Involvement of glucosinolates and phenolics in the promotion of broccoli seedling growth through the modulation of primary and secondary metabolism.

Secondary metabolites play an essential role in plant defense. However, the role of glucosinolates and phenols in brassica crop yield in the context of environmentally friendly agricultural practices has not been established. Our study investigated the effects of a Brassica extract, rich in these metabolites, on the physiology and metabolism of broccoli (Brassica oleracea L. var. italica) seedlings and the subsequent development of the plants in adult stages. The results showed an increase in growth in the extract-treated seedlings, which was associated with an alteration of primary and secondary metabolism. In particular, there was an increase in the levels of amino acids, phenolic compounds and hormones, while the levels of glucosinolates decreased. Lipid peroxidation diminished in treated plants, indicating improved membrane integrity. Treated plants subsequently grown in hydroponically showed increased water use efficiency, transpiration, and internal carbon, which contributed to the improved growth of these plants. Overall, our findings underscore the potential of the glucosinolates and phenols ratio as essential to improve crop growth and stress tolerance, as well as revealed the interest of studying the mechanisms involved in the possible uptake and integration of GSLs by broccoli seedlings after external application.

Mechanisms of neutralization of toxSAS from toxin-antitoxin modules.

Toxic small alarmone synthetase (toxSAS) enzymes constitute a family of bacterial effectors present in toxin-antitoxin and secretion systems. toxSASs act through either translation inhibition mediated by pyrophosphorylation of transfer RNA (tRNA) CCA ends or synthesis of the toxic alarmone adenosine pentaphosphate ((pp)pApp) and adenosine triphosphate (ATP) depletion, exemplified by FaRel2 and FaRel, respectively. However, structural bases of toxSAS neutralization are missing. Here we show that the pseudo-Zn2+ finger domain (pZFD) of the ATfaRel2 antitoxin precludes access of ATP to the pyrophosphate donor site of the FaRel2 toxin, without affecting recruitment of the tRNA pyrophosphate acceptor. By contrast, (pp)pApp-producing toxSASs are inhibited by Tis1 antitoxin domains though occlusion of the pyrophosphate acceptor-binding site. Consequently, the auxiliary pZFD of AT2faRel is dispensable for FaRel neutralization. Collectively, our study establishes the general principles of toxSAS inhibition by structured antitoxin domains, with the control strategy directly coupled to toxSAS substrate specificity.

Exploring Phenolic Compounds in Crop By-Products for Cosmetic Efficacy.

Phenolic compounds represent a group of secondary metabolites that serve essential functions in plants. Beyond their positive impact on plants, these phenolic metabolites, often referred to as polyphenols, possess a range of biological properties that can promote skin health. Scientific research indicates that topically using phenolics derived from plants can be advantageous, but their activity and stability highly depend on storage of the source material and the extraction method. These compounds have the ability to relieve symptoms and hinder the progression of different skin diseases. Because they come from natural sources and have minimal toxicity, phenolic compounds show potential in addressing the causes and effects of skin aging, skin diseases, and various types of skin damage, such as wounds and burns. Hence, this review provides extensive information on the particular crops from which by-product phenolic compounds can be sourced, also emphasizing the need to conduct research according to proper plant material storage practices and the choice of the best extracting method, along with an examination of their specific functions and the mechanisms by which they act to protect skin.

Transcriptomic Profiling of Pleural Effusions: Differences in Malignant and Infectious Fluids.

