Retinoid X receptor promotes hematopoietic stem cell fitness and quiescence and preserves hematopoietic homeostasis.

Hematopoietic stem cells (HSCs) balance self-renewal and differentiation to maintain hematopoietic fitness throughout life. In steady-state conditions, HSC exhaustion is prevented by the maintenance of most HSCs in a quiescent state, with cells entering the cell cycle only occasionally. HSC quiescence is regulated by retinoid and fatty-acid ligands of transcriptional factors of the nuclear retinoid X receptor (RXR) family. Here, we show that dual deficiency for hematopoietic RXRa and RXRb induces HSC exhaustion, myeloid cell/megakaryocyte differentiation, and myeloproliferative-like disease. RXRa and RXRb maintain HSC quiescence, survival, and chromatin compaction; moreover, transcriptome changes in RXRa;RXRb-deficient HSCs include premature acquisition of an aging-like HSC signature, MYC pathway upregulation, and RNA intron retention. Fitness loss and associated RNA transcriptome and splicing alterations in RXRa;RXRb-deficient HSCs are prevented by Myc haploinsufficiency. Our study reveals the critical importance of RXRs for the maintenance of HSC fitness and their protection from premature aging.We thank the members of the J.A.C. and M.R. laboratories for extensive discussions and critiques of the manuscript. We thank Daniel Metzger (Université de Strasbourg, France) for Rxrbf/f 418 mice, Juan Carlos Zúñiga-Pflücker (Sunnybrook Health Sciences Centre, Canada) for OP9-NL1 cells, Daniel Jiménez-Carretero (CNIC) for t-SNE analysis, the CRG (Barcelona, Spain) Genomics Unit for ATACseq sequencing, and S. Bartlett (CNIC) for editorial assistance. We also thank the staff of the CNIC Cellomics and Animal facilities for technical support. This study was supported by grants from the Spanish Ministerio de Ciencia e Innovación (MICIN) (SAF2017-90604-REDT-NurCaMein, RTI2018- 095928-B100, and PID2021-122552OB-I00), La Marató de TV3 Foundation (201605-32), and the Comunidad de Madrid (MOIR-B2017/BMD-3684) to M.R and from the Formación de Profesorado Universitario (FPU17/01731) program (MICIN) to J.P. The project also received funding from the US National Institutes of Health (R01 DK124115, P01 HL158688, R01 HL147536, R01 CA237016 and U54 DK126108 to J.A.C). 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 (grant CEX2020-001041-S funded by MICIN/AEI/10.13039/501100011033).S

RXRs control serous macrophage neonatal expansion and identity and contribute to ovarian cancer progression

Tissue-resident macrophages (TRMs) populate all tissues and play key roles in homeostasis, immunity and repair. TRMs express a molecular program that is mostly shaped by tissue cues. However, TRM identity and the mechanisms that maintain TRMs in tissues remain poorly understood. We recently found that serous-cavity TRMs (LPMs) are highly enriched in RXR transcripts and RXR-response elements. Here, we show that RXRs control mouse serous-macrophage identity by regulating chromatin accessibility and the transcriptional regulation of canonical macrophage genes. RXR deficiency impairs neonatal expansion of the LPM pool and reduces the survival of adult LPMs through excess lipid accumulation. We also find that peritoneal LPMs infiltrate early ovarian tumours and that RXR deletion diminishes LPM accumulation in tumours and strongly reduces ovarian tumour progression in mice. Our study reveals that RXR signalling controls the maintenance of the serous macrophage pool and that targeting peritoneal LPMs may improve ovarian cancer outcomes.This work was supported by a HFSP fellowship to M.C-A. (LT000110/2015-L/1), grants from the Spanish Ministerio de Ciencia e Innovación (MCI) (SAF2015-64287R, SAF2017-90604-REDT-NurCaMein, RTI2018-095928-B100), La Marató de TV3 Foundation (201605-32) and Comunidad de Madrid (MOIR-B2017/BMD-3684) to M.R, and the Formación de Profesorado Universitario (FPU17/01731) programme (MCI) to J.P. The CNIC is supported by the MCI and the Pro CNIC Foundation and is a Severo Ochoa Center of Excellence (SEV-2015-0505).S

Cytoskeleton Rearrangements during the Execution Phase of Apoptosis

Apoptosis is a regulated energy‐dependent process for the elimination of unnecessary or damaged cells during embryonic development, tissue homeostasis and many pathological conditions. Apoptosis is characterized by specific morphological and biochemical features in which caspase activation has a pivotal role. During apoptosis, cells undergo characteristic morphological reorganizations in which the cytoskeleton participates actively. Traditionally, this cytoskeleton rearrangement has been assigned mainly to actinomyosin ring contraction, with microtubule and intermediate filaments both reported to be depolymerized at early stages of apoptosis. However, recent results have shown that microtubules are reformed during the execution phase of apoptosis forming an apoptotic microtubule network (AMN). Current hypothesis proposes that AMN is required to maintain plasma membrane integrity and cell morphology during the execution phase of apoptosis. AMN disruption provokes apoptotic cell collapse, secondary necrosis and the subsequent release of toxic molecules which can damage surrounding cells and promote inflammation. Therefore, AMN formation in physiological or pathological apoptosis is essential for tissue homeostasis

