Función tolorógina, origen y diferenciación de las células dendríticas plasmacitoides residentes en el timo humano

Tesis doctoral inédita. Universidad Autónoma de Madrid, Facultad de Ciencias, Departamento de Biología Molecular. Fecha de lectura: 17-12-201

Effective innate immune response in natural HIV-1 controllers. Can mimicking lead to novel preventive and cure strategies against HIV-1?

Purpose of reviewHIV-1 controller individuals represents a model that can be useful for the development of novel vaccines and therapies. Initial studies pointed to the involvement of improved adaptive immunity, however, new emerging evidence suggests the contribution of innate cells to effective antiviral responses in spontaneous controllers. Therefore, understanding the alterations on innate cell subsets might be crucial to develop new effective therapeutic strategies.Recent findingsAmong different innate immune cells, dendritic cell (DC) and natural killer (NK) cell are essential for effective antiviral responses. DC from controllers display improved innate detection of HIV-1 transcripts, higher induction of interferons, higher antigen presenting capacities and increased metabolism and higher capacities to induce polyfunctional CD8+T-cell responses. Such properties have been mimicked by Toll-like receptor ligands and applied to DC-based immunotherapies in humans and in animal models. NK cells from controllers display higher expression of activating receptors promoting increased antibody-dependent cellular cytotoxicity (ADCC) and natural cytotoxicity activities. Neutralizing antibodies in combination with interleukin-15 superagonist or interferon-α can increase ADCC and cytotoxicity in NK cells from HIV-1 progressors.SummaryMimicking DC and NK cell innate profiles in controllers has become a promising strategy to step forward a novel efficient immunotherapy against the HIV-1 infectionE.M.G. was supported by the Ramo´n y Cajal Program (RYC2018-024374-I), the MINECO/FEDER RETOS program (RTI2018-097485-A-I00), Comunidad de Madrid Talento Program (2017-T1/BMD-5396), Gilead becas de investigacio´n (GLD19/00168), by Centro de Investigacio´n Biome´dica en Red (CIBERINF) de Enfermedades Infecciosas (CB21/13/00107), La Caixa Banking Foundation (H20-00218) and by REDINCOV grant from Fundacio´ La Marato´ TV3. M.C.M. was supported by La Caixa Banking Foundation (H20-00218) and Gilead becas de investigacio´n (GLD19/00168

Aperiodic two-layer energy management system for community microgrids based on blockchain strategy

Regulatory changes in different countries regarding self-consumption and growing public concern about the environment are encouraging the establishment of community microgrids. These community microgrids integrate a large number of small-scale distributed energy resources and offers a solution to enhance power system reliability and resilience. This work proposes a geographically-based split of the community microgrids into clusters of members that tend to have similar consumption and generation profiles, mimicking the most typical layout of cities. Assuming a community microgrid divided into clusters, a two-layer architecture is developed to facilitate the greater penetration of distributed energy resources in an efficient way. The first layer, referred as the market layer, is responsible for creating local energy markets with the aim of maximising the economic benefits for community microgrid members. The second layer is responsible for the network reconfiguration, which is based on the energy balance within each cluster. This layer complies with the IEC 61850 communication standard, in order to control commercial sectionalizing and tie switches. This allows the community microgrid network to be reconfigured to minimise energy exchanges with the main grid, without requiring interaction with the distributed system operator. To implement this two-layer energy management strategy, an aperiodic market approach based on Blockchain technology, and the additional functionality offered by Smart Contracts is adopted. This embraces the concept of energy communities since it decentralizes the control and eliminates intermediaries. The use of aperiodic control techniques helps to overcome the challenges of using Blockchain technology in terms of storage, computational requirements and member privacy. The scalability and modularity of the Smart Contract-based system allow each cluster of members to be designed by tailoring the system to their specific needs. The implementation of this strategy is based on low-cost off-the-shelf devices, such as Raspberry Pi 4 Model B boards, which operate as Blockchain nodes of community microgrid members. Finally, the strategy has been validated by emulating two use cases based on the IEEE 123-node system network model highlighting the benefits of the proposal.Comunidad de Madri

Extracellular vesicles from Listeria monocytogenes-infected dendritic cells alert the innate immune response.