Background and Objectives: Different cellular and molecular processes are involved in the production of malignant and infectious pleural effusions. However, the underlying mechanisms responsible for these differences or their consequences remain incompletely understood. The objective of this study was to identify differences in gene expression in pleural exudates of malignant and infectious aetiology and establish the possible different biological processes involved in both situations. Materials and Methods: RNA transcriptomic analysis was performed on 46 pleural fluid samples obtained during diagnostic thoracocenteses from 46 patients. There were 35 exudates (19 malignant and 16 infectious effusions) and 11 transudates that were used as a reference control group. Differential gene expression analysis for both exudative groups was identified. An enrichment score using the Human Kegg Orthology database was used for establishing the biological processes associated with malignant and infectious pleural effusions. Results: When comparing malignant exudates with infectious effusions, 27 differentially expressed genes with statistical significance were identified. Network analysis showed ten different biological processes for malignant and for infectious pleural effusions. In malignant fluids, processes related to protein synthesis and processing predominate. In infectious exudates, biological processes in connection with ATP production prevail. Conclusions: This study demonstrates differentially expressed genes in malignant and infectious pleural effusions, which could have important implications in the search for diagnostic or prognostic biomarkers. In addition, for the first time, biological processes involved in these two causes of pleural exudates have been described.

Brassica oleracea L. var. italica Aquaporin Reconstituted Proteoliposomes as Nanosystems for Resveratrol Encapsulation.

Aquaporins (AQPs), membrane proteins responsible for facilitating water transport, found in plant membrane vesicles (MV), have been related to the functionality and stability of MV. We focused on AQPs obtained from broccoli, as they show potential for biotechnological applications. To gain further insight into the role of AQPs in MV, we describe the heterologous overexpression of two broccoli AQPs (BoPIP1;2 and BoPIP2;2) in Pichia pastoris, resulting in their purification with high yield (0.14 and 0.99 mg per gram cells for BoPIP1;2 and BoPIP2;2). We reconstituted AQPs in liposomes to study their functionality, and the size of proteoliposomes did not change concerning liposomes. BoPIP2;2 facilitated water transport, which was preserved for seven days at 4 °C and at room temperature but not at 37 °C. BoPIP2;2 was incorporated into liposomes to encapsulate a resveratrol extract, resulting in increased entrapment efficiency (EE) compared to conventional liposomes. Molecular docking was utilized to identify binding sites in PIP2s for resveratrol, highlighting the role of aquaporins in the improved EE. Moreover, interactions between plant AQP and human integrin were shown, which may increase internalization by the human target cells. Our results suggest AQP-based alternative encapsulation systems can be used in specifically targeted biotechnological applications.

Deciphering the effect of salinity and boron stress on broccoli plants reveals that membranes phytosterols and PIP aquaporins facilitate stress adaptation.

Abiotic stresses, such as salinity and boron toxicity/deficiency, are prevalent in arid and semi-arid regions where broccoli is largely cultivated. This study aimed to investigate the physiological response of broccoli leaves to these stresses, focusing on parameters such as growth, relative water content, stomatal conductance, and mineral concentration after 15 days of treatment application. The effects of individual and combined stresses of salinity and boron (deficiency and toxicity) were examined. Additionally, the study explored the molecular aspects of PIP aquaporins in relation to their presence in the plasma membrane and their interaction with the lipid environment. The results showed that the combined stress of salinity and boron deficiency resulted in a significant reduction in plant biomass, suggesting a specific adaptation to this stress combination. Changes in stomatal conductance and mineral nutrient levels indicated that the adaptation mechanisms were associated with water and boron concentration in the leaves. The expression patterns of PIP aquaporins varied among the different stress treatments, either individually or in combination. Furthermore, the presence of aquaporins in the plasma membrane and microsomal fraction highlighted the potential regulatory roles of trafficking along with the membrane composition, particularly the concentration of phytosterols. The results underscore the importance of water transport by aquaporins and their interaction with the sterol composition in the membranes, in facilitating salinity-boron stress adaptation mechanisms.

Exploring membrane vesicles in citrus fruits: a comparative analysis of conventional and organic farming approaches.

