Desaturase gene expression in specific fatty acid desaturase mutants from Arabidopsis thaliana: evidence of compensatory pathways

Panel (S7-P10) presentado en la Reunión de la Sociedad Española de Fisiología Vegetal (SEFV) (18ª. Zaragoza. 8-11 septiembre 2009).Synthesis of glycerolipids in plants takes place almost exclusively in plastids and in the endoplasmic reticulum (ER) through two pathways, the “prokaryotic pathway” and the “eukaryotic pathway”. In the prokaryotic pathway, glycerolipids are synthesized entirely in plastids. On the other hand, in the eukaryotic pathway, phospholipids are synthesized in the ER while MGD, DGD and SL are synthesized in plastids from PC produced in the ER. In both glycerolipid synthetic pathways, fatty acid desaturases are key enzymes that introduce double bounds in fatty acid chains. They produce polyunsaturated fatty acids (PUFAs) which are important in maintaining membrane fluidity and function [1]. PUFAs also serve as precursors of plant hormones like jasmonates [2] that are involved in defence signalling against pathogen attack, wound response, plant development and adaptation to environmental stress. The relative amount of glycerolipid synthesized and desaturated by the two pathways may vary in different tissues and in different plant species. Arabidopsis thaliana is an example of 16:3 plants, in which both pathways are almost equally involved in total glycerolipid synthesis [3]. On the other hand, in 18:3 plants (such as Glycine max) the leading pathway is the eukaryotic one. Although several lines of evidence indicate the existence of regulatory mechanisms that coordinate the activity of both pathways for glycerolipid synthesis in plants, molecular and biochemical components of this regulatory mechanism as well as how this coordination takes place are still unknown. As a first step to understand how the communication between the chloroplast and the reticulum takes place, we have studied the changes in gene expression profiles of fatty acid desaturases in response to specific mutations in the desaturase pathway from Arabidopsis thaliana. We have analysed five different mutants, fad2; fad5; fad6; act1 and the triple mutant fad3/fad7/fad8. These mutations affected to desaturases operating in both glycerolipid synthetic pathways. The analysis of the effect of each mutation in the expression profiles of the rest of the desaturase genes has allowed us to observe transcriptional responses associated to the existence of compensatory pathways between the chloroplast and the reticulum to minimize the effects of the mutation. It has also helped us to identify the key role of the palmitoyl desaturase, fad5, in controlling the flux through the prokaryotic pathway. [1] Wallis & Browse (2002) Prog. in Lipid Res. 41: 254-278 [2] Weber et al. (1997) Proc. Natl. Acad. Sci. 94: 10473-10478 [3] Browse et al. (1986) Biochem. J. 235: 25-31This study was supported by the Spanish Ministry of Science and Education (project AGL2008-00733), , and the Aragón Government (PIP140/08).Peer reviewe

Evolution of the public problem of depopulation in Spain: longitudinal analysis of the media agenda

According to the sociology of public problems, the construction, visibility, and stabilization of a public problem require the mobilization of collectives of citizens interested in the issue, which act as an active entity in demanding actions and policies. One such issue is the depopulation of rural areas in Spain, which has shifted from a geographically localized problem to a matter of state policy. This article investigates the influence of framing in this process, analyzing a corpus of 5,980 headlines from newspapers in the Aragon, Castile-La Mancha, and Castile and Leon regions of Spain as well as from two national media outlets, corresponding to the period 2012–2021. Through the application of statistical analyses of lexical frequency in stages, the most significant terms in the evolution of the media’s coverage of the issue have been identified, which has made it possible to observe the appearance and displacement of concepts and their relationship to the most important milestones of social and political mobilization. In addition, its power to stir up sentiment in socio-political discourse has been explored. The consolidation of depopulation as a stable element in the Spanish media agenda, going beyond the regional sphere and having a presence dissociated from specific events through time, in contrast to what occurred some years ago, stands out. Finding a media-friendly frame -empty Spain [España vacía] and emptied Spain [España vaciada]- may have been a key element in this.