Communication through cell-cell contacts and extracellular vesicles (EVs) enables immune cells to coordinate their responses against diverse types of pathogens. The function exerted by EVs in this context depends on the proteins and nucleic acids loaded into EVs, which elicit specific responses involved in the resolution of infection. Several mechanisms control protein and nucleic acid loading into EVs; in this regard, acetylation has been described as a mechanism of cellular retention during protein sorting to exosomes. HDAC6 is a deacetylase involved in the control of cytoskeleton trafficking, organelle polarity and cell migration, defense against Listeria monocytogenes (Lm) infection and other immune related functions. Here, we show that the protein content of dendritic cells (DCs) and their secreted EVs (DEVs) vary during Lm infection, is enriched in proteins related to antiviral functions compared to non-infected cells and depends on HDAC6 expression. Analyses of the post-translational modifications revealed an alteration of the acetylation and ubiquitination profiles upon Lm infection both in DC lysates and DEVs. Functionally, EVs derived from infected DCs upregulate anti-pathogenic genes (e.g. inflammatory cytokines) in recipient immature DCs, which translated into protection from subsequent infection with vaccinia virus. Interestingly, absence of Listeriolysin O in Lm prevents DEVs from inducing this anti-viral state. In summary, these data underscore a new mechanism of communication between bacteria-infected DC during infection as they alert neighboring, uninfected DCs to promote antiviral responses.This study was supported by grant PDI-2020-120412RB-I00, PDC2021-121797-I00, BIO2015-67580-P and PGC2018-097019-BI00 from the Spanish Ministry of Economy and Competitiveness (MINECO), grant S2017/BMD-3671-INFLAMUNE-CM from the Comunidad de Madrid, a grant from the Ramón Areces Foundation “Ciencias de la Vida y la Salud” (XIX Concurso-2018), “la Caixa” Banking Foundation (grants HR17-00016 and HR17-00247), BIOIMID (PIE13/041) and PRB3 (IPT17/0019 - ISCIII-SGEFI/ ERDF, ProteoRed) from Instituto de Salud Carlos III, CIBER Cardiovascular (CB16/11/00272), and Fondo de Investigación Sanitaria del Instituto de Salud Carlos III and co-funding by Fondo Europeo de Desarrollo Regional FEDER). IF-D is supported by a Fellowship from the Spanish Ministry of Science, Innovation, and Universities (FPU15/02539). DC-F is supported by a Fellowship from “la Caixa” Foundation (LCF/BQ/DR19/11740010). The CNIC is supported by the Instituto de Salud Carlos III (ISCIII), the Ministerio de Ciencia e Innovación (MCIN) and the Pro CNIC Foundation, and is a Severo Ochoa Center of Excellence (CEX2020- 001041-S). Funding agencies did not intervene in the design of the studies, with no copyright over the study.S

Antiretroviral therapy duration and immunometabolic state determine efficacy of ex vivo dendritic cell-based treatment restoring functional HIV-specific CD8+ T cells in people living with HIV.