BACKGROUND: Recently, vesicles derived from plant cell membranes have received attention for their potential use as active biomolecules and nanocarriers, and obtaining them from organic crops may be an interesting option because different farming systems can affect production, plant secondary metabolism and biochemistry of cell membranes. The present study aimed to determine how organic and conventional farming affects the mineral nutrition, gas exchange, CO2 fixation and biochemical composition of lemon fruits, which could have an impact on the different fractions of cell membranes in pulp and juice. RESULTS: Organic trees had higher intrinsic water use efficiency (WUEi) but conventional trees had higher stomatal conductance (gs) and nitrogen use efficiency (NUtE). Also, organic lemons had significantly higher levels of some micronutrients (Ca, Cu, Fe and Zn). Second, the main differences in the membrane vesicles showed that organic pulp vesicles had a higher antioxidant activity and more oleic acid, whereas both types of vesicles from conventional lemons had more linoleic acid. CONCLUSION: In conclusion, organic farming did not alter carbon fixation parameters but impacted nitrogen fixation and water uptake, and resulted in higher micronutrient levels in lemons. These mineral nutritional changes could be related to the higher production of membranes that showed suitable morphological traits and a high antioxidant activity, positively correlated with a high amount of oleic acid, which could have stronger cell protection characteristics. © 2023 The Authors. Journal of The Science of Food and Agriculture published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Modification of Gene Expression of Tomato Plants through Foliar Flavonoid Application in Relation to Enhanced Growth.

The exogenous application of phenolic compounds is increasingly recognized as a valuable strategy for promoting growth and mitigating the adverse effects of abiotic stress. However, the biostimulant effect under optimal conditions has not been thoroughly explored. In this study, we investigated the impact of foliar application of flavonoids, specifically CropBioLife (CBL), on tomato plants grown under controlled conditions. Our study focused on determining growth parameters, such as cell size, and assessing the concentration of hormones. Principal component analysis (PCA) from all physiological variables was determined. Additionally, we utilized high-throughput mRNA-sequencing technology and bioinformatic methodologies to robustly analyze the transcriptomes of tomato leaves regulated by flavonoids. The findings revealed that CBL primarily influenced cell enlargement by 60%, leading to increased growth. Furthermore, CBL-treated plants exhibited higher concentrations of the hormone zeatin, but lower concentrations of IAA (changes of 50%). Moreover, RNA-seq analysis indicated that CBL-treated plants required increased mineral transport and water uptake, as evidenced by gene expression patterns. Genes related to pathways such as fatty acid degradation, phenylpropanoid biosynthesis, and ABC transporters showed regulatory mechanisms governing internal flavonoid biosynthesis, transport, and tissue concentration, ultimately resulting in higher flavonoid concentrations in tomato leaves.

Biochemical and X-ray analyses of the players involved in the faRel2/aTfaRel2 toxin-antitoxin operon.

The aTfaRel2/faRel2 operon from Coprobacillus sp. D7 encodes a bicistronic type II toxin-antitoxin (TA) module. The FaRel2 toxin is a toxic small alarmone synthetase (toxSAS) that inhibits translation through the pyrophosphorylation of uncharged tRNAs at the 3'-CCA end. The toxin is neutralized by the antitoxin ATfaRel2 through the formation of an inactive TA complex. Here, the production, biophysical analysis and crystallization of ATfaRel2 and FaRel2 as well as of the ATfaRel2-FaRel2 complex are reported. ATfaRel2 is monomeric in solution. The antitoxin crystallized in space group P21212 with unit-cell parameters a = 53.3, b = 34.2, c = 37.6 Å, and the best crystal diffracted to a resolution of 1.24 Å. Crystals of FaRel2 in complex with APCPP, a nonhydrolysable ATP analogue, belonged to space group P21, with unit-cell parameters a = 31.5, b = 60.6, c = 177.2 Å, ß = 90.6°, and diffracted to 2.6 Šresolution. The ATfaRel2-FaRel2Y128F complex forms a heterotetramer in solution composed of two toxins and two antitoxins. This complex crystallized in two space groups: F4132, with unit-cell parameters a = b = c = 227.1 Å, and P212121, with unit-cell parameters a = 51.7, b = 106.2, c = 135.1 Å. The crystals diffracted to 1.98 and 2.1 Šresolution, respectively.

Immune response after SARS-CoV-2 vaccination in patients with inflammatory immune-mediated diseases receiving immunosuppressive treatment.