Retinoid X receptor promotes hematopoietic stem cell fitness and quiescence and preserves hematopoietic homeostasis

Hematopoietic stem cells (HSCs) balance self-renewal and differentiation to maintain hematopoietic fitness throughout life. In steady-state conditions, HSC exhaustion is prevented by the maintenance of most HSCs in a quiescent state, with cells entering the cell cycle only occasionally. HSC quiescence is regulated by retinoid and fatty-acid ligands of transcriptional factors of the nuclear retinoid X receptor (RXR) family. Herein, we show that dual deficiency for hematopoietic RXRα and RXRβ induces HSC exhaustion, myeloid cell/megakaryocyte differentiation, and myeloproliferative-like disease. RXRα and RXRβ maintain HSC quiescence, survival, and chromatin compaction; moreover, transcriptome changes in RXRα;RXRβ-deficient HSCs include premature acquisition of an aging-like HSC signature, MYC pathway upregulation, and RNA intron retention. Fitness loss and associated RNA transcriptome and splicing alterations in RXRα;RXRβ-deficient HSCs are prevented by Myc haploinsufficiency. Our study reveals the critical importance of RXRs for the maintenance of HSC fitness and their protection from premature aging

Urine NMR-based TB metabolic fingerprinting for the diagnosis of TB in children

Tuberculosis (TB) is a major cause of morbidity and mortality in children, and early diagnosis and treatment are crucial to reduce long-term morbidity and mortality. In this study, we explore whether urine nuclear magnetic resonance (NMR)-based metabolomics could be used to identify differences in the metabolic response of children with different diagnostic certainty of TB. We included 62 children with signs and symptoms of TB and 55 apparently healthy children. Six of the children with presumptive TB had bacteriologically confirmed TB, 52 children with unconfirmed TB, and 4 children with unlikely TB. Urine metabolic fingerprints were identified using high- and low-field proton NMR platforms and assessed with pattern recognition techniques such as principal components analysis and partial least squares discriminant analysis. We observed differences in the metabolic fingerprint of children with bacteriologically confirmed and unconfirmed TB compared to children with unlikely TB (p = 0.041 and p = 0.013, respectively). Moreover, children with unconfirmed TB with X-rays compatible with TB showed differences in the metabolic fingerprint compared to children with non-pathological X-rays (p = 0.009). Differences in the metabolic fingerprint in children with different diagnostic certainty of TB could contribute to a more accurate characterisation of TB in the paediatric population. The use of metabolomics could be useful to improve the prediction of TB progression and diagnosis in children

Urine NMR-based TB metabolic fingerprinting for the diagnosis of TB in children

Tuberculosis (TB) is a major cause of morbidity and mortality in children, and early diagnosis and treatment are crucial to reduce long-term morbidity and mortality. In this study, we explore whether urine nuclear magnetic resonance (NMR)-based metabolomics could be used to identify differences in the metabolic response of children with different diagnostic certainty of TB. We included 62 children with signs and symptoms of TB and 55 apparently healthy children. Six of the children with presumptive TB had bacteriologically confirmed TB, 52 children with unconfirmed TB, and 4 children with unlikely TB. Urine metabolic fingerprints were identified using high- and low-field proton NMR platforms and assessed with pattern recognition techniques such as principal components analysis and partial least squares discriminant analysis. We observed differences in the metabolic fingerprint of children with bacteriologically confirmed and unconfirmed TB compared to children with unlikely TB (p = 0.041 and p = 0.013, respectively). Moreover, children with unconfirmed TB with X-rays compatible with TB showed differences in the metabolic fingerprint compared to children with non-pathological X-rays (p = 0.009). Differences in the metabolic fingerprint in children with different diagnostic certainty of TB could contribute to a more accurate characterisation of TB in the paediatric population. The use of metabolomics could be useful to improve the prediction of TB progression and diagnosis in children

El Receptor X de Retinoides como regulador clave de la hematopoiesis: desde las células madre a los macrófagos residentes de tejidos