Dysfunction of CD8+ T cells in people living with HIV-1 (PLWH) receiving anti-retroviral therapy (ART) has restricted the efficacy of dendritic cell (DC)-based immunotherapies against HIV-1. Heterogeneous immune exhaustion and metabolic states of CD8+ T cells might differentially associate with dysfunction. However, specific parameters associated to functional restoration of CD8+ T cells after DC treatment have not been investigated. We studied association of restoration of functional HIV-1-specific CD8+ T cell responses after stimulation with Gag-adjuvant-primed DC with ART duration, exhaustion, metabolic and memory cell subsets profiles. HIV-1-specific CD8+ T cell responses from a larger proportion of PLWH on long-term ART (more than 10 years; LT-ARTp) improved polyfunctionality and capacity to eliminate autologous p24+ infected CD4+ T cells in vitro. In contrast, functional improvement of CD8+ T cells from PLWH on short-term ART (less than a decade; ST-ARTp) after DC treatment was limited. This was associated with lower frequencies of central memory CD8+ T cells, increased co-expression of PD1 and TIGIT and reduced mitochondrial respiration and glycolysis induction upon TCR activation. In contrast, CD8+ T cells from LT-ARTp showed increased frequencies of TIM3+ PD1- cells and preserved induction of glycolysis. Treatment of dysfunctional CD8+ T cells from ST-ARTp with combined anti-PD1 and anti-TIGIT antibodies plus a glycolysis promoting drug restored their ability to eliminate infected CD4+ T cells. Together, our study identifies specific immunometabolic parameters for different PLWH subgroups potentially useful for future personalized DC-based HIV-1 vaccines. NIH (R21AI140930), MINECO/FEDER RETOS (RTI2018-097485-A-I00) and CIBERINF grants.NIH (R21AI140930), MINECO/FEDER RETOS (RTI2018-097485-A-I00) and CIBERINF grants. We would like to thank the NIH AIDS Reagent Pro- gram, Division of AIDS, NIAID, NIH for providing HIV-1 PTE Gag Peptide Pool from NIAID, DAIDS (cat #11057) for the study. We would also like to thank Alvaro Serrano Navarro, for his help on adapting the lin- ear mixed model previously described by Martin- C ofreces N. et al83 to our data. Graphical schematic rep- resentations were created with BioRender.com. EMG was supported by the NIH R21 program (R21AI140930), the Ramón y Cajal Program (RYC2018- 024374-I), the MINECO/FEDER RETOS program (RTI2018-097485-A-I00), by Comunidad de Madrid Talento Program (2017-T1/BMD-5396) and by Gilead becas de investigaci on (GLD19/00168). EMG and IDS are supported by Centro de Investigación Biomédica en Red (CIBERINF) de Enfermedades Infecciosas (CB21/ 13/00107). MCM was supported by NIH R21 program (R21AI140930), “La Caixa Banking Foundation (H20- 00218) and Gilead becas de investigaci on (GLD19/ 00168). MJB is supported by the Miguel Servet program funded by the Spanish Health Institute Carlos III (CP17/00179), the MINECO/FEDER RETOS program (RTI2018-101082-B-100), and Fundació La Marat o TV3 (201805-10FMTV3). EMG and MJB are both funded by “La Caixa Banking Foundation (H20-00218) and by REDINCOV grant from Fundació La Marat o TV3. FSM was supported by SAF2017-82886-R and PDI-2020- 120412RB-I00 grants from the Ministerio de Ciencia e Innovaci on, and HR17-00016 grant from “La Caixa Banking Foundation. HF was funded by PI21/01583 grant from Ministerio de Ciencia e Innovación, Instituto de Salud Carlos III. MJC was supported by PID2019- 104406RB-I00 from Ministerio de Ciencia e Innovación. ISC was funded by the CM21/00157 Rio- Hortega grant. IT was supported by grant for the pro- motion of research studies master-UAM 2021.S

MICa/b-dependent activation of natural killer cells by CD64+ inflammatory type 2 dendritic cells contributes to autoimmunity