BACKGROUND: Real world data on the response to the SARS-CoV-2 vaccine in patients with immunomediated diseases (IMIDs) treated with immunesuppressants is of great interest because vaccine response may be impaired. The main aim was to study the humoral and cellular immune response after SARS-CoV-2 vaccination in patients with IMIDs treated with immunosuppressants. The secondary aim was to describe the frequency of SARS-CoV-2 infections after vaccination in these patients. MATERIAL AND METHODS: This is an observational study including 86 patients with IMIDs. All patients were treated with biologic or targeted synthetic disease-modifying antirheumatic drugs [b/tsDMARDs: TNF inhibitors (TNFi), rituximab, anti-interleukin 6 receptor (anti-IL6R) or JAK inhibitors (JAKi)]. Demographic and clinical information were collected. After 4-6 weeks of 2nd and 3rd vaccine doses, humoral response was assessed using the Thermo Scientific ELiA SARS-CoV-2-Sp1 IgG Test. Also, in patients with serum SARS-CoV-2 antibody levels under 100UI/ml, cellular response was analyzed using the QuantiFERON SARS-CoV-2 Starter Pack. RESULTS: A total of 86 patients under b/tsDMARDs and 38 healthy controls were included. Most patients received TNFi (45 with TNFi, 31 with rituximab, 5 with anti-IL6R and 5 with JAKi). SARS-CoV-2 antibodies (Ab) were present in an 86% of patients with IMIDs and in 100% healthy controls (p = 0.017). However, 12 (14%) patients had undetectable SARS-CoV-2 Ab levels, all treated with rituximab. In addition, SARS-CoV-2 Ab (IU/ml) were statistically lower in patients (Mdn (IQR): 59.5 (17-163) in patients vs 625 (405-932) in controls, p < 0.001). Patients treated with rituximab had lower Ab levels than those treated with TNFi and controls (p < 0.001). The cellular response to SARS-CoV-2 vaccine was evaluated in 30 patients. Eleven patients had a positive cellular response, being more frequent in patients treated with rituximab (p = 0.03). SARS-CoV-2 infection was reported in 43% of patients and 34% of controls after vaccination. Only 6 (7%) patients required hospitalization, most of whom treated with rituximab (67%). CONCLUSION: SARS-CoV-2 antibody levels were lower in patients than in controls, especially in patients treated with rituximab. A cellular response can be detected despite having a poor humoral response. Severe infections in vaccinated patients with IMIDs are rare, and are observed mainly in patients treated with rituximab.

Thromboembolic and hemorrhagic complications in patients with prosthetic heart valves cared for in a tertiary care center. What have we learned?

BACKGROUND: Heart valve replacement surgery with mechanical or biological prostheses entails a risk of thromboembolism and bleeding complications. OBJECTIVE: To determine the complications related to complementary anticoagulation therapy and the probability of risk. METHODS: One-hundred and sixty-three patients who underwent heart valve replacement between 2002 and 2016 with either mechanical or biological prostheses, and who received vitamin K antagonists after hospital discharge, were studied. Anticoagulation therapy was categorized into optimal and non-optimal according to INR values prior to the development of complications. Patients with comorbidities and other risk factors for thrombosis and/or bleeding were excluded. RESULTS: In total, 68.7 % of patients received mechanical prostheses, and 31.3 %, biological prostheses (p ≤ 0.001); 25.2 % experienced the complications that motivated the study (p ≤ 0.001), which were hemorrhagic in 48.8 %, thromboembolic in 26.8 %, and of both types in 24.4 % (relative risk = 4.229). Among the patients with complications, 95.1 % received mechanical prostheses, and 4.9 %, biological (p = 0.005); non-optimal INR was identified in 49.7 % (p ≤ 0.001). CONCLUSIONS: Given the high risk of thromboembolic and hemorrhagic complications, valve prostheses must be carefully chosen, and care priorities should include prevention and follow-up, especially in those patients who require anticoagulation therapy.