Targeted release of probiotics from enteric microparticulated formulations

The development of advanced probiotic delivery systems, which preserve bacteria from degradation of the gastrointestinal tract and achieve a targeted release mediated by pH-independent swelling, is of great interest to improve the eficient delivery of probiotic bacteria to the target tissue. Gram-positive and Gram-negative bacteria models (Lactobacillus acidophilus (Moro) Hansen and Mocquot (ATCC®4356™) and Escherichia coli S17, respectively) have been successfully encapsulated for the first time in pH-independent microparticulate polymethacrylates (i.e., Eudraguard biotic) used for the targeted delivery of nutraceuticals to the colon. These bacteria have also been encapsulated within the mucoadhesive polymethacrylate Eudragit RS 100 widely used as targeted release formulation for active pharmaceutical ingredients. The enteric microparticles remained unaltered under simulated gastric conditions and released the contained viable microbial cargo under simulated intestinal conditions. Buoyancies of 90.2% and 57.3% for Eudragit and Eudraguard microparticles, respectively, and long-term stability (5 months) for the encapsulated microorganisms were found. Cytotoxicity of the microparticles formulated with both polymers was evaluated (0.5-20 mg/mL) on Caco-2 cells, showing high cytocompatibility. These results underline the suitability of the synthesized materials for the successful delivery of probiotic formulations to the target organ, highlighting for the first time the potential use of Eudraguard biotic as an effective enteric coating for the targeted delivery of probiotics

Near infrared dye-labelled polymeric micro- and nanomaterials:: In vivo imaging and evaluation of their local persistence

The use of micro- and nanomaterials as carriers of therapeutic molecules can enhance the efficiency of treatments while avoiding side effects thanks to the development of controlled drug delivery systems. The binding of a dye to a drug or to a drug carrier has opened up a wide range of possibilities for an effective in vivo optical tracing of drug biodistribution by using non-invasive real-time technologies prior to their potential use as therapeutic vectors. Here, we describe the fluorescent tagging of polymeric micro- and nanomaterials based on poly(lactic-co-glycolic) acid and on the thermoresponsive poly(N-isopropylacrylamide) with the fluorescent probe IR-820 which was chemically modified for its covalent coupling to the materials. The chemical modification of the dye and the polymers yielded micro- and nanoparticulated labelled materials to be potentially used as drug depots of different therapeutic molecules. In vitro biological studies revealed their reduced cytotoxicity. A spatiotemporal in vivo micro- and nanoparticle tracking allowed the evaluation of the biodistribution of materials showing their local persistence and high biocompatibility after pathological studies. These results underline the suitability of these materials for the local, sustained, not harmful and/or on-demand drug delivery and the remarkable importance of evaluating the biodistribution of materials and tissue persistence for their use as local drug depots

The effect of hollow gold nanoparticles on stem cell migration. Potencial application in tissue regeneration.

Every year trauma together with bone, joints and cartilage-associated diseases usually involve structural damage, resulting in a severe pain and disability for millions of people worldwide[1]. In regenerative medicine, cellular, tissue and organ-based approaches are developed to restore biological functions that have been lost[2],[3]. Therefore, tissue repair and regenerative medicine have attracted the interest of the scientific community, providing promising results in preclinical models and clinical pilot studies.pre-print3341 K

Matryoshka-type gastro-resistant microparticles for the oral treatment of Mycobacterium tuberculosis

Aim: Production of Matryoshka-type gastroresistant microparticles containing antibiotic-loaded poly lactic-co-glycolic acid (PLGA) nanoparticles (NP) against Mycobacterium tuberculosis. Materials & methods: The emulsification and evaporation methods were followed for the synthesis of PLGA–NPs and methacrylic acid-ethyl acrylate-based coatings to protect rifampicin from degradation under simulated gastric conditions. Results & conclusion: The inner antibiotic-loaded NPs here reported can be released under simulated intestinal conditions whereas their coating protects them from degradation under simulated gastric conditions. The encapsulation does not hinder the antituberculosis action of the encapsulated antibiotic rifampicin. A sustained antibiotic release could be obtained when using the drug-loaded encapsulated NPs. Compared with the administration of the free drug, a more effective elimination of M. tuberculosis was observed when applying the NPs against infected macrophages. The antibiotic-loaded PLGA–NPs were also able to cross an in vitro model of intestinal barrier. Matryoshka-type gastroresistant microparticles containing antibiotic-loaded poly lactic-co-glycolic acid nanoparticles against M. tuberculosis were produced to protect the antibiotic from degradation under simulated gastric conditions. The antibiotic-loaded poly lactic-co-glycolic acid nanoparticles were able to cross an in vitro model of intestinal barrier, being more effective in the elimination of M. tuberculosis when applied against infected macrophages compared with the use of the free drug

Cymantrenyl-nucleobases: synthesis, anticancer, antitrypanosomal and antimicrobial activity studies