Tesis Doctoral inédita leída en la Universidad Autónoma de Madrid, Facultad de Medicina, Departamento de Bioquímica. Fecha de Lectura: 12-04-2023Esta tesis tiene embargado el acceso al texto completo hasta el 12-10-2024La hematopoyesis es el proceso por el cual se producen las células de la sangre. Las células madre hematopoyéticas (CMHs) se sitúan en la cima de la jerarquía hematopoyética y de ellas emanan el resto de células de la sangre. La preservación de la homeostasis del sistema hematopoyético requiere que las CMHs se mantengan mayormente quiescentes. La proliferación y el agotamiento de las CMHs se asocian a un declive gradual de su eficacia biológica y capacidad regenerativa. Por ende, los procesos moleculares que controlan la funcionalidad de las CMHs deben estar regulados con precisión para así poder mantener una correcta diferenciación de las CMHs hacia eritrocitos, megacariocitos, linfocitos y células mieloides. Los macrófagos son células mieloides que actúan como centinelas del sistema inmune. Los macrófagos se encuentran en todos los tejidos del organismo donde detectan y destruyen patógenos, además de realizar otras funciones relevantes para la homeostasis tisular. Los macrófagos residentes de tejidos (MRTs) se diseminan durante la hematopoyesis embrionaria poblando todos los tejidos. Muchos MRTs tienen la habilidad de mantenerse localmente mediante auto-renovación. Sin embargo, en función del tejido y de la edad, la población de MRTs de origen embrionario puede ser reemplazada por monocitos derivados de la médula ósea que diferencian hasta MRTs. Avances recientes demuestran que los MRTs se valen de factores de transcripción determinantes del linaje (FTDLs) específicos y nicho-dependientes para dirigir su identidad genética y epigenética. Sin embargo, todavía hay MRTs cuyas señales clave del nicho y FTDLs son desconocidos. En este trabajo describimos que los Receptores X de Retinoides (RXRs) son factors de transcripción clave para la hematopoyesis, en particular para las CMHs y los MRTs de origen embrionario. Específicamente, demostramos que las CMHs deficientes en RXRs son menos quiescentes que células de fenotipo salvaje, lo que reduce su funcionalidad y capacidad regenerativa. La caracterización de los cambios transcripcionales y epigenéticos en las CMHs carentes de RXRs reveló una apertura de la cromatina, un aumento en la expresión de genes con firmas genéticas relacionadas con CMHs envejecidas y con la vía de señalización de MYC y un aumento en la retención de intrones. Además, demostramos que la depleción de los RXRs en las CMHs causa una hematopoyesis dirigida hacia la línea mieloide y un consecuente desorden mieloproliferativo letal asociado a la edad. Este papel de los RXRs en el desarrollo de células mieloides nos llevó a estudiar su función en los MRTs. Demostramos que los RXRs son dispensables para la mielopoyesis embrionaria, sin embargo, encontramos que son necesarios para mantener las poblaciones de ciertos MRTs adultos de origen embrionario: macrófagos de cavidades serosas (peritoneo, pleura y pericárdio), alveolares y del hígado. Específicamente, demostramos que los RXRs son esenciales para la expansión neonatal y para la posterior supervivencia y el manejo de lípidos en la etapa adulta de los macrófagos peritoneales de origen embrionario. También demostramos que los RXRs son necesarios para la maduración, identidad y función de los macrófagos alveolares. Para concluir, nuestros descubrimientos sugieren que DLL4 y TGF dos moléculas señalizadoras del nicho pulmonar, son necesarias para la inducción de la expresión de los RXRs y de genes clave para la identidad de los macrófagos alveolares. En su conjunto, los datos reunidos en esta tesis demuestran el papel crucial y célula-específico de los RXRs, desde preservar la quiescencia en las CMHs, hasta mantener la hematopoyesis balanceada, pasando por controlar proliferación y supervivencia en macrófagos peritoneales e identidad en los macrófagos alveolaresHematopoiesis is the process by which blood cells are produced. Hematopoietic stem cells (HSCs) locate at the top of hematopoietic hierarchy, and, from them, the rest of blood cells are derived. The preservation of the hematopoietic system homeostasis requires that HSCs remain mostly quiescent. HSC proliferation and exhaustion are associated with a gradual decline in HSC fitness and regeneration capacity. As a result, the molecular processes governing HSC functionality have to be tightly regulated in order to maintain a correct HSC differentiation towards erythrocytes, megakaryocytes, lymphocytes and myeloid cells. Macrophages are myeloid cells that act as sentinels of the immune system. Macrophages are found in every tissue of the organism where they detect and destruct foreign pathogens and play other relevant functions for tissue homeostasis. Tissue Resident Macrophages (TRMs) are seeded during embryonic hematopoiesis. Many of these TRMs have the ability to be maintained locally by self-renewal. However, according to the tissue and age, the population of embryo-derived TRMs can be displaced by bone marrow-derived monocytes that differentiate into undistinguishable TRMs. Recent advances demonstrate that TRMs require a specific and niche-dependent lineage-determining transcription factors (LDTFs) expression in order to orchestrate their genetic and epigenetic identity. Still, there are TRMs with unknown key niche signals and LDTFs. We showed that the Retinoid X Receptors (RXRs) are key TFs for hematopoiesis, including HSCs and embryo-derived TRMs. Specifically, we demonstrated that RXR-deficient HSCs are less quiescent, functionally-defective and lack regeneration capacity. Characterization of the transcriptomic and epigenetic changes in RXR-depleted HSCs revealed chromatin openness and gene upregulation, with acquisition of aged HSC, MYC pathway and alternative splicing signatures. Moreover, HSC RXR depletion led to a myeloid-biased hematopoiesis and, consequently, a lethal age-associated myeloproliferative disorder. This RXR role for myeloid cell development prompted us to study their function in TRMs. While que showed that RXRs are dispensable for embryonic myelopoiesis, we found that they are needed to maintain the populations of certain adult embryo-derived TRMs: serous cavity (peritoneum, pleura and pericardium), alveolar and liver macrophages. Particularly, we demonstrated that RXRs are essential for the neonatal expansion, and adult survival and lipid handling of embryo-derived peritoneal macrophages. We also demonstrated that RXRs are required for alveolar macrophage (AM) maturation, identity and function. To conclude, our findings suggest that the lung niche signals DLL4 and TGF are needed for AM RXRs and core gene expression. Altogether, the data gathered in this thesis demonstrate the crucial and cell-specific role of RXRs, from preserving HSC dormancy, to maintaining a balanced hematopoiesis, and even for controlling proliferation and survival in peritoneal macrophages, and identity in alveolar macrophage