ANTECEDENTES: El reemplazo valvular por prótesis mecánicas o biológicas implica riesgo de tromboembolismo y complicaciones hemorrágicas. OBJETIVO: Determinar las complicaciones relacionadas con la terapia de anticoagulación complementaria y la probabilidad de riesgo en pacientes portadores de prótesis valvulares del corazón. MÉTODOS: Se estudiaron 163 pacientes entre 2002 y 2016, portadores de prótesis mecánicas y biológicas, quienes recibieron antagonistas de la vitamina K posterior al egreso hospitalario. La terapia de anticoagulación se categorizó en óptima y no óptima conforme a los valores de INR previos a las complicaciones. Fueron excluidos los pacientes con comorbilidades y otros factores de riesgo de trombosis y/o sangrado. RESULTADOS: a 68.7 % de los pacientes se les colocó prótesis mecánica y a 31.3 %, biológica (p ≤ 0.001); 25.2 % presentó las complicaciones motivo de estudio (p ≤ 0.001), hemorrágicas en 48.8 %, tromboembólicas en 26.8 % y de ambos tipos en 24.4 % (riesgo relativo = 4.229); a 95.1 % de los pacientes con complicaciones se les colocó prótesis mecánica y a 4.9 %, biológica (p = 0.005); 49.7 % presentó INR no óptimo (p ≤ 0.001). CONCLUSIONES: Ante riesgo alto de complicaciones tromboembólicas y hemorrágicas, la elección de las prótesis valvulares, la prevención y el seguimiento son prioridades, principalmente en quienes requieren terapia de anticoagulación.

Complicaciones tromboembólicas y hemorrágicas en pacientes portadores de prótesis valvulares del corazón en un centro de tercer nivel. ¿Qué hemos aprendido? / Thromboembolic and hemorrhagic complications in patients with prosthetic heart valves at third level care center. What have we learned?

Resumen Antecedentes: El reemplazo valvular por prótesis mecánicas o biológicas implica riesgo de tromboembolismo y complicaciones hemorrágicas. Objetivo: Determinar las complicaciones relacionadas con la terapia de anticoagulación complementaria y la probabilidad de riesgo en pacientes portadores de prótesis valvulares del corazón. Métodos: Se estudiaron 163 pacientes entre 2002 y 2016, portadores de prótesis mecánicas y biológicas, quienes recibieron antagonistas de la vitamina K posterior al egreso hospitalario. La terapia de anticoagulación se categorizó en óptima y no óptima conforme a los valores de INR previos a las complicaciones. Fueron excluidos los pacientes con comorbilidades y otros factores de riesgo de trombosis y/o sangrado. Resultados: a 68.7 % de los pacientes se les colocó prótesis mecánica y a 31.3 %, biológica (p ≤ 0.001); 25.2 % presentó las complicaciones motivo de estudio (p ≤ 0.001), hemorrágicas en 48.8 %, tromboembólicas en 26.8 % y de ambos tipos en 24.4 % (riesgo relativo = 4.229); a 95.1 % de los pacientes con complicaciones se les colocó prótesis mecánica y a 4.9 %, biológica (p = 0.005); 49.7 % presentó INR no óptimo (p ≤ 0.001). Conclusiones: Ante riesgo alto de complicaciones tromboembólicas y hemorrágicas, la elección de las prótesis valvulares, la prevención y el seguimiento son prioridades, principalmente en quienes requieren terapia de anticoagulación.

Abstract Background: Heart valve replacement surgery with mechanical or biological prostheses entails a risk of thromboembolism and bleeding complications. Objective: To determine the complications related to complementary anticoagulation therapy and the probability of risk. Methods: One-hundred and sixty-three patients who underwent heart valve replacement between 2002 and 2016 with either mechanical or biological prostheses, and who received vitamin K antagonists after hospital discharge, were studied. Anticoagulation therapy was categorized into optimal and non-optimal according to INR values prior to the development of complications. Patients with comorbidities and other risk factors for thrombosis and/or bleeding were excluded. Results: In total, 68.7 % of patients received mechanical prostheses, and 31.3 %, biological prostheses (p ≤ 0.001); 25.2 % experienced the complications that motivated the study (p ≤ 0.001), which were hemorrhagic in 48.8 %, thromboembolic in 26.8 %, and of both types in 24.4 % (relative risk = 4.229). Among the patients with complications, 95.1 % received mechanical prostheses, and 4.9 %, biological (p = 0.005); non-optimal INR was identified in 49.7 % (p ≤ 0.001). Conclusions: Given the high risk of thromboembolic and hemorrhagic complications, valve prostheses must be carefully chosen, and care priorities should include prevention and follow-up, especially in those patients who require anticoagulation therapy.