Artículo escrito por un elevado número de autores, solo se referencian el que aparece en primer lugar, el nombre del grupo de colaboración, si le hubiere, y los autores pertenecientes a la UAMPrimary Sjögren's syndrome (pSS) is an inflammatory autoimmune disorder largely mediated by type I and II interferon (IFN). The potential contribution of innate immune cells, such as natural killer (NK) cells and dendritic cells (DC), to the pSS pathology remains understudied. Here, we identified an enriched CD16+ CD56hi NK cell subset associated with higher cytotoxic function, as well as elevated proportions of inflammatory CD64+ conventional dendritic cell (cDC2) subtype that expresses increased levels of MICa/b, the ligand for the activating receptor NKG2D, in pSS individuals. Circulating cDC2 from pSS patients efficiently induced activation of cytotoxic NK cells ex vivo and were found in proximity to CD56+ NK cells in salivary glands (SG) from pSS patients. Interestingly, transcriptional activation of IFN signatures associated with the RIG-I/DDX60 pathway, IFN I receptor, and its target genes regulate the expression of NKG2D ligands on cDC2 from pSS patients. Finally, increased proportions of CD64hi RAE-1+ cDC2 and NKG2D+CD11b+CD27+ NK cells were present in vivo in the SG after poly I:C injection. Our study provides novel insight into the contribution and interplay of NK and cDC2 in pSS pathology and identifies new potential therapy targetsRTI2018-097485-A-I00, PID2021-127899OB-I0

DataSheet_1_Extracellular vesicles from Listeria monocytogenes-infected dendritic cells alert the innate immune response.pdf [Dataset]

Supplementary Figure 1. Isolated EVs present typical size and topology. Supplementary Figure 2. Protein profiling from total cell lysates and their derived EVs from WT and KO-HDAC6 BMDCs. Supplementary Figure 3. Enrichment in acetylated and ubiquitinated DC proteins upon Lm infection. Supplementary Figure 4. Ubiquitination in K-48 and K-63 state in T lymphoblast total cell lysates and their derived EVs. Supplementary Figure 5. Pore filtration methods restrain Lm and do not induce strong antipathogenic responses. Supplementary Figure 6. IFN-β is detected following Lm infection. Table S1. List of antibodies used for Western-blot and Flow Cytometry and the used dilution. Table S2. List of primers, with their corresponding sequence, used for qPCR. Table S3: Protein quantification in total cell lysates Table S4: IPA analysis of total cell lysates: canonical pathways and diseases and functions category Table S5: Protein quantification in EVs Table S6: IPA analysis of EVs: diseases and functions category Table S7: Ubiquitinated and acetylated peptides in total cell lysates and EVs Table S8: Enrichment analysis of ubiquitinated and acetylated proteinsPeer reviewe

Deregulated cellular circuits driving immunoglobulins and complement consumption associate with the severity of COVID-19 patients

SARS-CoV-2 infection causes an abrupt response by the host immune system, which is largely responsible for the outcome of COVID-19. We investigated whether the specific immune responses in the peripheral blood of 276 patients were associated with the severity and progression of COVID-19. At admission, dramatic lymphopenia of T, B, and NK cells is associated with severity. Conversely, the proportion of B cells, plasmablasts, circulating follicular helper T cells (cTfh) and CD56–CD16+ NK-cells increased. Regarding humoral immunity, levels of IgM, IgA, and IgG were unaffected, but when degrees of severity were considered, IgG was lower in severe patients. Compared to healthy donors, complement C3 and C4 protein levels were higher in mild and moderate, but not in severe patients, while the activation peptide of C5 (C5a) increased from the admission in every patient, regardless of their severity. Moreover, total IgG, the IgG1 and IgG3 isotypes, and C4 decreased from day 0 to day 10 in patients who were hospitalized for more than two weeks, but not in patients who were discharged earlier. Our study provides important clues to understand the immune response observed in COVID-19 patients, associating severity with an imbalanced humoral response, and identifying new targets for therapeutic interventionThe study was funded by grants SAF2017- 82886-R to FS-M from the Ministerio de Economía y Competitividad, and from “La Caixa Banking Foundation” (HR17-00016) to FS-M. Grant PI018/01163 to CMC and grant PI19/00549 to AA were funded by Fondo de Investigaciones Sanitarias, Ministerio de Sanidad y Consumo, Spain. SAF2017-82886-R, PI018/01163 and PI19/00549 grants were also co-funded by European Regional Development Fund, ERDF/FEDER. This work has been funded by grants Fondo Supera COVID (CRUE-Banco de Santander) to FSM, and “Ayuda Covid 2019” from Comunidad de Madri