Molecular control of tissue-resident macrophage identity by nuclear receptors.

Macrophages are key immune cells that reside in almost all tissues of the body, where they exert pleiotropic functions in homeostasis and disease. Development and identity of macrophages in each organ are governed by tissue-dependent signaling pathways and transcription factors that ultimately define specific tissue-resident macrophage phenotypes and functions. In recent years, nuclear receptors, a class of ligand-activated transcription factors, have been found to play important roles in macrophage specification in several tissues. Nuclear receptors are thus important targets for therapies aimed at controlling the numbers and functions of tissue-resident macrophages. This review outlines current knowledge about the critical roles of nuclear receptors in tissue-resident macrophage development, specification, and maintenance.This work was supported by grants from the Spanish Ministerio de Ciencia e Innovacion (MCI) (SAF2017-90604-REDT-NurCaMein, RTI2018-095928-BI00), La Marato de TV3 Foundation (201605-32), and the Comunidad de Madrid (MOIRB2017/BMD-3684) to M.R, and the Formacion de Profesorado Universitario (FPU17/01731) program (MCI) to J.P. The CNIC is supported by the MCI and the Pro CNIC Foundation and is a Severo Ochoa Center of Excellence (SEV-2015-0505).S

Performance of QuantiFERON- TB Gold Plus assays in children and adolescents at risk of tuberculosis: a cross-sectional multicentre study

The QuantiFERON-TB Gold Plus (QFT-Plus) assay, which features two antigen-stimulated tubes (TB1 and TB2) instead of a single tube used in previous-generation interferon-gamma release assays (IGRAs), was launched in 2016. Despite this, data regarding the assay's performance in the paediatric setting remain scarce. This study aimed to determine the performance of QFT-Plus in a large cohort of children and adolescents at risk of tuberculosis (TB) in a low-burden setting. Methods: Cross-sectional, multicentre study at healthcare institutions participating in the Spanish Paediatric TB Research Network, including patients <18 years who had a QFT-Plus performed between September 2016 and June 2020. Results: Of 1726 patients (52.8% male, median age: 8.4 years), 260 (15.1%) underwent testing during contact tracing, 288 (16.7%) on clinical/radiological suspicion of tuberculosis disease (TBD), 649 (37.6%) during new-entrant migrant screening and 529 (30.6%) prior to initiation of immunosuppressive treatment. Overall, the sensitivity of QFT-Plus for TBD (n=189) and for latent tuberculosis infection (LTBI, n=195) was 83.6% and 68.2%, respectively. The agreement between QFT-Plus TB1 and TB2 antigen tubes was excellent (98.9%, κ=0.961). Only five (2.5%) patients with TBD had discordance between TB1 and TB2 results (TB1+/TB2-, n=2; TB1-/TB2+, n=3). Indeterminate assay results (n=54, 3.1%) were associated with young age, lymphopenia and elevated C reactive protein concentrations. Conclusions: Our non-comparative study indicates that QFT-Plus does not have greater sensitivity than previous-generation IGRAs in children in both TBD and LTBI. In TBD, the addition of the second antigen tube, TB2, does not enhance the assay's performance substantially.Sin financiación9.106 JCR (2021) Q1, 9/65 Respiratory System2.309 SJR (2021) Q1, 8/144 Pulmonary and Respiratory MedicineNo data IDR 2020UE