Detection of specific RBD+ IgG+ memory B cells by flow cytometry in healthcare workers and patients with inborn errors of immunity after BNT162b2 m RNA COVID-19 vaccination.

Introduction: Inborn errors of immunity (IEI) are a heterogeneous group of diseases caused by intrinsic defects of the immune system. Estimating the immune competence of immunocompromised patients for an infection risk assessment or after SARS-CoV-2 vaccination constituted a challenge. Methods: The aim of this study was to determine the humoral responses of patients with IEI through a comprehensive analysis of specific receptor-binding domain-positive (RBD+) IgG+ memory B cells (MBCs) by flow cytometry, together with routine S-specific IgG antibodies and QuantiFERON SARS-CoV-2 (T-cell response), before the vaccine and 3 weeks after a second dose. Results and discussion: We first analyzed the percentage of specific RBD+ IgG+ MBCs in healthy healthcare workers. Within the control group, there was an increase in the percentage of specific IgG+ RBD+ MBCs 21 days after the second dose, which was consistent with S-specific IgG antibodies.Thirty-one patients with IEI were included for the pre- and post-vaccination study; IgG+ RBD+ MBCs were not evaluated in 6 patients due to an absence of B cells in peripheral blood. We detected various patterns among the patients with IEI with circulating B cells (25, 81%): an adequate humoral response was observed in 12/25, consider by the detection of positive S-specific IgG antibodies and the presence of specific IgG+ RBD+ MBCs, presenting a positive T-cell response; in 4/25, very low S-specific IgG antibody counts correlated with undetectable events in the IgG+ RBD+ MBC compartment but with positive cellular response. Despite the presence of S-specific IgG antibodies, we were unable to detect a relevant percentage of IgG+ RBD+ MBCs in 5/25; however, all presented positive T-cell response. Lastly, we observed a profound failure of B and T-cell response in 3 (10%) patients with IEI, with no assessment of S-specific IgG antibodies, IgG+ RBD+ MBCs, and negative cellular response. The identification of specific IgG+ RBD+ MBCs by flow cytometry provides information on different humoral immune response outcomes in patients with IEI and aids the assessment of immune competence status after SARS-CoV-2 mRNA vaccine (BNT162b2), together with S-specific IgG antibodies and T-cell responses.

Confronting Secondary Metabolites with Water Uptake and Transport in Plants under Abiotic Stress.

Phenolic compounds and glucosinolates are secondary plant metabolites that play fundamental roles in plant resistance to abiotic stress. These compounds have been found to increase in stress situations related to plant adaptive capacity. This review assesses the functions of phenolic compounds and glucosinolates in plant interactions involving abiotic stresses such as drought, salinity, high temperature, metals toxicity, and mineral deficiency or excess. Furthermore, their relation with water uptake and transport mediated through aquaporins is reviewed. In this way, the increases of phenolic compounds and glucosinolate synthesis have been related to primary responses to abiotic stress and induction of resistance. Thus, their metabolic pathways, root exudation, and external application are related to internal cell and tissue movement, with a lack of information in this latter aspect.

Sapwood mycobiome varies across host, plant compartment and environments in Nothofagus forests from Northern Patagonia.