The synthesis of four cymantrene-5-fluorouracil derivatives (1-4) and two cymantreneadenine derivatives (5 and 6) is reported. All compounds were characterized by spectroscopic methods and the crystal structure of two derivatives (1 and 6) together with the previously described cymantrene-adenine compound C was determined by X-ray crystallography. While the compounds 1 and 6 crystallized in the triclinic P-1 space group, compound C crystallized in the monoclinic P21/m space group. The newly synthesized compounds 1-6 were tested together with the two previously described cymantrene derivatives B and C for their in vitro antiproliferative activity against seven cancer cell lines (MCF-7, MCF-7/DX, MDA-MB-231, SKOV-3, A549, HepG2 and U-87-MG), three bacterial strains (Staphylococcus aureus (methicillin-sensitive and methicillin-resistant strains), Staphylococcus epidermidis and Escherichia coli, including clinical isolates of S. aureus and S. epidermidis, as well as against the protozoan parasite Trypanosoma brucei. The most cytotoxic compounds were derivatives 2 and C for A549 and SKOV-3 cancer cell lines, respectively, with 50% growth inhibition (IC50) values of about 7 µM. The anticancer activity of the cymantrene compounds was determined to be due to their ability to induce oxidative stress and to trigger apoptosis and autophagy in cancer cells. Three derivatives (1, 4 and 5) displayed promising antitrypanosomal activity with GI50 values in the low micromolar range (3-4 µM). The introduction of the 5-fluorouracil moiety in 1 enhanced the trypanocidal activity compared to the activity previously reported for the corresponding uracil derivative. The antibacterial activity of cymantrene compounds 1 and C was within the range of 8-64 µg/mL and seemed to be the result of induced cell shrinking

A genome-wide association study follow-up suggests a possible role for PPARG in systemic sclerosis susceptibility

Introduction: A recent genome-wide association study (GWAS) comprising a French cohort of systemic sclerosis (SSc) reported several non-HLA single-nucleotide polymorphisms (SNPs) showing a nominal association in the discovery phase. We aimed to identify previously overlooked susceptibility variants by using a follow-up strategy.<p></p> Methods: Sixty-six non-HLA SNPs showing a P value <10-4 in the discovery phase of the French SSc GWAS were analyzed in the first step of this study, performing a meta-analysis that combined data from the two published SSc GWASs. A total of 2,921 SSc patients and 6,963 healthy controls were included in this first phase. Two SNPs, PPARG rs310746 and CHRNA9 rs6832151, were selected for genotyping in the replication cohort (1,068 SSc patients and 6,762 healthy controls) based on the results of the first step. Genotyping was performed by using TaqMan SNP genotyping assays. Results: We observed nominal associations for both PPARG rs310746 (PMH = 1.90 × 10-6, OR, 1.28) and CHRNA9 rs6832151 (PMH = 4.30 × 10-6, OR, 1.17) genetic variants with SSc in the first step of our study. In the replication phase, we observed a trend of association for PPARG rs310746 (P value = 0.066; OR, 1.17). The combined overall Mantel-Haenszel meta-analysis of all the cohorts included in the present study revealed that PPARG rs310746 remained associated with SSc with a nominal non-genome-wide significant P value (PMH = 5.00 × 10-7; OR, 1.25). No evidence of association was observed for CHRNA9 rs6832151 either in the replication phase or in the overall pooled analysis.<p></p> Conclusion: Our results suggest a role of PPARG gene in the development of SSc

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

Cross-disease Meta-analysis of Genome-wide Association Studies for Systemic Sclerosis and Rheumatoid Arthritis Reveals IRF4 as a New Common Susceptibility Locus

Objectives: Systemic sclerosis (SSc) and rheumatoid arthritis (RA) are autoimmune diseases that share clinical and immunological characteristics. To date, several shared SSc- RA loci have been identified independently. In this study, we aimed to systematically search for new common SSc-RA loci through an inter-disease meta-GWAS strategy. Methods: We performed a meta-analysis combining GWAS datasets of SSc and RA using a strategy that allowed identification of loci with both same-direction and opposingdirection allelic effects. The top single-nucleotide polymorphisms (SNPs) were followed-up in independent SSc and RA case-control cohorts. This allowed us to increase the sample size to a total of 8,830 SSc patients, 16,870 RA patients and 43,393 controls. Results: The cross-disease meta-analysis of the GWAS datasets identified several loci with nominal association signals (P-value < 5 x 10-6), which also showed evidence of association in the disease-specific GWAS scan. These loci included several genomic regions not previously reported as shared loci, besides risk factors associated with both diseases in previous studies. The follow-up of the putatively new SSc-RA loci identified IRF4 as a shared risk factor for these two diseases (Pcombined = 3.29 x 10-12). In addition, the analysis of the biological relevance of the known SSc-RA shared loci pointed to the type I interferon and the interleukin 12 signaling pathways as the main common etiopathogenic factors. Conclusions: Our study has identified a novel shared locus, IRF4, for SSc and RA and highlighted the usefulness of cross-disease GWAS meta-analysis in the identification of common risk loci