genetic correlations suggest no strong opposition to current breeding objectives in Spanish Holsteins.This research was funded by RTA2015-00022-C03 (METALGEN) project from the Spanish National Plan of Research, development and innovation 2013-2020 and the Department of Economic Development and Competitiveness (Basque Government, Vitoria-Gasteiz, Spain). Farmers and breeders' associations (EFRIFE, AFNA, GIFE, AFCA) are acknowledge for data sharing and infrastructure. Data were analyzed using the CESGA Supercomputing infrastructure (Galicia, Spain). The editor and three reviewers are acknowledged for thorough and insightful comments. The authors have not stated any conflicts of interest.Peer reviewe

Structural equation models to disentangle the biological relationship between microbiota and complex traits: Methane production in dairy cattle as a case of study

The advent of metagenomics in animal breeding poses the challenge of statistically modelling the relationship between the microbiome, the host genetics and relevant complex traits. A set of structural equation models (SEMs) of a recursive type within a Markov chain Monte Carlo (MCMC) framework was proposed here to jointly analyse the host–metagenome–phenotype relationship. A non‐recursive bivariate model was set as benchmark to compare the recursive model. The relative abundance of rumen microbes (RA), methane concentration (CH4) and the host genetics was used as a case of study. Data were from 337 Holstein cows from 12 herds in the north and north‐west of Spain. Microbial composition from each cow was obtained from whole metagenome sequencing of ruminal content samples using a MinION device from Oxford Nanopore Technologies. Methane concentration was measured with Guardian® NG infrared gas monitor from Edinburgh Sensors during cow's visits to the milking automated system. A quarterly average from the methane eructation peaks for each cow was computed and used as phenotype for CH4. Heritability of CH4 was estimated at 0.12 ± 0.01 in both the recursive and bivariate models. Likewise, heritability estimates for the relative abundance of the taxa overlapped between models and ranged between 0.08 and 0.48. Genetic correlations between the microbial composition and CH4 ranged from −0.76 to 0.65 in the non‐recursive bivariate model and from −0.68 to 0.69 in the recursive model. Regardless of the statistical model used, positive genetic correlations with methane were estimated consistently for the seven genera pertaining to the Ciliophora phylum, as well as for those genera belonging to the Euryarchaeota (Methanobrevibacter sp.), Chytridiomycota (Neocallimastix sp.) and Fibrobacteres (Fibrobacter sp.) phyla. These results suggest that rumen's whole metagenome recursively regulates methane emissions in dairy cows and that both CH4 and the microbiota compositions are partially controlled by the host genotype.National plan of research, development and innovation 2013‐2020, Grant/ Award Number: RTA2015‐0022‐CO3 (METALGEN)UCR::Vicerrectoría de Docencia::Ciencias Agroalimentarias::Facultad de Ciencias Agroalimentarias::Escuela de ZootecniaUCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias Agroalimentarias::Centro de Investigación en Nutrición Animal (CINA

High-Risk Sexual Practices Contribute to HIV-1 Double Infection Among Men Who Have Sex with Men in Madrid.

Data on the prevalence of double infection (DI) in HIV individuals are lacking in Spain. To fill this gap, we analyzed the prevalence of DI in a cohort of men who have sex with men (MSM) and examined factors contributing to DI. We selected 81 MSM attending Centro Sanitario Sandoval, a sexually transmitted diseases clinic in Madrid. We obtained by ultra-deep sequencing the proviral sequences in gag and env genes and performed a phylogenetic analysis for the identification of DI. Clinical, behavioral, host, and viral factors were studied for its association with DI. We detected six individuals with DI and one case of superinfection with a global prevalence of 8.6%. The genetic distance among the subtype B viruses in monoinfected individuals (24.4%) was lower than the distance between the two viruses in subtype B DI individuals (29.5%). Individuals with a high number of sexual contacts (>25 partners/year) had an 8.66 times higher risk of DI (p = .017). In this MSM cohort the prevalence of HIV DI was estimated at 8.6%. DI was strongly associated with the number of sexual partners. Because of the pathogenic consequences of HIV DI, this high prevalence should promote public health programs targeted at high-risk population such as MSM for the control of HIV infection and DI. HIV DI should be considered for a better clinical management of these individuals.Work in Centro Nacional de Microbiologia (Instituto de Salud Carlos III) was supported by grant SAF-2016-77894-R from Ministerio de Economia y Competitividad (MINECO, Spain), grant PI 13/02269 from Fondo de Investigación Sanitaria (FIS-Instituto de Salud Carlos III, Spain) and in part by the RIS-RETIC grants RD12/0017/0028 and RD16CIII/0002/0005 funded by the Intituto de Salud Carlos III-European Regional Development Fund. M.P. has a contract of RIS-RETIC RD16CIII/0002/0005.S

Circulating endotoxin levels correlates with the refractory state in CF patients.

<p>(<i>A</i>) Levels of LPS were tested by LAL kit endpoint-QCL1000 (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0029577#s4" target="_blank">Materials and Methods</a>) in plasma from controls (HC, n = 11), CF (n = 14), COPD (n = 8) and sepsis (n = 8) patients (*, <i>p</i><0.01). Circulating monocytes from healthy controls (HC, n = 11), CF patients (CF, n = 14), COPD patients (n = 8) and sepsis patients (n = 8) were isolated and cultured in presence of 10 ng/mL of LPS for 24 h (<i>B</i>), and 3 h (<i>C</i> and <i>D</i>). Next, levels of TNFα production were analysed by ELISA in the supernatant of cultures (<i>B</i>) or cells were harvested and mRNA of IL23p19 and IL10 were quantified by real time Q-PCR (<i>C</i> and <i>D</i>), (<i>B–D, insets</i>). Linear regression analysis was applied in each case to obtain best linear equation with CF only (R² is given).</p