Global forests are increasingly being threatened by altered climatic conditions and increased attacks by pests and pathogens. The complex ecological interactions among pathogens, microbial communities, tree hosts and the environment are important drivers of forest dynamics. Little is known about the ecology of forest pathology and related microbial communities in temperate forests of the southern hemisphere. In this study, we used next-generation sequencing to characterize sapwood-inhabiting fungal communities in North Patagonian Nothofagus forests and assessed patterns of diversity of taxa and ecological guilds across climatic, site and host variables (health condition and compartment) as a contribution to Nothofagus autecology. The diversity patterns inferred through the metabarcoding analysis were similar to those obtained through culture-dependent approaches. However, we detected additional heterogeneity and greater richness with culture-free methods. Host species was the strongest driver of fungal community structure and composition, while host health status was the weakest. The relative impacts of site, season, plant compartment and health status were different for each tree species; these differences can be interpreted as a matter of water availability. For Nothofagus dombeyi, which is distributed across a wide range of climatic conditions, site was the strongest driver of community composition. The microbiome of N. pumilio varied more with season and temperature, a relevant factor for forest conservation in the present climate change scenario. Both species carry a number of potential fungal pathogens in their sapwood, whether they exhibit symptoms or not. Our results provide insight into the diversity of fungi associated with the complex pathobiome of the dominant Nothofagus species in southern South America.

Los bosques del mundo están cada vez más amenazados por las condiciones climáticas alteradas y el aumento de los ataques de plagas y patógenos. Las complejas interacciones ecológicas entre los patógenos, las comunidades microbianas, los árboles hospedantes y el medio ambiente son impulsores importantes de la dinámica forestal. Poco se sabe sobre la ecología de la patología forestal y las comunidades microbianas relacionadas en los bosques templados del hemisferio sur. En este estudio, utilizamos la secuenciación Illumina para caracterizar las comunidades de hongos que habitan en la albura en los bosques de Nothofagus de la Patagonia Norte y evaluamos los patrones de diversidad de taxones y gremios ecológicos a través de variables climáticas, de sitio y de hospedante (identidad, condición de salud y compartimento) como una contribución a la autoecología de los Nothofagus. Los patrones de diversidad inferidos a través del análisis metabarcoding fueron similares a los obtenidos a través de enfoques dependientes de cultivo. Sin embargo, detectamos mayor heterogeneidad y mayor riqueza con métodos independientes de cultivo. La especie hospedante fue el modelador más fuerte de la estructura y composición de la comunidad fúngica, mientras que el estado de salud del hospedante fue el más débil. El impacto relativo del sitio, la estación, el compartimento y el estado de salud fueron diferentes para cada especie de árbol; estas diferencias pueden interpretarse en clave de disponibilidad de agua. Para N. dombeyi, que se distribuye a lo largo de una amplia gama de condiciones climáticas, el sitio fue el principal modelador de la composición de la comunidad. El micobioma de Nothofagus pumilio varió más con la estación y la temperatura, un factor relevante para la conservación de los bosques en el escenario actual de cambio climático. Ambas especies portan una serie de patógenos fúngicos potenciales en su albura, ya sea que muestren síntomas o no. Nuestros resultados brindan una idea de la diversidad de hongos asociados con el complejo patobioma de las especies dominantes de Nothofagus en el sur de América del Sur.

Potential of Sulforaphane and Broccoli Membrane Vesicles as Regulators of M1/M2 Human Macrophage Activity.

Macrophages have emerged as important therapeutic targets in many human diseases. The aim of this study was to analyze the effect of broccoli membrane vesicles and sulphoraphane (SFN), either free or encapsulated, on the activity of human monocyte-derived M1 and M2 macrophage primary culture. Our results show that exposure for 24 h to SFN 25 µM, free and encapsulated, induced a potent reduction on the activity of human M1 and M2 macrophages, downregulating proinflammatory and anti-inflammatory cytokines and phagocytic capability on C. albicans. The broccoli membrane vesicles do not represent inert nanocarriers, as they have low amounts of bioactive compounds, being able to modulate the cytokine production, depending on the inflammatory state of the cells. They could induce opposite effects to that of higher doses of SFN, reflecting its hormetic effect. These data reinforce the potential use of broccoli compounds as therapeutic agents not only for inflammatory diseases, but they also open new clinical possibilities for applications in other diseases related to immunodeficiency, autoimmunity, or in cancer therapy. Considering the variability of their biological effects in different scenarios, a proper therapeutic strategy with Brassica bioactive compounds should be designed for each pathology.