CSL–MAML-dependent Notch1 signaling controls T lineage–specific IL-7Rα gene expression in early human thymopoiesis and leukemia

Notch1 activation is essential for T-lineage specification of lymphomyeloid progenitors seeding the thymus. Progression along the T cell lineage further requires cooperative signaling provided by the interleukin 7 receptor (IL-7R), but the molecular mechanisms responsible for the dynamic and lineage-specific regulation of IL-7R during thymopoiesis are unknown. We show that active Notch1 binds to a conserved CSL-binding site in the human IL7R gene promoter and critically regulates IL7R transcription and IL-7R α chain (IL-7Rα) expression via the CSL–MAML complex. Defective Notch1 signaling selectively impaired IL-7Rα expression in T-lineage cells, but not B-lineage cells, and resulted in a compromised expansion of early human developing thymocytes, which was rescued upon ectopic IL-7Rα expression. The pathological implications of these findings are demonstrated by the regulation of IL-7Rα expression downstream of Notch1 in T cell leukemias. Thus, Notch1 controls early T cell development, in part by regulating the stage- and lineage-specific expression of IL-7Rα

Spatiotemporal Characteristics of the Largest HIV-1 CRF02_AG Outbreak in Spain: Evidence for Onward Transmissions

Background and Aim: The circulating recombinant form 02_AG (CRF02_AG) is the predominant clade among the human immunodeficiency virus type-1 (HIV-1) non-Bs with a prevalence of 5.97% (95% Confidence Interval-CI: 5.41–6.57%) across Spain. Our aim was to estimate the levels of regional clustering for CRF02_AG and the spatiotemporal characteristics of the largest CRF02_AG subepidemic in Spain.Methods: We studied 396 CRF02_AG sequences obtained from HIV-1 diagnosed patients during 2000–2014 from 10 autonomous communities of Spain. Phylogenetic analysis was performed on the 391 CRF02_AG sequences along with all globally sampled CRF02_AG sequences (N = 3,302) as references. Phylodynamic and phylogeographic analysis was performed to the largest CRF02_AG monophyletic cluster by a Bayesian method in BEAST v1.8.0 and by reconstructing ancestral states using the criterion of parsimony in Mesquite v3.4, respectively.Results: The HIV-1 CRF02_AG prevalence differed across Spanish autonomous communities we sampled from (p < 0.001). Phylogenetic analysis revealed that 52.7% of the CRF02_AG sequences formed 56 monophyletic clusters, with a range of 2–79 sequences. The CRF02_AG regional dispersal differed across Spain (p = 0.003), as suggested by monophyletic clustering. For the largest monophyletic cluster (subepidemic) (N = 79), 49.4% of the clustered sequences originated from Madrid, while most sequences (51.9%) had been obtained from men having sex with men (MSM). Molecular clock analysis suggested that the origin (tMRCA) of the CRF02_AG subepidemic was in 2002 (median estimate; 95% Highest Posterior Density-HPD interval: 1999–2004). Additionally, we found significant clustering within the CRF02_AG subepidemic according to the ethnic origin.Conclusion: CRF02_AG has been introduced as a result of multiple introductions in Spain, following regional dispersal in several cases. We showed that CRF02_AG transmissions were mostly due to regional dispersal in Spain. The hot-spot for the largest CRF02_AG regional subepidemic in Spain was in Madrid associated with MSM transmission risk group. The existence of subepidemics suggest that several spillovers occurred from Madrid to other areas. CRF02_AG sequences from Hispanics were clustered in a separate subclade suggesting no linkage between the local and Hispanic subepidemics