Single-cell Atlas of common variable immunodeficiency shows germinal center-associated epigenetic dysregulation in B-cell responses.

Common variable immunodeficiency (CVID), the most prevalent symptomatic primary immunodeficiency, displays impaired terminal B-cell differentiation and defective antibody responses. Incomplete genetic penetrance and ample phenotypic expressivity in CVID suggest the participation of additional pathogenic mechanisms. Monozygotic (MZ) twins discordant for CVID are uniquely valuable for studying the contribution of epigenetics to the disease. Here, we generate a single-cell epigenomics and transcriptomics census of naïve-to-memory B cell differentiation in a CVID-discordant MZ twin pair. Our analysis identifies DNA methylation, chromatin accessibility and transcriptional defects in memory B-cells mirroring defective cell-cell communication upon activation. These findings are validated in a cohort of CVID patients and healthy donors. Our findings provide a comprehensive multi-omics map of alterations in naïve-to-memory B-cell transition in CVID and indicate links between the epigenome and immune cell cross-talk. Our resource, publicly available at the Human Cell Atlas, gives insight into future diagnosis and treatments of CVID patients.

Membrane Vesicles for Nanoencapsulated Sulforaphane Increased Their Anti-Inflammatory Role on an In Vitro Human Macrophage Model.

At present, there is a growing interest in finding new non-toxic anti-inflammatory drugs to treat inflammation, which is a key pathology in the development of several diseases with considerable mortality. Sulforaphane (SFN), a bioactive compound derived from Brassica plants, was shown to be promising due to its anti-inflammatory properties and great potential, though its actual clinical use is limited due to its poor stability and bioavailability. In this sense, the use of nanocarriers could solve stability-related problems. In the current study, sulforaphane loaded into membrane vesicles derived from broccoli plants was studied to determine the anti-inflammatory potential in a human-macrophage-like in vitro cell model under both normal and inflammatory conditions. On the one hand, the release of SFN from membrane vesicles was modeled in vitro, and two release phases were stabilized, one faster and the other slower due to the interaction between SFN and membrane proteins, such as aquaporins. Furthermore, the anti-inflammatory action of sulforaphane-loaded membrane vesicles was demonstrated, as a decrease in interleukins crucial for the development of inflammation, such as TNF-α, IL-1ß and IL-6, was observed. Furthermore, these results also showed that membrane vesicles by themselves had anti-inflammatory properties, opening the possibility of new lines of research to study these vesicles, not only as carriers but also as active compounds.

Bioactive peptides from broccoli stems strongly enhance regenerative keratinocytes by stimulating controlled proliferation.

CONTEXT: As the interest on the research of plant derived bioactive peptides (BPs) for nutraceutical, cosmeceutical and medical applications is increasing, in this work, the application of peptide derived from broccoli to keratinocytes was studied. OBJECTIVE: We focussed on the characterization of different peptides hydrolysates from broccoli stems [extracted from total protein (E) and from membrane protein (MF)], and their activity when applied to human keratinocytes. MATERIALS AND METHODS: Peptide mixtures from broccoli stems (E and MF) were characterized by proteomics. They were applied to HaCaT cells in order to study cytotoxicity in a concentration range between 20 and 0.15625 µg of protein/mL and wound healing was studied after 24 and 48 h of treatment application. Also, proteomic and gene expression of keratinocytes were analysed. RESULTS: Depending on the source, proteins varied in peptide and amino acid composition. An increased proliferation of keratinocytes was shown after the application of the E peptides mixtures, reaching 190% with the lowest concentrations, but enhanced wound healing repair with E and MF appeared, reaching 59% of wound closure after 48 h. At the gene expression and protein levels of keratinocytes, the upregulation of anti-oncogene p53 and keratinization factors were observed. DISCUSSION: These results suggest that peptide mixtures obtained from broccoli augmented cell proliferation and prevented the carcinogenic, uncontrolled growth of the cells, with different mechanisms depending on the protein source. CONCLUSIONS: The results encourage the opening of new lines of research involving the use of Brassica peptides for pharmaceutic or cosmetic use.